WorldWideScience

Sample records for chlamydomonas lacking photosystem

  1. Carbon dioxide fixation and photoevolution of hydrogen and oxygen in a mutant of Chlamydomonas lacking Photosystem I

    Energy Technology Data Exchange (ETDEWEB)

    Greenbaum, E.; Lee, J.W.; Tevault, C.V. [Oak Ridge National Lab., TN (United States)] [and others

    1995-09-01

    Sustained photoassimilation of atmospheric CO{sub 2} and simultaneous photoevolution of molecular hydrogen and oxygen has been observed in a Photosystem I deficient mutant B4 of Chlamydomonas reinhardtii that contains only Photosystem II. The data indicate that Photosystem II alone is capable of spanning the potential difference between water oxidation/oxygen evolution and ferredoxin reduction. The rates of both CO{sub 2} fixation and hydrogen and oxygen evolution are similar in the mutant to that of the wild-type C. reinhardtii 137c containing both photosystems. The wild-type had stable photosynthetic activity, measured as CO{sub 2} fixation, under both air and anaerobic conditions, while the mutant was stable only under anaerobic conditions. The results are discussed in terms of the fundamental mechanisms and energetics of photosynthesis and possible implications for the evolution of oxygenic photosynthesis.

  2. Functional analysis of Photosystem I light-harvesting complexes (Lhca) gene products of Chlamydomonas reinhardtii

    NARCIS (Netherlands)

    Mozzo, Milena; Mantelli, Manuela; Passarini, Francesca; Caffarri, Stefano; Croce, Roberta; Bassi, Roberto

    2010-01-01

    The outer antenna system of Chlamydomonas reinhardtii Photosystem I is composed of nine gene products, but due to difficulty in purification their individual properties are not known. In this work, the functional properties of the nine Lhca antennas of Chlamydomonas, have been investigated upon expr

  3. Two equilibration pools of chlorophylls in the Photosystem I core antenna of Chlamydomonas reinhardtii.

    Science.gov (United States)

    Gibasiewicz, Krzysztof; Ramesh, V M; Lin, Su; Redding, Kevin; Woodbury, Neal W; Webber, Andrew N

    2007-04-01

    Femtosecond transient absorption spectroscopy was applied for a comparative study of excitation decay in several different Photosystem I (PSI) core preparations from the green alga Chlamydomonas reinhardtii. For PSI cores with a fully interconnected network of chlorophylls, the excitation energy was equilibrated over a pool of chlorophylls absorbing at approximately 683 nm, independent of excitation wavelength [Gibasiewicz et al. J Phys Chem B 105:11498-11506, 2001; J Phys Chem B 106:6322-6330, 2002]. In preparations with impaired connectivity between chlorophylls, we have found that the spectrum of chlorophylls connected to the reaction center (i.e., with approximately 20 ps decay time) over which the excitation is equilibrated becomes excitation-wavelength-dependent. Excitation at 670 nm is finally equilibrated over chlorophylls absorbing at approximately 675 nm, whereas excitation at 695 nm or 700 nm is equilibrated over chlorophylls absorbing at approximately 683 nm. This indicates that in the vicinity of the reaction center there are two spectrally different and spatially separated pools of chlorophylls that are equally capable of effective excitation energy transfer to the reaction center. We propose that they are related to the two groups of central PSI core chlorophylls lying on the opposite sides of reaction center.

  4. Absence of lutein, violaxanthin and neoxanthin affects the functional chlorophyll antenna size of photosystem-II but not that of photosystem-I in the green alga Chlamydomonas reinhardtii.

    Science.gov (United States)

    Polle, J E; Niyogi, K K; Melis, A

    2001-05-01

    Chlamydomonas reinhardtii double mutant npq2 lor1 lacks the beta, epsilon-carotenoids lutein and loroxanthin as well as all beta,beta-epoxycarotenoids derived from zeaxanthin (e.g. violaxanthin and neoxanthin). Thus, the only carotenoids present in the thylakoid membranes of the npq2 lor1 cells are beta-carotene and zeaxanthin. The effect of these mutations on the photochemical apparatus assembly and function was investigated. In cells of the mutant strain, the content of photosystem-II (PSII) and photosystem-I (PSI) was similar to that of the wild type, but npq2 lor1 had a significantly smaller PSII light-harvesting Chl antenna size. In contrast, the Chl antenna size of PSI was not truncated in the mutant. SDS-PAGE and Western blot analysis qualitatively revealed the presence of all LHCII and LHCI apoproteins in the thylakoid membrane of the mutant. The results showed that some of the LHCII and most of the LHCI were assembled and functionally connected with PSII and PSI, respectively. Photon conversion efficiency measurements, based on the initial slope of the light-saturation curve of photosynthesis and on the yield of Chl a fluorescence in vivo, showed similar efficiencies. However, a significantly greater light intensity was required for the saturation of photosynthesis in the mutant than in the wild type. It is concluded that zeaxanthin can successfully replace lutein and violaxanthin in most of the functional light-harvesting antenna of the npq2 lor1 mutant.

  5. Refactoring the Six-Gene Photosystem II Core in the Chloroplast of the Green Algae Chlamydomonas reinhardtii.

    Science.gov (United States)

    Gimpel, Javier A; Nour-Eldin, Hussam H; Scranton, Melissa A; Li, Daphne; Mayfield, Stephen P

    2016-07-15

    Oxygenic photosynthesis provides the energy to produce all food and most of the fuel on this planet. Photosystem II (PSII) is an essential and rate-limiting component of this process. Understanding and modifying PSII function could provide an opportunity for optimizing photosynthetic biomass production, particularly under specific environmental conditions. PSII is a complex multisubunit enzyme with strong interdependence among its components. In this work, we have deleted the six core genes of PSII in the eukaryotic alga Chlamydomonas reinhardtii and refactored them in a single DNA construct. Complementation of the knockout strain with the core PSII synthetic module from three different green algae resulted in reconstitution of photosynthetic activity to 85, 55, and 53% of that of the wild-type, demonstrating that the PSII core can be exchanged between algae species and retain function. The strains, synthetic cassettes, and refactoring strategy developed for this study demonstrate the potential of synthetic biology approaches for tailoring oxygenic photosynthesis and provide a powerful tool for unraveling PSII structure-function relationships.

  6. Site Energies of Active and Inactive Pheophytins in the Reaction Center of Photosystem II from Chlamydomonas Reinhardtii

    Energy Technology Data Exchange (ETDEWEB)

    Acharya, K.; Neupane, B.; Zazubovich, V.; Sayre, R. T.; Picorel, R.; Seibert, M.; Jankowiak, R.

    2012-03-29

    It is widely accepted that the primary electron acceptor in various Photosystem II (PSII) reaction center (RC) preparations is pheophytin {alpha} (Pheo {alpha}) within the D1 protein (Pheo{sub D1}), while Pheo{sub D2} (within the D2 protein) is photochemically inactive. The Pheo site energies, however, have remained elusive, due to inherent spectral congestion. While most researchers over the past two decades placed the Q{sub y}-states of Pheo{sub D1} and Pheo{sub D2} bands near 678-684 and 668-672 nm, respectively, recent modeling [Raszewski et al. Biophys. J. 2005, 88, 986-998; Cox et al. J. Phys. Chem. B 2009, 113, 12364-12374] of the electronic structure of the PSII RC reversed the assignment of the active and inactive Pheos, suggesting that the mean site energy of Pheo{sub D1} is near 672 nm, whereas Pheo{sub D2} ({approx}677.5 nm) and Chl{sub D1} ({approx}680 nm) have the lowest energies (i.e., the Pheo{sub D2}-dominated exciton is the lowest excited state). In contrast, chemical pigment exchange experiments on isolated RCs suggested that both pheophytins have their Q{sub y} absorption maxima at 676-680 nm [Germano et al. Biochemistry 2001, 40, 11472-11482; Germano et al. Biophys. J. 2004, 86, 1664-1672]. To provide more insight into the site energies of both Pheo{sub D1} and Pheo{sub D2} (including the corresponding Q{sub x} transitions, which are often claimed to be degenerate at 543 nm) and to attest that the above two assignments are most likely incorrect, we studied a large number of isolated RC preparations from spinach and wild-type Chlamydomonas reinhardtii (at different levels of intactness) as well as the Chlamydomonas reinhardtii mutant (D2-L209H), in which the active branch Pheo{sub D1} is genetically replaced with chlorophyll {alpha} (Chl {alpha}). We show that the Q{sub x}-/Q{sub y}-region site energies of Pheo{sub D1} and Pheo{sub D2} are {approx}545/680 nm and {approx}541.5/670 nm, respectively, in good agreement with our previous assignment

  7. Site energies of active and inactive pheophytins in the reaction center of Photosystem II from Chlamydomonas reinhardtii.

    Science.gov (United States)

    Acharya, K; Neupane, B; Zazubovich, V; Sayre, R T; Picorel, R; Seibert, M; Jankowiak, R

    2012-03-29

    It is widely accepted that the primary electron acceptor in various Photosystem II (PSII) reaction center (RC) preparations is pheophytin a (Pheo a) within the D1 protein (Pheo(D1)), while Pheo(D2) (within the D2 protein) is photochemically inactive. The Pheo site energies, however, have remained elusive, due to inherent spectral congestion. While most researchers over the past two decades placed the Q(y)-states of Pheo(D1) and Pheo(D2) bands near 678-684 and 668-672 nm, respectively, recent modeling [Raszewski et al. Biophys. J. 2005, 88, 986 - 998; Cox et al. J. Phys. Chem. B 2009, 113, 12364 - 12374] of the electronic structure of the PSII RC reversed the assignment of the active and inactive Pheos, suggesting that the mean site energy of Pheo(D1) is near 672 nm, whereas Pheo(D2) (~677.5 nm) and Chl(D1) (~680 nm) have the lowest energies (i.e., the Pheo(D2)-dominated exciton is the lowest excited state). In contrast, chemical pigment exchange experiments on isolated RCs suggested that both pheophytins have their Q(y) absorption maxima at 676-680 nm [Germano et al. Biochemistry 2001, 40, 11472 - 11482; Germano et al. Biophys. J. 2004, 86, 1664 - 1672]. To provide more insight into the site energies of both Pheo(D1) and Pheo(D2) (including the corresponding Q(x) transitions, which are often claimed to be degenerate at 543 nm) and to attest that the above two assignments are most likely incorrect, we studied a large number of isolated RC preparations from spinach and wild-type Chlamydomonas reinhardtii (at different levels of intactness) as well as the Chlamydomonas reinhardtii mutant (D2-L209H), in which the active branch Pheo(D1) is genetically replaced with chlorophyll a (Chl a). We show that the Q(x)-/Q(y)-region site energies of Pheo(D1) and Pheo(D2) are ~545/680 nm and ~541.5/670 nm, respectively, in good agreement with our previous assignment [Jankowiak et al. J. Phys. Chem. B 2002, 106, 8803 - 8814]. The latter values should be used to model excitonic

  8. Mutations of photosystem II D1 protein that empower efficient phenotypes of Chlamydomonas reinhardtii under extreme environment in space.

    Science.gov (United States)

    Giardi, Maria Teresa; Rea, Giuseppina; Lambreva, Maya D; Antonacci, Amina; Pastorelli, Sandro; Bertalan, Ivo; Johanningmeier, Udo; Mattoo, Autar K

    2013-01-01

    Space missions have enabled testing how microorganisms, animals and plants respond to extra-terrestrial, complex and hazardous environment in space. Photosynthetic organisms are thought to be relatively more prone to microgravity, weak magnetic field and cosmic radiation because oxygenic photosynthesis is intimately associated with capture and conversion of light energy into chemical energy, a process that has adapted to relatively less complex and contained environment on Earth. To study the direct effect of the space environment on the fundamental process of photosynthesis, we sent into low Earth orbit space engineered and mutated strains of the unicellular green alga, Chlamydomonas reinhardtii, which has been widely used as a model of photosynthetic organisms. The algal mutants contained specific amino acid substitutions in the functionally important regions of the pivotal Photosystem II (PSII) reaction centre D1 protein near the QB binding pocket and in the environment surrounding Tyr-161 (YZ) electron acceptor of the oxygen-evolving complex. Using real-time measurements of PSII photochemistry, here we show that during the space flight while the control strain and two D1 mutants (A250L and V160A) were inefficient in carrying out PSII activity, two other D1 mutants, I163N and A251C, performed efficient photosynthesis, and actively re-grew upon return to Earth. Mimicking the neutron irradiation component of cosmic rays on Earth yielded similar results. Experiments with I163N and A251C D1 mutants performed on ground showed that they are better able to modulate PSII excitation pressure and have higher capacity to reoxidize the QA (-) state of the primary electron acceptor. These results highlight the contribution of D1 conformation in relation to photosynthesis and oxygen production in space.

  9. Mutation of Photosystem II D1 protein that empower efficient phenotypes of Chlamydomonas Reinhardtii under extreme environment in space

    Science.gov (United States)

    Oxygenic photosynthesis involves capture and conversion of light energy into chemical energy, a process fundamental to life including plant productivity on Earth. Photosynthetic electron transport is catalyzed by two photochemical reaction centres in series, photosystem II (PS II) and photosytem I (...

  10. Enhanced excision repair and lack of PSII activity contribute to higher UV survival of Chlamydomonas reinhardtii cells in dark.

    Science.gov (United States)

    Chaudhari, Vishalsingh R; Vyawahare, Aniket; Bhattacharjee, Swapan K; Rao, Basuthkar J

    2015-03-01

    Plant cells are known to differentiate their responses to stress depending up on the light conditions. We observed that UVC sensitive phenotype of light grown asynchronous Chlamydomonas reinhardtii culture (Light culture: LC) can be converted to relatively resistant form by transfer to dark condition (Dark culture: DC) before UVC exposure. The absence of photosystem II (PSII) function, by either atrazine treatment in wild type or in D1 (psbA) null mutant, conferred UV protection even in LC. We provide an indirect support for involvement of reactive oxygen species (ROS) signalling by showing higher UV survival on exposures to mild dose of H2O2 or Methyl Viologen. Circadian trained culture also showed a rhythmic variation in UV sensitivity in response to alternating light-dark (12 h:12 h) entrainment, with maximum UV survival at the end of 12 h dark and minimum at the end of 12 h light. This rhythm failed to maintain in "free running" conditions, making it a non-circadian phenotype. Moreover, atrazine strongly inhibited rhythmic UV sensitivity and conferred a constitutively high resistance, without affecting internal circadian rhythm marker expression. Dampening of UV sensitivity rhythm in Thymine-dimer excision repair mutant (cc-888) suggested the involvement of DNA repair in this phenomenon. DNA excision repair (ER) assays in cell-free extracts revealed that dark incubated cells exhibit higher ER compared to those growing in light, underscoring the role of ER in conferring differential UV sensitivity in dark versus light incubation. We suggest that multiple factors such as ROS changes triggered by differences in PSII activity, concomitant with differential ER efficiency collectively contribute to light-dark (12 h: 12 h) rhythmicity in C. reinhardtii UV sensitivity.

  11. Truncated photosystem chlorophyll antenna size in the green microalga Chlamydomonas reinhardtii upon deletion of the TLA3-CpSRP43 gene.

    Science.gov (United States)

    Kirst, Henning; Garcia-Cerdan, Jose Gines; Zurbriggen, Andreas; Ruehle, Thilo; Melis, Anastasios

    2012-12-01

    The truncated light-harvesting antenna size3 (tla3) DNA insertional transformant of Chlamydomonas reinhardtii is a chlorophyll-deficient mutant with a lighter green phenotype, a lower chlorophyll (Chl) per cell content, and higher Chl a/b ratio than corresponding wild-type strains. Functional analyses revealed a higher intensity for the saturation of photosynthesis and greater light-saturated photosynthetic activity in the tla3 mutant than in the wild type and a Chl antenna size of the photosystems that was only about 40% of that in the wild type. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis and western-blot analyses showed that the tla3 strain was deficient in the Chl a/b light-harvesting complex. Molecular and genetic analyses revealed a single plasmid insertion in chromosome 4 of the tla3 nuclear genome, causing deletion of predicted gene g5047 and plasmid insertion within the fourth intron of downstream-predicted gene g5046. Complementation studies defined that gene g5047 alone was necessary and sufficient to rescue the tla3 mutation. Gene g5047 encodes a C. reinhardtii homolog of the chloroplast-localized SRP43 signal recognition particle, whose occurrence and function in green microalgae has not hitherto been investigated. Biochemical analysis showed that the nucleus-encoded and chloroplast-localized CrCpSRP43 protein specifically operates in the assembly of the peripheral components of the Chl a/b light-harvesting antenna. This work demonstrates that cpsrp43 deletion in green microalgae can be employed to generate tla mutants with a substantially diminished Chl antenna size. The latter exhibit improved solar energy conversion efficiency and photosynthetic productivity under mass culture and bright sunlight conditions.

  12. Light-intensity-dependent expression of Lhc gene family encoding light-harvesting chlorophyll-a/b proteins of photosystem II in Chlamydomonas reinhardtii.

    Science.gov (United States)

    Teramoto, Haruhiko; Nakamori, Akira; Minagawa, Jun; Ono, Taka-aki

    2002-09-01

    Excessive light conditions repressed the levels of mRNAs accumulation of multiple Lhc genes encoding light-harvesting chlorophyll-a/b (LHC) proteins of photosystem (PS)II in the unicellular green alga, Chlamydomonas reinhardtii. The light intensity required for the repression tended to decrease with lowering temperature or CO(2) concentration. The responses of six LhcII genes encoding the major LHC (LHCII) proteins and two genes (Lhcb4 and Lhcb5) encoding the minor LHC proteins of PSII (CP29 and CP26) were similar. The results indicate that the expression of these Lhc genes is coordinately repressed when the energy input through the antenna systems exceeds the requirement for CO(2) assimilation. The Lhc mRNA level repressed under high-light conditions was partially recovered by adding the electron transport inhibitor 3-(3,4-dichlorophenyl)-1,1-dimethylurea, suggesting that redox signaling via photosynthetic electron carriers is involved in the gene regulation. However, the mRNA level was still considerably lower under high-light than under low-light conditions even in the presence of 3-(3,4-dichlorophenyl)-1,1-dimethylurea. Repression of the Lhc genes by high light was prominent even in the mutants deficient in the reaction center(s) of PSII or both PSI and PSII. The results indicate that two alternative processes are involved in the repression of Lhc genes under high-light conditions, one of which is independent of the photosynthetic reaction centers and electron transport events.

  13. Alteration of proteins and pigments influence the function of photosystem I under iron deficiency from Chlamydomonas reinhardtii.

    Directory of Open Access Journals (Sweden)

    Venkateswarlu Yadavalli

    Full Text Available BACKGROUND: Iron is an essential micronutrient for all organisms because it is a component of enzyme cofactors that catalyze redox reactions in fundamental metabolic processes. Even though iron is abundant on earth, it is often present in the insoluble ferric [Fe (III] state, leaving many surface environments Fe-limited. The haploid green alga Chlamydomonas reinhardtii is used as a model organism for studying eukaryotic photosynthesis. This study explores structural and functional changes in PSI-LHCI supercomplexes under Fe deficiency as the eukaryotic photosynthetic apparatus adapts to Fe deficiency. RESULTS: 77K emission spectra and sucrose density gradient data show that PSI and LHCI subunits are affected under iron deficiency conditions. The visible circular dichroism (CD spectra associated with strongly-coupled chlorophyll dimers increases in intensity. The change in CD signals of pigments originates from the modification of interactions between pigment molecules. Evidence from sucrose gradients and non-denaturing (green gels indicates that PSI-LHCI levels were reduced after cells were grown for 72 h in Fe-deficient medium. Ultrafast fluorescence spectroscopy suggests that red-shifted pigments in the PSI-LHCI antenna were lost during Fe stress. Further, denaturing gel electrophoresis and immunoblot analysis reveals that levels of the PSI subunits PsaC and PsaD decreased, while PsaE was completely absent after Fe stress. The light harvesting complexes were also susceptible to iron deficiency, with Lhca1 and Lhca9 showing the most dramatic decreases. These changes in the number and composition of PSI-LHCI supercomplexes may be caused by reactive oxygen species, which increase under Fe deficiency conditions. CONCLUSIONS: Fe deficiency induces rapid reduction of the levels of photosynthetic pigments due to a decrease in chlorophyll synthesis. Chlorophyll is important not only as a light-harvesting pigment, but also has a structural role

  14. Effect of chromium oxide (III) nanoparticles on the production of reactive oxygen species and photosystem II activity in the green alga Chlamydomonas reinhardtii

    Energy Technology Data Exchange (ETDEWEB)

    Costa, Cristina Henning da [Department of Sanitary and Environmental Engineering, Federal University of Santa Catarina, Campus Universitário, CEP: 88040-970, Florianópolis, SC (Brazil); Perreault, François [School of Sustainable Engineering and the Built Environment, Arizona State University, Tempe, AZ 85287-3005 (United States); Oukarroum, Abdallah [Department of Chemistry, University of Quebec in Montréal, 2101, Jeanne Mance Street, Station Centre-Ville, Montréal, QC H2X 2J6 (Canada); Melegari, Sílvia Pedroso [Department of Sanitary and Environmental Engineering, Federal University of Santa Catarina, Campus Universitário, CEP: 88040-970, Florianópolis, SC (Brazil); Center of Marine Studies, Federal University of Parana, Beira-mar Avenue, 83255-976, Pontal do Parana, PR (Brazil); Popovic, Radovan [Department of Chemistry, University of Quebec in Montréal, 2101, Jeanne Mance Street, Station Centre-Ville, Montréal, QC H2X 2J6 (Canada); Matias, William Gerson, E-mail: william.g.matias@ufsc.br [Department of Sanitary and Environmental Engineering, Federal University of Santa Catarina, Campus Universitário, CEP: 88040-970, Florianópolis, SC (Brazil)

    2016-09-15

    With the growth of nanotechnology and widespread use of nanomaterials, there is an increasing risk of environmental contamination by nanomaterials. However, the potential implications of such environmental contamination are hard to evaluate since the toxicity of nanomaterials if often not well characterized. The objective of this study was to evaluate the toxicity of a chromium-based nanoparticle, Cr{sub 2}O{sub 3}-NP, used in a wide diversity of industrial processes and commercial products, on the unicellular green alga Chlamydomonas reinhardtii. The deleterious impacts of Cr{sub 2}O{sub 3}-NP were characterized using cell density measurements, production of reactive oxygen species (ROS), esterase enzymes activity, and photosystem II electron transport as indicators of toxicity. Cr{sub 2}O{sub 3}-NP exposure inhibited culture growth and significantly lowered cellular Chlorophyll a content. From cell density measurements, EC50 values of 2.05 ± 0.20 and 1.35 ± 0.06 g L{sup −1} Cr{sub 2}O{sub 3}-NP were obtained after 24 and 72 h of exposure, respectively. In addition, ROS levels were increased to 160.24 ± 2.47% and 59.91 ± 0.15% of the control value after 24 and 72 h of exposition to 10 g L{sup −1} Cr{sub 2}O{sub 3}-NP. At 24 h of exposure, the esterase activity increased to 160.24% of control value, revealing a modification of the short-term metabolic response of algae to Cr{sub 2}O{sub 3}-NP exposure. In conclusion, the metabolism of C. reinhardtii was the most sensitive to Cr{sub 2}O{sub 3}-NP after 24 h of treatment. - Highlights: • Cr{sub 2}O{sub 3} nanoparticles are unstable and form large aggregates in the medium. • EC50 for growth inhibition of C. reinhardtii is 1.35 g L{sup −1} at 72 h. • Cr{sub 2}O{sub 3} nanoparticles increase ROS levels at 10 g L{sup −1}. • Cr{sub 2}O{sub 3} nanoparticles affect photosynthetic electron transport.

  15. Altered Fermentative Metabolism in Chlamydomonas reinhardtii Mutants Lacking Pyruvate Formate Lyase and Both Pyruvate Formate Lyase and Alcohol Dehydrogenase

    Energy Technology Data Exchange (ETDEWEB)

    Catalanotti, C.; Dubini, A.; Subramanian, V.; Yang, W. Q.; Magneschi, L.; Mus, F.; Seibert, M.; Posewitz, M. C.; Grossman, A. R.

    2012-02-01

    Chlamydomonas reinhardtii, a unicellular green alga, often experiences hypoxic/anoxic soil conditions that activate fermentation metabolism. We isolated three Chlamydomonas mutants disrupted for the pyruvate formate lyase (PFL1) gene; the encoded PFL1 protein catalyzes a major fermentative pathway in wild-type Chlamydomonas cells. When the pfl1 mutants were subjected to dark fermentative conditions, they displayed an increased flux of pyruvate to lactate, elevated pyruvate decarboxylation, ethanol accumulation, diminished pyruvate oxidation by pyruvate ferredoxin oxidoreductase, and lowered H2 production. The pfl1-1 mutant also accumulated high intracellular levels of lactate, succinate, alanine, malate, and fumarate. To further probe the system, we generated a double mutant (pfl1-1 adh1) that is unable to synthesize both formate and ethanol. This strain, like the pfl1 mutants, secreted lactate, but it also exhibited a significant increase in the levels of extracellular glycerol, acetate, and intracellular reduced sugars and a decrease in dark, fermentative H2 production. Whereas wild-type Chlamydomonas fermentation primarily produces formate and ethanol, the double mutant reroutes glycolytic carbon to lactate and glycerol. Although the metabolic adjustments observed in the mutants facilitate NADH reoxidation and sustained glycolysis under dark, anoxic conditions, the observed changes could not have been predicted given our current knowledge of the regulation of fermentation metabolism.

  16. Refactoring the six-gene photosystem II core in the chloroplast of the green algae Chlamydomonas reinhardtii

    DEFF Research Database (Denmark)

    Gimpel, Javier A.; Nour-Eldin, Hussam Hassan; Scranton, Melissa A.

    2016-01-01

    Oxygenic photosynthesis provides the energy to produce all food and most of the fuel on this planet. Photosystem II (PSII) is an essential and rate-limiting component of this process. Understanding and modifying PSII function could provide an opportunity for optimizing photosynthetic biomass....... Complementation of the knockout strain with the core PSII synthetic module from three different green algae resulted in reconstitution of photosynthetic activity to 85, 55, and 53% of that of the wild-type, demonstrating that the PSII core can be exchanged between algae species and retain function. The strains...

  17. Transport and arrangement of the outer-dynein-arm docking complex in the flagella of Chlamydomonas mutants that lack outer dynein arms.

    Science.gov (United States)

    Wakabayashi, K; Takada, S; Witman, G B; Kamiya, R

    2001-04-01

    The outer dynein arms of Chlamydomonas flagella are attached to a precise site on the outer doublet microtubules and repeat at a regular interval of 24 nm. This binding is mediated by the outer dynein arm docking complex (ODA-DC), which is composed of three protein subunits. In this study, antibodies against the 83- and 62-kD subunits (DC83 and DC62) of the ODA-DC were used to analyze its state of association with outer arm components within the cytoplasm, and its localization in the axonemes of oda mutants. Immunoprecipitation indicates that DC83 and DC62 are preassembled within the cytoplasm, but that they are not associated with outer arm dynein. Both proteins are lost or greatly diminished in oda1 and oda3, mutants in the structural genes of DC62 and DC83, respectively, demonstrating that their association is necessary for their stable presence in the cytoplasm. Immunoelectron microscopy indicates that DC83 repeats at 24-nm intervals along the length of the doublet microtubules of oda6, which lacks outer arms; thus, outer arm periodicity may be determined by the ODA-DC. Flagellar regeneration and temporary dikaryon experiments indicate that the ODA-DC can be rapidly transported into the flagellum and assembled on the doublet microtubules independently of the outer arms and independently of flagellar growth. Unexpectedly, the intensity of ODA-DC labeling decreased toward the distal ends of axonemes of oda6 but not wild-type cells, suggesting that the outer arms reciprocally contribute to the assembly/stability of the ODA-DC.

  18. Altered Fermentative Metabolism in Chlamydomonas reinhardtii Mutants Lacking Pyruvate Formate Lyase and Both Pyruvate Formate Lyase and Alcohol Dehydrogenase[W

    Science.gov (United States)

    Catalanotti, Claudia; Dubini, Alexandra; Subramanian, Venkataramanan; Yang, Wenqiang; Magneschi, Leonardo; Mus, Florence; Seibert, Michael; Posewitz, Matthew C.; Grossman, Arthur R.

    2012-01-01

    Chlamydomonas reinhardtii, a unicellular green alga, often experiences hypoxic/anoxic soil conditions that activate fermentation metabolism. We isolated three Chlamydomonas mutants disrupted for the pyruvate formate lyase (PFL1) gene; the encoded PFL1 protein catalyzes a major fermentative pathway in wild-type Chlamydomonas cells. When the pfl1 mutants were subjected to dark fermentative conditions, they displayed an increased flux of pyruvate to lactate, elevated pyruvate decarboxylation, ethanol accumulation, diminished pyruvate oxidation by pyruvate ferredoxin oxidoreductase, and lowered H2 production. The pfl1-1 mutant also accumulated high intracellular levels of lactate, succinate, alanine, malate, and fumarate. To further probe the system, we generated a double mutant (pfl1-1 adh1) that is unable to synthesize both formate and ethanol. This strain, like the pfl1 mutants, secreted lactate, but it also exhibited a significant increase in the levels of extracellular glycerol, acetate, and intracellular reduced sugars and a decrease in dark, fermentative H2 production. Whereas wild-type Chlamydomonas fermentation primarily produces formate and ethanol, the double mutant reroutes glycolytic carbon to lactate and glycerol. Although the metabolic adjustments observed in the mutants facilitate NADH reoxidation and sustained glycolysis under dark, anoxic conditions, the observed changes could not have been predicted given our current knowledge of the regulation of fermentation metabolism. PMID:22353371

  19. Origin of pronounced differences in 77 K fluorescence of the green alga Chlamydomonas reinhardtii in state 1 and 2

    NARCIS (Netherlands)

    Ünlü, Caner; Polukhina, Iryna; Amerongen, van Herbert

    2016-01-01

    In response to changes in the reduction state of the plastoquinone pool in its thylakoid membrane, the green alga Chlamydomonas reinhardtti is performing state transitions: remodelling of its thylakoid membrane leads to a redistribution of excitations over photosystems I and II (PSI and PSII). Th

  20. Dynamic regulation of photosynthesis in Chlamydomonas reinhardtii.

    Science.gov (United States)

    Minagawa, Jun; Tokutsu, Ryutaro

    2015-05-01

    Plants and algae have acquired the ability to acclimatize to ever-changing environments to survive. During photosynthesis, light energy is converted by several membrane protein supercomplexes into electrochemical energy, which is eventually used to assimilate CO2 . The efficiency of photosynthesis is modulated by many environmental factors, including temperature, drought, CO2 concentration, and the quality and quantity of light. Recently, our understanding of such regulators of photosynthesis and the underlying molecular mechanisms has increased considerably. The photosynthetic supercomplexes undergo supramolecular reorganizations within a short time after receiving environmental cues. These reorganizations include state transitions that balance the excitation of the two photosystems: qE quenching, which thermally dissipates excess energy at the level of the light-harvesting antenna, and cyclic electron flow, which supplies the increased ATP demanded by CO2 assimilation and the pH gradient to activate qE quenching. This review focuses on the recent findings regarding the environmental regulation of photosynthesis in model organisms, paying particular attention to the unicellular green alga Chlamydomonas reinhardtii, which offer a glimpse into the dynamic behavior of photosynthetic machinery in nature.

  1. Excitation energy transfer in Chlamydomonas reinhardtii deficient in the PSI core or the PSII core under conditions mimicking state transitions.

    Science.gov (United States)

    Wlodarczyk, Lucyna M; Dinc, Emine; Croce, Roberta; Dekker, Jan P

    2016-06-01

    The efficient use of excitation energy in photosynthetic membranes is achieved by a dense network of pigment-protein complexes. These complexes fulfill specific functions and interact dynamically with each other in response to rapidly changing environmental conditions. Here, we studied how in the intact cells of Chlamydomonas reinhardtii (C.r.) the lack of the photosystem I (PSI) core or the photosystem II (PSII) core affects these interactions. To that end the mutants F15 and M18 (both PSI-deficient) and FUD7 (PSII-deficient) were incubated under conditions known to promote state transitions in wild-type. The intact cells were then instantly frozen to 77K and the full-spectrum time-resolved fluorescence emission of the cells was measured by means of streak camera. In the PSI-deficient mutants excitation energy transfer (EET) towards light-harvesting complexes of PSI (Lhca) occurs in less than 0.5 ns, and fluorescence from Lhca decays in 3.1 ns. Decreased trapping by PSII and increased fluorescence of Lhca upon state 1 (S1)→state 2 (S2) transition appears in the F15 and less in the M18 mutant. In the PSII-deficient mutant FUD7, quenched (0.5 ns) and unquenched (2 ns) light-harvesting complexes of PSII (LHCII) are present in both states, with the quenched form more abundant in S2 than in S1. Moreover, EET of 0.4 ns from the remaining LHCII to PSI increases upon S1→S2 transition. We relate the excitation energy kinetics observed in F15, M18 and FUD7 to the remodeling of the photosynthetic apparatus in these mutants under S1 and S2 conditions.

  2. Light harvesting in photosystem II

    NARCIS (Netherlands)

    Amerongen, van H.; Croce, R.

    2013-01-01

    Water oxidation in photosynthesis takes place in photosystem II (PSII). This photosystem is built around a reaction center (RC) where sunlight-induced charge separation occurs. This RC consists of various polypeptides that bind only a few chromophores or pigments, next to several other cofactors. It

  3. Phosphorylation controls the localization and activation of the lumenal carbonic anhydrase in Chlamydomonas reinhardtii.

    Directory of Open Access Journals (Sweden)

    Amaya Blanco-Rivero

    Full Text Available BACKGROUND: Cah3 is the only carbonic anhydrase (CA isoform located in the thylakoid lumen of Chlamydomonas reinhardtii. Previous studies demonstrated its association with the donor side of the photosystem II (PSII where it is required for the optimal function of the water oxidizing complex. However this enzyme has also been frequently proposed to perform a critical function in inorganic carbon acquisition and CO(2 fixation and all mutants lacking Cah3 exhibit very poor growth after transfer to low CO(2 conditions. RESULTS/CONCLUSIONS: In the present work we demonstrate that after transfer to low CO(2, Cah3 is phosphorylated and that phosphorylation is correlated to changes in its localization and its increase in activity. When C. reinhardtii wild-type cells were acclimated to limiting CO(2 conditions, the Cah3 activity increased about 5-6 fold. Under these conditions, there were no detectable changes in the level of the Cah3 polypeptide. The increase in activity was specifically inhibited in the presence of Staurosporine, a protein kinase inhibitor, suggesting that the Cah3 protein was post-translationally regulated via phosphorylation. Immunoprecipitation and in vitro dephosphorylation experiments confirm this hypothesis. In vivo phosphorylation analysis of thylakoid polypeptides indicates that there was a 3-fold increase in the phosphorylation signal of the Cah3 polypeptide within the first two hours after transfer to low CO(2 conditions. The increase in the phosphorylation signal was correlated with changes in the intracellular localization of the Cah3 protein. Under high CO(2 conditions, the Cah3 protein was only associated with the donor side of PSII in the stroma thylakoids. In contrast, in cells grown at limiting CO(2 the protein was partly concentrated in the thylakoids crossing the pyrenoid, which did not contain PSII and were surrounded by Rubisco molecules. SIGNIFICANCE: This is the first report of a CA being post

  4. Oxidation-reduction signalling components in regulatory pathways of state transitions and photosystem stoichiometry adjustment in chloroplasts.

    Science.gov (United States)

    Puthiyaveetil, Sujith; Ibrahim, Iskander M; Allen, John F

    2012-02-01

    State transitions and photosystem stoichiometry adjustment are two oxidation-reduction (redox)-regulated acclimatory responses in photosynthesis. State transitions are short-term adaptations that, in chloroplasts, involve reversible post-translational modification by phosphorylation of light-harvesting complex II (LHC II). Photosystem stoichiometry adjustments are long-term responses involving transcriptional regulation of reaction centre genes. Both responses are initiated by changes in light quality and are regulated by the redox state of plastoquinone (PQ). The LHC II kinase involved in the state 2 transition is a serine/threonine kinase known as STT7 in Chlamydomonas, and as STN7 in Arabidopsis. The phospho-LHC II phosphatase that produces the state 1 transition is a PP2C-type protein phosphatase currently termed both TAP38 and PPH1. In plants and algae, photosystem stoichiometry adjustment is governed by a modified two-component sensor kinase of cyanobacterial origin - chloroplast sensor kinase (CSK). CSK is a sensor of the PQ redox state. Chloroplast sigma factor 1 (SIG1) and plastid transcription kinase (PTK) are the functional partners of CSK in chloroplast gene regulation. We suggest a signalling pathway for photosystem stoichiometry adjustment. The signalling pathways of state transitions and photosystem stoichiometry adjustments are proposed to be distinct, with the two pathways sensing PQ redox state independently of each other.

  5. Two types of chloroplast gene promoters in Chlamydomonas reinhardtii.

    Science.gov (United States)

    Klein, U; De Camp, J D; Bogorad, L

    1992-04-15

    Structures of the promoters of Chlamydomonas reinhardtii plastid atpB and 16S rRNA-encoding genes were analyzed in vivo. Chimeric constructs, containing the Chlamydomonas chloroplast atpB or 16S rRNA-encoding gene promoter coupled to the Escherichia coli uidA (beta-glucuronidase, GUS) reporter gene and bordered by C. reinhardtii chloroplast sequences, were stably introduced into the chloroplast of Chlamydomonas by microprojectile bombardment. Activity of the promoters in the chloroplast of GUS gene-positive transformants was assayed by measuring the abundance of GUS transcripts and determining the relative rates of GUS transcription in vivo. Deletion analyses of the 16S rRNA gene and atpB promoter fragments showed that the two promoters differ structurally. The 16S rRNA gene promoter resembles the bacterial sigma 70 type with typical -10 and -35 elements. The atpB promoter, on the other hand, lacks a conserved motif in the -35 region but contains, in the -10 region, a characteristic octameric palindrome (TATAATAT) that is conserved in the promoter sequences of some other C. reinhardtii chloroplast genes. For maximum activity, the atpB promoter requires sequences of approximately 22 base pairs upstream and approximately 60 base pairs downstream of the transcription start site.

  6. Modulation of Chlamydomonas reinhardtii flagellar motility by redox poise

    Science.gov (United States)

    Wakabayashi, Ken-ichi; King, Stephen M.

    2006-01-01

    Redox-based regulatory systems are essential for many cellular activities. Chlamydomonas reinhardtii exhibits alterations in motile behavior in response to different light conditions (photokinesis). We hypothesized that photokinesis is signaled by variations in cytoplasmic redox poise resulting from changes in chloroplast activity. We found that this effect requires photosystem I, which generates reduced NADPH. We also observed that photokinetic changes in beat frequency and duration of the photophobic response could be obtained by altering oxidative/reductive stress. Analysis of reactivated cell models revealed that this redox poise effect is mediated through the outer dynein arms (ODAs). Although the global redox state of the thioredoxin-related ODA light chains LC3 and LC5 and the redox-sensitive Ca2+-binding subunit of the docking complex DC3 did not change upon light/dark transitions, we did observe significant alterations in their interactions with other flagellar components via mixed disulfides. These data indicate that redox poise directly affects ODAs and suggest that it may act in the control of flagellar motility. PMID:16754958

  7. New steps toward understanding the regulation of photosystem i biogenesis in chlamydomonas reinhardtii

    OpenAIRE

    Douchi, Damien

    2016-01-01

    Cette thèse décrit l’identification de MAC1, une protéine nouvellement identifiée encodée par le génome nucléaire, impliquée dans la stabilisation de l’ARN messager de psaC, sous-unité du photosystème I. L’identification de ce gène a été possible suite à l’isolation d’un mutant d’insertion dont le génotypage par PCR inverse qui a caractérisé le gène affecté. Ce gène code pour MAC1 qui est une protéine de 100 kDa environ contenant des répétitions de type « Tetratricopeptide repeats », TPR ou H...

  8. Selenoprotein-Transgenic Chlamydomonas reinhardtii

    Directory of Open Access Journals (Sweden)

    Jiazuan Ni

    2013-02-01

    Full Text Available Selenium (Se deficiency is associated with the occurrence of many diseases. However, excessive Se supplementation, especially with inorganic Se, can result in toxicity. Selenoproteins are the major forms of Se in vivo to exert its biological function. Expression of those selenoproteins, especially with the application of a newly developed system, is thus very important for studying the mechanism of Se in nutrition. The use of Chlamydomonas reinhardtii (C. reinhardtii as a biological vector to express an heterogeneous protein is still at the initial stages of development. In order to investigate the possibility of using this system to express selenoproteins, human 15-KDa selenoprotein (Sep15, a small but widely distributed selenoprotein in mammals, was chosen for the expression platform test. Apart from the wild-type human Sep15 gene fragment, two Sep15 recombinants were constructed containing Sep15 open reading frame (ORF and the selenocysteine insertion sequence (SECIS element from either human Sep15 or C. reinhardtii selenoprotein W1, a highly expressed selenoprotein in this alga. Those Sep15-containing plasmids were transformed into C. reinhardtii CC-849 cells. Results showed that Sep15 fragments were successfully inserted into the nuclear genome and expressed Sep15 protein in the cells. The transgenic and wild-type algae demonstrated similar growth curves in low Se culture medium. To our knowledge, this is the first report on expressing human selenoprotein in green alga.

  9. LHCSR3 affects de-coupling and re-coupling of LHCII to PSII during state transitions in Chlamydomonas reinhardtii

    Science.gov (United States)

    Roach, Thomas; Na, Chae Sun

    2017-01-01

    Photosynthetic organisms have to tolerate rapid changes in light intensity, which is facilitated by non-photochemical quenching (NPQ) and involves modification of energy transfer from light-harvesting complexes (LHC) to the photosystem reaction centres. NPQ includes dissipating excess light energy to heat (qE) and the reversible coupling of LHCII to photosystems (state transitions/qT), which are considered separate NPQ mechanisms. In the model alga Chlamydomonas reinhardtii the LHCSR3 protein has a well characterised role in qE. Here, it is shown in the npq4 mutant, deficient in LHCSR3, that energy coupling to photosystem II (PSII) more akin to qT is also disrupted, but no major differences in LHC phosphorylation or LHC compositions were found in comparison to wild-type cells. The qT of wild-type cells possessed two kinetically distinguishable phases, with LHCSR3 participating in the more rapid (<2 min) phase. This LHCSR3-mediated qT was sensitive to physiological levels of H2O2, which accelerated qE induction, revealing a way that may help C. reinhardtii tolerate a sudden increase in light intensity. Overall, a clear mechanistic overlap between qE and qT is shown. PMID:28233792

  10. An improved ARS2-derived nuclear reporter enhances the efficiency and ease of genetic engineering in Chlamydomonas

    DEFF Research Database (Denmark)

    Specht, Elizabeth A; Nour-Eldin, Hussam Hassan; Hoang, Kevin T D

    2015-01-01

    The model alga Chlamydomonas reinhardtii has been used to pioneer genetic engineering techniques for high-value protein and biofuel production from algae. To date, most studies of transgenic Chlamydomonas have utilized the chloroplast genome due to its ease of engineering, with a sizeable suite o...... nuclear promoters. This enhanced arylsulfatase reporter construct improves the efficiency and ease of genetic engineering within the Chlamydomonas nuclear genome, with potential application to other algal strains.......The model alga Chlamydomonas reinhardtii has been used to pioneer genetic engineering techniques for high-value protein and biofuel production from algae. To date, most studies of transgenic Chlamydomonas have utilized the chloroplast genome due to its ease of engineering, with a sizeable suite...... of reporters and well-characterized expression constructs. The advanced manipulation of algal nuclear genomes has been hampered by limited strong expression cassettes, and a lack of high-throughput reporters. We have improved upon an endogenous reporter gene - the ARS2 gene encoding an arylsulfatase enzyme...

  11. Antenna complexes protect Photosystem I from Photoinhibition

    Directory of Open Access Journals (Sweden)

    Hienerwadel Rainer

    2009-06-01

    Full Text Available Abstract Background Photosystems are composed of two moieties, a reaction center and a peripheral antenna system. In photosynthetic eukaryotes the latter system is composed of proteins belonging to Lhc family. An increasing set of evidences demonstrated how these polypeptides play a relevant physiological function in both light harvesting and photoprotection. Despite the sequence similarity between antenna proteins associated with the two Photosystems, present knowledge on their physiological role is mostly limited to complexes associated to Photosystem II. Results In this work we analyzed the physiological role of Photosystem I antenna system in Arabidopsis thaliana both in vivo and in vitro. Plants depleted in individual antenna polypeptides showed a reduced capacity for photoprotection and an increased production of reactive oxygen species upon high light exposure. In vitro experiments on isolated complexes confirmed that depletion of antenna proteins reduced the resistance of isolated Photosystem I particles to high light and that the antenna is effective in photoprotection only upon the interaction with the core complex. Conclusion We show that antenna proteins play a dual role in Arabidopsis thaliana Photosystem I photoprotection: first, a Photosystem I with an intact antenna system is more resistant to high light because of a reduced production of reactive oxygen species and, second, antenna chlorophyll-proteins are the first target of high light damages. When photoprotection mechanisms become insufficient, the antenna chlorophyll proteins act as fuses: LHCI chlorophylls are degraded while the reaction center photochemical activity is maintained. Differences with respect to photoprotection strategy in Photosystem II, where the reaction center is the first target of photoinhibition, are discussed.

  12. The Chlamydomonas genome project: a decade on

    Science.gov (United States)

    Blaby, Ian K.; Blaby-Haas, Crysten; Tourasse, Nicolas; Hom, Erik F. Y.; Lopez, David; Aksoy, Munevver; Grossman, Arthur; Umen, James; Dutcher, Susan; Porter, Mary; King, Stephen; Witman, George; Stanke, Mario; Harris, Elizabeth H.; Goodstein, David; Grimwood, Jane; Schmutz, Jeremy; Vallon, Olivier; Merchant, Sabeeha S.; Prochnik, Simon

    2014-01-01

    The green alga Chlamydomonas reinhardtii is a popular unicellular organism for studying photosynthesis, cilia biogenesis and micronutrient homeostasis. Ten years since its genome project was initiated, an iterative process of improvements to the genome and gene predictions has propelled this organism to the forefront of the “omics” era. Housed at Phytozome, the Joint Genome Institute’s (JGI) plant genomics portal, the most up-to-date genomic data include a genome arranged on chromosomes and high-quality gene models with alternative splice forms supported by an abundance of RNA-Seq data. Here, we present the past, present and future of Chlamydomonas genomics. Specifically, we detail progress on genome assembly and gene model refinement, discuss resources for gene annotations, functional predictions and locus ID mapping between versions and, importantly, outline a standardized framework for naming genes. PMID:24950814

  13. Light-harvesting complex II (LHCII) and its supramolecular organization in Chlamydomonas reinhardtii.

    Science.gov (United States)

    Drop, Bartlomiej; Webber-Birungi, Mariam; Yadav, Sathish K N; Filipowicz-Szymanska, Alicja; Fusetti, Fabrizia; Boekema, Egbert J; Croce, Roberta

    2014-01-01

    LHCII is the most abundant membrane protein on earth. It participates in the first steps of photosynthesis by harvesting sunlight and transferring excitation energy to the core complex. Here we have analyzed the LHCII complex of the green alga Chlamydomonas reinhardtii and its association with the core of Photosystem II (PSII) to form multiprotein complexes. Several PSII supercomplexes with different antenna sizes have been purified, the largest of which contains three LHCII trimers (named S, M and N) per monomeric core. A projection map at a 13Å resolution was obtained allowing the reconstruction of the 3D structure of the supercomplex. The position and orientation of the S trimer are the same as in plants; trimer M is rotated by 45° and the additional trimer (named here as LHCII-N), which is taking the position occupied in plants by CP24, is directly associated with the core. The analysis of supercomplexes with different antenna sizes suggests that LhcbM1, LhcbM2/7 and LhcbM3 are the major components of the trimers in the PSII supercomplex, while LhcbM5 is part of the "extra" LHCII pool not directly associated with the supercomplex. It is also shown that Chlamydomonas LHCII has a slightly lower Chlorophyll a/b ratio than the complex from plants and a blue shifted absorption spectrum. Finally the data indicate that there are at least six LHCII trimers per dimeric core in the thylakoid membranes, meaning that the antenna size of PSII of C. reinhardtii is larger than that of plants.

  14. MEETING: Chlamydomonas Annotation Jamboree - October 2003

    Energy Technology Data Exchange (ETDEWEB)

    Grossman, Arthur R

    2007-04-13

    Shotgun sequencing of the nuclear genome of Chlamydomonas reinhardtii (Chlamydomonas throughout) was performed at an approximate 10X coverage by JGI. Roughly half of the genome is now contained on 26 scaffolds, all of which are at least 1.6 Mb, and the coverage of the genome is ~95%. There are now over 200,000 cDNA sequence reads that we have generated as part of the Chlamydomonas genome project (Grossman, 2003; Shrager et al., 2003; Grossman et al. 2007; Merchant et al., 2007); other sequences have also been generated by the Kasuza sequence group (Asamizu et al., 1999; Asamizu et al., 2000) or individual laboratories that have focused on specific genes. Shrager et al. (2003) placed the reads into distinct contigs (an assemblage of reads with overlapping nucleotide sequences), and contigs that group together as part of the same genes have been designated ACEs (assembly of contigs generated from EST information). All of the reads have also been mapped to the Chlamydomonas nuclear genome and the cDNAs and their corresponding genomic sequences have been reassembled, and the resulting assemblage is called an ACEG (an Assembly of contiguous EST sequences supported by genomic sequence) (Jain et al., 2007). Most of the unique genes or ACEGs are also represented by gene models that have been generated by the Joint Genome Institute (JGI, Walnut Creek, CA). These gene models have been placed onto the DNA scaffolds and are presented as a track on the Chlamydomonas genome browser associated with the genome portal (http://genome.jgi-psf.org/Chlre3/Chlre3.home.html). Ultimately, the meeting grant awarded by DOE has helped enormously in the development of an annotation pipeline (a set of guidelines used in the annotation of genes) and resulted in high quality annotation of over 4,000 genes; the annotators were from both Europe and the USA. Some of the people who led the annotation initiative were Arthur Grossman, Olivier Vallon, and Sabeeha Merchant (with many individual

  15. Loss of CpSRP54 function leads to a truncated light-harvesting antenna size in Chlamydomonas reinhardtii.

    Science.gov (United States)

    Jeong, Jooyeon; Baek, Kwangryul; Kirst, Henning; Melis, Anastasios; Jin, EonSeon

    2017-01-01

    The Chlamydomonas reinhardtii truncated light-harvesting antenna 4 (tla4) DNA transposon mutant has a pale green phenotype, a lower chlorophyll (Chl) per cell and a higher Chl a/b ratio in comparison with the wild type. It required a higher light intensity for the saturation of photosynthesis and displayed a greater per chlorophyll light-saturated rate of oxygen evolution than the wild type. The Chl antenna size of the photosystems in the tla4 mutant was only about 65% of that measured in the wild type. Molecular genetic analysis revealed that a single plasmid DNA insertion disrupted two genes on chromosome 11 of the mutant. A complementation study identified the "chloroplast signal recognition particle 54" gene (CpSRP54), as the lesion causing the tla4 phenotype. Disruption of this gene resulted in partial failure to assemble and, therefore, lower levels of light-harvesting Chl-binding proteins in the C. reinhardtii thylakoids. A comparative in silico 3-D structure-modeling analysis revealed that the M-domain of the CpSRP54 of C. reinhardtii possesses a more extended finger loop structure, due to different amino acid composition, as compared to that of the Arabidopsis CpSRP54. The work demonstrated that CpSRP54 deletion in microalgae can serve to generate tla mutants with a markedly smaller photosystem Chl antenna size, improved solar energy conversion efficiency, and photosynthetic productivity in high-density cultures under bright sunlight conditions.

  16. Excitation energy transfer in the photosystem I

    Energy Technology Data Exchange (ETDEWEB)

    Webber, Andrew N

    2012-09-25

    Photosystem I is a multimeric pigment protein complex in plants, green alage and cyanobacteria that functions in series with Photosystem II to use light energy to oxidize water and reduce carbon dioxide. The Photosystem I core complex contains 96 chlorophyll a molecules and 22 carotenoids that are involved in light harvesting and electron transfer. In eucaryotes, PSI also has a peripheral light harvesting complex I (LHCI). The role of specific chlorophylls in excitation and electron transfer are still unresolved. In particular, the role of so-called bridging chlorophylls, located between the bulk antenna and the core electron transfer chain, in the transfer of excitation energy to the reaction center are unknown. During the past funding period, site directed mutagenesis has been used to create mutants that effect the physical properties of these key chlorophylls, and to explore how this alters the function of the photosystem. Studying these mutants using ultrafast absorption spectroscopy has led to a better understanding of the process by which excitation energy is transferred from the antenna chlorophylls to the electron transfer chain chlorophylls, and what the role of connecting chlorophylls and A_0 chlorophylls is in this process. We have also used these mutants to investigate whch of the central group of six chlorophylls are involved in the primary steps of charge separation and electron transfer.

  17. Light stress and photoprotection in Chlamydomonas reinhardtii.

    Science.gov (United States)

    Erickson, Erika; Wakao, Setsuko; Niyogi, Krishna K

    2015-05-01

    Plants and algae require light for photosynthesis, but absorption of too much light can lead to photo-oxidative damage to the photosynthetic apparatus and sustained decreases in the efficiency and rate of photosynthesis (photoinhibition). Light stress can adversely affect growth and viability, necessitating that photosynthetic organisms acclimate to different environmental conditions in order to alleviate the detrimental effects of excess light. The model unicellular green alga, Chlamydomonas reinhardtii, employs diverse strategies of regulation and photoprotection to avoid, minimize, and repair photo-oxidative damage in stressful light conditions, allowing for acclimation to different and changing environments.

  18. Real-time monitoring of genetically modified Chlamydomonas reinhardtii during the Foton M3 space mission

    Science.gov (United States)

    Lambreva, M.; Rea, G.; Antonacci, A.; Serafini, A.; Damasso, M.; Pastorelli, S.; Margonelli, A.; Johanningmeier, U.; Bertalan, I.; Pezzotti, G.; Giardi, M. T.

    2008-09-01

    Long-term space exploration, colonization or habitation requires biological life support systems capable to cope with the deleterious space environment. The use of oxygenic photosynthetic microrganisms is an intriguing possibility mainly for food, O2 and nutraceutical compounds production. The critical points of utilizing plants- or algae-based life support systems are the microgravity and the ionizing radiation, which can influence the performance of these organisms. The aim of the present study was to assess the effects of space environment on the photosynthetic activity of various microrganisms and to select space stresstolerant strains. Photosystem II D1 protein sitedirected and random mutants of the unicellular green alga Chlamydomonas reinhardtii [1] were used as a model system to test and select the amino acid substitutions capable to account for space stress tolerance. We focussed our studies also on the accumulation of the Photosystem II photoprotective carotenoids (the xantophylls violaxanthin, anteraxanthin and zeaxanthin), powerful antioxidants that epidemiological studies demonstrated to be human vision protectors. For this purpose some mutants modified at the level of enzymes involved in the biosynthesis of xanthophylls were included in the study [2]. To identify the consequences of the space environment on the photosynthetic apparatus the changes in the Photosystem II efficiency were monitored in real time during the ESA-Russian Foton- M3 mission in September 2007. For the space flight a high-tech, multicell fluorescence detector, Photo-II, was designed and built by the Centre for Advanced Research in Space Optics in collaboration with Kayser-Italy, Biosensor and DAS. Photo-II is an automatic device developed to measure the chlorophyll fluorescence and to provide a living conditions for several different algae strains (Fig.1). Twelve different C. reinhardti strains were analytically selected and two replications for each strain were brought to space

  19. [An experiment with Chlamydomonas reinhardtii on the Kosmos-2044 biosatellite].

    Science.gov (United States)

    Gavrilova, O V; Gabova, A V; Goriainova, L N; Filatova, E V

    1992-01-01

    Space experiment with Chlamydomonas reinhardtii demonstrated that the microgravity effects were noted in Chlamydomonas at both cellular and population levels: in space the cell size is increased, stage of active growth of the culture is extended, it contains the juvenile vegetative motile cells in greater quantities. Ultrastructural analysis indicated that in microgravity the changes in shape, structure and distribution of intracellular organelles and in volume ratio of organelles and cytoplasma are absent. Chlamydomonas data are in line with the results of the Infusoria and Chlorella experiments.

  20. Controlling expression of genes in the unicellular alga Chlamydomonas reinhardtii with a vitamin-repressible riboswitch.

    Science.gov (United States)

    Ramundo, Silvia; Rochaix, Jean-David

    2015-01-01

    Chloroplast genomes of land plants and algae contain generally between 100 and 150 genes. These genes are involved in plastid gene expression and photosynthesis and in various other tasks. The function of some chloroplast genes is still unknown and some of them appear to be essential for growth and survival. Repressible and reversible expression systems are highly desirable for functional and biochemical characterization of these genes. We have developed a genetic tool that allows one to regulate the expression of any coding sequence in the chloroplast genome of the unicellular alga Chlamydomonas reinhardtii. Our system is based on vitamin-regulated expression of the nucleus-encoded chloroplast Nac2 protein, which is specifically required for the expression of any plastid gene fused to the psbD 5'UTR. With this approach, expression of the Nac2 gene in the nucleus and, in turn, that of the chosen chloroplast gene artificially driven by the psbD 5'UTR, is controlled by the MetE promoter and Thi4 riboswitch, which can be inactivated in a reversible way by supplying vitamin B12 and thiamine to the growth medium, respectively. This system opens interesting possibilities for studying the assembly and turnover of chloroplast multiprotein complexes such as the photosystems, the ribosome, and the RNA polymerase. It also provides a way to overcome the toxicity often associated with the expression of proteins of biotechnological interest in the chloroplast.

  1. Retrograde Signaling and Photoprotection in a gun4 Mutant of Chlamydomonas reinhardtii

    Institute of Scientific and Technical Information of China (English)

    Cinzia Formighieri; Mauro Ceol; Giulia Bonente; Jean-David Rochaix; Roberto Bassi

    2012-01-01

    GUN4 is a regulatory subunit of Mg-chelatase involved in the control of tetrapyrrole synthesis in plants and cyanobacteria.Here,we report the first characterization of a gun4 insertion mutant of the unicellular green alga Chlamydomonas reinhardtii.The mutant contains 50% of chlorophyll as compared to wild-type and accumulates ProtolX.In contrast to the increase in LHCtranscription,the accumulation of most LHC proteins is drastically diminished,implying posttranscriptional down-regulation in the absence of transcriptional coordination.We found that 803 genes change their expression level in gun4 as compared to wild-type,by RNA-Seq,and this wide-ranging effect on transcription is apparent under physiological conditions.Besides LHCs,we identified transcripts encoding enzymes of the tetrapyrrole pathway and factors involved in signal transduction,transcription,and chromatin remodeling.Moreover,we observe perturbations in electron transport with a strongly decreased PSI-to-PSII ratio.This is accompanied by an enhanced activity of the plastid terminal oxidase (PTOX) that could have a physiological role in decreasing photosystem Ⅱ excitation pressure.

  2. Process development for hydrogen production with Chlamydomonas reinhardtii based on growth and product formation kinetics.

    Science.gov (United States)

    Lehr, Florian; Morweiser, Michael; Rosello Sastre, Rosa; Kruse, Olaf; Posten, Clemens

    2012-11-30

    Certain strains of microalgae are long known to produce hydrogen under anaerobic conditions. In Chlamydomonas reinhardtii the oxygen-sensitive hydrogenase enzyme recombines electrons from the chloroplast electron transport chain with protons to form molecular hydrogen directly inside the chloroplast. A sustained hydrogen production can be obtained under low sulfur conditions in C. reinhardtii, reducing the net oxygen evolution by reducing the photosystem II activity and thereby overcoming the inhibition of the hydrogenases. The development of specially adapted hydrogen production strains led to higher yields and optimized biological process preconditions. So far sustainable hydrogen production required a complete exchange of the growth medium to establish sulfur-deprived conditions after biomass growth. In this work we demonstrate the transition from the biomass growth phase to the hydrogen production phase in a single batch culture only by exact dosage of sulfur. This eliminates the elaborate and energy intensive solid-liquid separation step and establishes a process strategy to proceed further versus large scale production. This strategy has been applied to determine light dependent biomass growth and hydrogen production kinetics to assess the potential of H₂ production with C. reinhardtii as a basis for scale up and further process optimization.

  3. Advances in the biotechnology of hydrogen production with the microalga Chlamydomonas reinhardtii.

    Science.gov (United States)

    Torzillo, Giuseppe; Scoma, Alberto; Faraloni, Cecilia; Giannelli, Luca

    2015-01-01

    Biological hydrogen production is being evaluated for use as a fuel, since it is a promising substitute for carbonaceous fuels owing to its high conversion efficiency and high specific energy content. The basic advantages of biological hydrogen production over other "green" energy sources are that it does not compete for agricultural land use, and it does not pollute, as water is the only by-product of the combustion. These characteristics make hydrogen a suitable fuel for the future. Among several biotechnological approaches, photobiological hydrogen production carried out by green microalgae has been intensively investigated in recent years. A select group of photosynthetic organisms has evolved the ability to harness light energy to drive hydrogen gas production from water. Of these, the microalga Chlamydomonas reinhardtii is considered one of the most promising eukaryotic H2 producers. In this model microorganism, light energy, H2O and H2 are linked by two excellent catalysts, the photosystem 2 (PSII) and the [FeFe]-hydrogenase, in a pathway usually referred to as direct biophotolysis. This review summarizes the main advances made over the past decade as an outcome of the discovery of the sulfur-deprivation process. Both the scientific and technical barriers that need to be overcome before H2 photoproduction can be scaled up to an industrial level are examined. Actual and theoretical limits of the efficiency of the process are also discussed. Particular emphasis is placed on algal biohydrogen production outdoors, and guidelines for an optimal photobioreactor design are suggested.

  4. Mutagenesis of Ser24 of cytochrome b559 α subunit affects PSⅡ activities in Chlamydomonas reinhardtii

    Institute of Scientific and Technical Information of China (English)

    MA JingJing; LI LiangBi; JING YuXiang; KUANG TingYun

    2007-01-01

    In order to study the functions of cytochrome b559 (Cyt b559) in photosystem two (PSⅡ) activity, mutant S24F of Chlamydomonas reinhardtii was constructed using site directed mutagenesis, in which Serine24 (Ser24) locating downstream of Histidine23 (His23) in c subunit of Cyt b559 was replaced by Phenylalanine (Phe). Physiological and biochemical analysis showed that mutant S24F could be grown photoautotrophically or photoheterotrophically. However, their growth rate was slower either on HSM or TAP medium than that of the control; Analysis of PSⅡ activity revealed that its oxygen evolution was about 71% of wild type (WT); The Photochemical efficiency of PSll (Fv/Fm) of S24F was reduced 0.23 compared with WT; S24F was more sensitive to strong light irradiance than the wild type; Furthermore,SDS-PAGE and Western-blotting analysis indicated that the expression levels of c subunit of Cyt b559,LHCⅡ and PsbO of S24F were a little less than those of the wild type. Overall, these data suggests that Ser24 plays a significant role in making Cyt b559 structure maintain PSⅡ complex activity of oxygen evolution although it is not directly bound to heme group.

  5. Integration of carbon assimilation modes with photosynthetic light capture in the green alga Chlamydomonas reinhardtii.

    Science.gov (United States)

    Berger, Hanna; Blifernez-Klassen, Olga; Ballottari, Matteo; Bassi, Roberto; Wobbe, Lutz; Kruse, Olaf

    2014-10-01

    The unicellular green alga Chlamydomonas reinhardtii is capable of using organic and inorganic carbon sources simultaneously, which requires the adjustment of photosynthetic activity to the prevailing mode of carbon assimilation. We obtained novel insights into the regulation of light-harvesting at photosystem II (PSII) following altered carbon source availability. In C. reinhardtii, synthesis of PSII-associated light-harvesting proteins (LHCBMs) is controlled by the cytosolic RNA-binding protein NAB1, which represses translation of particular LHCBM isoform transcripts. This mechanism is fine-tuned via regulation of the nuclear NAB1 promoter, which is activated when linear photosynthetic electron flow is restricted by CO(2)-limitation in a photoheterotrophic context. In the wild-type, accumulation of NAB1 reduces the functional PSII antenna size, thus preventing a harmful overexcited state of PSII, as observed in a NAB1-less mutant. We further demonstrate that translation control as a newly identified long-term response to prolonged CO(2)-limitation replaces LHCII state transitions as a fast response to PSII over-excitation. Intriguingly, activation of the long-term response is perturbed in state transition mutant stt7, suggesting a regulatory link between the long- and short-term response. We depict a regulatory circuit operating on distinct timescales and in different cellular compartments to fine-tune light-harvesting in photoheterotrophic eukaryotes.

  6. Effect of aluminum on cellular division and photosynthetic electron transport in Euglena gracilis and Chlamydomonas acidophila.

    Science.gov (United States)

    Perreault, François; Dewez, David; Fortin, Claude; Juneau, Philippe; Diallo, Amirou; Popovic, Radovan

    2010-04-01

    The present study investigated aluminum's effect on cellular division and the photosynthetic processes in Euglena gracilis and Chlamydomonas acidophila at pH 3.0, at which Al is present mostly as Al(3+), AlSO(4) (+), and Al(SO(4))(2) (-). These algal species were exposed to 100, 188, and 740 microM Al, and after 24 h cell-bound Al was significantly different from control only for the highest concentration tested. However, very different effects of Al on algal cellular division, biomass per cell, and photosynthetic activity were found. Aluminum stimulated cell division but decreased at some level biomass per cell in C. acidophila. Primary photochemistry of photosynthesis, as Photosystem II quantum yield, and energy dissipation via nonphotochemical activity were slightly affected. However, for E. gracilis, under the same conditions, Al did not show a stimulating effect on cellular division or photosynthetic activity. Primary photochemical activity was diminished, and energy dissipation via nonphotochemical pathways was strongly increased. Therefore, when Al is highly available in aquatic ecosystems, these effects may indicate very different response mechanisms that are dependent on algal species.

  7. Paternal inheritance of mitochondria in Chlamydomonas.

    Science.gov (United States)

    Nakamura, Soichi

    2010-03-01

    To analyze mitochondrial DNA (mtDNA)inheritance, differences in mtDNA between Chlamydomonas reinhardtii and Chlamydomonas smithii, respiration deficiency and antibiotic resistance were used to distinguish mtDNA origins. The analyses indicated paternal inheritance. However, these experiments raised questions regarding whether paternal inheritance occurred normally.Mitochondrial nucleoids were observed in living zygotes from mating until 3 days after mating and then until progeny formation. However, selective disappearance of nucleoids was not observed. Subsequently, experimental serial backcrosses between the two strains demonstrated strict paternal inheritance. The fate of mt+ and mt- mtDNA was followed using the differences in mtDNA between the two strains. The slow elimination of mt+ mtDNA through zygote maturation in darkness was observed, and later the disappearance of mt+ mtDNA was observed at the beginning of meiosis. To explain the different fates of mtDNA, methylation status was investigated; however, no methylation was detected. Variously constructed diploid cells showed biparental inheritance. Thus, when the mating process occurs normally, paternal inheritance occurs. Mutations disrupting mtDNA inheritance have not yet been isolated. Mutations that disrupt maternal inheritance of chloroplast DNA (cpDNA) do not disrupt inheritance of mtDNA. The genes responsible for mtDNA inheritance are different from those of chloroplasts.

  8. Retrograde bilin signaling enables Chlamydomonas greening and phototrophic survival

    Science.gov (United States)

    Duanmu, Deqiang; Casero, David; Dent, Rachel M.; Gallaher, Sean; Yang, Wenqiang; Rockwell, Nathan C.; Martin, Shelley S.; Pellegrini, Matteo; Niyogi, Krishna K.; Merchant, Sabeeha S.; Grossman, Arthur R.; Lagarias, J. Clark

    2013-01-01

    The maintenance of functional chloroplasts in photosynthetic eukaryotes requires real-time coordination of the nuclear and plastid genomes. Tetrapyrroles play a significant role in plastid-to-nucleus retrograde signaling in plants to ensure that nuclear gene expression is attuned to the needs of the chloroplast. Well-known sites of synthesis of chlorophyll for photosynthesis, plant chloroplasts also export heme and heme-derived linear tetrapyrroles (bilins), two critical metabolites respectively required for essential cellular activities and for light sensing by phytochromes. Here we establish that Chlamydomonas reinhardtii, one of many chlorophyte species that lack phytochromes, can synthesize bilins in both plastid and cytosol compartments. Genetic analyses show that both pathways contribute to iron acquisition from extracellular heme, whereas the plastid-localized pathway is essential for light-dependent greening and phototrophic growth. Our discovery of a bilin-dependent nuclear gene network implicates a widespread use of bilins as retrograde signals in oxygenic photosynthetic species. Our studies also suggest that bilins trigger critical metabolic pathways to detoxify molecular oxygen produced by photosynthesis, thereby permitting survival and phototrophic growth during the light period. PMID:23345435

  9. A brief introduction to the model microswimmer {\\it Chlamydomonas reinhardtii}

    CERN Document Server

    Jeanneret, Raphaël; Polin, Marco

    2016-01-01

    The unicellular biflagellate green alga {\\it Chlamydomonas reinhardtii} has been an important model system in biology for decades, and in recent years it has started to attract growing attention also within the biophysics community. Here we provide a concise review of some of the aspects of {\\it Chlamydomonas} biology and biophysics most immediately relevant to physicists that might be interested in starting to work with this versatile microorganism.

  10. Consequences of Modification of Photosystem Stoichiometry and Amount in Cyanobacteria

    Energy Technology Data Exchange (ETDEWEB)

    Vermaas, Willem [Arizona State Univ., Tempe, AZ (United States)

    2016-12-13

    The proposed research seeks to address two interconnected, important questions that impact photosynthetic processes and that reflect key differences between the photosynthetic systems of cyanobacteria and plants or algae. The first question is what are the reasons and consequences of the high photosystem I / photosystem II (PS I/PS II) ratio in many cyanobacteria, vs. a ratio that is close to unity in many plants and algae. The corresponding hypothesis is that most of PS I functions in cyclic electron transport, and that reduction in PS I will result primarily in a shortage of ATP rather than reducing power. This hypothesis will be tested by reducing the amount of PS I by changing the promoter region of the psaAB operon in the cyanobacterium Synechocystis sp. PCC 6803 and generating a range of mutants with different PS I content and thereby different PS I/PS II ratios, with some of the mutants having a PS II/PS I ratio closer to that in plants. The resulting mutants will be probed in terms of their growth rates, electron transfer rates, and P700 redox kinetics. A second question relates to a Mehler-type reaction catalyzed by two flavoproteins, Flv1 and Flv3, that accept electrons from PS I and that potentially function as an electron safety valve leading to no useful purpose of the photosynthesis-generated electrons. The hypothesis to be tested is that Flv1 and Flv3 use the electrons for useful purposes such as cyclic electron flow around PS I. This hypothesis will be tested by analysis of a mutant strain lacking flv3, the gene for one of the flavoproteins. This research is important for a more detailed understanding of the consequences of photosystem stoichiometry and amounts in a living system. Such an understanding is critical for not only insights in the regulatory systems of the organism but also to guide the development of biological or bio-hybrid systems for solar energy conversion into fuels.

  11. An improved ARS2-derived nuclear reporter enhances the efficiency and ease of genetic engineering in Chlamydomonas.

    Science.gov (United States)

    Specht, Elizabeth A; Nour-Eldin, Hussam Hassan; Hoang, Kevin T D; Mayfield, Stephen P

    2015-03-01

    The model alga Chlamydomonas reinhardtii has been used to pioneer genetic engineering techniques for high-value protein and biofuel production from algae. To date, most studies of transgenic Chlamydomonas have utilized the chloroplast genome due to its ease of engineering, with a sizeable suite of reporters and well-characterized expression constructs. The advanced manipulation of algal nuclear genomes has been hampered by limited strong expression cassettes, and a lack of high-throughput reporters. We have improved upon an endogenous reporter gene - the ARS2 gene encoding an arylsulfatase enzyme - that was first cloned and characterized decades ago but has not been used extensively. The new construct, derived from ARS2 cDNA, expresses significantly higher levels of reporter protein and transforms more efficiently, allowing qualitative and quantitative screening using a rapid, inexpensive 96-well assay. The improved arylsulfatase expression cassette was used to screen a new transgene promoter from the ARG7 gene, and found that the ARG7 promoter can express the ARS2 reporter as strongly as the HSP70-RBCS2 chimeric promoter that currently ranks as the best available promoter, thus adding to the list of useful nuclear promoters. This enhanced arylsulfatase reporter construct improves the efficiency and ease of genetic engineering within the Chlamydomonas nuclear genome, with potential application to other algal strains.

  12. Genetic tools and techniques for Chlamydomonas reinhardtii.

    Science.gov (United States)

    Mussgnug, Jan H

    2015-07-01

    The development of tools has always been a major driving force for the advancement of science. Optical microscopes were the first instruments that allowed discovery and descriptive studies of the subcellular features of microorganisms. Although optical and electron microscopes remained at the forefront of microbiological research tools since their inventions, the advent of molecular genetics brought about questions which had to be addressed with new "genetic tools". The unicellular green microalgal genus Chlamydomonas, especially the most prominent species C. reinhardtii, has become a frequently used model organism for many diverse fields of research and molecular genetic analyses of C. reinhardtii, as well as the available genetic tools and techniques, have become increasingly sophisticated throughout the last decades. The aim of this review is to provide an overview of the molecular key features of C. reinhardtii and summarize the progress related to the development of tools and techniques for genetic engineering of this organism, from pioneering DNA transformation experiments to state-of-the-art techniques for targeted nuclear genome editing and high-throughput screening approaches.

  13. High quality metabolomic data for Chlamydomonas reinhardtii

    Directory of Open Access Journals (Sweden)

    Fiehn Oliver

    2008-04-01

    Full Text Available Abstract The green eukaryote alga Chlamydomonas reinhardtii is a unicellular model to study control of metabolism in a photosynthetic organism. We here present method improvements for metabolite profiling based on GC-TOF mass spectrometry focusing on three parameters: quenching and cell disruption, extract solvent composition and metabolite annotation. These improvements facilitate using smaller cell numbers and hence, smaller culture volumes which enable faster and more precise sampling techniques that eventually lead to a higher number of samples that can be processed, e.g. for time course experiments. Quenching of metabolism was achieved by mixing 1 ml of culture to 1 ml of -70°C cold 70% methanol. After centrifugation, cells were lyophilized and disrupted by milling using 2-6E6 lyophilized cells, around 500-fold less than previously reported. Glass beads were compared to metal balls for milling, and five different extraction solvents were tested. Additionally, all peaks were annotated in an automated way using the GC-TOF database BinBase instead of manual investigation of a single reference chromatogram. Median precision of analysis was used to decide for the eventual procedure which was applied to a proof-of-principle study of time dependent changes of metabolism under standard conditions.

  14. Radial spoke proteins of Chlamydomonas flagella

    Science.gov (United States)

    Yang, Pinfen; Diener, Dennis R.; Yang, Chun; Kohno, Takahiro; Pazour, Gregory J.; Dienes, Jennifer M.; Agrin, Nathan S.; King, Stephen M.; Sale, Winfield S.; Kamiya, Ritsu; Rosenbaum, Joel L.; Witman, George B.

    2007-01-01

    Summary The radial spoke is a ubiquitous component of ‘9+2’ cilia and flagella, and plays an essential role in the control of dynein arm activity by relaying signals from the central pair of microtubules to the arms. The Chlamydomonas reinhardtii radial spoke contains at least 23 proteins, only 8 of which have been characterized at the molecular level. Here, we use mass spectrometry to identify 10 additional radial spoke proteins. Many of the newly identified proteins in the spoke stalk are predicted to contain domains associated with signal transduction, including Ca2+-, AKAP- and nucleotide-binding domains. This suggests that the spoke stalk is both a scaffold for signaling molecules and itself a transducer of signals. Moreover, in addition to the recently described HSP40 family member, a second spoke stalk protein is predicted to be a molecular chaperone, implying that there is a sophisticated mechanism for the assembly of this large complex. Among the 18 spoke proteins identified to date, at least 12 have apparent homologs in humans, indicating that the radial spoke has been conserved throughout evolution. The human genes encoding these proteins are candidates for causing primary ciliary dyskinesia, a severe inherited disease involving missing or defective axonemal structures, including the radial spokes. PMID:16507594

  15. Drosophila roadblock and Chlamydomonas Lc7

    Science.gov (United States)

    Bowman, Aaron B.; Patel-King, Ramila S.; Benashski, Sharon E.; McCaffery, J. Michael; Goldstein, Lawrence S.B.; King, Stephen M.

    1999-01-01

    Eukaryotic organisms utilize microtubule-dependent motors of the kinesin and dynein superfamilies to generate intracellular movement. To identify new genes involved in the regulation of axonal transport in Drosophila melanogaster, we undertook a screen based upon the sluggish larval phenotype of known motor mutants. One of the mutants identified in this screen, roadblock (robl), exhibits diverse defects in intracellular transport including axonal transport and mitosis. These defects include intra-axonal accumulations of cargoes, severe axonal degeneration, and aberrant chromosome segregation. The gene identified by robl encodes a 97–amino acid polypeptide that is 57% identical (70% similar) to the 105–amino acid Chlamydomonas outer arm dynein–associated protein LC7, also reported here. Both robl and LC7 have homology to several other genes from fruit fly, nematode, and mammals, but not Saccharomyces cerevisiae. Furthermore, we demonstrate that members of this family of proteins are associated with both flagellar outer arm dynein and Drosophila and rat brain cytoplasmic dynein. We propose that roadblock/LC7 family members may modulate specific dynein functions. PMID:10402468

  16. tla1, a DNA insertional transformant of the green alga Chlamydomonas reinhardtii with a truncated light-harvesting chlorophyll antenna size.

    Science.gov (United States)

    Polle, Juergen E W; Kanakagiri, Sarada-Devi; Melis, Anastasios

    2003-05-01

    DNA insertional mutagenesis and screening of the green alga Chlamydomonas reinhardtii was employed to isolate tla1, a stable transformant having a truncated light-harvesting chlorophyll antenna size. Molecular analysis showed a single plasmid insertion into an open reading frame of the nuclear genome corresponding to a novel gene ( Tla1) that encodes a protein of 213 amino acids. Genetic analysis showed co-segregation of plasmid and tla1 phenotype. Biochemical analyses showed the tla1 mutant to be chlorophyll deficient, with a functional chlorophyll antenna size of photosystem I and photosystem II being about 50% and 65% of that of the wild type, respectively. It contained a correspondingly lower amount of light-harvesting proteins than the wild type and had lower steady-state levels of Lhcb mRNA. The tla1 strain required a higher light intensity for the saturation of photosynthesis and showed greater solar conversion efficiencies and a higher photosynthetic productivity than the wild type under mass culture conditions. Results are discussed in terms of the tla1 mutation, its phenotype, and the role played by the Tla1 gene in the regulation of the photosynthetic chlorophyll antenna size in C. reinhardtii.

  17. Hydrogen production by Chlamydomonas reinhardtii: an elaborate interplay of electron sources and sinks

    Energy Technology Data Exchange (ETDEWEB)

    Hemschemeier, A; Happe, T. [Ruhr Univ Bochum, AG Photobiotechnol, Lehrstuhl Biochem Pflanzen, Fak Biol, D-44780 Bochum (Germany); Fouchard, S [Univ Nantes, CNRS, Lab GEPEA, UMR 6144, F-44602 St Nazaire (France); Cournac, L; Peltier, G. [CEA Cadarache, DSV, IBEB, SBVME, Lab Bioenerget et Biotechnol Bacteries et Microalgues, CEA-CNRS-Univ Aix Marseille, UMR 6191, F-13108 St Paul Les Durance (France)

    2008-07-01

    The unicellular green alga Chlamydomonas reinhardtii possesses a [FeFe]-hydrogenase HydA1 (EC 1.12.7.2), which is coupled to the photosynthetic electron transport chain. Large amounts of H{sub 2} are produced in a light-dependent reaction for several days when C. reinhardtii cells are deprived of sulfur. Under these conditions, the cells drastically change their physiology from aerobic photosynthetic growth to an anaerobic resting state. The understanding of the underlying physiological processes is not only important for getting further insights into the adaptability of photosynthesis, but will help to optimize the biotechnological application of algae as H{sub 2} producers. Two of the still most disputed questions regarding H{sub 2} generation by C. reinhardtii concern the electron source for H{sub 2} evolution and the competition of the hydrogenase with alternative electron sinks. We analyzed the H{sub 2} metabolism of S-depleted C. reinhardtii cultures utilizing a special mass spectrometer setup and investigated the influence of photosystem II (PSII)- or ribulose-bisphosphate-carboxylase/oxygenase (Rubisco)-deficiency. We show that electrons for H{sub 2}-production are provided both by PSII activity and by a non-photochemical plastoquinone reduction pathway, which is dependent on previous PSII activity. In a Rubisco-deficient strain, which produces H{sub 2} also in the presence of sulfur, H{sub 2} generation seems to be the only significant electron sink for PSII activity and rescues this strain at least partially from a light-sensitive phenotype.The latter indicates that the down-regulation of assimilatory pathways in S-deprived C. reinhardtii cells is one of the important prerequisites for a sustained H{sub 2} evolution. (authors)

  18. [The Carnot efficiency and plant photosystems].

    Science.gov (United States)

    Jennings, R C; Santabarbara, S; Belgio, E; Zucchelli, G

    2014-01-01

    The concept that the Carnot efficiency places an upper limit of 0.60-0.75 on the thermodynamic efficiency of photosynthetic primary photochemistry is examined using the PSI-LHCI preparation. The maximal quantum efficiency was determined approximately 0.99 which yielded a thermodynamic efficiency of 0.96, a value far above that predicted on the basis of the Carnot efficiency. The commonly presented reasoning leading to the Carnot efficiency idea was therefore critically examined. It is concluded that the crucial assumption that the pigment system, under illumination, is in equilibrium with the incident light field, at a black body temperature of Tr, is erroneous, as the temperature of the excited state pigments was experimentally shown to be that of the sample solvent (thermal bath), 280 K in this case. It is concluded that the classical reasoning used to describe the thermodynamics of heat systems is not applicable to "photonic" systems such as plant photosystems.

  19. Material science lesson from the biological photosystem

    Science.gov (United States)

    Kim, Younghye; Lee, Jun Ho; Ha, Heonjin; Im, Sang Won; Nam, Ki Tae

    2016-08-01

    Inspired by photosynthesis, artificial systems for a sustainable energy supply are being designed. Each sequential energy conversion process from light to biomass in natural photosynthesis is a valuable model for an energy collection, transport and conversion system. Notwithstanding the numerous lessons of nature that provide inspiration for new developments, the features of natural photosynthesis need to be reengineered to meet man's demands. This review describes recent strategies toward adapting key lessons from natural photosynthesis to artificial systems. We focus on the underlying material science in photosynthesis that combines photosystems as pivotal functional materials and a range of materials into an integrated system. Finally, a perspective on the future development of photosynthesis mimetic energy systems is proposed.

  20. Real-time monitoring of genetically modified Chlamydomonas reinhardtii during the Foton M3 space mission and ground irradiation experiment

    Science.gov (United States)

    Lambreva, Maya; Rea, Giuseppina; Antonacci, Amina; Serafini, Agnese; Damasso, Mario; Margonelli, Andrea; Johanningmeier, Udo; Bertalan, Ivo; Pezzotti, Gianni; Giardi, Maria Teresa

    Long-term space exploration, colonization or habitation requires biological life support systems capable to cope with the deleterious space environment. The use of oxygenic photosynthetic microrganisms is an intriguing possibility mainly for food, O2 and nutraceutical compounds production. The critical points of utilizing plantsor algae-based life support systems are the microgravity and the ionizing radiation, which can influence the performance of these organisms. The aim of the present study was to assess the effects of space environment on the photosynthetic activity of various microrganisms and to select space stress-tolerant strains. Site-directed and random mutants of the unicellular green alga Chlamydomonas reinhardtii of Photosystem II D1 protein were used as a model system to test and select the amino acid substitutions capable to account for space stress tolerance. We focussed our studies also on the accumulation of the Photosystem II photoprotective carotenoids (the xantophylls violaxanthin, anteraxanthin and zeaxanthin), powerful antioxidants that epidemiological studies demonstrated to be human vision protectors. Metabolite profiling by quantitative HPLC methods revealed the organisms and the stress conditions capable to accumulate the highest pigment levels. In order to develop a project for a rationale metabolic engineering of algal secondary metabolites overproduction, we are performing expression analyses on the carotenoid biosynthetic pathway under physiological and mimicked space conditions. To identify the consequences of the space environment on the photosynthetic apparatus the changes in the Photosystem II efficiency were monitored in real time during the ESA-Russian Foton-M3 mission in September 2007. For the space flight a high-tech, multicell fluorescence biosensor, Photo-II, was designed and built by the Centre for Advanced Research in Space Optics in collaboration with Kayser-Italy, Biosensor and DAS. Photo-II is an automatic device

  1. Relationships between H{sub 2} photoproduction and different electron transport pathways in sulfur-deprived Chlamydomonas reinhardtii

    Energy Technology Data Exchange (ETDEWEB)

    Antal, Taras K.; Volgusheva, Alena A.; Kukarskih, Galina P.; Krendeleva, Tatyana E.; Rubin, Andrej B. [Faculty of Biology, Moscow State University, Vorobyevi Gory 119992, Moscow (Russian Federation)

    2009-11-15

    In this study the relationships between photosystem (PS) II dependent and independent pathways of H{sub 2} photoproduction, cyclic electron transport around PS I, chloro- and mitorespiration, and transmembrane {delta}pH were examined by inhibitor analysis in S deprived Chlamydomonas reinhardtii. The rate of non-photochemical reduction of plastoquinones in photosynthetic membranes was significantly diminished under starvation which may explain the minor contribution of the PS II independent pathway of H{sub 2} photoproduction in starved cells. The suppressive effect of the herbicide 3-(3,4-dichlorophenyl)-1,1-dimethylurea on the long-term H{sub 2} photoproduction was shown to be entirely attributed to the inhibition of electron transport in PS II, whereas non-specific interactions did not take place. Ferredoxin-quinone reductase - dependent cyclic electron transport around PS I slowed down H{sub 2} photoproduction more than two fold. This result was related to the competition between ferredoxin-quinone reductase and hydrogenase for the reduced ferredoxin and to the decrease in transmembrane {delta}pH induced by the cyclic electron flow. The {delta}pH gradient was shown to down regulate the PS II independent pathway of H{sub 2} photoproduction in starved cells. (author)

  2. Origin of pronounced differences in 77 K fluorescence of the green alga Chlamydomonas reinhardtii in state 1 and 2.

    Science.gov (United States)

    Ünlü, Caner; Polukhina, Iryna; van Amerongen, Herbert

    2016-04-01

    In response to changes in the reduction state of the plastoquinone pool in its thylakoid membrane, the green alga Chlamydomonas reinhardtti is performing state transitions: remodelling of its thylakoid membrane leads to a redistribution of excitations over photosystems I and II (PSI and PSII). These transitions are accompanied by marked changes in the 77 K fluorescence spectrum, which form the accepted signature of state transitions. The changes are generally thought to reflect a redistribution of light-harvesting complexes (LHCs) over PSII (fluorescing below 700 nm) and PSI (fluorescing above 700 nm). Here we studied the picosecond fluorescence properties of C. reinhardtti over a broad range of wavelengths with very low excitation intensities (0.2 nJ per laser pulse). Cells were directly used for time-resolved fluorescence measurements at 77 K without further treatment, such as medium exchange with glycerol. It is observed that upon going from state 1 (relatively more fluorescence below 700 nm) to state 2 (relatively more fluorescence above 700 nm), a large part of the fluorescence of LHC/PSII becomes substantially quenched in concurrence with LHC detachment from PSII, whereas the absolute amount of PSI fluorescence hardly changes. These results are in agreement with the recent proposal that the amount of LHC moving from PSII to PSI upon going from state 1 to state 2 is rather limited (Unlu et al. Proc Natl Acad Sci USA 111 (9):3460-3465, 2014).

  3. The chloroplast atpA gene cluster in Chlamydomonas reinhardtii. Functional analysis of a polycistronic transcription unit.

    Science.gov (United States)

    Drapier, D; Suzuki, H; Levy, H; Rimbault, B; Kindle, K L; Stern, D B; Wollman, F A

    1998-06-01

    Most chloroplast genes in vascular plants are organized into polycistronic transcription units, which generate a complex pattern of mono-, di-, and polycistronic transcripts. In contrast, most Chlamydomonas reinhardtii chloroplast transcripts characterized to date have been monocistronic. This paper describes the atpA gene cluster in the C. reinhardtii chloroplast genome, which includes the atpA, psbI, cemA, and atpH genes, encoding the alpha-subunit of the coupling-factor-1 (CF1) ATP synthase, a small photosystem II polypeptide, a chloroplast envelope membrane protein, and subunit III of the CF0 ATP synthase, respectively. We show that promoters precede the atpA, psbI, and atpH genes, but not the cemA gene, and that cemA mRNA is present only as part of di-, tri-, or tetracistronic transcripts. Deletions introduced into the gene cluster reveal, first, that CF1-alpha can be translated from di- or polycistronic transcripts, and, second, that substantial reductions in mRNA quantity have minimal effects on protein synthesis rates. We suggest that posttranscriptional mRNA processing is common in C. reinhardtii chloroplasts, permitting the expression of multiple genes from a single promoter.

  4. Mastoparan-Induced Cell Death Signalling in Chlamydomonas Reinhardtii

    NARCIS (Netherlands)

    Yordanova, Z.P.; Kapchina-Toteva, V.M.; Woltering, E.J.; Cristescu, S.M.; Harren, F.J.M.; Yakimova, E.T.

    2009-01-01

    The present study was focused on the elucidation of stress-induced cell death signaling events in the unicellular alga Chlamydomonas reinhardtii exposed to treatment with wasp venom mastoparan. By applying pharmacological approach with specific inhibitors, we have investigated the involvement of eth

  5. On the ultrafast kinetics of the energy and electron transfer reactions in photosystem I

    Energy Technology Data Exchange (ETDEWEB)

    Slavov, Chavdar Lyubomirov

    2009-07-09

    The subject of the current work is one of the main participants in the light-dependent phase of oxygenic photosynthesis, Photosystem I (PS I). This complex carries an immense number of cofactors: chlorophylls (Chl), carotenoids, quinones, etc, which together with the protein entity exhibit several exceptional properties. First, PS I has an ultrafast light energy trapping kinetics with a nearly 100% quantum efficiency. Secondly, both of the electron transfer branches in the reaction center are suggested to be active. Thirdly, there are some so called 'red' Chls in the antenna system of PS I, absorbing light with longer wavelengths than the reaction center. These 'red' Chls significantly modify the trapping kinetics of PS I. The purpose of this thesis is to obtain better understanding of the above-mentioned, specific features of PS I. This will not merely cast more light on the mechanisms of energy and electron transfer in the complex, but also will contribute to the future developments of optimized artificial light-harvesting systems. In the current work, a number of PS I complexes isolated from different organisms (Thermosynechococcus elongatus, Chlamydomonas reinhardtii, Arabidopsis thaliana) and possessing distinctive features (different macroorganisation, monomers, trimers, monomers with a semibelt of peripheral antenna attached; presence of 'red' Chls) is investigated. The studies are primarily focused on the electron transfer kinetics in each of the cofactor branches in the PS I reaction center, as well as on the effect of the antenna size and the presence of 'red' Chls on the trapping kinetics of PS I. These aspects are explored with the help of several ultrafast optical spectroscopy methods: (i) time-resolved fluorescence ? single photon counting and synchroscan streak camera; and (ii) ultrafast transient absorption. Physically meaningful information about the molecular mechanisms of the energy trapping in PS I is

  6. Interruption of the Calvin cycle inhibits the repair of Photosystem II from photodamage.

    Science.gov (United States)

    Takahashi, Shunichi; Murata, Norio

    2005-07-15

    In photosynthetic organisms, impairment of the activities of enzymes in the Calvin cycle enhances the extent of photoinactivation of Photosystem II (PSII). We investigated the molecular mechanism responsible for this phenomenon in the unicellular green alga Chlamydomonas reinhardtii. When the Calvin cycle was interrupted by glycolaldehyde, which is known to inhibit phosphoribulokinase, the extent of photoinactivation of PSII was enhanced. The effect of glycolaldehyde was very similar to that of chloramphenicol, which inhibits protein synthesis de novo in chloroplasts. The interruption of the Calvin cycle by the introduction of a missense mutation into the gene for the large subunit of ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) also enhanced the extent of photoinactivation of PSII. In such mutant 10-6C cells, neither glycolaldehyde nor chloramphenicol has any additional effect on photoinactivation. When wild-type cells were incubated under weak light after photodamage to PSII, the activity of PSII recovered gradually and reached a level close to the initial level. However, recovery was inhibited in wild-type cells by glycolaldehyde and was also inhibited in 10-6C cells. Radioactive labelling and Northern blotting demonstrated that the interruption of the Calvin cycle suppressed the synthesis de novo of chloroplast proteins, such as the D1 and D2 proteins, but did not affect the levels of psbA and psbD mRNAs. Our results suggest that the photoinactivation of PSII that is associated with the interruption of the Calvin cycle is attributable primarily to the inhibition of the protein synthesis-dependent repair of PSII at the level of translation in chloroplasts.

  7. Spectral hole burning studies of photosystem II

    Energy Technology Data Exchange (ETDEWEB)

    Chang, H.C.

    1995-11-01

    Low temperature absorption and hole burning spectroscopies were applied to the D1-D2-cyt b{sub 559} and the CP47 and CP43 antenna protein complexes of Photosystem H from higher plants. Low temperature transient and persistent hole-burning data and theoretical calculations on the kinetics and temperature dependence of the P680 hole profile are presented and provide convincing support for the linker model. Implicit in the linker model is that the 684-nm-absorbing Chl a serve to shuttle energy from the proximal antenna complex to reaction center. The stoichiometry of isolated Photosystem H Reaction Center (PSII RC) in several different preparations is also discussed. The additional Chl a are due to 684-nm-absorbing Chl a, some contamination by the CP47 complex, and non-native Chl a absorbing near 670 nm. In the CP47 protein complex, attention is focused on the lower energy chlorophyll a Q{sub y}-states. High pressure hole-burning studies of PSII RC revealed for the first time a strong pressure effect on the primary electron transfer dynamics. The 4.2 K lifetime of P680*, the primary donor state, increases from 2.0 ps to 7.0 ps as pressure increases from 0.1 to 267 MPa. Importantly, this effect is irreversible (plastic) while the pressure induced effect on the low temperature absorption and non-line narrowed P680 hole spectra are reversible (elastic). Nonadiabatic rate expressions, which take into account the distribution of energy gap values, are used to estimate the linear pressure shift of the acceptor state energy for both the superexchange and two-step mechanisms for primary charge separation. It was found that the pressure dependence could be explained with a linear pressure shift of {approximately} 1 cm{sup -1}/MPa in magnitude for the acceptor state. The results point to the marriage of hole burning and high pressures as having considerable potential for the study of primary transport dynamics in reaction centers and antenna complexes.

  8. Interaction of ascorbate with photosystem I.

    Science.gov (United States)

    Trubitsin, Boris V; Mamedov, Mahir D; Semenov, Alexey Yu; Tikhonov, Alexander N

    2014-11-01

    Ascorbate is one of the key participants of the antioxidant defense in plants. In this work, we have investigated the interaction of ascorbate with the chloroplast electron transport chain and isolated photosystem I (PSI), using the EPR method for monitoring the oxidized centers [Formula: see text] and ascorbate free radicals. Inhibitor analysis of the light-induced redox transients of P700 in spinach thylakoids has demonstrated that ascorbate efficiently donates electrons to [Formula: see text] via plastocyanin. Inhibitors (DCMU and stigmatellin), which block electron transport between photosystem II and Pc, did not disturb the ascorbate capacity for electron donation to [Formula: see text]. Otherwise, inactivation of Pc with CN(-) ions inhibited electron flow from ascorbate to [Formula: see text]. This proves that the main route of electron flow from ascorbate to [Formula: see text] runs through Pc, bypassing the plastoquinone (PQ) pool and the cytochrome b 6 f complex. In contrast to Pc-mediated pathway, direct donation of electrons from ascorbate to [Formula: see text] is a rather slow process. Oxidized ascorbate species act as alternative oxidants for PSI, which intercept electrons directly from the terminal electron acceptors of PSI, thereby stimulating photooxidation of P700. We investigated the interaction of ascorbate with PSI complexes isolated from the wild type cells and the MenB deletion strain of cyanobacterium Synechocystis sp. PCC 6803. In the MenB mutant, PSI contains PQ in the quinone-binding A1-site, which can be substituted by high-potential electron carrier 2,3-dichloro-1,4-naphthoquinone (Cl2NQ). In PSI from the MenB mutant with Cl2NQ in the A1-site, the outflow of electrons from PSI is impeded due to the uphill electron transfer from A1 to the iron-sulfur cluster FX and further to the terminal clusters FA/FB, which manifests itself as a decrease in a steady-state level of [Formula: see text]. The addition of ascorbate promoted photooxidation

  9. Proton transfer pathways in Photosystem II

    Science.gov (United States)

    Ishikita, Hiroshi

    2014-03-01

    Using quantum mechanics/molecular mechanics calculations and the 1.9-Å crystal structure of Photosystem II (Umena, Y., Kawakami, K., Shen, J.-R., and Kamiya, N. (2011) Nature 473, 55-60), we investigated the H-bonding environment of the redox active tyrosine, TyrD and obtained insights that help explain its slow redox kinetics and the stability of TyrD radical. The water molecule distal to TyrD, 4 Å away from the phenolic O of TyrD (OTyrD) , corresponds to the presence of the tyrosyl radical state. The water molecule proximal to TyrD, in H-bonding distance to OTyrD, corresponds to the presence of the unoxidised tyrosine. The H+ released upon oxidation of TyrD is transferred to the proximal water, which shifts to the distal position, triggering a concerted proton transfer pathway involving D2-Arg180 and a series of waters, through which the proton reaches the aqueous phase at D2-His61. The water movement linked to the ejection of the proton from the hydrophobic environment near TyrD makes oxidation slow and quasi-irreversible, explaining the great stability of the TyrD radical. A symmetry-related proton pathway associated with TyrZ is pointed out and this is associated with one of the Cl- sites. This may represent a proton pathway functional in the water oxidation cycle.

  10. Multistep organic synthesis of modular photosystems

    Science.gov (United States)

    2012-01-01

    Summary Quite extensive synthetic achievements vanish in the online supporting information of publications on functional systems. Underappreciated, their value is recognized by experts only. As an example, we here focus in on the recent synthesis of multicomponent photosystems with antiparallel charge-transfer cascades in co-axial hole- and electron-transporting channels. The synthetic steps are described one-by-one, starting with commercial starting materials and moving on to key intermediates, such as asparagusic acid, an intriguing natural product, as well as diphosphonate “feet”, and panchromatic naphthalenediimides (NDIs), to finally reach the target molecules. These products are initiators and propagators for self-organizing surface-initiated polymerization (SOSIP), a new method introduced to secure facile access to complex architectures. Chemoorthogonal to the ring-opening disulfide exchange used for SOSIP, hydrazone exchange is then introduced to achieve stack exchange, which is a “switching” technology invented to drill giant holes into SOSIP architectures and fill them with functional π-stacks of free choice. PMID:23015840

  11. Reverse genetics in Chlamydomonas: a platform for isolating insertional mutants

    OpenAIRE

    de Montaigu Amaury; Magneschi Leonardo; Catalanotti Claudia; Yang Wenqiang; Mus Florence; Pootakham Wirulda; Gonzalez-Ballester David; Higuera Jose J; Prior Matthew; Galván Aurora; Fernandez Emilio; Grossman Arthur R

    2011-01-01

    Abstract A method was developed to identify insertional mutants of Chlamydomonas reinhardtii disrupted for selected target genes. The approach relies on the generation of thousands of transformants followed by PCR-based screenings that allow for identification of strains harboring the introduced marker gene within specific genes of interest. Our results highlight the strengths and limitations of two independent screens that differed in the nature of the marker DNA used (PCR-amplified fragment...

  12. An optimized, chemically regulated gene expression system for Chlamydomonas.

    Directory of Open Access Journals (Sweden)

    Paola Ferrante

    Full Text Available BACKGROUND: Chlamydomonas reinhardtii is a model system for algal and cell biology and is used for biotechnological applications, such as molecular farming or biological hydrogen production. The Chlamydomonas metal-responsive CYC6 promoter is repressed by copper and induced by nickel ions. However, induction by nickel is weak in some strains, poorly reversible by chelating agents like EDTA, and causes, at high concentrations, toxicity side effects on Chlamydomonas growth. Removal of these bottlenecks will encourage the wide use of this promoter as a chemically regulated gene expression system. METHODOLOGY: Using a codon-optimized Renilla luciferase as a reporter gene, we explored several strategies to improve the strength and reversibility of CYC6 promoter induction. Use of the first intron of the RBCS2 gene or of a modified TAP medium increases the strength of CYC6 induction up to 20-fold. In the modified medium, induction is also obtained after addition of specific copper chelators, like TETA. At low concentrations (up to 10 microM TETA is a more efficient inducer than Ni, which becomes a very efficient inducer at higher concentrations (50 microM. Neither TETA nor Ni show toxicity effects at the concentrations used. Unlike induction by Ni, induction by TETA is completely reversible by micromolar copper concentrations, thus resulting in a transient "wave" in luciferase activity, which can be repeated in subsequent growth cycles. CONCLUSIONS: We have worked out a chemically regulated gene expression system that can be finely tuned to produce temporally controlled "waves" in gene expression. The use of cassettes containing the CYC6 promoter, and of modified growth media, is a reliable and economically sustainable system for the temporally controlled expression of foreign genes in Chlamydomonas.

  13. Actin is required for IFT regulation in Chlamydomonas reinhardtii.

    Science.gov (United States)

    Avasthi, Prachee; Onishi, Masayuki; Karpiak, Joel; Yamamoto, Ryosuke; Mackinder, Luke; Jonikas, Martin C; Sale, Winfield S; Shoichet, Brian; Pringle, John R; Marshall, Wallace F

    2014-09-01

    Assembly of cilia and flagella requires intraflagellar transport (IFT), a highly regulated kinesin-based transport system that moves cargo from the basal body to the tip of flagella [1]. The recruitment of IFT components to basal bodies is a function of flagellar length, with increased recruitment in rapidly growing short flagella [2]. The molecular pathways regulating IFT are largely a mystery. Because actin network disruption leads to changes in ciliary length and number, actin has been proposed to have a role in ciliary assembly. However, the mechanisms involved are unknown. In Chlamydomonas reinhardtii, conventional actin is found in both the cell body and the inner dynein arm complexes within flagella [3, 4]. Previous work showed that treating Chlamydomonas cells with the actin-depolymerizing compound cytochalasin D resulted in reversible flagellar shortening [5], but how actin is related to flagellar length or assembly remains unknown. Here we utilize small-molecule inhibitors and genetic mutants to analyze the role of actin dynamics in flagellar assembly in Chlamydomonas reinhardtii. We demonstrate that actin plays a role in IFT recruitment to basal bodies during flagellar elongation and that when actin is perturbed, the normal dependence of IFT recruitment on flagellar length is lost. We also find that actin is required for sufficient entry of IFT material into flagella during assembly. These same effects are recapitulated with a myosin inhibitor, suggesting that actin may act via myosin in a pathway by which flagellar assembly is regulated by flagellar length.

  14. Regulation by glutathionylation of isocitrate lyase from Chlamydomonas reinhardtii.

    Science.gov (United States)

    Bedhomme, Mariette; Zaffagnini, Mirko; Marchand, Christophe H; Gao, Xing-Huang; Moslonka-Lefebvre, Mathieu; Michelet, Laure; Decottignies, Paulette; Lemaire, Stéphane D

    2009-12-25

    Post-translational modification of protein cysteine residues is emerging as an important regulatory and signaling mechanism. We have identified numerous putative targets of redox regulation in the unicellular green alga Chlamydomonas reinhardtii. One enzyme, isocitrate lyase (ICL), was identified both as a putative thioredoxin target and as an S-thiolated protein in vivo. ICL is a key enzyme of the glyoxylate cycle that allows growth on acetate as a sole source of carbon. The aim of the present study was to clarify the molecular mechanism of the redox regulation of Chlamydomonas ICL using a combination of biochemical and biophysical methods. The results clearly show that purified C. reinhardtii ICL can be inactivated by glutathionylation and reactivated by glutaredoxin, whereas thioredoxin does not appear to regulate ICL activity, and no inter- or intramolecular disulfide bond could be formed under any of the conditions tested. Glutathionylation of the protein was investigated by mass spectrometry analysis, Western blotting, and site-directed mutagenesis. The enzyme was found to be protected from irreversible oxidative inactivation by glutathionylation of its catalytic Cys(178), whereas a second residue, Cys(247), becomes artifactually glutathionylated after prolonged incubation with GSSG. The possible functional significance of this post-translational modification of ICL in Chlamydomonas and other organisms is discussed.

  15. Observation of Coherence in the Photosystem II Reaction Center

    CERN Document Server

    Fuller, Franklin D; Senlik, S Seckin; Wilcox, Daniel E; Ogilvie, Jennifer P

    2013-01-01

    Photosynthesis powers life on our planet. The basic photosynthetic architecture comprises antenna complexes to harvest solar energy and reaction centers to convert the energy into a stable charge separated state. In oxygenic photosynthesis, the initial charge separation event occurs in the photosystem II reaction center; the only known natural enzyme that uses solar energy to split water. Energy transfer and charge separation in photosynthesis are rapid and have high quantum efficiencies. Recently, nonlinear spectroscopic experiments have suggested that electronic coherence may play a role in energy transfer efficiency in antenna complexes. Here we report the observation of coherence in the photosystem II reaction center by two dimensional electronic spectroscopy. The frequencies of the observed coherences match exciton difference frequencies and/or known vibrational modes of the photosystem II reaction center. These observations raise questions about the possible role of electronic and/or vibrational coheren...

  16. Representative Pathways of Excitation Migration in Photosystem I

    CERN Document Server

    Park, S; Lu, D; Schulten, K; Park, Sanghyun; Sener, Melih K.; Lu, Deyu; Schulten, Klaus

    2002-01-01

    Photosystem I is a protein-pigment complex that performs photosynthesis in plants, green algae, and cyanobacteria. It contains an aggregate of chlorophylls that absorbs light and delivers the resulting electronic excitation to the special pair of chlorophylls where the excitation energy is used for producing charge separation across the cell membrane. The seemingly random arrangement of chlorophylls in photosystem I poses the question which pathways excitation migration follows towards the special pair after absorption of light at any of its chlorophylls. We employ a master equation to describe the process of excitation migration in photosystem I, and find representative paths of excitation migration based on the mean first-passage time from each chlorophyll to the special pair. The resulting paths, beginning at each of the chlorophylls and ending at the special pair, provide a complete yet distilled picture of the excitation migration towards the special pair.

  17. Three Acyltransferases and Nitrogen-responsive Regulator Are Implicated in Nitrogen Starvation-induced Triacylglycerol Accumulation in Chlamydomonas*

    Science.gov (United States)

    Boyle, Nanette R.; Page, Mark Dudley; Liu, Bensheng; Blaby, Ian K.; Casero, David; Kropat, Janette; Cokus, Shawn J.; Hong-Hermesdorf, Anne; Shaw, Johnathan; Karpowicz, Steven J.; Gallaher, Sean D.; Johnson, Shannon; Benning, Christoph; Pellegrini, Matteo; Grossman, Arthur; Merchant, Sabeeha S.

    2012-01-01

    Algae have recently gained attention as a potential source for biodiesel; however, much is still unknown about the biological triggers that cause the production of triacylglycerols. We used RNA-Seq as a tool for discovering genes responsible for triacylglycerol (TAG) production in Chlamydomonas and for the regulatory components that activate the pathway. Three genes encoding acyltransferases, DGAT1, DGTT1, and PDAT1, are induced by nitrogen starvation and are likely to have a role in TAG accumulation based on their patterns of expression. DGAT1 and DGTT1 also show increased mRNA abundance in other TAG-accumulating conditions (minus sulfur, minus phosphorus, minus zinc, and minus iron). Insertional mutants, pdat1-1 and pdat1-2, accumulate 25% less TAG compared with the parent strain, CC-4425, which demonstrates the relevance of the trans-acylation pathway in Chlamydomonas. The biochemical functions of DGTT1 and PDAT1 were validated by rescue of oleic acid sensitivity and restoration of TAG accumulation in a yeast strain lacking all acyltransferase activity. Time course analyses suggest than a SQUAMOSA promoter-binding protein domain transcription factor, whose mRNA increases precede that of lipid biosynthesis genes like DGAT1, is a candidate regulator of the nitrogen deficiency responses. An insertional mutant, nrr1-1, accumulates only 50% of the TAG compared with the parental strain in nitrogen-starvation conditions and is unaffected by other nutrient stresses, suggesting the specificity of this regulator for nitrogen-deprivation conditions. PMID:22403401

  18. Assembly of the light-harvesting chlorophyll antenna in the green alga Chlamydomonas reinhardtii requires expression of the TLA2-CpFTSY gene.

    Science.gov (United States)

    Kirst, Henning; García-Cerdán, Jose Gines; Zurbriggen, Andreas; Melis, Anastasios

    2012-02-01

    The truncated light-harvesting antenna2 (tla2) mutant of Chlamydomonas reinhardtii showed a lighter-green phenotype, had a lower chlorophyll (Chl) per-cell content, and higher Chl a/b ratio than corresponding wild-type strains. Physiological analyses revealed a higher intensity for the saturation of photosynthesis and greater P(max) values in the tla2 mutant than in the wild type. Biochemical analyses showed that the tla2 strain was deficient in the Chl a-b light-harvesting complex, and had a Chl antenna size of the photosystems that was only about 65% of that in the wild type. Molecular and genetic analyses showed a single plasmid insertion in the tla2 strain, causing a chromosomal DNA rearrangement and deletion/disruption of five nuclear genes. The TLA2 gene, causing the tla2 phenotype, was cloned by mapping the insertion site and upon complementation with each of the genes that were deleted. Successful complementation was achieved with the C. reinhardtii TLA2-CpFTSY gene, whose occurrence and function in green microalgae has not hitherto been investigated. Functional analysis showed that the nuclear-encoded and chloroplast-localized CrCpFTSY protein specifically operates in the assembly of the peripheral components of the Chl a-b light-harvesting antenna. In higher plants, a cpftsy null mutation inhibits assembly of both the light-harvesting complex and photosystem complexes, thus resulting in a seedling-lethal phenotype. The work shows that cpftsy deletion in green algae, but not in higher plants, can be employed to generate tla mutants. The latter exhibit improved solar energy conversion efficiency and photosynthetic productivity under mass culture and bright sunlight conditions.

  19. Role of bicarbonate at the acceptor side of photosystem II

    NARCIS (Netherlands)

    Rensen, van J.J.S.

    2002-01-01

    Besides being the substrate for the carboxylation reaction of photosynthesis, CO2 (bicarbonate) is required for the activity of Photosystem II (water plastoquinone oxido-reductase). It plays a role on the electron donor side as well as the electron acceptor side. In this contribution, attention will

  20. Functional properties of the oxygen evolving complex of photosystem II.

    NARCIS (Netherlands)

    Vliet, van P.H.

    1996-01-01

    This Thesis presents the results of a study by electron paramagnetic resonance (EPR) and measurements of oxygen evolution of the Oxygen Evolving Complex of Photosystem 11 (PS-II) in PS-II enriched membranes from spinach.The experimental part of this Thesis is preceded by a general introduction (Chap

  1. Photosystem I-​based Biophotovoltaics on Nanostructured Hematite

    NARCIS (Netherlands)

    Ocakoglu, K.; Krupnik, T.; van den Bosch, B.; Harputlu, E.; Gullo, M.P.; Olmos, J.D.J.; Yildirimcan, S.; Gupta, R.K.; Yakuphanoglu, F.; Barbieri, A.; Reek, J.N.H.; Kargul, J.

    2014-01-01

    The electronic coupling between a robust red algal photosystem I (PSI) associated with its light harvesting antenna (LHCI) and nanocrystalline n-​type semiconductors, TiO2 and hematite (α-​Fe2O3) is utilized for fabrication of the biohybrid dye-​sensitized solar cells (DSSC)​. PSI-​LHCI is immobiliz

  2. Recent progress in the crystallographic studies of photosystem II

    NARCIS (Netherlands)

    Guskov, Albert; Gabdulkhakov, Azat; Broser, Matthias; Glöckner, Carina; Hellmich, Julia; Kern, Jan; Frank, Joachim; Müh, Frank; Saenger, Wolfram; Zouni, Athina

    2010-01-01

    The photosynthetic oxygen-evolving photosystem II (PSII) is the only known biochemical system that is able to oxidize water molecules and thereby generates almost all oxygen in the Earth's atmosphere. The elucidation of the structural and mechanistic aspects of PSII keeps scientists all over the wor

  3. Photosystem I from the unusual cyanobacterium Gloeobacter violaceus

    NARCIS (Netherlands)

    Mangels, D.; Kruip, J.; Berry, S.; Rögner, M.; Boekema, E.J.; Koenig, F.

    2002-01-01

    Photosystem I (PS I) from the primitive cyanobacterium Gloeobacter violaceus has been purified and characterised. Despite the fact that the isolated complexes have the same subunit composition as complexes from other cyanobacteria, the amplitude of flash-induced absorption difference spectra indicat

  4. PMS : Photosystem I electron donor or fluorescence quencher

    NARCIS (Netherlands)

    Wientjes, Emilie; Croce, Roberta

    2012-01-01

    Light energy harvested by the pigments in Photosystem I (PSI) is used for charge separation in the reaction center (RC), after which the positive charge resides on a special chlorophyll dimer called P700. In studies on the PSI trapping kinetics, P700(+) is usually chemically reduced to re-open the R

  5. Multiple-endpoint assay provides a detailed mechanistic view of responses to herbicide exposure in Chlamydomonas reinhardtii

    Energy Technology Data Exchange (ETDEWEB)

    Nestler, Holger [Eawag, Swiss Federal Institute of Aquatic Science and Technology, Ueberlandstrasse 133, 8600 Duebendorf (Switzerland); ETH Zurich, Swiss Federal Institute of Technology, Institute of Biogeochemistry and Pollutant Dynamics, Universitaetstrasse 16, 8092 Zurich (Switzerland); Groh, Ksenia J.; Schoenenberger, Rene; Behra, Renata [Eawag, Swiss Federal Institute of Aquatic Science and Technology, Ueberlandstrasse 133, 8600 Duebendorf (Switzerland); Schirmer, Kristin [Eawag, Swiss Federal Institute of Aquatic Science and Technology, Ueberlandstrasse 133, 8600 Duebendorf (Switzerland); ETH Zurich, Swiss Federal Institute of Technology, Institute of Biogeochemistry and Pollutant Dynamics, Universitaetstrasse 16, 8092 Zurich (Switzerland); EPF Lausanne, School of Architecture, Civil and Environmental Engineering, 1015 Lausanne (Switzerland); Eggen, Rik I.L. [Eawag, Swiss Federal Institute of Aquatic Science and Technology, Ueberlandstrasse 133, 8600 Duebendorf (Switzerland); ETH Zurich, Swiss Federal Institute of Technology, Institute of Biogeochemistry and Pollutant Dynamics, Universitaetstrasse 16, 8092 Zurich (Switzerland); Suter, Marc J.-F., E-mail: suter@eawag.ch [Eawag, Swiss Federal Institute of Aquatic Science and Technology, Ueberlandstrasse 133, 8600 Duebendorf (Switzerland); ETH Zurich, Swiss Federal Institute of Technology, Institute of Biogeochemistry and Pollutant Dynamics, Universitaetstrasse 16, 8092 Zurich (Switzerland)

    2012-04-15

    The release of herbicides into the aquatic environment raises concerns about potential detrimental effects on ecologically important non-target species, such as unicellular algae, necessitating ecotoxicological risk assessment. Algal toxicity tests based on growth, a commonly assessed endpoint, are integrative, and hence do not provide information about underlying toxic mechanisms and effects. This limitation may be overcome by measuring more specific biochemical and physiological endpoints. In the present work, we developed and applied a novel multiple-endpoint assay, and analyzed the effects of the herbicides paraquat, diuron and norflurazon, each representing a specific mechanism of toxic action, on the single celled green alga Chlamydomonas reinhardtii. The endpoints added to assessment of growth were pigment content, maximum and effective photosystem II quantum yield, ATP content, esterase and oxidative activity. All parameters were measured at 2, 6 and 24 h of exposure, except for growth and pigment content, which were determined after 6 and 24 h only. Effective concentrations causing 50% of response (EC50s) and lowest observable effect concentrations (LOECs) were determined for all endpoints and exposure durations where possible. The assay provided a detailed picture of the concentration- and time-dependent development of effects elicited by the analyzed herbicides, thus improving the understanding of the underlying toxic mechanisms. Furthermore, the response patterns were unique to the respective herbicide and reflected the different mechanisms of toxicity. The comparison of the endpoint responses and sensitivities revealed that several physiological and biochemical parameters reacted earlier or stronger to disturbances than growth. Overall, the presented multiple-endpoint assay constitutes a promising basis for investigating stressor and toxicant effects in green algae.

  6. Mutagenesis and phenotypic selection as a strategy toward domestication of Chlamydomonas reinhardtii strains for improved performance in photobioreactors.

    Science.gov (United States)

    Bonente, Giulia; Formighieri, Cinzia; Mantelli, Manuela; Catalanotti, Claudia; Giuliano, Giovanni; Morosinotto, Tomas; Bassi, Roberto

    2011-09-01

    Microalgae have a valuable potential for biofuels production. As a matter of fact, algae can produce different molecules with high energy content, including molecular hydrogen (H(2)) by the activity of a chloroplastic hydrogenase fueled by reducing power derived from water and light energy. The efficiency of this reaction, however, is limited and depends from an intricate relationships between oxygenic photosynthesis and mitochondrial respiration. The way toward obtaining algal strains with high productivity in photobioreactors requires engineering of their metabolism at multiple levels in a process comparable to domestication of crops that were derived from their wild ancestors through accumulation of genetic traits providing improved productivity under conditions of intensive cultivation as well as improved nutritional/industrial properties. This holds true for the production of any biofuels from algae: there is the need to isolate multiple traits to be combined and produce organisms with increased performances. Among the different limitations in H(2) productivity, we identified three with a major relevance, namely: (i) the light distribution through the mass culture; (ii) the strong sensitivity of the hydrogenase to even very low oxygen concentrations; and (iii) the presence of alternative pathways, such as the cyclic electron transport, competing for reducing equivalents with hydrogenase and H(2) production. In order to identify potentially favorable mutations, we generated a collection of random mutants in Chlamydomonas reinhardtii which were selected through phenotype analysis for: (i) a reduced photosynthetic antenna size, and thus a lower culture optical density; (ii) an altered photosystem II activity as a tool to manipulate the oxygen concentration within the culture; and (iii) State 1-State 2 transition mutants, for a reduced cyclic electron flow and maximized electrons flow toward the hydrogenase. Such a broad approach has been possible thanks to the

  7. Efficiency of energy funneling in the photosystem II supercomplex of higher plants

    CERN Document Server

    Kreisbeck, Christoph

    2015-01-01

    The investigation of energy transfer properties in photosynthetic multi-protein networks gives insight into their underlying design principles.Here, we discuss excitonic energy transfer mechanisms of the photosystem II (PS-II) C$_2$S$_2$M$_2$ supercomplex, which is the largest isolated functional unit of the photosynthetic apparatus of higher plants.Despite the lack of a decisive energy gradient in C$_2$S$_2$M$_2$, we show that the energy transfer is directed by relaxation to low energy states. C$_2$S$_2$M$_2$ is not organized to form pathways with strict energetic downhill transfer, which has direct consequences on the transfer efficiency, transfer pathways and transfer limiting steps. The exciton dynamics is sensitive to small structural changes, which, for instance, are induced by the reorganization of vibrational coordinates. In order to incorporate the reorganization process in our numerical simulations, we go beyond rate equations and use the hierarchically coupled equation of motion approach (HEOM). Wh...

  8. Synthesizing and salvaging NAD: lessons learned from Chlamydomonas reinhardtii.

    Directory of Open Access Journals (Sweden)

    Huawen Lin

    2010-09-01

    Full Text Available The essential coenzyme nicotinamide adenine dinucleotide (NAD+ plays important roles in metabolic reactions and cell regulation in all organisms. Bacteria, fungi, plants, and animals use different pathways to synthesize NAD+. Our molecular and genetic data demonstrate that in the unicellular green alga Chlamydomonas NAD+ is synthesized from aspartate (de novo synthesis, as in plants, or nicotinamide, as in mammals (salvage synthesis. The de novo pathway requires five different enzymes: L-aspartate oxidase (ASO, quinolinate synthetase (QS, quinolate phosphoribosyltransferase (QPT, nicotinate/nicotinamide mononucleotide adenylyltransferase (NMNAT, and NAD+ synthetase (NS. Sequence similarity searches, gene isolation and sequencing of mutant loci indicate that mutations in each enzyme result in a nicotinamide-requiring mutant phenotype in the previously isolated nic mutants. We rescued the mutant phenotype by the introduction of BAC DNA (nic2-1 and nic13-1 or plasmids with cloned genes (nic1-1 and nic15-1 into the mutants. NMNAT, which is also in the de novo pathway, and nicotinamide phosphoribosyltransferase (NAMPT constitute the nicotinamide-dependent salvage pathway. A mutation in NAMPT (npt1-1 has no obvious growth defect and is not nicotinamide-dependent. However, double mutant strains with the npt1-1 mutation and any of the nic mutations are inviable. When the de novo pathway is inactive, the salvage pathway is essential to Chlamydomonas for the synthesis of NAD+. A homolog of the human SIRT6-like gene, SRT2, is upregulated in the NS mutant, which shows a longer vegetative life span than wild-type cells. Our results suggest that Chlamydomonas is an excellent model system to study NAD+ metabolism and cell longevity.

  9. Individual Flagellar Waveform Affects Collective Behavior of Chlamydomonas reinhardtii.

    Science.gov (United States)

    Kage, Azusa; Mogami, Yoshihiro

    2015-08-01

    Bioconvection is a form of collective motion that occurs spontaneously in the suspension of swimming microorganisms. In a previous study, we quantitatively described the "pattern transition," a phase transition phenomenon that so far has exclusively been observed in bioconvection of the unicellular green alga Chlamydomonas. We suggested that the transition could be induced by changes in the balance between the gravitational and shear-induced torques, both of which act to determine the orientation of the organism in the shear flow. As both of the torques should be affected by the geometry of the Chlamydomonas cell, alteration in the flagellar waveform might change the extent of torque generation by altering overall geometry of the cell. Based on this working hypothesis, we examined bioconvection behavior of two flagellar mutants of Chlamydomonas reinhardtii, ida1 and oda2, making reference to the wild type. Flagella of ida1 beat with an abnormal waveform, while flagella of oda2 show a normal waveform but lower beat frequency. As a result, both mutants had swimming speed of less than 50% of the wild type. ida1 formed bioconvection patterns with smaller spacing than those of wild type and oda2. Two-axis view revealed the periodic movement of the settling blobs of ida1, while oda2 showed qualitatively similar behavior to that of wild type. Unexpectedly, ida1 showed stronger negative gravitaxis than did wild type, while oda2 showed relatively weak gravitaxis. These findings suggest that flagellar waveform, not swimming speed or beat frequency, strongly affect bioconvection behavior in C. reinhardtii.

  10. Reverse genetics in Chlamydomonas: a platform for isolating insertional mutants

    Directory of Open Access Journals (Sweden)

    de Montaigu Amaury

    2011-07-01

    Full Text Available Abstract A method was developed to identify insertional mutants of Chlamydomonas reinhardtii disrupted for selected target genes. The approach relies on the generation of thousands of transformants followed by PCR-based screenings that allow for identification of strains harboring the introduced marker gene within specific genes of interest. Our results highlight the strengths and limitations of two independent screens that differed in the nature of the marker DNA used (PCR-amplified fragment containing the plasmid-free marker versus entire linearized plasmid with the marker and in the strategies used to maintain and store transformants.

  11. Genome-wide analysis of alternative splicing in Chlamydomonas reinhardtii

    Directory of Open Access Journals (Sweden)

    Thomas Julie

    2010-02-01

    Full Text Available Abstract Background Genome-wide computational analysis of alternative splicing (AS in several flowering plants has revealed that pre-mRNAs from about 30% of genes undergo AS. Chlamydomonas, a simple unicellular green alga, is part of the lineage that includes land plants. However, it diverged from land plants about one billion years ago. Hence, it serves as a good model system to study alternative splicing in early photosynthetic eukaryotes, to obtain insights into the evolution of this process in plants, and to compare splicing in simple unicellular photosynthetic and non-photosynthetic eukaryotes. We performed a global analysis of alternative splicing in Chlamydomonas reinhardtii using its recently completed genome sequence and all available ESTs and cDNAs. Results Our analysis of AS using BLAT and a modified version of the Sircah tool revealed AS of 498 transcriptional units with 611 events, representing about 3% of the total number of genes. As in land plants, intron retention is the most prevalent form of AS. Retained introns and skipped exons tend to be shorter than their counterparts in constitutively spliced genes. The splice site signals in all types of AS events are weaker than those in constitutively spliced genes. Furthermore, in alternatively spliced genes, the prevalent splice form has a stronger splice site signal than the non-prevalent form. Analysis of constitutively spliced introns revealed an over-abundance of motifs with simple repetitive elements in comparison to introns involved in intron retention. In almost all cases, AS results in a truncated ORF, leading to a coding sequence that is around 50% shorter than the prevalent splice form. Using RT-PCR we verified AS of two genes and show that they produce more isoforms than indicated by EST data. All cDNA/EST alignments and splice graphs are provided in a website at http://combi.cs.colostate.edu/as/chlamy. Conclusions The extent of AS in Chlamydomonas that we observed is much

  12. Artificially acquired chlorophyll b is highly acceptable to the thylakoid-lacking cyanobacterium, Gloeobacter violaceus PCC 7421.

    Science.gov (United States)

    Araki, Mie; Akimoto, Seiji; Mimuro, Mamoru; Tsuchiya, Tohru

    2014-08-01

    Unicellular cyanobacterium Gloeobacter violaceus is an only known oxygenic photosynthetic organism that lacks thylakoid membrane. Molecular phylogenetic analyses indicate that G. violaceus is an early-branching cyanobacterium within cyanobacterial clade. Therefore, the photosynthetic system of G. violaceus is considered to be partly similar to that of the ancestral cyanobacteria that would lack thylakoid membrane. G. violaceus possesses chlorophyll (Chl) a as the only chlorophyll species like most cyanobacteria. It was proposed that the ancestral oxygenic photosynthetic organism had not only Chl a and phycobilins but also Chl b. However, no organism which contains both Chl a and Chl b and lacks thylakoid membrane has been found in nature. Therefore, we introduced the chlorophyllide a oxygenase gene responsible for Chl b biosynthesis into G. violaceus. In the resultant transformant, Chl b accumulated at approximately 11% of total Chl independent of growth phase. Photosystem I complexes isolated from the transformant contained Chl b at 9.9% of total Chl. The presence of Chl b in the photosystem I complexes did not inhibit trimer formation. Furthermore, time-resolved fluorescence spectrum demonstrated that Chl b transferred energy to Chl a in the photosystem I complexes and did not disturb the energy transfer among the Chl a molecules. These results show that G. violaceus is tolerant to artificially produced Chl b and suggest the flexibility of photosystem for Chl composition in the ancestral oxygenic photosynthetic organism.

  13. Structural response of photosystem 2 to iron deficiency : Characterization of a new photosystem 2-IdiA complex from the cyanobacterium Thermosynechococcus elongatus BP-1

    NARCIS (Netherlands)

    Lax, Julia E. -M.; Arteni, Ana A.; Boekema, Egbert J.; Pistorius, Elfriede K.; Michel, Klaus-Peter; Roegner, Matthias

    2007-01-01

    Iron deficiency triggers various processes in cyanobacterial cells of which the synthesis of an additional antenna system (IsiA) around photosystem (PS) 1 is well documented [T.S. Bibby, J. Nield, J. Barber, Iron deficiency induces the formation of an antenna ring around trimeric photosystem I in cy

  14. The microalga Chlamydomonas reinhardtii CW-15 as a solar cell for hydrogen peroxide photoproduction. Comparison between free and immobilized cells and thylakoids for energy conversion efficiency

    Energy Technology Data Exchange (ETDEWEB)

    Scholz, W.; Galvan, F.; Rosa, F.F. de la [Instituto de Bioquimica Vegetal y Fotosintesis, Universidad de Sevilla y CSIC, Sevilla (Spain)

    1995-11-28

    Immobilized cells and thylakoid vesicles of the microalga Chlamydomonas reinhardtii CW-15 have been developed as a solar cell because of their capabilities of producing hydrogen peroxide. This compound is an efficient and clean fuel used for rocket propulsion, motors and for heating. Hydrogen peroxide is produced by the photosystem in a catalyst cycle in which a redox mediator (methyl viologen) is reduced by electrons obtained from water by the photosynthetic apparatus of the microalga and it is re-oxidized by the oxygen dissolved in the solution. The photoproduction has been investigated using a discontinuous system with whole cells, or thylakoid vesicles, free or immobilized on alginate. The stimulation by azide as an inhibitor of catalase has also been analyzed. Under determined optimum conditions, the photoproduction by Ca-alginate entrapped cells, with a rate of 33 {mu}mol H{sub 2}O{sub 2}/mg Chl.h, was maintained for several hours with an energy conversion efficiency of 0.25%

  15. Microoxic Niches within the Thylakoid Stroma of Air-Grown Chlamydomonas reinhardtii Protect [FeFe]-Hydrogenase and Support Hydrogen Production under Fully Aerobic Environment1[OPEN

    Science.gov (United States)

    Liran, Oded; Milrad, Yuval; Eilenberg, Haviva; Weiner, Iddo

    2016-01-01

    Photosynthetic hydrogen production in the microalga Chlamydomonas reinhardtii is catalyzed by two [FeFe]-hydrogenase isoforms, HydA1 and HydA2, both irreversibly inactivated upon a few seconds exposure to atmospheric oxygen. Until recently, it was thought that hydrogenase is not active in air-grown microalgal cells. In contrast, we show that the entire pool of cellular [FeFe]-hydrogenase remains active in air-grown cells due to efficient scavenging of oxygen. Using membrane inlet mass spectrometry, 18O2 isotope, and various inhibitors, we were able to dissect the various oxygen uptake mechanisms. We found that both chlororespiration, catalyzed by plastid terminal oxidase, and Mehler reactions, catalyzed by photosystem I and Flavodiiron proteins, significantly contribute to oxygen uptake rate. This rate is considerably enhanced with increasing light, thus forming local anaerobic niches at the proximity of the stromal face of the thylakoid membrane. Furthermore, we found that in transition to high light, the hydrogen production rate is significantly enhanced for a short duration (100 s), thus indicating that [FeFe]-hydrogenase functions as an immediate sink for surplus electrons in aerobic as well as in anaerobic environments. In summary, we show that an anaerobic locality in the chloroplast preserves [FeFe]-hydrogenase activity and supports continuous hydrogen production in air-grown microalgal cells. PMID:27443604

  16. Light inhibition of mitochondrial respiration in a mutant of Chlamydomonas reinhardtii devoid of ribulose-1,5-bisphosphate carboxylase/oxygenase activity.

    Science.gov (United States)

    Gans, P; Rebeille, F

    1988-01-01

    The effect of light on mitochondrial respiration has been investigated in Chlamydomonas reinhardtii rcl-u-1-10-6C, a mutant devoid of ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) activity. No CO2 uptake was observed in the light, confirming that there was no Rubisco activity, but the CO2 evolution rate was diminished by 65 to 80%. This inhibition was ascribable to a decrease in the tricarboxylic acid cycle (Krebs cycle) activity. At the same time, O2 evolution associated with stimulation of the O2 uptake appears. Darkness or addition of DCMU fully reversed the effect of light, indicating that the inhibitory process is linked to photosystem activities. Levels of pyridine nucleotides (NAD(H) and NADP(H)) and adenine nucleotides (ATP and ADP), the most probable mediators of the interaction between photosynthesis and respiration, were measured in dark and in light. During a dark to light transition the level of NADPH increased significantly whereas the NAD(H) pool remained almost fully oxidized. The level of ADP was always extremely low. These results suggest that the inhibition of Krebs cycle activity is due to a competition for cytosolic ADP between chloroplastic photophosphorylations and oxidative phosphorylations.

  17. Development of the light-harvesting chlorophyll antenna in the green alga Chlamydomonas reinhardtii is regulated by the novel Tla1 gene.

    Science.gov (United States)

    Tetali, Sarada D; Mitra, Mautusi; Melis, Anastasios

    2007-03-01

    The Chlamydomonas reinhardtii tla1 (truncated light-harvesting chlorophyll antenna size) mutant was generated upon DNA insertional mutagenesis and shown to specifically possess a smaller than wild type (WT) chlorophyll antenna size in both photosystems. Molecular and genetic analysis revealed that the exogenous plasmid DNA was inserted at the end of the 5' UTR and just prior to the ATG start codon of a hitherto unknown nuclear gene (termed Tla1), which encodes a protein of 213 amino acids. The Tla1 gene in the mutant is transcribed with a new 5' UTR sequence, derived from the 3' end of the transforming plasmid. This replacement of the native 5' UTR and promoter regions resulted in enhanced transcription of the tla1 gene in the mutant but inhibition in the translation of the respective tla1 mRNA. Transformation of the tla1 mutant with WT Tla1 genomic DNA successfully rescued the mutant. These results are evidence that polymorphism in the 5' UTR of the Tla1 transcripts resulted in the tla1 phenotype and that expression of the Tla1 gene is a prerequisite for the development/assembly of the Chl antenna in C. reinhardtii. A blast search with the Tla1 deduced amino acid sequence

  18. A role for the membrane in regulating Chlamydomonas flagellar length.

    Directory of Open Access Journals (Sweden)

    William Dentler

    Full Text Available Flagellar assembly requires coordination between the assembly of axonemal proteins and the assembly of the flagellar membrane and membrane proteins. Fully grown steady-state Chlamydomonas flagella release flagellar vesicles from their tips and failure to resupply membrane should affect flagellar length. To study vesicle release, plasma and flagellar membrane surface proteins were vectorially pulse-labeled and flagella and vesicles were analyzed for biotinylated proteins. Based on the quantity of biotinylated proteins in purified vesicles, steady-state flagella appeared to shed a minimum of 16% of their surface membrane per hour, equivalent to a complete flagellar membrane being released every 6 hrs or less. Brefeldin-A destroyed Chlamydomonas Golgi, inhibited the secretory pathway, inhibited flagellar regeneration, and induced full-length flagella to disassemble within 6 hrs, consistent with flagellar disassembly being induced by a failure to resupply membrane. In contrast to membrane lipids, a pool of biotinylatable membrane proteins was identified that was sufficient to resupply flagella as they released vesicles for 6 hrs in the absence of protein synthesis and to support one and nearly two regenerations of flagella following amputation. These studies reveal the importance of the secretory pathway to assemble and maintain full-length flagella.

  19. Molecular techniques to interrogate and edit the Chlamydomonas nuclear genome.

    Science.gov (United States)

    Jinkerson, Robert E; Jonikas, Martin C

    2015-05-01

    The success of the green alga Chlamydomonas reinhardtii as a model organism is to a large extent due to the wide range of molecular techniques that are available for its characterization. Here, we review some of the techniques currently used to modify and interrogate the C. reinhardtii nuclear genome and explore several technologies under development. Nuclear mutants can be generated with ultraviolet (UV) light and chemical mutagens, or by insertional mutagenesis. Nuclear transformation methods include biolistic delivery, agitation with glass beads, and electroporation. Transforming DNA integrates into the genome at random sites, and multiple strategies exist for mapping insertion sites. A limited number of studies have demonstrated targeted modification of the nuclear genome by approaches such as zinc-finger nucleases and homologous recombination. RNA interference is widely used to knock down expression levels of nuclear genes. A wide assortment of transgenes has been successfully expressed in the Chlamydomonas nuclear genome, including transformation markers, fluorescent proteins, reporter genes, epitope tagged proteins, and even therapeutic proteins. Optimized expression constructs and strains help transgene expression. Emerging technologies such as the CRISPR/Cas9 system, high-throughput mutant identification, and a whole-genome knockout library are being developed for this organism. We discuss how these advances will propel future investigations.

  20. Purification and photobiochemical profile of photosystem 1 from a high-salt tolerant, oleaginous Chlorella (Trebouxiophycaea, Chlorophyta).

    Science.gov (United States)

    McConnell, Michael D; Lowry, David; Rowan, Troy N; van Dijk, Karin; Redding, Kevin E

    2015-06-01

    The eukaryotic green alga Chlamydomonas reinhardtii has been studied extensively within the biofuel industry as a model organism, as researchers look towards algae to provide chemical feedstocks (i.e., lipids) for the production of liquid transportation fuels. C. reinhardtii, however, is unsuitable for high-level production of such precursors due to its relatively poor lipid accumulation and fresh-water demand. In this study we offer insight into the primary light harvesting and electron transfer reactions that occur during phototropic growth in a high-salt tolerant strain of Chlorella (a novel strain introduced here as NE1401), a single-celled eukaryotic algae also in the phylum Chlorophyta. Under nutrient starvation many eukaryotic algae increase dramatically the amount of lipids stored in lipid bodies within their cell interiors. Microscopy and lipid analyses indicate that Chlorella sp. NE1401 may become a superior candidate for algal biofuels production. We have purified highly active Photosystem 1 (PS1) complexes to study in vitro, so that we may understand further the photobiochemisty of this promising biofuel producer and how its characteristics compare and contrast with that of the better understood C. reinhardtii. Our findings suggest that the PS1 complex from Chlorella sp. NE1401 demonstrates similar characteristics to that of C. reinhardtii with respect to light-harvesting and electron transfer reactions. We also illustrate that the relative extent of the light state transition performed by Chlorella sp. NE1401 is smaller compared to C. reinhardtii, although they are triggered by the same dynamic light stresses.

  1. Partially Functional Outer-Arm Dynein in a Novel Chlamydomonas Mutant Expressing a Truncated γ Heavy Chain▿

    Science.gov (United States)

    Liu, Zhongmei; Takazaki, Hiroko; Nakazawa, Yuki; Sakato, Miho; Yagi, Toshiki; Yasunaga, Takuo; King, Stephen M.; Kamiya, Ritsu

    2008-01-01

    The outer dynein arm of Chlamydomonas flagella contains three heavy chains (α, β, and γ), each of which exhibits motor activity. How they assemble and cooperate is of considerable interest. Here we report the isolation of a novel mutant, oda2-t, whose γ heavy chain is truncated at about 30% of the sequence. While the previously isolated γ chain mutant oda2 lacks the entire outer arm, oda2-t retains outer arms that contain α and β heavy chains, suggesting that the N-terminal sequence (corresponding to the tail region) is necessary and sufficient for stable outer-arm assembly. Thin-section electron microscopy and image analysis localize the γ heavy chain to a basal region of the outer-arm image in the axonemal cross section. The motility of oda2-t is lower than that of the wild type and oda11 (lacking the α heavy chain) but higher than that of oda2 and oda4-s7 (lacking the motor domain of the β heavy chain). Thus, the outer-arm dynein lacking the γ heavy-chain motor domain is partially functional. The availability of mutants lacking individual heavy chains should greatly facilitate studies on the structure and function of the outer-arm dynein. PMID:18487347

  2. Chloramphenicol Mediates Superoxide Production in Photosystem II and Enhances Its Photodamage in Isolated Membrane Particles

    Science.gov (United States)

    Rehman, Ateeq Ur; Kodru, Sandeesha; Vass, Imre

    2016-01-01

    Chloramphenicol (CAP) is an inhibitor of protein synthesis, which is frequently used to decouple photodamage and protein synthesis dependent repair of Photosystem II during the process of photoinhibition. It has been reported earlier that CAP is able to mediate superoxide production by transferring electrons from the acceptor side of Photosystem I to oxygen. Here we investigated the interaction of CAP with Photosystem II electron transport processes by oxygen uptake and variable chlorophyll fluorescence measurements. Our data show that CAP can accept electrons at the acceptor side of Photosystem II, most likely from Pheophytin, and deliver them to molecular oxygen leading to superoxide production. In addition, the presence of CAP enhances photodamage of Photosystem II electron transport in isolated membrane particles, which effect is reversible by superoxide dismutase. It is concluded that CAP acts as electron acceptor in Photosystem II and mediates its superoxide dependent photodamage. This effect has potential implications for the application of CAP in photoinhibitory studies in intact systems. PMID:27092170

  3. The LC7 Light Chains of Chlamydomonas Flagellar Dyneins Interact with Components Required for Both Motor Assembly and Regulation

    Science.gov (United States)

    DiBella, Linda M.; Sakato, Miho; Patel-King, Ramila S.; Pazour, Gregory J.; King, Stephen M.

    2004-01-01

    Members of the LC7/Roadblock family of light chains (LCs) have been found in both cytoplasmic and axonemal dyneins. LC7a was originally identified within Chlamydomonas outer arm dynein and associates with this motor's cargo-binding region. We describe here a novel member of this protein family, termed LC7b that is also present in the Chlamydomonas flagellum. Levels of LC7b are reduced ∼20% in axonemes isolated from strains lacking inner arm I1 and are ∼80% lower in the absence of the outer arms. When both dyneins are missing, LC7b levels are diminished to <10%. In oda9 axonemal extracts that completely lack outer arms, LC7b copurifies with inner arm I1, whereas in ida1 extracts that are devoid of I1 inner arms it associates with outer arm dynein. We also have observed that some LC7a is present in both isolated axonemes and purified 18S dynein from oda1, suggesting that it is also a component of both the outer arm and inner arm I1. Intriguingly, in axonemal extracts from the LC7a null mutant, oda15, which assembles ∼30% of its outer arms, LC7b fails to copurify with either dynein, suggesting that it interacts with LC7a. Furthermore, both the outer arm γ heavy chain and DC2 from the outer arm docking complex completely dissociate after salt extraction from oda15 axonemes. EDC cross-linking of purified dynein revealed that LC7b interacts with LC3, an outer dynein arm thioredoxin; DC2, an outer arm docking complex component; and also with the phosphoprotein IC138 from inner arm I1. These data suggest that LC7a stabilizes both the outer arms and inner arm I1 and that both LC7a and LC7b are involved in multiple intradynein interactions within both dyneins. PMID:15304520

  4. Phosphoprotein SAK1 is a regulator of acclimation to singlet oxygen in Chlamydomonas reinhardtii.

    Science.gov (United States)

    Wakao, Setsuko; Chin, Brian L; Ledford, Heidi K; Dent, Rachel M; Casero, David; Pellegrini, Matteo; Merchant, Sabeeha S; Niyogi, Krishna K

    2014-05-23

    Singlet oxygen is a highly toxic and inevitable byproduct of oxygenic photosynthesis. The unicellular green alga Chlamydomonas reinhardtii is capable of acclimating specifically to singlet oxygen stress, but the retrograde signaling pathway from the chloroplast to the nucleus mediating this response is unknown. Here we describe a mutant, singlet oxygen acclimation knocked-out 1 (sak1), that lacks the acclimation response to singlet oxygen. Analysis of genome-wide changes in RNA abundance during acclimation to singlet oxygen revealed that SAK1 is a key regulator of the gene expression response during acclimation. The SAK1 gene encodes an uncharacterized protein with a domain conserved among chlorophytes and present in some bZIP transcription factors. The SAK1 protein is located in the cytosol, and it is induced and phosphorylated upon exposure to singlet oxygen, suggesting that it is a critical intermediate component of the retrograde signal transduction pathway leading to singlet oxygen acclimation.DOI: http://dx.doi.org/10.7554/eLife.02286.001.

  5. Phosphoprotein SAK1 is a regulator of acclimation to singlet oxygen in Chlamydomonas reinhardtii

    Science.gov (United States)

    Wakao, Setsuko; Chin, Brian L; Ledford, Heidi K; Dent, Rachel M; Casero, David; Pellegrini, Matteo; Merchant, Sabeeha S; Niyogi, Krishna K

    2014-01-01

    Singlet oxygen is a highly toxic and inevitable byproduct of oxygenic photosynthesis. The unicellular green alga Chlamydomonas reinhardtii is capable of acclimating specifically to singlet oxygen stress, but the retrograde signaling pathway from the chloroplast to the nucleus mediating this response is unknown. Here we describe a mutant, singlet oxygen acclimation knocked-out 1 (sak1), that lacks the acclimation response to singlet oxygen. Analysis of genome-wide changes in RNA abundance during acclimation to singlet oxygen revealed that SAK1 is a key regulator of the gene expression response during acclimation. The SAK1 gene encodes an uncharacterized protein with a domain conserved among chlorophytes and present in some bZIP transcription factors. The SAK1 protein is located in the cytosol, and it is induced and phosphorylated upon exposure to singlet oxygen, suggesting that it is a critical intermediate component of the retrograde signal transduction pathway leading to singlet oxygen acclimation. DOI: http://dx.doi.org/10.7554/eLife.02286.001 PMID:24859755

  6. Prospects of using unicellular algae protein in biological life-support systems. [Chlorella, Chlamydomonas, Spirulina, Euglena

    Energy Technology Data Exchange (ETDEWEB)

    Antonyan, A.A.; Abakumova, I.A.; Meleshko, G.I.; Vlasova, T.F.

    The concentration, amino acid composition and biological value of proteins of unicellular algae belonging to various taxonomic groups (Chlorella, Chlamydomonas, Spirulina, Euglena) were investigated. With respect to their characteristics, these algae hold promise as components of biological life-support systems (BLSS). Indices characterizing the protein and biomass quality and biological value were calculated. Such indices as A/E (where A is an essential amino acid and E is the sum total of amino acids), anti-E/T (where anti-E is nitrogen of essential amino acids and T is its sum total), amino acid number, factor of digestibility in vitro were high enough and close to the respective parameters of the reference protein. Animal experiments showed high biological value of the algal biomass and the lack of its toxic or other adverse effects. It is suggested that the differences in the protein composition associated with various algal forms and cultivation conditions can be used to produce balanced diets by varying the portion of each form of the photoautotropic component of BLSS.

  7. A Hard Day's Night: Diatoms continue recycling Photosystem II in the dark

    Directory of Open Access Journals (Sweden)

    Gang Li

    2016-11-01

    Full Text Available Marine diatoms are photosynthetic, and thrive in environments where light fluctuates. Like all oxygenic photosynthetic organisms diatoms face a light-dependent inactivation of the Photosystem II complexes that photooxidize water to generate biosynthetic reductant. To maintain photosynthesis this photoinactivation must be countered by slow and metabolically expensive protein turnover, which is light dependent in cyanobacteria and in plants. We tracked daily cycles of the content, synthesis and degradation of Photosystem II, in a small and in a large marine diatom, under low and high growth light levels. We show that, unlike plants, diatoms maintain extensive cycling of Photosystem II proteins even in the dark. Photosystem II protein cycling saturates at low light, and continued cycling in dark periods, using energy from respiration, allows the diatoms to catch up to excess photoinactivation accumulated over the preceding illuminated period. The large diatom suffers only limited photoinactivation of Photosystem II, but cycling of Photosystem II protein exceeds Photosystem II inactivation, so the large diatom recycles functional Photosystem II units before they are inactivated. Through the diel cycle the contents of active Photosystem II centers and Photosystem II proteins change predictably, but are not correlated, generating large changes in the fraction of total PSII that is active at a given time or growth condition. We propose that dark and steady cycling of Photosystem II proteins is driven by the tight integration of chloroplastic and mitochondrial metabolism in diatoms. This ability for baseline, continuous Photosystem II repair could contribute to the success of diatoms in mixed water environments that carry them from illumination to darkness and back.

  8. Chlamydomonas fla mutants reveal a link between deflagellation and intraflagellar transport

    Directory of Open Access Journals (Sweden)

    Quarmby Lynne

    2003-08-01

    Full Text Available Abstract Background Cilia and flagella are often lost in anticipation of mitosis or in response to stress. There are two ways that a cell can lose its flagella: resorption or deflagellation. Deflagellation involves active severing of the axoneme at the base of the flagellum; this process is defective in Chlamydomonas fa mutants. In contrast, resorption has been thought to occur as a consequence of constitutive disassembly at the tip in the absence of continued assembly, which requires intraflagellar transport (IFT. Chlamydomonas fla mutants are unable to build and maintain flagella due to defects in IFT. Results fla10 cells, which are defective in kinesin-II, the anterograde IFT motor, resorb their flagella at the restrictive temperature (33°C, as previously reported. We find that in standard media containing ~300 microM calcium, fla10 cells lose flagella by deflagellation at 33°C. This temperature-induced deflagellation of a fla mutant is not predicted by the IFT-based model for flagellar length control. Other fla mutants behave similarly, losing their flagella by deflagellation instead of resorption, if adequate calcium is available. These data suggest a new model whereby flagellar resorption involves active disassembly at the base of the flagellum via a mechanism with components in common with the severing machinery of deflagellation. As predicted by this model, we discovered that deflagellation stimuli induce resorption if deflagellation is blocked either by mutation in a FA gene or by lack of calcium. Further support for this model comes from our discovery that fla10-fa double mutants resorb their flagella more slowly than fla10 mutants. Conclusions Deflagellation of the fla10 mutant at the restrictive temperature is indicative of an active disassembly signal, which can manifest as either resorption or deflagellation. We propose that when IFT is halted by either an inactivating mutation or a cellular signal, active flagellar disassembly

  9. Wiring of Photosystem II to Hydrogenase for Photoelectrochemical Water Splitting.

    Science.gov (United States)

    Mersch, Dirk; Lee, Chong-Yong; Zhang, Jenny Zhenqi; Brinkert, Katharina; Fontecilla-Camps, Juan C; Rutherford, A William; Reisner, Erwin

    2015-07-08

    In natural photosynthesis, light is used for the production of chemical energy carriers to fuel biological activity. The re-engineering of natural photosynthetic pathways can provide inspiration for sustainable fuel production and insights for understanding the process itself. Here, we employ a semiartificial approach to study photobiological water splitting via a pathway unavailable to nature: the direct coupling of the water oxidation enzyme, photosystem II, to the H2 evolving enzyme, hydrogenase. Essential to this approach is the integration of the isolated enzymes into the artificial circuit of a photoelectrochemical cell. We therefore developed a tailor-made hierarchically structured indium-tin oxide electrode that gives rise to the excellent integration of both photosystem II and hydrogenase for performing the anodic and cathodic half-reactions, respectively. When connected together with the aid of an applied bias, the semiartificial cell demonstrated quantitative electron flow from photosystem II to the hydrogenase with the production of H2 and O2 being in the expected two-to-one ratio and a light-to-hydrogen conversion efficiency of 5.4% under low-intensity red-light irradiation. We thereby demonstrate efficient light-driven water splitting using a pathway inaccessible to biology and report on a widely applicable in vitro platform for the controlled coupling of enzymatic redox processes to meaningfully study photocatalytic reactions.

  10. Robust expression of a bioactive mammalian protein in Chlamydomonas chloroplast

    Energy Technology Data Exchange (ETDEWEB)

    Mayfield, Stephen P

    2015-01-13

    Methods and compositions are disclosed to engineer chloroplast comprising heterologous mammalian genes via a direct replacement of chloroplast Photosystem II (PSII) reaction center protein coding regions to achieve expression of recombinant protein above 5% of total protein. When algae is used, algal expressed protein is produced predominantly as a soluble protein where the functional activity of the peptide is intact. As the host algae is edible, production of biologics in this organism for oral delivery of proteins/peptides, especially gut active proteins, without purification is disclosed.

  11. New thioredoxin targets in the unicellular photosynthetic eukaryote Chlamydomonas reinhardtii

    Science.gov (United States)

    Lemaire, Stéphane D.; Guillon, Blanche; Le Maréchal, Pierre; Keryer, Eliane; Miginiac-Maslow, Myroslawa; Decottignies, Paulette

    2004-01-01

    Proteomics were used to identify the proteins from the eukaryotic unicellular green alga Chlamydomonas reinhardtii that can be reduced by thioredoxin. These proteins were retained specifically on a thioredoxin affinity column made of a monocysteinic thioredoxin mutant able to form mixed disulfides with its targets. Of a total of 55 identified targets, 29 had been found previously in higher plants or Synechocystis, but 26 were new targets. Biochemical tests were performed on three of them, showing a thioredoxin-dependent activation of isocitrate lyase and isopropylmalate dehydrogenase and a thioredoxin-dependent deactivation of catalase that is redox insensitive in Arabidopsis. In addition, we identified a Ran protein, a previously uncharacterized nuclear target in a photosynthetic organism. The metabolic and evolutionary implications of these findings are discussed. PMID:15123830

  12. A deoxyribonuclease from Chlamydomonas reinhardii. 1. Purification and properties.

    Science.gov (United States)

    Tait, G C; Harris, W J

    1977-05-16

    A deoxyribonuclease has been purified more than 2000-fold from the green algae, Chlamydomonas reinhardii. The enzyme is most active on denatured DNA. Optimum activity is at pH 8.5, in 80 mM Tris-HCl buffer and 2 mM CaCl2. Other divalent cations can replace Ca2+ with varying lower efficiency. EDTA and inorganic phosphate are strongly inhibitory, while ATP and high concentrations of 2-mercaptoethanol are slightly inhibitory. The molecular weight is approximately 35 000, the Stokes radius is 2.7 nm, and the sedimentation coefficient 2.8 S. It is a single polypeptide chain, and the frictional ratio of 1.27 suggests it is only slightly asymetrical. The isoelectric point is 9.5. This enzyme has been termed exonuclease 1.

  13. Cilia and ciliopathies:From Chlamydomonas and beyond

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    The biological function of motile cilia/flagella has long been recognized. The non-motile primary cilium, once regarded as a vestigial organelle, however, has been found recently to play unexpected roles in mammalian physiology and development. Defects in cilia have profound impact on human health. Diseases related to cilia, collectively called ciliopathies include male infertility, primary cilia dyskinesia, renal cyst formation, blindness, polydactyly, obesity, hypertension, and even mental retardation. Our current understanding of cilia and ciliopathies has been fueled by basic research employing various model organisms including Chlamydomonas, a unicellular green alga. This review article provides a general introduction to the cell biology of cilia and an overview of various cilia-related diseases.

  14. Supercomplexes of photosystem I and antenna proteins in cyanobacteria and green plants

    NARCIS (Netherlands)

    Kouril, R.; Zygadlo, A.; Jensen, P. E.; Yeremenko, N.; Matthijs, H.C.P.; D'Haene, S.; Dekker, J.P.; Boekema, E.J.; Matthijs, H.C.P.

    2005-01-01

    Photosystem I (PSI) and Photosystem II (PSII) can form supercomplexes with various types of antenna proteins. One such protein is Light-harvesting complex II (LHCII). In plants it is bound in its trimeric form to dimeric PSII. In spinach and Arabidopsis thaliana most dimers bind 2-4 LHCII trimers. W

  15. Effects of Chlorophyll Availability on Fluorescence Components of Photosystems in the ORF469-Deletion Mutant of Cyanobacterium

    Institute of Scientific and Technical Information of China (English)

    1999-01-01

    PCR-amplified ORF469 fragment from Synechocystis sp.PCC 6803 was cloned into pUC118 and a construct was made in which part of ORF469 was deleted and replaced by erythromycin resistance cassette.Transformation of wild type strain of Synechocystis sp.PCC 6803 with this construct yielded a mutant in which ORF469 was deleted.In the resulting mutant, the light-independent pathway of chlorophyll biosynthesis was inactivated and availability of chlorophyll was fully dependent on light.When propagated the mutant in dark, the chlorophyll was non-detectable and protochlorophyllide with 645 nm fluorescence emission peak was accumulated.Meanwhile, the fluorescence emission peaks (excited at 435 nm) of thylakoids at 685 nm, 695 nm and 725 nm, which represented relative chlorophyll-binding proteins, disappeared.Upon return of dark-grown ORF469 mutant to the light, greening occurred and chlorophyll was synthesized to assembly fluorescence emission components in photosystems.Newly synthesized chlorophyll combined the fluorescence component of 685 nm at first, then 725 nm and 695 nm at last, which indicates a pecking order for biogenesis of chlorophyll-binding proteins when availability of chlorophyll is limited.The mutant lacking ORF469 in Synechocystis sp.PCC 6803 was suggested as an excellent cyanobacterial system for studies on the interactions between chlorophyll and chlorophyll-binding proteins in photosystems.

  16. Temperature dependence and polarization of fluorescence from Photosystem I in the cyanobacterium Synechocystis sp. PCC 6803.

    Science.gov (United States)

    Wittmershaus, B P; Woolf, V M; Vermaas, W F

    1992-02-01

    To determine the fluorescence properties of cyanobacterial Photosystem I (PS I) in relatively intact systems, fluorescence emission from 20 to 295 K and polarization at 77 K have been measured from phycobilisomes-less thylakoids of Synechocystis sp. PCC 6803 and a mutant strain lacking Photosystem II (PS II). At 295 K, the fluorescence maxima are 686 nm in the wild type from PS I and PS II and at 688 nm from PS I in the mutant. This emission is characteristic of bulk antenna chlorophylls (Chls). The 690-nm fluorescence component of PS I is temperature independent. For wild-type and mutant, 725-nm fluorescence increases by a factor of at least 40 from 295 to 20 K. We model this temperature dependence assuming a small number of Chls within PS I, emitting at 725 nm, with an energy level below that of the reaction center, P700. Their excitation transfer rate to P700 decreases with decreasing temperature increasing the yield of 725-nm fluorescence.Fluorescence excitation spectra of polarized emission from low-energy Chls were measured at 77 and 295 K on the mutant lacking PS II. At excitation wavelengths longer than 715 nm, 760-nm emission is highly polarized indicating either direct excitation of the emitting Chls with no participation in excitation transfer or total alignment of the chromophores. Fluorescence at 760 nm is unpolarized for excitation wavelengths shorter than 690 nm, inferring excitation transfer between Chls before 760-nm fluorescence occurs.Our measurements illustrate that: 1) a single group of low-energy Chls (F725) of the core-like PS I complex in cyanobacteria shows a strongly temperature-dependent fluorescence and, when directly excited, nearly complete fluorescence polarization, 2) these properties are not the result of detergent-induced artifacts as we are examining intact PS I within the thylakoid membrane of S. 6803, and 3) the activation energy for excitation transfer from F725 Chls to P700 is less than that of F735 Chls in green plants; F725

  17. Diversity of viral photosystem-I psaA genes.

    Science.gov (United States)

    Hevroni, Gur; Enav, Hagay; Rohwer, Forest; Béjà, Oded

    2015-08-01

    Marine photosynthesis is one of the major contributors to the global carbon cycle and the world's oxygen supply. This process is largely driven by cyanobacteria, namely Synechococcus and Prochlorococcus. Genes encoding photosystem-II (PSII) reaction center proteins are found in many cyanophage genomes, and are expressed during the infection of their hosts. On the basis of metagenomics, cyanophage photosystem-I (PSI) gene cassettes were recently discovered with two gene arrangements psaJF→C→A→B→K→E→D and psaD→C→A→B. It was suggested that the horizontal transfer of PSII and PSI genes is increasing phage fitness. To better understand their diversity, we designed degenerate primers to cover a wide diversity of organisms, and using PCR we targeted the psaC→A arrangement, which is unique to cyanophages cassettes. We examined viral concentrates from four islands in the Pacific Ocean and found samples containing the psaC→A arrangement. Analyses of the amplified viral psaA gene revealed six subgroups varying in their level of similarity and %G+C content, suggesting that the diversity of cyanophage PSI genes is greater than originally thought.

  18. Genetic and biochemical analysis of the TLA1 gene in Chlamydomonas reinhardtii.

    Science.gov (United States)

    Mitra, Mautusi; Melis, Anastasios

    2010-02-01

    The Chlamydomonas reinhardtii genomic DNA database contains a predicted open reading frame (ORF-P) without an apparent stop-codon and unknown coding sequence, located in close proximity and immediately upstream of the TLA1 gene (GenBank Accession No. AF534570). The latter was implicated in the regulation of the light-harvesting chlorophyll antenna size of photosynthesis (Tetali et al. Planta 225:813-829, 2007). To provide currently lacking information on ORF-P and its potential participation in TLA1 gene expression, thus in the regulation of the chlorophyll antenna size, genetic and biochemical analyses were undertaken. The coding and UTR regions of the ORF-P were defined and delineated from those of the adjacent TLA1 gene. ORF-P is shown to encode a protein with a distinct RING-like zinc finger domain that is present in numerous eukaryotic proteins, believed to play a role in cellular ubiquitination, leading to regulation of cellular processes like signaling, growth, transcription, and DNA repair. It is further shown that the two genes share a 74-bp overlap between the 3' UTR region of ORF-P and the 5' UTR region of TLA1. However, they possess distinct start and stop codons and separate coding sequences, and transcribed as separate mRNAs without any trans-splicing between them. Complementation experiments showed that the TLA1 gene alone is sufficient to rescue the truncated chlorophyll antenna size phenotype of the tla1 mutant. Protein sequence alignments in C. reinhardtii and the colorless microalga Polytomella parva suggested that TLA1 defines the relationship between nucleus and organelle in microalgae, indirectly affecting the development of the chlorophyll antenna size.

  19. Ergodicity, configurational entropy and free energy in pigment solutions and plant photosystems: influence of excited state lifetime.

    Science.gov (United States)

    Jennings, Robert C; Zucchelli, Giuseppe

    2014-01-01

    We examine ergodicity and configurational entropy for a dilute pigment solution and for a suspension of plant photosystem particles in which both ground and excited state pigments are present. It is concluded that the pigment solution, due to the extreme brevity of the excited state lifetime, is non-ergodic and the configurational entropy approaches zero. Conversely, due to the rapid energy transfer among pigments, each photosystem is ergodic and the configurational entropy is positive. This decreases the free energy of the single photosystem pigment array by a small amount. On the other hand, the suspension of photosystems is non-ergodic and the configurational entropy approaches zero. The overall configurational entropy which, in principle, includes contributions from both the single excited photosystems and the suspension which contains excited photosystems, also approaches zero. Thus the configurational entropy upon photon absorption by either a pigment solution or a suspension of photosystem particles is approximately zero.

  20. Late steps in cytoplasmic maturation of assembly-competent axonemal outer arm dynein in Chlamydomonas require interaction of ODA5 and ODA10 in a complex.

    Science.gov (United States)

    Dean, Anudariya B; Mitchell, David R

    2015-10-15

    Axonemal dyneins are multisubunit enzymes that must be preassembled in the cytoplasm, transported into cilia by intraflagellar transport, and bound to specific sites on doublet microtubules, where their activity facilitates microtubule sliding-based motility. Outer dynein arms (ODAs) require assembly factors to assist their preassembly, transport, and attachment to cargo (specific doublet A-tubule sites). In Chlamydomonas, three assembly factors--ODA5, ODA8, and ODA10--show genetic interactions and have been proposed to interact in a complex, but we recently showed that flagellar ODA8 does not copurify with ODA5 or ODA10. Here we show that ODA5 and ODA10 depend on each other for stability and coexist in a complex in both cytoplasmic and flagellar extracts. Immunofluorescence and immuno-electron microscopy reveal that ODA10 in flagella localizes strictly to a proximal region of doublet number 1, which completely lacks ODAs in Chlamydomonas. Studies of the in vitro binding of ODAs to axonemal doublets reveal a role for the ODA5/ODA10 assembly complex in cytoplasmic maturation of ODAs into a form that can bind to doublet microtubules.

  1. The Chlamydomonas Genome Reveals the Evolution of Key Animal and Plant Functions

    Energy Technology Data Exchange (ETDEWEB)

    Merchant, Sabeeha S

    2007-04-09

    Chlamydomonas reinhardtii is a unicellular green alga whose lineage diverged from land plants over 1 billion years ago. It is a model system for studying chloroplast-based photosynthesis, as well as the structure, assembly, and function of eukaryotic flagella (cilia), which were inherited from the common ancestor of plants and animals, but lost in land plants. We sequenced the 120-megabase nuclear genome of Chlamydomonas and performed comparative phylogenomic analyses, identifying genes encoding uncharacterized proteins that are likely associated with the function and biogenesis of chloroplasts or eukaryotic flagella. Analyses of the Chlamydomonas genome advance our understanding of the ancestral eukaryotic cell, reveal previously unknown genes associated with photosynthetic and flagellar functions, and establish links between ciliopathy and the composition and function of flagella.

  2. Lipidomic Analysis of Chlamydomonas reinhardtii under Nitrogen and Sulfur Deprivation.

    Directory of Open Access Journals (Sweden)

    Dawei Yang

    Full Text Available Chlamydomonas reinhardtii accumulates lipids under complete nutrient starvation conditions while overall growth in biomass stops. In order to better understand biochemical changes under nutrient deprivation that maintain production of algal biomass, we used a lipidomic assay for analyzing the temporal regulation of the composition of complex lipids in C. reinhardtii in response to nitrogen and sulfur deprivation. Using a chip-based nanoelectrospray direct infusion into an ion trap mass spectrometer, we measured a diversity of lipid species reported for C. reinhardtii, including PG phosphatidylglycerols, PI Phosphatidylinositols, MGDG monogalactosyldiacylglycerols, DGDG digalactosyldiacylglycerols, SQDG sulfoquinovosyldiacylglycerols, DGTS homoserine ether lipids and TAG triacylglycerols. Individual lipid species were annotated by matching mass precursors and MS/MS fragmentations to the in-house LipidBlast mass spectral database and MS2Analyzer. Multivariate statistics showed a clear impact on overall lipidomic phenotypes on both the temporal and the nutrition stress level. Homoserine-lipids were found up-regulated at late growth time points and higher cell density, while triacyclglycerols showed opposite regulation of unsaturated and saturated fatty acyl chains under nutritional deprivation.

  3. Response of Chlamydomonas reinhardtii to naphthenic acid exposure

    Energy Technology Data Exchange (ETDEWEB)

    Goff, K.; Wilson, K. [Saskatchewan Univ., Saskatoon, SK (Canada); Headley, J. [Environment Canada, Ottawa, ON (Canada)

    2010-07-01

    This study examined the feasibility of using a model organism for the algal bioremediation of oil sands process water (OSPW), a highly toxic mixture of sediments, bitumen, ions, and organic and inorganic compounds. Naphthenic acids (NAs) are a contaminant class of particular concern. Bioremediation techniques may mitigate toxicity of OSPW in general, and NAs in particular. Although most studies on the biodegradation of NAs focus on the role of bacteria, fungi, and emergent macrophytes, studies have indicated that algae may also play a key role through direct degradation, biosequestration, or photosynthetic aeration of waters to promote other biological reactions. Chlamydomonas frigida is of particular interest, but no cultures are currently available. Therefore, this study used C. reinhardtii, a well-characterized model organism, to begin analysis of potential algal bioremediation of OSPW. Cultures of C. reinhardtii were grown heterotrophically in nutrient media spiked with a dilution series of NAs. Culture densities were measured to compile growth curves over time, changes in rate of growth, and survivability. Negative ion electrospray mass spectrometry was used to determine the concentration of NAs in solution in relation to growth rate and culture density. The study determined the tolerance of C. reinhardtii to NAs. A mechanism for this tolerance was then proposed.

  4. Insecticides induced biochemical changes in freshwater microalga Chlamydomonas mexicana.

    Science.gov (United States)

    Kumar, Muthukannan Satheesh; Kabra, Akhil N; Min, Booki; El-Dalatony, Marwa M; Xiong, Jiuqiang; Thajuddin, Nooruddin; Lee, Dae Sung; Jeon, Byong-Hun

    2016-01-01

    The effect of insecticides (acephate and imidacloprid) on a freshwater microalga Chlamydomonas mexicana was investigated with respect to photosynthetic pigments, carbohydrate and protein contents, fatty acids composition and induction of stress indicators including proline, superoxide dismutase (SOD) and catalase (CAT). C. mexicana was cultivated with 1, 5, 10, 15, 20 and 25 mg L(-1) of acephate and imidacloprid. The microalga growth increased with increasing concentrations of both insecticides up to 15 mg L(-1), beyond which the growth declined compared to control condition (without insecticides). C. mexicana cultivated with 15 mg L(-1) of both insecticides for 12 days was used for further analysis. The accumulation of photosynthetic pigments (chlorophyll and carotenoids), carbohydrates and protein was decreased in the presence of both insecticides. Acephate and imidacloprid induced the activities of superoxide dismutase (SOD) and catalase (CAT) and increased the concentration of proline in the microalga, which play a defensive role against various environmental stresses. Fatty acid analysis revealed that the fraction of polyunsaturated fatty acids decreased on exposure to both insecticides. C. mexicana also promoted 25 and 21% removal of acephate and imidacloprid, respectively. The biochemical changes in C. mexicana on exposure to acephate and imidacloprid indicate that the microalga undergoes an adaptive change in response to the insecticide-induced oxidative stress.

  5. Establishing Chlamydomonas reinhardtii as an industrial biotechnology host.

    Science.gov (United States)

    Scaife, Mark A; Nguyen, Ginnie T D T; Rico, Juan; Lambert, Devinn; Helliwell, Katherine E; Smith, Alison G

    2015-05-01

    Microalgae constitute a diverse group of eukaryotic unicellular organisms that are of interest for pure and applied research. Owing to their natural synthesis of value-added natural products microalgae are emerging as a source of sustainable chemical compounds, proteins and metabolites, including but not limited to those that could replace compounds currently made from fossil fuels. For the model microalga, Chlamydomonas reinhardtii, this has prompted a period of rapid development so that this organism is poised for exploitation as an industrial biotechnology platform. The question now is how best to achieve this? Highly advanced industrial biotechnology systems using bacteria and yeasts were established in a classical metabolic engineering manner over several decades. However, the advent of advanced molecular tools and the rise of synthetic biology provide an opportunity to expedite the development of C. reinhardtii as an industrial biotechnology platform, avoiding the process of incremental improvement. In this review we describe the current status of genetic manipulation of C. reinhardtii for metabolic engineering. We then introduce several concepts that underpin synthetic biology, and show how generic parts are identified and used in a standard manner to achieve predictable outputs. Based on this we suggest that the development of C. reinhardtii as an industrial biotechnology platform can be achieved more efficiently through adoption of a synthetic biology approach.

  6. Expression of human soluble TRAIL in Chlamydomonas reinhardtii chloroplast

    Institute of Scientific and Technical Information of China (English)

    YANG Zongqi; LI yinü; CHEN Feng; LI Dong; ZHANG Zhifang; LIU Yanxin; ZHENG Dexian; WANG Yong; SHEN Guifang

    2006-01-01

    Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) induces selectively apoptosis in various tumor cells and virus-infected cells, but rarely in normal cells. A chloroplast expression vector, p64TRAIL, containing the cDNA coding for the soluble TRAIL (sTRAIL), was constructed with clpP-trnL-petB-chlL-rpl23-rpl2 as Chlamydomonas reinhardtii plastid homologous recombinant fragments and spectinomycin-resistant aadA gene as a select marker. The plasmid p64TRAIL was transferred into the chloroplast genome of C. reinhardtii by the biolistic method. Three independently transformed lines were obtained by 100 mg/L spectinomycin selection. PCR amplification, Southern blot analysis of the sTRAIL coding region DNA and cultivation cells in the dark all showed that the exogenous DNA had been integrated into chloroplast genome of C. reinhardtii. Western blot analysis showed that human soluble TRAIL was expressed in C. reinhardtii chloroplast. The densitometric analysis of Western blot indicated that the expressed human sTRAIL protein in the chloroplasts of C. reinhardtii accounted for about 0.43%-0.67% of the total soluble proteins.These experimental results demonstrated the possibility of using transgenic chloroplasts of green alga as bioreactors for production of biopharmaceuticals.

  7. Identification of the Elusive Pyruvate Reductase of Chlamydomonas reinhardtii Chloroplasts

    Science.gov (United States)

    Burgess, Steven J.; Taha, Hussein; Yeoman, Justin A.; Iamshanova, Oksana; Chan, Kher Xing; Boehm, Marko; Behrends, Volker; Bundy, Jacob G.; Bialek, Wojciech; Murray, James W.; Nixon, Peter J.

    2016-01-01

    Under anoxic conditions the green alga Chlamydomonas reinhardtii activates various fermentation pathways leading to the creation of formate, acetate, ethanol and small amounts of other metabolites including d-lactate and hydrogen. Progress has been made in identifying the enzymes involved in these pathways and their subcellular locations; however, the identity of the enzyme involved in reducing pyruvate to d-lactate has remained unclear. Based on sequence comparisons, enzyme activity measurements, X-ray crystallography, biochemical fractionation and analysis of knock-down mutants, we conclude that pyruvate reduction in the chloroplast is catalyzed by a tetrameric NAD+-dependent d-lactate dehydrogenase encoded by Cre07.g324550. Its expression during aerobic growth supports a possible function as a ‘lactate valve’ for the export of lactate to the mitochondrion for oxidation by cytochrome-dependent d-lactate dehydrogenases and by glycolate dehydrogenase. We also present a revised spatial model of fermentation based on our immunochemical detection of the likely pyruvate decarboxylase, PDC3, in the cytoplasm. PMID:26574578

  8. Chlorophyll composition and photochemical activity of photosystems detached from chloroplast grana and stroma lamellae.

    Science.gov (United States)

    Gasanov, R A; French, C S

    1973-07-01

    A stroma fraction that has photosystem 1 activity and grana lamellae fractions that have activities for both photosystems were isolated by differential centrifugation of a needle valve homogenate. Subsequent fractions, corresponding to photosystems 1 (F-1D) and 2 (F-2D) were isolated by digitonin treatment of the grana lamellae (P-10K) and compared with respect to their chlorophyll composition and electron transport activities.Fraction F-2D from grana lamellae having photosystem 2 activity is primarily active in photosystem 2 and contains only the four major forms of chlorophyll a with a predominance of chlorophyll a 677 nm. This fraction differs from the original grana membranes in the absence of the longwavelength form of chlorophyll a and in the widening of the absorption band of chlorophyll a 682 nm from 10.9 to 15.6 nm.Photosystem 1 particles from grana and stroma both have high photosystem 1 activity but differ from each other in the proportions of the four major forms of chlorophyll a. The short-wavelength forms of chlorophyll a and also chlorophyll b 650 nm in particles from grana lamellae comprise relatively more total area than these same forms in the particles from stroma. In addition, the fraction corresponding to photosystem 1 from grana lamellae is not shifted to the long-wavelength side of the main absorption maximum, as compared to the photosystem 2 particles from grana and the original grana membrane fraction; this is usually observed in fractions that have photosystem 1 activity. Furthermore, the longest wavelength form of chlorophyll a in the photosystem 1 particles from grana is at 700 nm, while in the same fraction from stroma, it is at 706 nm.The half-width of the four main forms of chlorophyll a and both forms of chlorophyll b in the photosystem 1 fraction from grana is narrower than that of the corresponding forms in the same fraction from stroma. This may indicate a different packing of pigment molecules that are aggregated on the surface

  9. Negative impact on growth and photosynthesis in the green alga Chlamydomonas reinhardtii in the presence of the estrogen 17α-ethynylestradiol.

    Science.gov (United States)

    Pocock, Tessa; Falk, Stefan

    2014-01-01

    It is well known that estrogenic compounds affect development of fertilized eggs of many species of birds, fish and amphibians through disrupted activity of carbonic anhydrase (CA). The most potent activity comes from the most commonly occurring synthetic sterol, 17α-Ethynylestradiol (EE2). Less is known about the responses of aquatic phytoplankton to these compounds. Here we show for the first time that, in comparision to the control, the addition of 7 µM EE2 reduced the growth rate of the green alga Chlamydomonas reinhardtii by 68% for cells grown at high CO2. When cells were grown in ambient air (low Ci) with a fully activated carbon concentrating mechanism through the induction of CA activity, the growth rates were reduced by as much as 119%. A reduced growth rate could be observed at EE2 concentrations as low as 10 pM. This was accompanied by a reduced maximum capacity for electron transport in photosystem II as determined by a lower FV/FM for low Ci-grown cells, which indicates the involvement of CAH3, a CA specifically located in the thylakoid lumen involved in proton pumping across the thylakoid membranes. These results were in agreement with an observed reduction in the chloroplastic affinity for Ci as shown by a strong increase in the Michaelis-Menten K0.5 for HCO3-. In itself, a lowering of the growth rate of a green alga by addition of the sterol EE2 warrants further investigation into the potential environmental impact by the release of treated waste water.

  10. Negative impact on growth and photosynthesis in the green alga Chlamydomonas reinhardtii in the presence of the estrogen 17α-ethynylestradiol.

    Directory of Open Access Journals (Sweden)

    Tessa Pocock

    Full Text Available It is well known that estrogenic compounds affect development of fertilized eggs of many species of birds, fish and amphibians through disrupted activity of carbonic anhydrase (CA. The most potent activity comes from the most commonly occurring synthetic sterol, 17α-Ethynylestradiol (EE2. Less is known about the responses of aquatic phytoplankton to these compounds. Here we show for the first time that, in comparision to the control, the addition of 7 µM EE2 reduced the growth rate of the green alga Chlamydomonas reinhardtii by 68% for cells grown at high CO2. When cells were grown in ambient air (low Ci with a fully activated carbon concentrating mechanism through the induction of CA activity, the growth rates were reduced by as much as 119%. A reduced growth rate could be observed at EE2 concentrations as low as 10 pM. This was accompanied by a reduced maximum capacity for electron transport in photosystem II as determined by a lower FV/FM for low Ci-grown cells, which indicates the involvement of CAH3, a CA specifically located in the thylakoid lumen involved in proton pumping across the thylakoid membranes. These results were in agreement with an observed reduction in the chloroplastic affinity for Ci as shown by a strong increase in the Michaelis-Menten K0.5 for HCO3-. In itself, a lowering of the growth rate of a green alga by addition of the sterol EE2 warrants further investigation into the potential environmental impact by the release of treated waste water.

  11. Wiring photosystem I for direct solar hydrogen production.

    Science.gov (United States)

    Lubner, Carolyn E; Grimme, Rebecca; Bryant, Donald A; Golbeck, John H

    2010-01-26

    The generation of H(2) by the use of solar energy is a promising way to supply humankind's energy needs while simultaneously mitigating environmental concerns that arise due to climate change. The challenge is to find a way to connect a photochemical module that harnesses the sun's energy to a catalytic module that generates H(2) with high quantum yields and rates. In this review, we describe a technology that employs a "molecular wire" to connect a terminal [4Fe-4S] cluster of Photosystem I directly to a catalyst, which can be either a Pt nanoparticle or the distal [4Fe-4S] cluster of an [FeFe]- or [NiFe]-hydrogenase enzyme. The keys to connecting these two moieties are surface-located cysteine residues, which serve as ligands to Fe-S clusters and which can be changed through site-specific mutagenesis to glycine residues, and the use of a molecular wire terminated in sulfhydryl groups to connect the two modules. The sulfhydryl groups at the end of the molecular wire form a direct chemical linkage to a suitable catalyst or can chemically rescue a [4Fe-4S] cluster, thereby generating a strong coordination bond. Specifically, the molecular wire can connect the F(B) iron-sulfur cluster of Photosystem I either to a Pt nanoparticle or, by using the same type of genetic modification, to the differentiated iron atom of the distal [4Fe-4S].(Cys)(3)(Gly) cluster of hydrogenase. When electrons are supplied by a sacrificial donor, this technology forms the cathode of a photochemical half-cell that evolves H(2) when illuminated. If such a device were connected to the anode of a photochemical half-cell that oxidizes water, an in vitro solar energy converter could be realized that generates only O(2) and H(2) in the light. A similar methodology can be used to connect Photosystem I to other redox proteins that have surface-located [4Fe-4S] clusters. The controlled light-driven production of strong reductants by such systems can be used to produce other biofuels or to provide

  12. Crystal structure of monomeric photosystem II from Thermosynechococcus elongatus at 3.6-a resolution.

    Science.gov (United States)

    Broser, Matthias; Gabdulkhakov, Azat; Kern, Jan; Guskov, Albert; Müh, Frank; Saenger, Wolfram; Zouni, Athina

    2010-08-20

    The membrane-embedded photosystem II core complex (PSIIcc) uses light energy to oxidize water in photosynthesis. Information about the spatial structure of PSIIcc obtained from x-ray crystallography was so far derived from homodimeric PSIIcc of thermophilic cyanobacteria. Here, we report the first crystallization and structural analysis of the monomeric form of PSIIcc with high oxygen evolution capacity, isolated from Thermosynechococcus elongatus. The crystals belong to the space group C222(1), contain one monomer per asymmetric unit, and diffract to a resolution of 3.6 A. The x-ray diffraction pattern of the PSIIcc-monomer crystals exhibit less anisotropy (dependence of resolution on crystal orientation) compared with crystals of dimeric PSIIcc, and the packing of the molecules within the unit cell is different. In the monomer, 19 protein subunits, 35 chlorophylls, two pheophytins, the non-heme iron, the primary plastoquinone Q(A), two heme groups, 11 beta-carotenes, 22 lipids, seven detergent molecules, and the Mn(4)Ca cluster of the water oxidizing complex could be assigned analogous to the dimer. Based on the new structural information, the roles of lipids and protein subunits in dimer formation of PSIIcc are discussed. Due to the lack of non-crystallographic symmetry and the orientation of the membrane normal of PSIIcc perpendicular ( approximately 87 degrees ) to the crystallographic b-axis, further information about the structure of the Mn(4)Ca cluster is expected to become available from orientation-dependent spectroscopy on this new crystal form.

  13. Radiolytic mapping of solvent-contact surfaces in Photosystem II of higher plants: experimental identification of putative water channels within the photosystem.

    Science.gov (United States)

    Frankel, Laurie K; Sallans, Larry; Bellamy, Henry; Goettert, Jost S; Limbach, Patrick A; Bricker, Terry M

    2013-08-09

    Photosystem II uses water as an enzymatic substrate. It has been hypothesized that this water is vectored to the active site for water oxidation via water channels that lead from the surface of the protein complex to the Mn4O5Ca metal cluster. The radiolysis of water by synchrotron radiation produces amino acid residue-modifying OH(•) and is a powerful technique to identify regions of proteins that are in contact with water. In this study, we have used this technique to oxidatively modify buried amino acid residues in higher plant Photosystem II membranes. Fourier transform ion cyclotron resonance mass spectrometry was then used to identify these oxidized amino acid residues that were located in several core Photosystem II subunits (D1, D2, CP43, and CP47). While, as expected, the majority of the identified oxidized residues (≈75%) are located on the solvent-exposed surface of the complex, a number of buried residues on these proteins were also modified. These residues form groups which appear to lead from the surface of the complex to the Mn4O5Ca cluster. These residues may be in contact with putative water channels in the photosystem. These results are discussed within the context of a number of largely computational studies that have identified putative water channels in Photosystem II.

  14. Mechanisms of light harvesting by photosystem II in plants

    CERN Document Server

    Amarnath, Kapil; Schneider, Anna R; Fleming, Graham R

    2015-01-01

    Light harvesting by photosystem II (PSII) in plants is highly efficient and acclimates to rapid changes in the intensity of sunlight. However, the mechanisms of PSII light harvesting have remained experimentally inaccessible. Using a structure-based model of excitation energy flow in 200 nanometer (nm) x 200 nm patches of the grana membrane, where PSII is located, we accurately simulated chlorophyll fluorescence decay data with no free parameters. Excitation movement through the light harvesting antenna is diffusive, but becomes subdiffusive in the presence of charge separation at reaction centers. The influence of membrane morphology on light harvesting efficiency is determined by the excitation diffusion length of 50 nm in the antenna. Our model provides the basis for understanding how nonphotochemical quenching mechanisms affect PSII light harvesting in grana membranes.

  15. Robustness and Optimality of Light Harvesting in Cyanobacterial Photosystem I

    CERN Document Server

    Sener, M K; Ritz, T; Park, S; Fromme, P; Schulten, K; Sener, Melih K.; Lu, Deyu; Ritz, Thorsten; Park, Sanghyun; Fromme, Petra; Schulten, Klaus

    2002-01-01

    As most biological species, photosynthetic lifeforms have evolved to function optimally, despite thermal disorder and with fault tolerance. It remains a challenge to understand how this is achieved. To address this challenge the function of the protein-pigment complex photosystem I (PSI) of the cyanobacterium Synechococcus elongatus is investigated theoretically. The recently obtained high resolution structure of this complex exhibits an aggregate of 96 chlorophylls that are electronically coupled to function as a light-harvesting antenna complex. This paper constructs an effective Hamiltonian for the chlorophyll aggregate to describe excitation transfer dynamics and spectral properties of PSI. For this purpose, a new kinetic expansion method, the sojourn expansion, is introduced. Our study shows that at room temperature fluctuations of site energies have little effect on the calculated excitation lifetime and quantum yield, which compare favorably with experimental results. The efficiency of the system is fo...

  16. Influence of calcium ion on photosystem Ⅱ oxygen evolution

    Institute of Scientific and Technical Information of China (English)

    杜林方; 孙逊; 潘用华; 林宏辉; 梁厚果

    1995-01-01

    Treatment of photosystem Ⅱ particles with NaCl-washings,low-pH washings or detergentOG-solubilizings inhibited oxygen evolution and the inhibition was reversed by addition of exogenous Ca2+.Dynamic analysis with Ca2+reconstitution revealed two Ca2+binding sites with different affinities in theNaCl-washed PS Ⅱ particles or in the low-pH-treated ones.Oxygen-evolving PS Ⅱ core complex also con-tained the high and low affinity Ca2+binding sites.Ca2+enhanced the intensity of fluorescence emission of PSⅡ core complex.These results suggest that calcium play two roles in PS Ⅱ,the low affinity Ca2+is associ-ated with energy transfer while the high affinity Ca2+is concerned with water splitting reaction.

  17. Enhanced photocurrent production by photosystem I with modified viologen derivatives

    Energy Technology Data Exchange (ETDEWEB)

    Yamanoi, Yoshinori, E-mail: yamanoi@chem.s.u-tokyo.ac.jp [Department of Chemistry, School of Science, University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033 (Japan); Terasaki, Nao [National Institute of Advanced Industrial Science and Technology (AIST), 807-1 Shuku-machi, Tosu, Saga 841-0052 (Japan); Miyachi, Mariko [Department of Chemistry, School of Science, University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033 (Japan); Inoue, Yasunori [Department of Applied Biological Science, Faculty of Science and Technology, Tokyo University of Science, Yamazaki 2641, Noda, Chiba 278-8510 (Japan); Nishihara, Hiroshi, E-mail: nisihara@chem.s.u-tokyo.ac.jp [Department of Chemistry, School of Science, University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033 (Japan)

    2012-06-01

    This paper describes the construction of a photogenerated electron transfer system based on photosystem I (PSI) and modified molecular wires. Gold electrode surfaces were functionalized with sodium 3-mercapto-1-propanesulfonate and the 2,2 Prime -dimethyl-4,4 Prime -bipyridine cation to form a self-assembled monolayer, which enabled the subsequent immobilization of PSI. A higher photocurrent was observed in the presence of sodium ascorbate as an electron donor under irradiation at 680 nm. - Highlights: Black-Right-Pointing-Pointer Molecular wire was prepared by stepwise alkylation of 2,2 Prime -dimethyl-4,4 Prime -bipridine. Black-Right-Pointing-Pointer Oxidation potentials of the modified molecular wire shifted negatively. Black-Right-Pointing-Pointer Modified gold electrode showed a higher current density than that of previous system.

  18. Phytotoxicity of four photosystem II herbicides to tropical seagrasses.

    Directory of Open Access Journals (Sweden)

    Florita Flores

    Full Text Available Coastal waters of the Great Barrier Reef (GBR are contaminated with agricultural pesticides, including the photosystem II (PSII herbicides which are the most frequently detected at the highest concentrations. Designed to control weeds, these herbicides are equally potent towards non-target marine species, and the close proximity of seagrass meadows to flood plumes has raised concerns that seagrasses may be the species most threatened by herbicides from runoff. While previous work has identified effects of PSII herbicides on the photophysiology, growth and mortality in seagrass, there is little comparative quantitative toxicity data for seagrass. Here we applied standard ecotoxicology protocols to quantify the concentrations of four priority PSII herbicides that inhibit photochemistry by 10, 20 and 50% (IC10, IC20 and IC50 over 72 h in two common seagrass species from the GBR lagoon. The photosystems of seagrasses Zosteramuelleri and Haloduleuninervis were shown to be generally more sensitive to the PSII herbicides Diuron, Atrazine, Hexazinone and Tebuthiuron than corals and tropical microalgae. The herbicides caused rapid inhibition of effective quantum yield (∆F/F m ', indicating reduced photosynthesis and maximum effective yields (Fv/Fm corresponding to chronic damage to PSII. The PSII herbicide concentrations which affected photosynthesis have been exceeded in the GBR lagoon and all of the herbicides inhibited photosynthesis at concentrations lower than current marine park guidelines. There is a strong likelihood that the impacts of light limitation from flood plumes and reduced photosynthesis from PSII herbicides exported in the same waters would combine to affect seagrass productivity. Given that PSII herbicides have been demonstrated to affect seagrass at environmental concentrations, we suggest that revision of environmental guidelines and further efforts to reduce PSII herbicide concentrations in floodwaters may both help protect

  19. Antenna entropy in plant photosystems does not reduce the free energy for primary charge separation.

    Science.gov (United States)

    Jennings, Robert C; Zucchelli, Giuseppe

    2014-12-01

    We have investigated the concept of the so-called "antenna entropy" of higher plant photosystems. Several interesting points emerge: 1. In the case of a photosystemwhich harbours an excited state, the “antenna entropy” is equivalent to the configurational (mixing) entropy of a thermodynamic canonical ensemble. The energy associated with this parameter has been calculated for a hypothetical isoenergetic photosystem, photosystem I and photosystem II, and comes out in the range of 3.5 - 8% of the photon energy considering 680 nm. 2. The “antenna entropy” seems to be a rather unique thermodynamic phenomenon, in as much as it does not modify the free energy available for primary photochemistry, as has been previously suggested. 3. It is underlined that this configurational (mixing) entropy, unlike heat dispersal in a thermal system, does not involve energy dilution. This points out an important difference between thermal and electronic energy dispersal.

  20. Remodeling of Membrane Lipids in Iron-starved Chlamydomonas*

    Science.gov (United States)

    Urzica, Eugen I.; Vieler, Astrid; Hong-Hermesdorf, Anne; Page, M. Dudley; Casero, David; Gallaher, Sean D.; Kropat, Janette; Pellegrini, Matteo; Benning, Christoph; Merchant, Sabeeha S.

    2013-01-01

    Chlamydomonas reinhardtii cells exposed to abiotic stresses (e.g. nitrogen, zinc, or phosphorus deficiency) accumulate triacylglycerols (TAG), which are stored in lipid droplets. Here, we report that iron starvation leads to formation of lipid droplets and accumulation of TAGs. This occurs between 12 and 24 h after the switch to iron-starvation medium. C. reinhardtii cells deprived of iron have more saturated fatty acid (FA), possibly due to the loss of function of FA desaturases, which are iron-requiring enzymes with diiron centers. The abundance of a plastid acyl-ACP desaturase (FAB2) is decreased to the same degree as ferredoxin. Ferredoxin is a substrate of the desaturases and has been previously shown to be a major target of the iron deficiency response. The increase in saturated FA (C16:0 and C18:0) is concomitant with the decrease in unsaturated FA (C16:4, C18:3, or C18:4). This change was gradual for diacylglyceryl-N,N,N-trimethylhomoserine (DGTS) and digalactosyldiacylglycerol (DGDG), whereas the monogalactosyldiacylglycerol (MGDG) FA profile remained stable during the first 12 h, whereas MGDG levels were decreasing over the same period of time. These changes were detectable after only 2 h of iron starvation. On the other hand, DGTS and DGDG contents gradually decreased until a minimum was reached after 24–48 h. RNA-Seq analysis of iron-starved C. reinhardtii cells revealed notable changes in many transcripts coding for enzymes involved in FA metabolism. The mRNA abundances of genes coding for components involved in TAG accumulation (diacylglycerol acyltransferases or major lipid droplet protein) were increased. A more dramatic increase at the transcript level has been observed for many lipases, suggesting that major remodeling of lipid membranes occurs during iron starvation in C. reinhardtii. PMID:23983122

  1. Remodeling of membrane lipids in iron-starved Chlamydomonas.

    Science.gov (United States)

    Urzica, Eugen I; Vieler, Astrid; Hong-Hermesdorf, Anne; Page, M Dudley; Casero, David; Gallaher, Sean D; Kropat, Janette; Pellegrini, Matteo; Benning, Christoph; Merchant, Sabeeha S

    2013-10-18

    Chlamydomonas reinhardtii cells exposed to abiotic stresses (e.g. nitrogen, zinc, or phosphorus deficiency) accumulate triacylglycerols (TAG), which are stored in lipid droplets. Here, we report that iron starvation leads to formation of lipid droplets and accumulation of TAGs. This occurs between 12 and 24 h after the switch to iron-starvation medium. C. reinhardtii cells deprived of iron have more saturated fatty acid (FA), possibly due to the loss of function of FA desaturases, which are iron-requiring enzymes with diiron centers. The abundance of a plastid acyl-ACP desaturase (FAB2) is decreased to the same degree as ferredoxin. Ferredoxin is a substrate of the desaturases and has been previously shown to be a major target of the iron deficiency response. The increase in saturated FA (C16:0 and C18:0) is concomitant with the decrease in unsaturated FA (C16:4, C18:3, or C18:4). This change was gradual for diacylglyceryl-N,N,N-trimethylhomoserine (DGTS) and digalactosyldiacylglycerol (DGDG), whereas the monogalactosyldiacylglycerol (MGDG) FA profile remained stable during the first 12 h, whereas MGDG levels were decreasing over the same period of time. These changes were detectable after only 2 h of iron starvation. On the other hand, DGTS and DGDG contents gradually decreased until a minimum was reached after 24-48 h. RNA-Seq analysis of iron-starved C. reinhardtii cells revealed notable changes in many transcripts coding for enzymes involved in FA metabolism. The mRNA abundances of genes coding for components involved in TAG accumulation (diacylglycerol acyltransferases or major lipid droplet protein) were increased. A more dramatic increase at the transcript level has been observed for many lipases, suggesting that major remodeling of lipid membranes occurs during iron starvation in C. reinhardtii.

  2. Ultraviolet modification of Chlamydomonas reinhardtii for carbon capture

    Directory of Open Access Journals (Sweden)

    Gopal NS

    2016-04-01

    Full Text Available Nikhil S Gopal,1 K Sudhakar2 1The Lawrenceville School, Lawrenceville, NJ, USA; 2Bioenergy Laboratory, Malauna Azad National Institute of Technology, Bhopal, India Purpose: Carbon dioxide (CO2 levels have been rising rapidly. Algae are single-cell organisms with highly efficient CO2 uptake mechanisms. Algae yield two to ten times more biomass versus terrestrial plants and can grow nearly anywhere. Large scale CO2 sequestration is not yet sustainable due to high amounts of nitrogen (N and phosphate (P needed to grow algae in media. Methods: Mutant strains of Chlamydomonas reinhardtii were created using ultraviolet light (2.2–3 K J/m2 and natural selection using media with 20%–80% lower N and P compared to standard Sueoka's high salt medium. Strains were selected based upon growth in media concentrations varying from 20% to 80% less N/P compared to control. Biomass was compared to wild-type control (CC-125 using direct counts, optical density dry weight, and mean doubling time. Results: Mean doubling time was 20 and 25 hours in the low N and N/P strains, respectively (vs 66 hours in control. Using direct counts, growth rates of mutant strains of low N and N/P cultures were not statistically different from control (P=0.37 and 0.70, respectively. Conclusion: Two new strains of algae, as well as wild-type control, were able to grow while using 20%–40% less N and P. Ultraviolet light-based modification of algae is an inexpensive and alternative option to genetic engineering techniques. This technique might make larger scale biosequestration possible. Keywords: biosequestration, ultraviolet, carbon sequestration, carbon capture, algae

  3. Physiological characterization of cadmium-exposed Chlamydomonas reinhardtii.

    Science.gov (United States)

    Bräutigam, Anja; Schaumlöffel, Dirk; Preud'homme, Hugues; Thondorf, Iris; Wesenberg, Dirk

    2011-12-01

    Chlamydomonas reinhardtii is a common model organism for investigation of metal stress. This green alga produces phytochelatins in the presence of metal ions. The influence of cadmium is of main interest, because it is a strong activator of phytochelatin synthase. Cell wall bound and intracellular cadmium content was determined after exposition to 70 µm CdCl(2), showing the main portion of the metal outside the cell. Nevertheless, imported cadmium was sufficient to cause significant changes in thiolpeptide metabolism and its transcriptional regulation. Modern analytical approaches enable new insights into phytochelatin (PC) distribution. A new rapid and precise UPLC-MS method allowed high-throughput PC quantification in algal samples after 1, 4, 24 and 48 h cadmium stress. Initially, canonic PCs were synthesized in C. reinhardtii during cadmium exposition, but afterwards CysPCs became the major thiolpeptides. Thus, after 48 h the concentration of the PC-isoforms CysPC(2-3) and CysGSH attained between 105 and 199 nmol g(-1) fresh weight (FW), whereas the PC(2-3) concentrations were only 15 nmol g(-1) FW. The relative quantification of γ-glutamyl transpeptidase (γ-GT) mRNA suggests the generation of CysPCs by glutamate cleavage from canonic PCs by γ-GT. Furthermore, a homology model of C. reinhardtii phytochelatin synthase was constructed to verify the use of crystal structures from Anabaena sp. phytochelatin synthase (PCS) for docking studies with canonical PCs and CysPCs. From the difference in energy scores, we hypothesize that CysPC may prevent the synthesis of canonical PCs by blocking the binding pocket. Finally, possible physiological reasons for the high abundance of CysPC compared with their canonic precursors are discussed.

  4. Flagellar coordination in Chlamydomonas cells held on micropipettes.

    Science.gov (United States)

    Rüffer, U; Nultsch, W

    1998-01-01

    The two flagella of Chlamydomonas are known to beat synchronously: During breaststroke beating they are generally coordinated in a bilateral way while in shock responses during undulatory beating coordination is mostly parallel [Rüffer and Nultsch, 1995: Botanica Acta 108:169-276]. Analysis of a great number of shock responses revealed that in undulatory beats also periods of bilateral coordination are found and that the coordination type may change several times during a shock response, without concomitant changes of the beat envelope and the beat period. In normal wt cells no coordination changes are found during breaststroke beating, but only short temporary asynchronies: During 2 or 3 normal beats of the cis flagellum, the trans flagellum performs 3 or 4 flat beats with a reduced beat envelope and a smaller beat period, resulting in one additional trans beat. Long periods with flat beats of the same shape and beat period are found in both flagella of the non-phototactic mutant ptx1 and in defective wt 622E cells. During these periods, the coordination is parallel, the two flagella beat alternately. A correlation between normal asynchronous trans beats and the parallel-coordinated beats in the presumably cis defective cells and also the undulatory beats is discussed. In the cis defective cells, a perpetual spontaneous change between parallel beats with small beat periods (higher beat frequency) and bilateral beats with greater beat periods (lower beat frequency) are observed and render questionable the existence of two different intrinsic beat frequencies of the two flagella cis and trans. Asynchronies occur spontaneously but may also be induced by light changes, either step-up or step-down, but not by both stimuli in turn as breaststroke flagellar photoresponses (BFPRs). Asynchronies are not involved in phototaxis. They are independent of the BFPRs, which are supposed to be the basis of phototaxis. Both types of coordination must be assumed to be regulated

  5. Adaptation prevents the extinction of Chlamydomonas reinhardtii under toxic beryllium

    Directory of Open Access Journals (Sweden)

    Beatriz Baselga-Cervera

    2016-03-01

    Full Text Available The current biodiversity crisis represents a historic challenge for natural communities: the environmental rate of change exceeds the population’s adaptation capability. Integrating both ecological and evolutionary responses is necessary to make reliable predictions regarding the loss of biodiversity. The race against extinction from an eco-evolutionary perspective is gaining importance in ecological risk assessment. Here, we performed a classical study of population dynamics—a fluctuation analysis—and evaluated the results from an adaption perspective. Fluctuation analysis, widely used with microorganisms, is an effective empirical procedure to study adaptation under strong selective pressure because it incorporates the factors that influence demographic, genetic and environmental changes. The adaptation of phytoplankton to beryllium (Be is of interest because human activities are increasing the concentration of Be in freshwater reserves; therefore, predicting the effects of human-induced pollutants is necessary for proper risk assessment. The fluctuation analysis was performed with phytoplankton, specifically, the freshwater microalgae Chlamydomonas reinhardtii, under acute Be exposure. High doses of Be led to massive microalgae death; however, by conducting a fluctuation analysis experiment, we found that C. reinhardtii was able to adapt to 33 mg/l of Be due to pre-existing genetic variability. The rescuing adapting genotype presented a mutation rate of 9.61 × 10−6 and a frequency of 10.42 resistant cells per million wild-type cells. The genetic adaptation pathway that was experimentally obtained agreed with the theoretical models of evolutionary rescue (ER. Furthermore, the rescuing genotype presented phenotypic and physiologic differences from the wild-type genotype, was 25% smaller than the Be-resistant genotype and presented a lower fitness and quantum yield performance. The abrupt distinctions between the wild-type and the Be

  6. Light saturation curves show competence of the water splitting complex in inactive Photosystem II reaction centers.

    Science.gov (United States)

    Nedbal, L; Gibas, C; Whitmarsh, J

    1991-12-01

    Photosystem II complexes of higher plants are structurally and functionally heterogeneous. While the only clearly defined structural difference is that Photosystem II reaction centers are served by two distinct antenna sizes, several types of functional heterogeneity have been demonstrated. Among these is the observation that in dark-adapted leaves of spinach and pea, over 30% of the Photosystem II reaction centers are unable to reduce plastoquinone to plastoquinol at physiologically meaningful rates. Several lines of evidence show that the impaired reaction centers are effectively inactive, because the rate of oxidation of the primary quinone acceptor, QA, is 1000 times slower than in normally active reaction centers. However, there are conflicting opinions and data over whether inactive Photosystem II complexes are capable of oxidizing water in the presence of certain artificial electron acceptors. In the present study we investigated whether inactive Photosystem II complexes have a functional water oxidizing system in spinach thylakoid membranes by measuring the flash yield of water oxidation products as a function of flash intensity. At low flash energies (less that 10% saturation), selected to minimize double turnovers of reaction centers, we found that in the presence of the artificial quinone acceptor, dichlorobenzoquinone (DCBQ), the yield of proton release was enhanced 20±2% over that observed in the presence of dimethylbenzoquinone (DMBQ). We argue that the extra proton release is from the normally inactive Photosystem II reaction centers that have been activated in the presence of DCBQ, demonstrating their capacity to oxidize water in repetitive flashes, as concluded by Graan and Ort (Biochim Biophys Acta (1986) 852: 320-330). The light saturation curves indicate that the effective antenna size of inactive reaction centers is 55±12% the size of active Photosystem II centers. Comparison of the light saturation dependence of steady state oxygen evolution

  7. Photosystem 0, a proposed ancestral photosystem without reducing power that synthesizes ATP during light-dark cycling

    CERN Document Server

    Muller, A W J

    2005-01-01

    Photosystem 0 is a proposed membrane-based mechanism for free energy gain as ATP from fluctuating light. The PS0 reaction centers have no reducing power: charge transport is only temporary. Light induces within the reaction centers metastable dipoles that generate a membrane potential that drives ATP synthesis while protons move through the ATPsynthase enzyme. After the decay of the dipole potential in the dark, the protons either (1) return across the membrane by conduction or (2) are pumped back by ATPsynthase, backwards active as ATPase at a higher H+/ATP ratio. PS0 constitutes a link to previously proposed free energy sources for early evolution that work on thermal cycling. Several contemporary photosynthetic phenomena may be relics of PS0.

  8. Constitution and energetics of photosystem I and photosystem II in the chlorophyll d-dominated cyanobacterium Acaryochloris marina.

    Science.gov (United States)

    Tomo, Tatsuya; Allakhverdiev, Suleyman I; Mimuro, Mamoru

    2011-01-01

    This mini review presents current topics of discussion about photosystem (PS) I and PS II of photosynthesis in the Acaryochloris marina. A. marina is a photosynthetic cyanobacterium in which chlorophyll (Chl) d is the major antenna pigment (>95%). However, Chl a is always present in a few percent. Chl d absorbs light with a wavelength up to 30 nm red-shifted from Chl a. Therefore, the chlorophyll species of the special pair in PS II has been a matter of debate because if Chl d was the special pair component, the overall energetics must be different in A. marina. The history of this field indicates that a purified sample is necessary for the reliable identification and characterization of the special pair. In view of the spectroscopic data and the redox potential of pheophytin, we discuss the nature of special pair constituents and the localization of the enigmatic Chl a.

  9. Effects of light, food availability and temperature stress on the function of photosystem II and photosystem I of coral symbionts.

    Directory of Open Access Journals (Sweden)

    Mia O Hoogenboom

    Full Text Available BACKGROUND: Reef corals are heterotrophic coelenterates that achieve high productivity through their photosynthetic dinoflagellate symbionts. Excessive seawater temperature destabilises this symbiosis and causes corals to "bleach," lowering their photosynthetic capacity. Bleaching poses a serious threat to the persistence of coral reefs on a global scale. Despite expanding research on the causes of bleaching, the mechanisms remain a subject of debate. METHODOLOGY/PRINCIPAL FINDINGS: This study determined how light and food availability modulate the effects of temperature stress on photosynthesis in two reef coral species. We quantified the activities of Photosystem II, Photosystem I and whole chain electron transport under combinations of normal and stressful growth temperatures, moderate and high light levels and the presence or absence of feeding of the coral hosts. Our results show that PS1 function is comparatively robust against temperature stress in both species, whereas PS2 and whole chain electron transport are susceptible to temperature stress. In the symbiotic dinoflagellates of Stylophora pistillata the contents of chlorophyll and major photosynthetic complexes were primarily affected by food availability. In Turbinaria reniformis growth temperature was the dominant influence on the contents of the photosynthetic complexes. In both species feeding the host significantly protected photosynthetic function from high temperature stress. CONCLUSIONS/SIGNIFICANCE: Our findings support the photoinhibition model of coral bleaching and demonstrate that PS1 is not a major site for thermal damage during bleaching events. Feeding mitigates bleaching in two scleractinian corals, so that reef responses to temperature stresses will likely be influenced by the coinciding availabilities of prey for the host.

  10. Rapid induction of lipid droplets in Chlamydomonas reinhardtii and Chlorella vulgaris by Brefeldin A.

    Directory of Open Access Journals (Sweden)

    Sangwoo Kim

    Full Text Available Algal lipids are the focus of intensive research because they are potential sources of biodiesel. However, most algae produce neutral lipids only under stress conditions. Here, we report that treatment with Brefeldin A (BFA, a chemical inducer of ER stress, rapidly triggers lipid droplet (LD formation in two different microalgal species, Chlamydomonas reinhardtii and Chlorella vulgaris. LD staining using Nile red revealed that BFA-treated algal cells exhibited many more fluorescent bodies than control cells. Lipid analyses based on thin layer chromatography and gas chromatography revealed that the additional lipids formed upon BFA treatment were mainly triacylglycerols (TAGs. The increase in TAG accumulation was accompanied by a decrease in the betaine lipid diacylglyceryl N,N,N-trimethylhomoserine (DGTS, a major component of the extraplastidic membrane lipids in Chlamydomonas, suggesting that at least some of the TAGs were assembled from the degradation products of membrane lipids. Interestingly, BFA induced TAG accumulation in the Chlamydomonas cells regardless of the presence or absence of an acetate or nitrogen source in the medium. This effect of BFA in Chlamydomonas cells seems to be due to BFA-induced ER stress, as supported by the induction of three homologs of ER stress marker genes by the drug. Together, these results suggest that ER stress rapidly triggers TAG accumulation in two green microalgae, C. reinhardtii and C. vulgaris. A further investigation of the link between ER stress and TAG synthesis may yield an efficient means of producing biofuel from algae.

  11. Effect of O2:CO2 Ratio on the Primary Metabolism of Chlamydomonas reinhardtii

    NARCIS (Netherlands)

    Kliphuis, A.M.J.; Martens, D.E.; Janssen, M.G.J.; Wijffels, R.H.

    2011-01-01

    High oxygen:carbon dioxide ratios may have a negative effect on growth and productivity of microalgae. To investigate the effect of O2 and CO2 concentrations and the ratio between these on the metabolism of Chlamydomonas reinhardtii we performed turbidostat experiments at different O2:CO2 ratios. Th

  12. Phytochelatins do not correlate with the level of Cd accumulation in Chlamydomonas spp.

    Science.gov (United States)

    Nishikawa, Kahoko; Onodera, Ai; Tominaga, Noriko

    2006-06-01

    Chlamydomonas acidophila KT-1 and Chlamydomonas acidophila DVB238 exhibit a strong heavy metal tolerance, but C. acidophila DVB238 can accumulate a much higher amount of Cadmium (Cd) than C. acidophila KT-1. Phytochelatins (PCs) are known to play an important role in the detoxification of several toxic heavy metals, but the relationship between PCs and Cd accumulation is not clear. PC metabolism and Cd accumulation were investigated by using three Chlamydomonas strains including Chlamydomonas reinhardtii C-9 as a standard alga. The results showed that the PC content did not correlate closely with the level of Cd accumulation, maintenance of a high GSH level seeming to be more important for Cd accumulation. The ultrastructure of C. acidophila KT-1 was extremely disrupted by a great increase in starch granules, which resulted in a moribund state, but hyper-accumulator C. acidophila DVB238 did not exhibit an increase in starch granules in its cells, in spite of Cd accumulation in its chloroplasts, cytosol and vacuoles. These results indicated that C. acidophila DVB238 probably has a developed detoxification system preventing such as destruction of the cells due to Cd toxicity.

  13. Mastoparan-induced programmed cell death in the unicellular alga Chlamydomonas reinhardtti

    NARCIS (Netherlands)

    Yordanova, Z.P.; Woltering, E.J.; Kapchina-Toteva, V.M.; Iakimova, E.T.

    2013-01-01

    The present study was focused on the elucidation of stress-induced cell death signaling events in the unicellular alga Chlamydomonas reinhardtii exposed to treatment with wasp venom mastoparan. By applying pharmacological approach with specific inhibitors, we have investigated the involvement of eth

  14. Environmental pH and the requirement for the extrinsic proteins of Photosystem II in the function of cyanobacterial photosynthesis

    Directory of Open Access Journals (Sweden)

    Jaz N Morris

    2016-08-01

    Full Text Available In one of the final stages of cyanobacterial Photosystem II (PS II assembly, binding of up to four extrinsic proteins to PS II stabilizes the oxygen-evolving complex (OEC. Growth of cyanobacterial mutants deficient in certain combinations of these thylakoid-lumen-associated polypeptides is sensitive to changes in environmental pH, despite the physical separation of the membrane-embedded PS II complex from the external environment. In this perspective, we discuss the effect of environmental pH on OEC function and photoautotrophic growth in cyanobacteria, with reference to pH-sensitive PS II mutants lacking extrinsic proteins. We consider the possibilities that, compared to pH 10.0, pH 7.5 increases susceptibility to PS II-generated reactive oxygen species (ROS, causing photoinhibition and reducing PS II assembly in some mutants and that perturbations to channels in the lumenal regions of PS II might alter the accessibility of water to the active site, and egress of oxygen and protons to the thylakoid lumen. Reduced levels of PS II in these mutants and reduced OEC activity arising from the disruption of substrate/product channels could reduce the trans-thylakoid pH gradient (ΔpH, leading to the impairement of photosynthesis. Growth of some PS II mutants at pH 7.5 can be rescued by elevating CO2 levels, suggesting that the pH-sensitive phenotype might primarily be an indirect result of back-pressure in the electron transport chain that results in heightened production of ROS by the impaired photosystem.

  15. X-Ray Fiber Diffraction Recordings from Oriented Demembranated Chlamydomonas Flagellar Axonemes.

    Science.gov (United States)

    Toba, Shiori; Iwamoto, Hiroyuki; Kamimura, Shinji; Oiwa, Kazuhiro

    2015-06-16

    The high homology of its axonemal components with humans and a large repertoire of axonemal mutants make Chlamydomonas a useful model system for experiments on the structure and function of eukaryotic cilia and flagella. Using this organism, we explored the spatial arrangement of axonemal components under physiological conditions by small-angle x-ray fiber diffraction. Axonemes were oriented in physiological solution by continuous shear flow and exposed to intense and stable x rays generated in the synchrotron radiation facility SPring-8, BL45XU. We compared diffraction patterns from axonemes isolated from wild-type and mutant strains lacking the whole outer arm (oda1), radial spoke (pf14), central apparatus (pf18), or the α-chain of the outer arm dynein (oda11). Diffraction of the axonemes showed a series of well-defined meridional/layer-line and equatorial reflections. Diffraction patterns from mutant axonemes exhibited a systematic loss/attenuation of meridional/layer-line reflections, making it possible to determine the origin of various reflections. The 1/24 and 1/12 nm(-1) meridional reflections of oda1 and oda11 were much weaker than those of the wild-type, suggesting that the outer dynein arms are the main contributor to these reflections. The weaker 1/32 and 1/13.7 nm(-1) meridional reflections from pf14 compared with the wild-type suggest that these reflections come mainly from the radial spokes. The limited contribution of the central pair apparatus to the diffraction patterns was confirmed by the similarity between the patterns of the wild-type and pf18. The equatorial reflections were complex, but a comparison with electron micrograph-based models allowed the density of each axonemal component to be estimated. Addition of ATP to rigor-state axonemes also resulted in subtle changes in equatorial intensity profiles, which could report nucleotide-dependent structural changes of the dynein arms. The first detailed description of axonemal reflections

  16. Dynamics of the Special Pair of Chlorophylls of Photosystem II.

    Science.gov (United States)

    Narzi, Daniele; Bovi, Daniele; De Gaetano, Pietro; Guidoni, Leonardo

    2016-01-13

    Cholophylls are at the basis of the photosynthetic energy conversion mechanisms in algae, plants, and cyanobacteria. In photosystem II, the photoproduced electrons leave a special pair of chlorophylls (namely, P(D1) and P(D2)) that becomes cationic. This oxidizing pair [P(D1),P(D2)](+), in turn, triggers a cascade of oxidative events, eventually leading to water splitting and oxygen evolution. In the present work, using quantum mechanics/molecular mechanics calculations, we investigate the electronic structure and the dynamics of the P(D1)P(D2) special pair in both its oxidized and reduced states. In agreement with previously reported static calculations, the symmetry between the two chlorophylls was found to be broken, the positive charge being preferentially located on P(D1). Nevertheless, this study reveals for the first time that large charge fluctuations occur along dynamics, temporarily inverting the charge preference for the two branches. Finally, a vibrational analysis pinpointed that such charge fluctuations are strongly coupled to specific modes of the special pair.

  17. Isolation and Characterization of Photosystem Ⅱ of Porphyra yezoensis Ueda

    Institute of Scientific and Technical Information of China (English)

    Zheng-Quan GAO; Guang-Ce WANG; Cheng-Kui TSENG

    2004-01-01

    The thylakoid membranes were isolated and purified from gametophyte ofPorphyra yezoensis Ueda (P. yezoensis) by sucrose density gradient ultracentrifugation. After P.yezoensis gametophyte thylakoid membranes were solubilized with SDS, the photosystem Ⅱ (PSII) particles were isolated and purified. The activity of PSII particles was determined with DCIP (2,6-dichloroindophenol) photoreduction reaction. The composition of purified PSII particles was detected by SDS-PAGE. As a result, seven proteins including 55 kD protein, 47 kD protein, 43 kD protein, 33 kD protein, 31 kD protein, 29 kD protein, and 18 kD protein were found. Compared with PSII particles of higher plants and other algae, they were identified as D1/D2 complex, CP47, CP43, 33 kD protein, D1, D2 and cyt c-550 respectively. Besides, other three new proteins of 20 kD, 16 kD and 14 kD respectively were found. Among these extrinsic proteins, the 16 kD and 14 kD proteins had not been reported previously, and the 20 kD protein was found for the first time in multicellular red algae.

  18. Structure/Function/Dynamics of Photosystem II Plastoquinone Binding Sites

    Science.gov (United States)

    Lambreva, Maya D.; Russo, Daniela; Polticelli, Fabio; Scognamiglio, Viviana; Antonacci, Amina; Zobnina, Veranika; Campi, Gaetano; Rea, Giuseppina

    2014-01-01

    Photosystem II (PSII) continuously attracts the attention of researchers aiming to unravel the riddle of its functioning and efficiency fundamental for all life on Earth. Besides, an increasing number of biotechnological applications have been envisaged exploiting and mimicking the unique properties of this macromolecular pigment-protein complex. The PSII organization and working principles have inspired the design of electrochemical water splitting schemes and charge separating triads in energy storage systems as well as biochips and sensors for environmental, agricultural and industrial screening of toxic compounds. An intriguing opportunity is the development of sensor devices, exploiting native or manipulated PSII complexes or ad hoc synthesized polypeptides mimicking the PSII reaction centre proteins as bio-sensing elements. This review offers a concise overview of the recent improvements in the understanding of structure and function of PSII donor side, with focus on the interactions of the plastoquinone cofactors with the surrounding environment and operational features. Furthermore, studies focused on photosynthetic proteins structure/function/dynamics and computational analyses aimed at rational design of high-quality bio-recognition elements in biosensor devices are discussed. PMID:24678671

  19. (Unraveling photosystems): Progress report, July 1, 1982-June 30, 1983

    Energy Technology Data Exchange (ETDEWEB)

    Bogorad, L.

    1983-01-01

    The overall objective of this program is to identify and characterize genes for components of the photosynthetic apparatus - especially genes for components of photosystem II (PS II). During the past year two atrazine resistant mutants of the blue-green alga Anacystis nidulans R-2 have been isolated and partially characterized. Thylakoid membranes have been prepared from these cells as well as from wild type cells and their rates of oxygen evolution measured in the presence and absence of atrazine. The I/sub 50/ for the two mutants was approximately 2 x 10/sup -8/M whereas for wild type cells it was 7.5 x 10/sup -10/M. Visible absorption and fluorescence emission spectra of the mutant and wild type cells have been compared. One of the mutants possesses spectra similar to the wild type while the second mutant does not. The latter mutant appears to have increased absorption peaks due to phycocyanin and allophycocyanin. DNA has been purified from both atrazine resistant mutants and used to transform wild type cells to atrazine resistance. Transformation of cells to the resistant phenotype is reproducible although the frequency of transformation varies. ''Mutant DNA'' subjected to restriction enzyme cleavage prior to use for transformation yields transformants but the frequency is generally about one order of magnitude lower than with uncut DNA. 54 refs., 3 figs.

  20. Functional Implications of Photosystem II Crystal Formation in Photosynthetic Membranes*

    Science.gov (United States)

    Tietz, Stefanie; Puthiyaveetil, Sujith; Enlow, Heather M.; Yarbrough, Robert; Wood, Magnus; Semchonok, Dmitry A.; Lowry, Troy; Li, Zhirong; Jahns, Peter; Boekema, Egbert J.; Lenhert, Steven; Niyogi, Krishna K.; Kirchhoff, Helmut

    2015-01-01

    The structural organization of proteins in biological membranes can affect their function. Photosynthetic thylakoid membranes in chloroplasts have the remarkable ability to change their supramolecular organization between disordered and semicrystalline states. Although the change to the semicrystalline state is known to be triggered by abiotic factors, the functional significance of this protein organization has not yet been understood. Taking advantage of an Arabidopsis thaliana fatty acid desaturase mutant (fad5) that constitutively forms semicrystalline arrays, we systematically test the functional implications of protein crystals in photosynthetic membranes. Here, we show that the change into an ordered state facilitates molecular diffusion of photosynthetic components in crowded thylakoid membranes. The increased mobility of small lipophilic molecules like plastoquinone and xanthophylls has implications for diffusion-dependent electron transport and photoprotective energy-dependent quenching. The mobility of the large photosystem II supercomplexes, however, is impaired, leading to retarded repair of damaged proteins. Our results demonstrate that supramolecular changes into more ordered states have differing impacts on photosynthesis that favor either diffusion-dependent electron transport and photoprotection or protein repair processes, thus fine-tuning the photosynthetic energy conversion. PMID:25897076

  1. Functional Implications of Photosystem II Crystal Formation in Photosynthetic Membranes.

    Science.gov (United States)

    Tietz, Stefanie; Puthiyaveetil, Sujith; Enlow, Heather M; Yarbrough, Robert; Wood, Magnus; Semchonok, Dmitry A; Lowry, Troy; Li, Zhirong; Jahns, Peter; Boekema, Egbert J; Lenhert, Steven; Niyogi, Krishna K; Kirchhoff, Helmut

    2015-05-29

    The structural organization of proteins in biological membranes can affect their function. Photosynthetic thylakoid membranes in chloroplasts have the remarkable ability to change their supramolecular organization between disordered and semicrystalline states. Although the change to the semicrystalline state is known to be triggered by abiotic factors, the functional significance of this protein organization has not yet been understood. Taking advantage of an Arabidopsis thaliana fatty acid desaturase mutant (fad5) that constitutively forms semicrystalline arrays, we systematically test the functional implications of protein crystals in photosynthetic membranes. Here, we show that the change into an ordered state facilitates molecular diffusion of photosynthetic components in crowded thylakoid membranes. The increased mobility of small lipophilic molecules like plastoquinone and xanthophylls has implications for diffusion-dependent electron transport and photoprotective energy-dependent quenching. The mobility of the large photosystem II supercomplexes, however, is impaired, leading to retarded repair of damaged proteins. Our results demonstrate that supramolecular changes into more ordered states have differing impacts on photosynthesis that favor either diffusion-dependent electron transport and photoprotection or protein repair processes, thus fine-tuning the photosynthetic energy conversion.

  2. Directed mutagenesis as a probe of photosystem II function

    Energy Technology Data Exchange (ETDEWEB)

    Bowlby, N.; Sithole, I.; McIntosh, L. (MSU-DOE Plant Research Laboratory, East Lansing, MI (USA)); James, J.; Babcock, G. (Michigan State Univ., East Lansing (USA))

    1990-05-01

    Several mutants of Synechocystis 6803 have been generated by site-directed mutagenesis of psbA and psbD in order to study the roles of the D1 and D2 proteins during H{sub 2}O oxidation. Of key importance to these studies is the analysis of PSII electron transfer components such as Y{sub Z}{sup +} and Y{sub D}{sup +}, and the identification of possible ligands to Mn and Ca. Detailed information can be gained through EPR spectroscopy, however, sensitivity is limited by the PSII concentration of the sample. A simple procedure for the purification of PSII from this cyanobacterium, in sufficient amounts for EPR studies, will be presented. Preliminary results from mutant analysis extend the characterization of the two tyrosine radicals and suggest an important structural role for Pro{sup 162} of D1 and Pro{sup 161} of D2 in the ability of the photosystem to form the radical species on the adjacent tyrosine residues Tyr{sup 161} and Tyr{sup 160} on D1 and D2 respectively.

  3. Protein film photoelectrochemistry of the water oxidation enzyme photosystem II.

    Science.gov (United States)

    Kato, Masaru; Zhang, Jenny Z; Paul, Nicholas; Reisner, Erwin

    2014-09-21

    Photosynthesis is responsible for the sunlight-powered conversion of carbon dioxide and water into chemical energy in the form of carbohydrates and the release of O2 as a by-product. Although many proteins are involved in photosynthesis, the fascinating machinery of Photosystem II (PSII) is at the heart of this process. This tutorial review describes an emerging technique named protein film photoelectrochemistry (PF-PEC), which allows for the light-dependent activity of PSII adsorbed onto an electrode surface to be studied. The technique is straightforward to use, does not require highly specialised and/or expensive equipment, is highly selective for the active fractions of the adsorbed enzyme, and requires a small amount of enzyme sample. The use of PF-PEC to study PSII can yield insights into its activity, stability, quantum yields, redox behaviour, and interfacial electron transfer pathways. It can also be used in PSII inhibition studies and chemical screening, which may prove useful in the development of biosensors. PSII PF-PEC cells also serve as proof-of-principle solar water oxidation systems; here, a comparison is made against PSII-inspired synthetic photocatalysts and materials for artificial photosynthesis.

  4. ApcD is necessary for efficient energy transfer from phycobilisomes to photosystem I and helps to prevent photoinhibition in the cyanobacterium Synechococcus sp. PCC 7002.

    Science.gov (United States)

    Dong, Chunxia; Tang, Aihui; Zhao, Jindong; Mullineaux, Conrad W; Shen, Gaozhong; Bryant, Donald A

    2009-09-01

    Phycobilisomes (PBS) are the major light-harvesting, protein-pigment complexes in cyanobacteria and red algae. PBS absorb and transfer light energy to photosystem (PS) II as well as PS I, and the distribution of light energy from PBS to the two photosystems is regulated by light conditions through a mechanism known as state transitions. In this study the quantum efficiency of excitation energy transfer from PBS to PS I in the cyanobacterium Synechococcus sp. PCC 7002 was determined, and the results showed that energy transfer from PBS to PS I is extremely efficient. The results further demonstrated that energy transfer from PBS to PS I occurred directly and that efficient energy transfer was dependent upon the allophycocyanin-B alpha subunit, ApcD. In the absence of ApcD, cells were unable to perform state transitions and were trapped in state 1. Action spectra showed that light energy transfer from PBS to PS I was severely impaired in the absence of ApcD. An apcD mutant grew more slowly than the wild type in light preferentially absorbed by phycobiliproteins and was more sensitive to high light intensity. On the other hand, a mutant lacking ApcF, which is required for efficient energy transfer from PBS to PS II, showed greater resistance to high light treatment. Therefore, state transitions in cyanobacteria have two roles: (1) they regulate light energy distribution between the two photosystems; and (2) they help to protect cells from the effects of light energy excess at high light intensities.

  5. Ultraviolet-B induced damage to photosystem II in intact filaments of Spirulina platensis

    Energy Technology Data Exchange (ETDEWEB)

    Kolli, B.K.; Tiwari, S.; Mohanty, P. [Jawaharlal Nehru Univ., New Delhi (India). Schoof of Life Sciences

    1998-05-01

    When Spirulina platensis filaments were exposed to 0.75 mW x m{sup -2} x s{sup -1} of ultraviolet-B radiation (the ultraviolet-B radiation under clear sky condition is {proportional_to}1.0 mW x m{sup -2} x s{sup -1}), an inhibition in photosystem II activity was observed, the inhibition being 90% after 90 min exposure. Upon exposure to ultraviolet-B, the room temperature emission characteristics of Spirulina cells were altered when excited with light primarily absorbed by chlorophyll a or phycobilisomes. When the cells were exposed for 3 h the emission at 685 nm (F{sub 685}), when excited at 440 nm (primarily chlorophyll a absorption), was enhanced compared to 715 nm (F{sub 715}) band of photosystem I suggesting a decrease in energy transfer from photosystem II to photosystem I. Similarly, when the cells were excited at 580 nm (primarily the phycobilisomes), the ratio of emission intensity at 685 nm (F{sub 685}) to that of 655 nm (F{sub 655}) was decreased in the exposed cells. This change in emission characteristics seems to be linked with the uncoupling of the energy transfer from all phycocyanin to chlorophyll a of photosystem II. A small shift in emission peak positions was also indicated when excited either at 440 nm or 580 nm. (orig./AJ)

  6. Target and specificity of a nuclear gene product that participates in mRNA 3'-end formation in Chlamydomonas chloroplasts.

    Science.gov (United States)

    Levy, H; Kindle, K L; Stern, D B

    1999-12-10

    Chloroplast mRNA maturation is catalyzed by nucleus-encoded processing enzymes. We previously described a recessive nuclear mutation (crp3) that affects 3'-end formation of several chloroplast mRNAs in Chlamydomonas reinhardtii (Levy, H., Kindle, K. L., and Stern, D. B. (1997) Plant Cell 9, 825-836). In the crp3 background, atpB mRNA lacking a 3'-inverted repeat normally required for stability accumulates as a discrete transcript. The mutation also affects the atpA gene cluster; polycistronic mRNAs with psbI or cemA 3'-ends accumulate to a lower level in the crp3 background. Here, we demonstrate that the crp3 mutation also alters 3'-end formation of psbI mRNA and cemA-containing mRNAs. A novel 3'-end is formed in monocistronic psbI transcripts, and this is the only terminus observed when the psbI 3'-untranslated region is fused to an aadA reporter gene. Accumulation of mRNAs with 3'-ends between cemA and atpH, which is immediately downstream, was reduced. However, this sequence was not recognized as a 3'-end formation element in chimeric genes. The crp3 mutation was able to confer stability to three different atpB 3'-stem-loop-disrupting mutations that lack sequence similarity, but are located at a similar distance from the translation termination codon. We propose that the wild-type CRP3 gene product is part of the general 3' --> 5' processing machinery.

  7. The energy balance of the biomass generation of Chlamydomonas acidophila under acidic and neutral conditions and Chlamydomonas reinhardtii; Die Energiebilanz der Biomasseneubildung von Chlamydomonas acidophila unter sauren und neutralen Bedingungen und von Chlamydomonas reinhardtii

    Energy Technology Data Exchange (ETDEWEB)

    Langner, Uwe

    2009-01-16

    In this study the influence of pH < 3 as an extreme environment has been investigated for the eukaryotic green alga Chlamydomonas (C.) acidophila. The limited number of trophic levels, consisting of bacteria, phytoplankton, zooplankton and macrophytes, is a special characteristic of extreme acidic water bodies. C. acidophila was isolated from an extreme acidic mining lake (RL 111) (Bissinger et al. 2000). A special feature of the examined algal species is its wide tolerance range of external pH values from 2 to 7 (Cassin 1874, Gerloff-Elias et al. 2005a). C. acidophila is a dominant species in the acidic mining lakes, it can grow up to chlorophyll maxima of 500 {mu}g L{sup -1} during the summer time (Nixdorf et al. 1998, 2003). The alga can be found elsewhere in extreme acidic water bodies around the world. The hydrochemistry of the acidic mining lakes in the central regions of Germany and Lusatia show clear differences compared to neutral water bodies. Some of the characteristics of acidic mining lakes are high metal and heavy metal (aluminum) concentrations, high ion concentrations, which lead to high conductivity, as well as low phosphate concentrations, ammonium as the only nitrogen source and CO{sub 2} as the only inorganic carbon source. Many eukaryotic microalgae in acidic water bodies including C. acidophila show a neutral cytosolic pH. This is provided by special adaptations of the organisms. Thus, organisms in extreme acidic environments have a positive cell surface charge, a very efficient H{sup +}-ATPase and high internal buffer capacities. These mechanisms work contrary to the proton influx and the acidification of the cytosol and are therefore proof for the physiological impact of osmoregulation by microalgae in extreme acidic environments (Sekler et al.1991, Weiss et al. 1999). Hence, these mechanisms also complicate the access to nutrients for the algal cell. The passive influx of positively charged ions such as potassium or ammonium is reduced by

  8. Phototaxis beyond turning: persistent accumulation and response acclimation of the microalga Chlamydomonas reinhardtii

    CERN Document Server

    Arrieta, Jorge; Chioccioli, Maurizio; Polin, Marco; Tuval, Idan

    2016-01-01

    Phototaxis is an important reaction to light displayed by a wide range of motile microorganisms. Flagellated eukaryotic microalgae in particular, like the model organism Chlamydomonas reinhardtii, steer either towards or away from light by a rapid and precisely timed modulation of their flagellar activity. Cell steering, however, is only the beginning of a much longer process which ultimately allows cells to determine their light exposure history. This process is not well understood. Here we present a first quantitative study of the long timescale phototactic motility of Chlamydomonas at both single cell and population levels. Our results reveal that the phototactic strategy adopted by these microorganisms leads to an efficient exposure to light, and that the phototactic response is modulated over typical timescales of tens of seconds. The adaptation dynamics for phototaxis and chlorophyll fluorescence show a striking quantitative agreement, suggesting that photosynthesis controls quantitatively how cells nav...

  9. Estimation of oxygen evolution by marine phytoplankton from measurement of the efficiency of Photosystem II electron flow.

    NARCIS (Netherlands)

    Geel, C.; Versluis, W.; Snel, J.F.H.

    1997-01-01

    The relation between photosynthetic oxygen evolution and Photosystem II electron transport was investigated for the marine algae t Phaeodactylum tricornutum, Dunaliella tertiolecta, Tetraselmis sp., t Isochrysis sp. and t Rhodomonas sp.. The rate of Photosystem II electron transport was estimated fr

  10. Lacking power impairs executive functions

    NARCIS (Netherlands)

    Smith, P.K.; Jostmann, N.B.; Galinsky, A.D.; Dijk, W.W. van

    2008-01-01

    Four experiments explored whether lacking power impairs executive functioning, testing the hypothesis that the cognitive presses of powerlessness increase vulnerability to performance decrements during complex executive tasks. In the first three experiments, low power impaired performance on executi

  11. Expression and molecular analysis of phbB gene in Chlamydomonas reinhardtii

    Institute of Scientific and Technical Information of China (English)

    WANG Chaogang; HU Zhangli; HU Wei; LEI Anping

    2004-01-01

    The expression vector containing phbB and ble genes was constructed and transformed into cell-walldeficient strain Chlamydomonas reinhardtii CC-849 by the glass-bead method. The transgenic alga was selected and maintained in the TAP agar plates containing 10 μg/mL Zeomycin. Transgenic alga, which could express phbB at the transcriptional level, was obtained and further confirmed with PCR, Southern blot and RT-PCR-DNA hybridization analysis.

  12. Three-dimensional structure of photosystem II from Thermosynechococcus elongates in complex with terbutryn

    Energy Technology Data Exchange (ETDEWEB)

    Gabdulkhakov, A. G., E-mail: azat@vega.protes.ru; Dontsova, M. V. [Russian Academy of Sciences, Institute of Protein Research (Russian Federation); Saenger, W. [Free University of Berlin, Institute for Chemistry and Biochemistry/Crystallography (Germany)

    2011-11-15

    Photosystem II is a key component of the photosynthetic pathway producing oxygen at the thylakoid membrane of cyanobacteria, green algae, and plants. The three-dimensional structure of photosystem II from the cyanobacterium Thermosynechococcus elongates in a complex with herbicide terbutryn (a photosynthesis inhibitor) was determined for the first time by X-ray diffraction and refined at 3.2 Angstrom-Sign resolution (R{sub factor} = 26.9%, R{sub free} = 29.9%, rmsd for bond lengths is 0.013 Angstrom-Sign , and rmsd for bond angles is 2.2 Degree-Sign ). The terbutryn molecule was located in the binding pocket of the mobile plastoquinone. The atomic coordinates of the refined structure of photosystem II in a complex with terbutryn were deposited in the Protein Data Bank.

  13. Ultrafast infrared observation of exciton equilibration from oriented single crystals of photosystem II

    Science.gov (United States)

    Kaucikas, Marius; Maghlaoui, Karim; Barber, Jim; Renger, Thomas; van Thor, Jasper J.

    2016-12-01

    In oxygenic photosynthesis, two photosystems work in series. Each of them contains a reaction centre that is surrounded by light-harvesting antennae, which absorb the light and transfer the excitation energy to the reaction centre where electron transfer reactions are driven. Here we report a critical test for two contrasting models of light harvesting by photosystem II cores, known as the trap-limited and the transfer-to-the trap-limited model. Oriented single crystals of photosystem II core complexes of Synechococcus elongatus are excited by polarized visible light and the transient absorption is probed with polarized light in the infrared. The dichroic amplitudes resulting from photoselection are maintained on the 60 ps timescale that corresponds to the dominant energy transfer process providing compelling evidence for the transfer-to-the-trap limitation of the overall light-harvesting process. This finding has functional implications for the quenching of excited states allowing plants to survive under high light intensities.

  14. Circular spectropolarimetric sensing of chiral photosystems in decaying leaves

    Science.gov (United States)

    Patty, C. H. Lucas; Visser, Luuk J. J.; Ariese, Freek; Buma, Wybren Jan; Sparks, William B.; van Spanning, Rob J. M.; Röling, Wilfred F. M.; Snik, Frans

    2017-03-01

    Circular polarization spectroscopy has proven to be an indispensable tool in photosynthesis research and (bio)molecular research in general. Oxygenic photosystems typically display an asymmetric Cotton effect around the chlorophyll absorbance maximum with a signal ≤ 1 % . In vegetation, these signals are the direct result of the chirality of the supramolecular aggregates. The circular polarization is thus directly influenced by the composition and architecture of the photosynthetic macrodomains, and is thereby linked to photosynthetic functioning. Although ordinarily measured only on a molecular level, we have developed a new spectropolarimetric instrument, TreePol, that allows for both laboratory and in-the-field measurements. Through spectral multiplexing, TreePol is capable of fast measurements with a sensitivity of ∼ 1 *10-4 and is therefore suitable of non-destructively probing the molecular architecture of whole plant leaves. We have measured the chiroptical evolution of Hedera helix leaves for a period of 22 days. Spectrally resolved circular polarization measurements (450-900 nm) on whole leaves in transmission exhibit a strong decrease in the polarization signal over time after plucking, which we accredit to the deterioration of chiral macro-aggregates. Chlorophyll a levels measured over the same period by means of UV-vis absorption and fluorescence spectroscopy showed a much smaller decrease. With these results we are able to distinguish healthy from deteriorating leaves. Hereby we indicate the potency of circular polarization spectroscopy on whole and intact leaves as a nondestructive tool for structural and plant stress assessment. Additionally, we underline the establishment of circular polarization signals as remotely accessible means of detecting the presence of extraterrestrial life.

  15. Survival of Human Metallothionein-2 Transplastomic Chlamydomonas reinhardtii to Ultraviolet B Exposure

    Institute of Scientific and Technical Information of China (English)

    Yu-Kun ZHANG; Gui-Fang SHEN; Bing-Gen RU

    2006-01-01

    Solar ultraviolet (UV) radiation has a great influence on green organisms, especially plankton like Chlamydomonas. A human metallothionein-2 gene, which is generally considered to have an anti-radiation function by its coding product, was transferred into the chloroplast genome of Chlamydomonas reinhardtii.To dynamically measure the UV effects on Chlamydomonas cells grown in liquid tris-acetate-phosphate medium, a new method was developed based on the relationship between the chlorophyll content of an algal culture and its absorbance at 570 nm after the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. In this experiment, both the wild-type and the transplastomic C. reinhardtii cells were cultivated in 96-well microplates containing liquid tris-acetate-phosphate medium in the absence or presence of zinc, copper,cadmium and cysteine. The transgenic C. reinhardtii showed a higher resistance than wild-type to UV-B exposure under all the examined conditions. Metals in the medium had positive impacts on both types of cells,but had significant influence only on the transplastomic cells. However, the high cell viability of the transgenic alga at the end of the 8 h UV-B treatment disappeared after a 20-h recovery culture. Cysteine did not protect cells from UV-B damage, but clearly enhanced the growth of both wild-type and transgenic C.reinhardtii.

  16. Polyclonal antibodies against the TLA1 protein also recognize with high specificity the D2 reaction center protein of PSII in the green alga Chlamydomonas reinhardtii.

    Science.gov (United States)

    Mitra, Mautusi; Dewez, David; García-Cerdán, Jose Gines; Melis, Anastasios

    2012-04-01

    The Chlamydomonas reinhardtii DNA-insertional transformant truncated light-harvesting antenna 1 (tla1) mutant, helped identify the novel TLA1 gene (GenBank Accession # AF534570-71) as an important genetic determinant in the chlorophyll antenna size of photosynthesis. Down-regulation in the amount of the TLA1 23 kDa protein in the cell resulted in smaller chlorophyll antenna size for both photosystems (in Tetali et al. Planta 225:813-829, 2007). Specific polyclonal antibodies, raised against the recombinant TLA1 protein, showed a cross-reaction with the predicted 23 kDa TLA1 protein in C. reinhardtii protein extracts, but also showed a strong cross-reaction with a protein band migrating to 28.5 kDa. Questions of polymorphism, or posttranslational modification of the TLA1 protein were raised as a result of the unexpected 28.5 kDa cross-reaction. Work in this paper aimed to elucidate the nature of the unexpected 28.5 kDa cross-reaction, as this was deemed to be important in terms of the functional role of the TLA1 protein in the regulation of the chlorophyll antenna size of photosynthesis. Immuno-precipitation of the 28.5 kDa protein, followed by LC-mass spectrometry, showed amino acid sequences ascribed to the psbD/D2 reaction center protein of PSII. The common antigenic determinant between TLA1 and D2 was shown to be a stretch of nine conserved amino acids V-F-L(V)LP-GNAL in the C-terminus of the two proteins, constituting a high antigenicity "GNAL" domain. Antibodies raised against the TLA1 protein containing this domain recognized both the TLA1 and the D2 protein. Conversely, antibodies raised against the TLA1 protein minus the GNAL domain specifically recognized the 23 kDa TLA1 protein and failed to recognize the 28.5 kDa D2 protein. D2 antibodies raised against an oligopeptide containing this domain also cross-reacted with the TLA1 protein. It is concluded that the 28.5 kDa cross-reaction of C. reinhardtii protein extracts with antiTLA1 antibodies is due to

  17. D1-arginine257 mutants (R257E, K, and Q) of Chlamydomonas reinhardtii have a lowered QB redox potential: analysis of thermoluminescence and fluorescence measurements

    Science.gov (United States)

    Rose, Stuart; Minagawa, Jun; Seufferheld, Manfredo; Padden, Sean; Svensson, Bengt; Kolling, Derrick R. J.; Crofts, Antony R.; Govindjee

    2009-01-01

    Arginine257 (R257), in the de-helix that caps the QB site of the D1 protein, has been shown by mutational studies to play a key role in the sensitivity of Photosystem II (PS II) to bicarbonate-reversible binding of the formate anion. In this article, the role of this residue has been further investigated through D1 mutations (R257E, R257Q, and R257K) in Chlamydomonas reinhardtii. We have investigated the activity of the QB site by studying differences from wild type on the steady-state turnover of PS II, as assayed through chlorophyll (Chl) a fluorescence yield decay after flash excitation. The effects of p-benzoquinone (BQ, which oxidizes reduced QB, QB−) and 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU, which blocks electron flow from QA− to QB) were measured. The equilibrium constants of the two-electron gate were obtained through thermoluminescence measurements. The thermoluminescence properties were changed in the mutants, especially when observed after pretreatment with 100 μM BQ. A theoretical analysis of the thermoluminescence data, based mainly on the recombination pathways model of Rappaport et al. (2005), led to the conclusion that the free-energy difference for the recombination of QB− with S2 was reduced by 20–40 mV in the three mutants (D1-R257K, D1-R257Q, and D1-R257E); this was interpreted to be due to a lowering of the redox potential of QB/QB−. Further, since the recombination of QA− with S2 was unaffected, we suggest that no significant change in redox potential of QA/QA− occurred in these three mutants. The maximum variable Chl a fluorescence yield is lowered in the mutants, in the order R257K > R257Q > R257E, compared to wild type. Our analysis of the binary oscillations in Chl a fluorescence following pretreatment of cells with BQ showed that turnover of the QB site was relatively unaffected in the three mutants. The mutant D1-R257E had the lowest growth rate and steady-state activity and showed the weakest binary oscillations

  18. Structure of photosystem II and substrate binding at room temperature

    Science.gov (United States)

    Gul, Sheraz; Fuller, Franklin; Koroidov, Sergey; Brewster, Aaron S.; Tran, Rosalie; Alonso-Mori, Roberto; Kroll, Thomas; Michels-Clark, Tara; Laksmono, Hartawan; Sierra, Raymond G.; Stan, Claudiu A.; Hussein, Rana; Zhang, Miao; Douthit, Lacey; Kubin, Markus; de Lichtenberg, Casper; Long Vo, Pham; Nilsson, Håkan; Cheah, Mun Hon; Shevela, Dmitriy; Saracini, Claudio; Bean, Mackenzie A.; Seuffert, Ina; Sokaras, Dimosthenis; Weng, Tsu-Chien; Pastor, Ernest; Weninger, Clemens; Fransson, Thomas; Lassalle, Louise; Bräuer, Philipp; Aller, Pierre; Docker, Peter T.; Andi, Babak; Orville, Allen M.; Glownia, James M.; Nelson, Silke; Sikorski, Marcin; Zhu, Diling; Hunter, Mark S.; Lane, Thomas J.; Aquila, Andy; Koglin, Jason E.; Robinson, Joseph; Liang, Mengning; Boutet, Sébastien; Lyubimov, Artem Y.; Uervirojnangkoorn, Monarin; Moriarty, Nigel W.; Liebschner, Dorothee; Afonine, Pavel V.; Waterman, David G.; Evans, Gwyndaf; Wernet, Philippe; Dobbek, Holger; Weis, William I.; Brunger, Axel T.; Zwart, Petrus H.; Adams, Paul D.; Zouni, Athina; Messinger, Johannes; Bergmann, Uwe; Sauter, Nicholas K.; Kern, Jan; Yachandra, Vittal K.; Yano, Junko

    2016-01-01

    Light-induced oxidation of water by photosystem II (PS II) in plants, algae and cyanobacteria has generated most of the dioxygen in the atmosphere. PS II, a membrane-bound multi-subunit pigment-protein complex, couples the one-electron photochemistry at the reaction center with the four-electron redox chemistry of water oxidation at the Mn4CaO5 cluster in the oxygen-evolving complex (OEC) (Fig. 1a, Extended Data Fig. 1). Under illumination, the OEC cycles through five intermediate S-states (S0 to S4)1, where S1 is the dark stable state and S3 is the last semi-stable state before O-O bond formation and O2 evolution2,3. A detailed understanding of the O-O bond formation mechanism remains a challenge, and elucidating the structures of the OEC in the different S-states, as well as the binding of the two substrate waters to the catalytic site4-6, is a prerequisite for this purpose. Here we report the use of femtosecond pulses from an X-ray free electron laser (XFEL) to obtain damage free, room temperature (RT) structures of dark-adapted (S1), two-flash illuminated (2F; S3-enriched), and ammonia-bound two-flash illuminated (2F-NH3; S3-enriched) PS II. Although the recent 1.95 Å structure of PS II7 at cryogenic temperature using an XFEL provided a damage-free view of the S1 state, RT measurements are required to study the structural landscape of proteins under functional conditions8,9, and also for in situ advancement of the S-states. To investigate the water-binding site(s), ammonia, a water analog, has been used as a marker, as it binds to the Mn4CaO5 cluster in the S2 and S3 states10. Since the ammonia-bound OEC is active, the ammonia-binding Mn site is not a substrate water site10-13. Thus, this approach, together with a comparison of the native dark and 2F states, is used to discriminate between proposed O-O bond formation mechanisms. PMID:27871088

  19. New tools for chloroplast genetic engineering allow the synthesis of human growth hormone in the green alga Chlamydomonas reinhardtii.

    Science.gov (United States)

    Wannathong, Thanyanan; Waterhouse, Janet C; Young, Rosanna E B; Economou, Chloe K; Purton, Saul

    2016-06-01

    In recent years, there has been an increasing interest in the exploitation of microalgae in industrial biotechnology. Potentially, these phototrophic eukaryotes could be used for the low-cost synthesis of valuable recombinant products such as bioactive metabolites and therapeutic proteins. The algal chloroplast in particular represents an attractive target for such genetic engineering, both because it houses major metabolic pathways and because foreign genes can be targeted to specific loci within the chloroplast genome, resulting in high-level, stable expression. However, routine methods for chloroplast genetic engineering are currently available only for one species-Chlamydomonas reinhardtii-and even here, there are limitations to the existing technology, including the need for an expensive biolistic device for DNA delivery, the lack of robust expression vectors, and the undesirable use of antibiotic resistance markers. Here, we describe a new strain and vectors for targeted insertion of transgenes into a neutral chloroplast locus that (i) allow scar-less fusion of a transgenic coding sequence to the promoter/5'UTR element of the highly expressed endogenous genes psaA or atpA, (ii) employ the endogenous gene psbH as an effective but benign selectable marker, and (iii) ensure the successful integration of the transgene construct in all transformant lines. Transformation is achieved by a simple and cheap method of agitation of a DNA/cell suspension with glass beads, with selection based on the phototrophic rescue of a cell wall-deficient ΔpsbH strain. We demonstrate the utility of these tools in the creation of a transgenic line that produces high levels of functional human growth hormone.

  20. Multiple stressor effects in Chlamydomonas reinhardtii – Toward understanding mechanisms of interaction between effects of ultraviolet radiation and chemical pollutants

    Energy Technology Data Exchange (ETDEWEB)

    Korkaric, Muris [Eawag, Swiss Federal Institute of Aquatic Science and Technology, Department of Environmental Toxicology, 8600, Duebendorf (Switzerland); ETH Zürich, Institute of Biogeochemistry and Pollutant Dynamics, 8092 Zürich (Switzerland); Behra, Renata; Fischer, Beat B. [Eawag, Swiss Federal Institute of Aquatic Science and Technology, Department of Environmental Toxicology, 8600, Duebendorf (Switzerland); Junghans, Marion [Swiss Center for Applied Ecotoxicology Eawag-EPFL, 8600, Duebendorf (Switzerland); Eggen, Rik I.L., E-mail: rik.eggen@eawag.ch [Eawag, Swiss Federal Institute of Aquatic Science and Technology, Department of Environmental Toxicology, 8600, Duebendorf (Switzerland); ETH Zürich, Institute of Biogeochemistry and Pollutant Dynamics, 8092 Zürich (Switzerland)

    2015-05-15

    Highlights: • Systematic study of multiple stressor effects of UVR and chemicals in C. reinhardtii. • UVR and chemicals did not act independently on algal photosynthesis and reproduction. • Multiple stressor effects of UVR and chemicals depended on chemical MOA. • Synergistic effect interactions not limited to oxidative stress inducing chemicals. • Multiple MOAs of UVR may limit applicability of current prediction models. - Abstract: The effects of chemical pollutants and environmental stressors, such as ultraviolet radiation (UVR), can interact when organisms are simultaneously exposed, resulting in higher (synergistic) or lower (antagonistic) multiple stressor effects than expected based on the effects of single stressors. Current understanding of interactive effects is limited due to a lack of mechanism-based multiple stressor studies. It has been hypothesized that effect interactions may generally occur if chemical and non-chemical stressors cause similar physiological effects in the organism. To test this hypothesis, we exposed the model green alga Chlamydomonas reinhardtii to combinations of UVR and single chemicals displaying modes of action (MOA) similar or dissimilar to the impact of UVR on photosynthesis. Stressor interactions were analyzed based on the independent action model. Effect interactions were found to depend on the MOA of the chemicals, and also on their concentrations, the exposure time and the measured endpoint. Indeed, only chemicals assumed to cause effects on photosynthesis similar to UVR showed interactions with UVR on photosynthetic yield: synergistic in case of Cd(II) and paraquat and antagonistic in case of diuron. No interaction on photosynthesis was observed for S-metolachlor, which acts dissimilarly to UVR. However, combined effects of S-metolachlor and UVR on algal reproduction were synergistic, highlighting the importance of considering additional MOA of UVR. Possible mechanisms of stressor effect interactions are

  1. Photosystem II inhibitor resistance in the Columbia Basin of Washington state

    Science.gov (United States)

    Potato and mint (peppermint and spearmint) are commonly produced in the irrigated regions of the Pacific Northwest and both crops rely heavily on photosystem II (PSII) inhibitor herbicides metribuzin (potato) and terbacil (mint) for weed management. Seed was collected in 2010 from Powell amaranth, r...

  2. Photosystem II electron flow as a measure for phytoplankton gross primary production.

    NARCIS (Netherlands)

    Geel, C.

    1997-01-01

    Saturating pulse fluorescence measurements, well known from studies of higher plants for determination of photosystem II (PS II) characteristics, were applied to cultures of the green alga Dunaliella teitiolecta (Chapter 2). The actual efficiency of PS II (φ PS II ), the maximal efficienc

  3. Continuum probe two-dimensional electronic spectroscopy of the photosystem II reaction center

    Directory of Open Access Journals (Sweden)

    Ogilvie J. P.

    2013-03-01

    Full Text Available We report two-dimensional electronic spectroscopy of the photosystem II reaction center, collected in the pump-probe geometry employing a continuum probe. This enables observation of ion bands that report on intermediates in the charge separation process.

  4. Photosystem I from plants as a bacterial cytochrome P450 surrogate electron donor

    DEFF Research Database (Denmark)

    Jensen, Kenneth; Johnston, Jonathan B.; Montellano, Paul R. Ortiz de;

    2012-01-01

    ) is a multifunctional electron carrier that in plants accepts electrons from photosystem I (PSI) and facilitates photoreduction of NADP+ to NADPH mediated by ferredoxin-NAD(P)H oxidoreductase (FdR). In bacteria, the electron flow is reversed and Fdx accepts electrons from NADPH via FdR and serves as the direct electron...

  5. Crystal structure of monomeric photosystem II from Thermosynechococcus elongatus at 3.6-a resolution

    NARCIS (Netherlands)

    Broser, Matthias; Gabdulkhakov, Azat; Kern, Jan; Guskov, Albert; Müh, Frank; Saenger, Wolfram; Zouni, Athina

    2010-01-01

    The membrane-embedded photosystem II core complex (PSIIcc) uses light energy to oxidize water in photosynthesis. Information about the spatial structure of PSIIcc obtained from x-ray crystallography was so far derived from homodimeric PSIIcc of thermophilic cyanobacteria. Here, we report the first c

  6. Structural basis of cyanobacterial photosystem II Inhibition by the herbicide terbutryn

    NARCIS (Netherlands)

    Broser, Matthias; Glöckner, Carina; Gabdulkhakov, Azat; Guskov, Albert; Buchta, Joachim; Kern, Jan; Müh, Frank; Dau, Holger; Saenger, Wolfram; Zouni, Athina

    2011-01-01

    Herbicides that target photosystem II (PSII) compete with the native electron acceptor plastoquinone for binding at the Q(B) site in the D1 subunit and thus block the electron transfer from Q(A) to Q(B). Here, we present the first crystal structure of PSII with a bound herbicide at a resolution of 3

  7. Evidence for a trimeric organization of the photosystem I complex from the thermophilic cyanobacterium Synechococcus sp.

    NARCIS (Netherlands)

    Boekema, E.J.; Dekker, J.P.; Heel, M.G. van; Rögner, M.; Saenger, W.; Witt, I.; Witt, H.T.

    1987-01-01

    A photosystem I (PS I) reaction center complex was isolated and purified from the cyanobacterium Synechococcus sp. The complex has a molecular mass of about 600 kDa and contains 120 Chl a molecules per photoactive Chl a1 (P-700). Electron micrographs show that the PS I complex has the shape of a dis

  8. Copper Toxicity Affects Photosystem II Electron Transport at the Secondary Quinone Acceptor, QB1

    Science.gov (United States)

    Mohanty, Narendranath; Vass, Imre; Demeter, Sándor

    1989-01-01

    The nature of Cu2+ inhibition of photosystem II (PSII) photochemistry in pea (Pisum sativum L.) thylakoids was investigated monitoring Hill activity and light emission properties of photosystem II. In Cu2+-inhibited thylakoids, diphenyl carbazide addition does not relieve the loss of Hill activity. The maximum yield of fluorescence induction restored by hydroxylamine in Tris-inactivated thylakoids is markedly reduced by Cu2+. This suggests that Cu2+ does not act on the donor side of PSII but on the reaction center of PSII or on components beyond. Thermoluminescence and delayed luminescence studies show that charge recombination between the positively charged intermediate in water oxidation cycle (S2) and negatively charged primary quinone acceptor of pSII (QA−) is largely unaffected by Cu2+. The S2QB− charge recombination, however, is drastically inhibited which parallels the loss of Hill activity. This indicates that Cu2+ inhibits photosystem II photochemistry primarily affecting the function of the secondary quinone electron acceptor, QB. We suggest that Cu2+ does not block electron flow between the primary and secondary quinone acceptor but modifies the QB site in such a way that it becomes unsuitable for further photosystem II photochemistry. PMID:16666731

  9. Identification of a 32-34-kilodalton polypeptide as a herbicide receptor protein in photosystem II.

    Science.gov (United States)

    Mullet, J E; Arntzen, C J

    1981-04-13

    Photosystem II particles which retained high rates of herbicide-sensitive activity were used to examine the site(s) of action of various herbicides. A polypeptide of 32-34 kdaltons was identified as the triazine-herbicide binding site based upon: (a) parallel loss of atrazine activity and the polypeptide during either trypsin treatment or selective detergent depletion of protein in the Photosystem II complex, and (b) covalent labeling of the polypeptide by a 14C-labeled photoaffinity triazine. In Photosystem II particles depleted of the 32-34-kdalton polypeptide, electron transport was still active and was slightly sensitive to DCMU and largely sensitive to dinoseb (urea and nitrophenol herbicides, respectively). On the basis of this result it is proposed that the general herbicide binding site common to atrazine, DCMU and dinoseb is formed by a minimum of two polypeptides which determine affinity and/or mediate herbicide-induced inhibition of electron transport on the acceptor side of Photosystem II.

  10. Filling the Green Gap of a Megadalton Photosystem I Complex by Conjugation of Organic Dyes

    NARCIS (Netherlands)

    Gordiichuk, Pavlo I; Rimmerman, Dolev; Paul, Avishek; Gautier, Daniel A; Gruszka, Agnieszka; Saller, Manfred; de Vries, Jan Willem; Wetzelaer, Gert-Jan A H; Manca, Marianna; Gomulya, Widianta; Matmor, Maayan; Gloukhikh, Ekaterina; Loznik, Mark; Ashkenasy, Nurit; Blom, Paul W M; Rögner, Matthias; Loi, Maria Antonietta; Richter, Shachar; Herrmann, Andreas

    2016-01-01

    Photosynthesis is Natur&s major process for converting solar into chemical energy. One of the key players' in this process is the multiprotein complex photosystem I (PSI) that through absorption of incident,photons enables electron transfer, which makes this, protein attractive for applications in b

  11. Multiple LHCII antennae can transfer energy efficiently to a single Photosystem I.

    Science.gov (United States)

    Bos, Inge; Bland, Kaitlyn M; Tian, Lijin; Croce, Roberta; Frankel, Laurie K; van Amerongen, Herbert; Bricker, Terry M; Wientjes, Emilie

    2017-02-22

    Photosystems I and II (PSI and PSII) work in series to drive oxygenic photosynthesis. The two photosystems have different absorption spectra, therefore changes in light quality can lead to imbalanced excitation of the photosystems and a loss in photosynthetic efficiency. In a short-term adaptation response termed state transitions, excitation energy is directed to the light-limited photosystem. In higher plants a special pool of LHCII antennae, which can be associated with either PSI or PSII, participates in these state transitions. It is known that one LHCII antenna can associate with the PsaH site of PSI. However, membrane fractions were recently isolated in which multiple LHCII antennae appear to transfer energy to PSI. We have used time-resolved fluorescence-streak camera measurements to investigate the energy transfer rates and efficiency in these membrane fractions. Our data show that energy transfer from LHCII to PSI is relatively slow. Nevertheless, the trapping efficiency in supercomplexes of PSI with ~2.4 LHCIIs attached is 94%. The absorption cross section of PSI can thus be increased with ~65% without having significant loss in quantum efficiency. Comparison of the fluorescence dynamics of PSI-LHCII complexes, isolated in a detergent or located in their native membrane environment, indicates that the environment influences the excitation energy transfer rates in these complexes. This demonstrates the importance of studying membrane protein complexes in their natural environment.

  12. Catalytic Oxygen Evolution by a Bioinorganic Model of the Photosystem II Oxygen-Evolving Complex

    Science.gov (United States)

    Howard, Derrick L.; Tinoco, Arthur D.; Brudvig, Gary W.; Vrettos, John S.; Allen, Bertha Connie

    2005-01-01

    Bioinorganic models of the manganese Mn4 cluster are important not only as aids in understanding the structure and function of the oxygen-evolving complex (OEC), but also in developing artificial water-oxidation catalysts. The mechanism of water oxidation by photosystem II (PSII) is thought to involve the formation of a high-valent terminal Mn-oxo…

  13. Efficient Heterologous Transformation of Chlamydomonas reinhardtii npq2 Mutant with the Zeaxanthin Epoxidase Gene Isolated and Characterized from Chlorella zofingiensis

    Directory of Open Access Journals (Sweden)

    Herminia Rodríguez

    2012-09-01

    Full Text Available In the violaxanthin cycle, the violaxanthin de-epoxidase and zeaxanthin epoxidase catalyze the inter-conversion between violaxanthin and zeaxanthin in both plants and green algae. The zeaxanthin epoxidase gene from the green microalga Chlorella zofingiensis (Czzep has been isolated. This gene encodes a polypeptide of 596 amino acids. A single copy of Czzep has been found in the C. zofingiensis genome by Southern blot analysis. qPCR analysis has shown that transcript levels of Czzep were increased after zeaxanthin formation under high light conditions. The functionality of Czzep gene by heterologous genetic complementation in the Chlamydomonas mutant npq2, which lacks zeaxanthin epoxidase (ZEP activity and accumulates zeaxanthin in all conditions, was analyzed. The Czzep gene was adequately inserted in the pSI105 vector and expressed in npq2. The positive transformants were able to efficiently convert zeaxanthin into violaxanthin, as well as to restore their maximum quantum efficiency of the PSII (Fv/Fm. These results show that Chlamydomonas can be an efficient tool for heterologous expression and metabolic engineering for biotechnological applications.

  14. Changes in Thermostability of Photosystem Ⅱ and Leaf Lipid Composition of Rice Mutant with Deficiency of Light-harvesting Chlorophyll Protein Complexes

    Institute of Scientific and Technical Information of China (English)

    Yunlai Tang; Mei Chen; Yinong Xu; Tingyun Kuang

    2007-01-01

    We studied the difference in thermostability of photosystem Ⅱ (PSⅡ) and leaf lipid composition between a T-DNA insertion mutant rice (Oryza sativa L.) VG28 and its wild type Zhonghua11. Native green gel and SDS-PAGE electrophoreses revealed that the mutant VG28 lacked all light-harvesting chlorophyll a/b protein complexes. Both the mutant and wild type were sensitive to high temperatures, and the maximal efficiency of PSⅡ photochemistry (Fv/Fm) and oxygen-evolving activity of PSⅡ in leaves significantly decreased with increasing temperature. However, the PSⅡ activity of the mutant was markedly more sensitive to high temperatures than that of the wild type. Lipid composition analysis showed that the mutant had less phosphatidylglycerol and sulfoquinovosyl diacylglycerol compared with the wild type. Fatty acid analysis revealed that the mutant had an obvious decrease in the content of unsaturation of membrane lipids on the thermostability of PSll are discussed.

  15. Self-consistent QM/MM methodologies for structural refinement of photosystem II and other macromolecules of biological interest

    Energy Technology Data Exchange (ETDEWEB)

    Batista, Enrique R [Los Alamos National Laboratory; Sproviero, Eduardo M [YALE UNIV; Newcomer, Michael [YALE UNIV; Gascon, Jose A [YALE UNIV; Batista, Victor S [YALE UNIV

    2008-01-01

    The combination of quantum mechanics and molecular mechanics (QM/MM) is one of the most promising approaches to study the structure, function, and properties of proteins and nucleic acids. However, there some instances in which the limitations of either the MM (lack of a proper electronic description) or QM (limited to a few number of atoms) methods prevent a proper description of the system. To address this issue, we review here our approach to fine-tune the structure of biological systems using post-QM/MM refinements. These protocols are based on spectroscopy data, and/or partitioning of the system to extend the QM description to a larger region of a protein. We illustrate these methodologies through applications to several biomolecules, which were pre-optimized at the QM/MM level and then further refined using postQM/MM refinement methodologies: mod(QM/MM), which refines the atomic charges of the residues included in the MM region accounting for polarization effects; mod(QM/MM)-opt that partition the MM region in smaller parts and optimizes each part in an iterative. self-consistent way, and the Polarized-Extended X-Ray Absorption Fine Structure (P-EXAFS) fitting procedure, which fine-tune the atomic coordinates to reproduce experimental polarized EXAFS spectra. The first two techniques were applied to the guanine quadruplex. while the P-EXAFS refinement was applied to the oxygen evolving complex of photosystem II.

  16. A structure-based model of energy transfer reveals the principles of light harvesting in photosystem II supercomplexes.

    Science.gov (United States)

    Bennett, Doran I G; Amarnath, Kapil; Fleming, Graham R

    2013-06-19

    Photosystem II (PSII) initiates photosynthesis in plants through the absorption of light and subsequent conversion of excitation energy to chemical energy via charge separation. The pigment binding proteins associated with PSII assemble in the grana membrane into PSII supercomplexes and surrounding light harvesting complex II trimers. To understand the high efficiency of light harvesting in PSII requires quantitative insight into energy transfer and charge separation in PSII supercomplexes. We have constructed the first structure-based model of energy transfer in PSII supercomplexes. This model shows that the kinetics of light harvesting cannot be simplified to a single rate limiting step. Instead, substantial contributions arise from both excitation diffusion through the antenna pigments and transfer from the antenna to the reaction center (RC), where charge separation occurs. Because of the lack of a rate-limiting step, fitting kinetic models to fluorescence lifetime data cannot be used to derive mechanistic insight on light harvesting in PSII. This model will clarify the interpretation of chlorophyll fluorescence data from PSII supercomplexes, grana membranes, and leaves.

  17. Engineering the chloroplast targeted malarial vaccine antigens in Chlamydomonas starch granules.

    Directory of Open Access Journals (Sweden)

    David Dauvillée

    Full Text Available BACKGROUND: Malaria, an Anopheles-borne parasitic disease, remains a major global health problem causing illness and death that disproportionately affects developing countries. Despite the incidence of malaria, which remains one of the most severe infections of human populations, there is no licensed vaccine against this life-threatening disease. In this context, we decided to explore the expression of Plasmodium vaccine antigens fused to the granule bound starch synthase (GBSS, the major protein associated to the starch matrix in all starch-accumulating plants and algae such as Chlamydomonas reinhardtii. METHODS AND FINDINGS: We describe the development of genetically engineered starch granules containing plasmodial vaccine candidate antigens produced in the unicellular green algae Chlamydomonas reinhardtii. We show that the C-terminal domains of proteins from the rodent Plasmodium species, Plasmodium berghei Apical Major Antigen AMA1, or Major Surface Protein MSP1 fused to the algal granule bound starch synthase (GBSS are efficiently expressed and bound to the polysaccharide matrix. Mice were either immunized intraperitoneally with the engineered starch particles and Freund adjuvant, or fed with the engineered particles co-delivered with the mucosal adjuvant, and challenged intraperitoneally with a lethal inoculum of P. Berghei. Both experimental strategies led to a significantly reduced parasitemia with an extension of life span including complete cure for intraperitoneal delivery as assessed by negative blood thin smears. In the case of the starch bound P. falciparum GBSS-MSP1 fusion protein, the immune sera or purified immunoglobulin G of mice immunized with the corresponding starch strongly inhibited in vitro the intra-erythrocytic asexual development of the most human deadly plasmodial species. CONCLUSION: This novel system paves the way for the production of clinically relevant plasmodial antigens as algal starch-based particles

  18. Rapid construction and screening of artificial microRNA systems in Chlamydomonas reinhardtii.

    Science.gov (United States)

    Hu, Jinlu; Deng, Xuan; Shao, Ning; Wang, Gaohong; Huang, Kaiyao

    2014-09-01

    The unicellular green algae Chlamydomonas reinhardtii is a classic model for the study of flagella/cilia and photosynthesis, and it has recently been exploited for producing biopharmaceuticals and biofuel. Due to the low frequency of homologous recombination, reverse genetic manipulation in Chlamydomonas relies mainly on miRNA- and siRNA-based knockdown methods. However, the difficulty in constructing artificial miRNA vectors, laborious screening of knockdown transformants, and undesired epigenetic silencing of exogenous miRNA constructs limit their application. We have established a one-step procedure to construct an artificial miRNA precursor by annealing eight oligonucleotides of approximately 40 nucleotides. In the final construct, the Gaussia princeps luciferase gene (G-Luc) is positioned between the promoter and the artificial miRNA precursor so that knockdown strains may quickly be screened by visualizing luciferase luminescence using a photon-counting camera. Furthermore, the luciferase activity of transformants correlates with the knockdown level of two test target proteins: the chloroplast protein VIPP1 (vesicle inducing protein in plastids 1) and the flagellar protein CDPK3 (calcium-dependent protein kinase 3). Adding an intron from RBCS2 (ribulose bisphosphate carboxylase/oxygenase small subunit 2) to the miRNA construct enhanced both the luciferase activity and the miRNA knockdown efficiency. A second miRNA vector incorporated the promoter of the nitrate reductase gene to allow inducible expression of the artificial miRNA. These vectors will facilitate application of the artificial miRNA and provide tools for studying the mechanism of epigenetics in Chlamydomonas, and may also be adapted for use in other model organisms.

  19. Acclimation of Antarctic Chlamydomonas to the sea-ice environment: a transcriptomic analysis.

    Science.gov (United States)

    Liu, Chenlin; Wang, Xiuliang; Wang, Xingna; Sun, Chengjun

    2016-07-01

    The Antarctic green alga Chlamydomonas sp. ICE-L was isolated from sea ice. As a psychrophilic microalga, it can tolerate the environmental stress in the sea-ice brine, such as freezing temperature and high salinity. We performed a transcriptome analysis to identify freezing stress responding genes and explore the extreme environmental acclimation-related strategies. Here, we show that many genes in ICE-L transcriptome that encoding PUFA synthesis enzymes, molecular chaperon proteins, and cell membrane transport proteins have high similarity to the gens from Antarctic bacteria. These ICE-L genes are supposed to be acquired through horizontal gene transfer from its symbiotic microbes in the sea-ice brine. The presence of these genes in both sea-ice microalgae and bacteria indicated the biological processes they involved in are possibly contributing to ICE-L success in sea ice. In addition, the biological pathways were compared between ICE-L and its closely related sister species, Chlamydomonas reinhardtii and Volvox carteri. In ICE-L transcripome, many sequences homologous to the plant or bacteria proteins in the post-transcriptional, post-translational modification, and signal-transduction KEGG pathways, are absent in the nonpsychrophilic green algae. These complex structural components might imply enhanced stress adaptation capacity. At last, differential gene expression analysis at the transcriptome level of ICE-L indicated that genes that associated with post-translational modification, lipid metabolism, and nitrogen metabolism are responding to the freezing treatment. In conclusion, the transcriptome of Chlamydomonas sp. ICE-L is very useful for exploring the mutualistic interaction between microalgae and bacteria in sea ice; and discovering the specific genes and metabolism pathways responding to the freezing acclimation in psychrophilic microalgae.

  20. Protocol: methodology for chromatin immunoprecipitation (ChIP in Chlamydomonas reinhardtii

    Directory of Open Access Journals (Sweden)

    Strenkert Daniela

    2011-11-01

    Full Text Available Abstract We report on a detailed chromatin immunoprecipitation (ChIP protocol for the unicellular green alga Chlamydomonas reinhardtii. The protocol is suitable for the analysis of nucleosome occupancy, histone modifications and transcription factor binding sites at the level of mononucleosomes for targeted and genome-wide studies. We describe the optimization of conditions for crosslinking, chromatin fragmentation and antibody titer determination and provide recommendations and an example for the normalization of ChIP results as determined by real-time PCR.

  1. Domain organization of photosystem II in membranes of the cyanobacterium Synechocystis PCC6803 investigated by electron microscopy

    NARCIS (Netherlands)

    Folea, I. Mihaela; Zhang, Pengpeng; Aro, Eva-Mari; Boekema, Egbert J.

    2008-01-01

    The supramolecular organization of photosystem II (PSII) complexes in the photosynthetic membrane of the cyanobacterium Synechocystis 6803 was studied by electron microscopy. After mild detergent solubilization, crystalline PSII arrays were extracted in which dimeric PSII particles associate in mult

  2. The Vitamin B12-Dependent Photoreceptor AerR Relieves Photosystem Gene Repression by Extending the Interaction of CrtJ with Photosystem Promoters

    Science.gov (United States)

    Fang, Mingxu

    2017-01-01

    ABSTRACT Purple nonsulfur bacteria adapt their physiology to a wide variety of environmental conditions often through the control of transcription. One of the main transcription factors involved in controlling expression of the Rhodobacter capsulatus photosystem is CrtJ, which functions as an aerobic repressor of photosystem genes. Recently, we reported that a vitamin B12 binding antirepressor of CrtJ called AerR is required for anaerobic expression of the photosystem. However, the mechanism whereby AerR regulates CrtJ activity is unclear. In this study, we used a combination of next-generation sequencing and biochemical methods to globally identify genes under control of CrtJ and the role of AerR in controlling this regulation. Our results indicate that CrtJ has a much larger regulon than previously known, with a surprising regulatory function under both aerobic and anaerobic photosynthetic growth conditions. A combination of in vivo chromatin immunoprecipitation-DNA sequencing (ChIP-seq) and ChIP-seq and exonuclease digestion (ChIP-exo) studies and in vitro biochemical studies demonstrate that AerR forms a 1:2 complex with CrtJ (AerR-CrtJ2) and that this complex binds to many promoters under photosynthetic conditions. The results of in vitro and in vivo DNA binding studies indicate that AerR-CrtJ2 anaerobically forms an extended interaction with the bacteriochlorophyll bchC promoter to relieve repression by CrtJ. This is contrasted by aerobic growth conditions where CrtJ alone functions as an aerobic repressor of bchC expression. These results indicate that the DNA binding activity of CrtJ is modified by interacting with AerR in a redox-regulated manner and that this interaction alters CrtJ’s function. PMID:28325764

  3. Dephosphorylation Pathway of D-myo-Inositol 1,4,5-trisphosphate in the Unicellular Green Alga Chlamydomonas eugametos

    NARCIS (Netherlands)

    Klerk, Hans; Himbergen, John A.J. van; Musgrave, Alan; Haastert, Peter J.M. van; Ende, Herman van den

    1994-01-01

    In vitro dephosphorylation of D-myo-inositol 1,4,5-trisphosphate [Ins(l,4,5)P-3] by vegetative cells, gametes and zygotes of the green alga Chlamydomonas eugametos was studied using a soluble cell fraction as enzyme source and labelled Ins(1,4,5)P-3 as substrate. This compound was dephosphorylated y

  4. An Indexed, Mapped Mutant Library Enables Reverse Genetics Studies of Biological Processes in Chlamydomonas reinhardtii.

    Science.gov (United States)

    Li, Xiaobo; Zhang, Ru; Patena, Weronika; Gang, Spencer S; Blum, Sean R; Ivanova, Nina; Yue, Rebecca; Robertson, Jacob M; Lefebvre, Paul A; Fitz-Gibbon, Sorel T; Grossman, Arthur R; Jonikas, Martin C

    2016-02-01

    The green alga Chlamydomonas reinhardtii is a leading unicellular model for dissecting biological processes in photosynthetic eukaryotes. However, its usefulness has been limited by difficulties in obtaining mutants in specific genes of interest. To allow generation of large numbers of mapped mutants, we developed high-throughput methods that (1) enable easy maintenance of tens of thousands of Chlamydomonas strains by propagation on agar media and by cryogenic storage, (2) identify mutagenic insertion sites and physical coordinates in these collections, and (3) validate the insertion sites in pools of mutants by obtaining >500 bp of flanking genomic sequences. We used these approaches to construct a stably maintained library of 1935 mapped mutants, representing disruptions in 1562 genes. We further characterized randomly selected mutants and found that 33 out of 44 insertion sites (75%) could be confirmed by PCR, and 17 out of 23 mutants (74%) contained a single insertion. To demonstrate the power of this library for elucidating biological processes, we analyzed the lipid content of mutants disrupted in genes encoding proteins of the algal lipid droplet proteome. This study revealed a central role of the long-chain acyl-CoA synthetase LCS2 in the production of triacylglycerol from de novo-synthesized fatty acids.

  5. Measurement of Lipid Droplet Accumulation Kinetics in Chlamydomonas reinhardtii Using Seoul-Fluor

    Directory of Open Access Journals (Sweden)

    Noo Li Jeon

    2013-10-01

    Full Text Available Alternative energy resources have become an important issue due to the limited stocks of petroleum-based fuel. Microalgae, a source of renewable biodiesel, use solar light to convert CO2 into lipid droplets (LDs. Quantification of LDs in microalgae is required for developing and optimizing algal bioprocess engineering. However, conventional quantification methods are both time and labor-intensive and difficult to apply in high-throughput screening systems. LDs in plant and mammalian cells can be visualized by staining with various fluorescence probes such as the Nile Red, BODIPY, and Seoul-Fluor (SF series. This report describes the optimization of LD staining in Chlamydomonas reinhardtii with SF probes via systematic variations of dye concentration, staining time, temperature, and pH. A protocol for quantitative measurement of accumulation kinetics of LDs in C. reinhardtii was developed using a spectrofluorimeter and the accuracy of LD size measurement was confirmed by transmission electron microscopy (TEM. Our results indicate that our spectrofluorimeter-based measurement approach can monitor kinetics of intracellular LDs (in control and nitrogen-source-starved Chlamydomonas reinhardtii accumulation that has not been possible in the case of conventional imaging-based methods. Our results presented here confirmed that an SF44 can be a powerful tool for in situ monitoring and tracking of intracellular LDs formation.

  6. Introducing Dunaliella LIP promoter containing light-inducible motifs improves transgenic expression in Chlamydomonas reinhardtii.

    Science.gov (United States)

    Baek, Kwangryul; Lee, Yew; Nam, Onyou; Park, Seunghye; Sim, Sang Jun; Jin, EonSeon

    2016-03-01

    Promoter of the light-inducible protein gene (LIP) of Dunaliella was recently isolated in our laboratory. The aim of this work is to find the light-inducible motif in the Dunaliella LIP promoter and verify its regulatory motif with a Gaussia luciferase reporter gene transformed in Chlamydomonas reinhardtii. 400 bp upstream to the translational start site of the Dunaliella LIP gene was gradually truncated and analyzed for the luciferase expression. Furthermore, this promoter comprising duplicated or triplicated light-responsive motifs was tested for its augmentation of light response. Two putative light-responsive motifs, GT-1 binding motif and sequences over-represented in light-repressed promoters (SORLIP) located in the 200 bp LIP promoter fragment were analyzed for their light responsibility. It is turned out that SORLIP was responsible for the light-inducible activity. With the copy number of SORLIP up to three showed stronger high light response compared with the native LIP promoter fragment. Therefore, we found a light-responsive DNA motif operating in Chlamydomonas and confirm a synthetic promoter including this motif displayed light inducibility in heterologously transformed green algae for the first time. This light-inducible expression system will be applied to various area of algal research including algal biotechnology.

  7. Proteomic Alterations of Antarctic Ice Microalga Chlamydomonas sp. Under Low-Temperature Stress

    Institute of Scientific and Technical Information of China (English)

    Guang-Feng Kan; Jin-Lai Miao; Cui-Juan Shi; Guang-You Li

    2006-01-01

    Antarctic ice microalga can survive and thrive in cold channels or pores in the Antarctic ice layer. In order to understand the adaptive mechanisms to low temperature, in the present study we compared two-dimensional polyacrylamide gel electrophoresis (2-DE) profiles of normal and low temperature-stressed Antarctic ice microalga Chlamydomonas sp. cells. In addition, new protein spots induced by low temperature were identified with peptide mass fingerprinting based on matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) and database searching. Well-resolved and reproducible 2-DE patterns of both normal and low temperature-stressed cells were acquired. A total of 626 spots was detected in control cells and 652 spots were detected in the corresponding low temperature-stressed cells. A total of 598 spots was matched between normal and stressed cells. Two newly synthesized proteins (a and b) in low temperature-stressed cells were characterized. Protein spot A (53 kDa, pI 6.0) was similar to isopropylmalate/homocitrate/citramalate synthases, which act in the transport and metabolism of amino acids. Protein spot b (25 kDa, pI 8.0) was related to glutathione S-transferase, which functions as a scavenger of active oxygen, free radicals, and noxious metabolites. The present study is valuable for the application of ice microalgae, establishing an ice microalga Chlamydomonas sp. proteome database, and screening molecular biomarkers for further studies.

  8. Characterization of Hydrocortisone Biometabolites and 18S rRNA Gene in Chlamydomonas reinhardtii Cultures

    Directory of Open Access Journals (Sweden)

    Seyed Bagher Mosavi-Azam

    2008-10-01

    Full Text Available A unicellular microalga, Chlamydomonas reinhardtii, was isolated from rice paddy-field soil and water samples and used in the biotransformation of hydrocortisone (1. This strain has not been previously tested for steroid bioconversion. Fermentation was carried out in BG-11 medium supplemented with 0.05% substrate at 25ºC for 14 days of incubation. The products obtained were chromatographically purified and characterized using spectroscopic methods. 11b,17b-Dihydroxyandrost-4-en-3-one (2, 11b-hydroxyandrost-4-en-3,17-dione (3, 11b,17a,20b,21-tetrahydroxypregn-4-en-3-one (4 and prednisolone (5 were the main products of the bioconversion. The observed bioreaction features were the side chain degradation of the substrate to give compounds 2 and 3 and the 20-ketone reduction and 1,2-dehydrogenation affording compounds 4 and 5, respectively. A time course study showed the accumulation of product 2 from the second day of the fermentation and of compounds 3, 4 and 5 from the third day. All the metabolites reached their maximum concentration in seven days. Microalgal 18S rRNA gene was also amplified by PCR. PCR products were sequenced to confirm their authenticity as 18S rRNA gene of microalgae. The result of PCR blasted with other sequenced microalgae in NCBI showed 100% homology to the 18S small subunit rRNA of two Chlamydomonas reinhardtii spp.

  9. The small molecule fenpropimorph rapidly converts chloroplast membrane lipids to triacylglycerols in Chlamydomonas reinhardtii

    Science.gov (United States)

    Kim, Hanul; Jang, Sunghoon; Kim, Sangwoo; Yamaoka, Yasuyo; Hong, Daewoong; Song, Won-Yong; Nishida, Ikuo; Li-Beisson, Yonghua; Lee, Youngsook

    2015-01-01

    Concern about global warming has prompted an intense interest in developing economical methods of producing biofuels. Microalgae provide a promising platform for biofuel production, because they accumulate high levels of lipids, and do not compete with food or feed sources. However, current methods of producing algal oil involve subjecting the microalgae to stress conditions, such as nitrogen deprivation, and are prohibitively expensive. Here, we report that the fungicide fenpropimorph rapidly causes high levels of neutral lipids to accumulate in Chlamydomonas reinhardtii cells. When treated with fenpropimorph (10 μg mL-1) for 1 h, Chlamydomonas cells accumulated at least fourfold the amount of triacylglycerols (TAGs) present in the untreated control cells. Furthermore, the quantity of TAGs present after 1 h of fenpropimorph treatment was over twofold higher than that formed after 9 days of nitrogen starvation in medium with no acetate supplement. Biochemical analysis of lipids revealed that the accumulated TAGs were derived mainly from chloroplast polar membrane lipids. Such a conversion of chloroplast polar lipids to TAGs is desirable for biodiesel production, because polar lipids are usually removed during the biodiesel production process. Thus, our data exemplified that a cost and time effective method of producing TAGs is possible using fenpropimorph or similar drugs. PMID:25759683

  10. Biofixation of Carbon dioxide by Chlamydomonas sp. in a Tubular Photobioreactor

    Directory of Open Access Journals (Sweden)

    H Hadiyanto

    2012-04-01

    Full Text Available The biogas production from anaerobic digestion is a potential fuel for power generators application, if biogas can be upgraded to the same standards as fossil natural gas by CO2, H2S, and other non-combustible component removal. Microalgae Chlamydomonas sp. has potency to biofix the carbon dioxide and can be used as an additional food ingredient. The variations of flow rate and carbon dioxide concentration in the process resulting different value of biomass production and carbon dioxide biofixation. Biomass production at 40% carbon dioxide concentration obtained 5.685 gr/dm3 at 10% carbon dioxide concentration obtained 4.892 gr/dm3. The greatest value of carbon dioxide absorption occurs at a 40% concentration amounting to 12.09%. The rate of growth and productivity of microalgae tend to rise in 10% and 20% (%v carbon dioxide concentration, but began started a constant at 30% and 40% (%v carbon dioxide concentration. Biomass production tends to increase in light conditions while a constant in dark conditions. This study used Chlamydomonas sp. as media culture and performed on bubble column and tubular reactor with 6 litres of culture medium at a temperature of 28oC and atmospheric pressure.

  11. Biofixation of Carbon dioxide by Chlamydomonas sp. in a Tubular Photobioreactor

    Directory of Open Access Journals (Sweden)

    H Hadiyanto

    2012-02-01

    Full Text Available The biogas production from anaerobic digestion is a potential fuel for power generators application, if biogas can be upgraded to the same standards as fossil natural gas by CO2, H2S, and other non-combustible component removal. Microalgae Chlamydomonas sp. has potency to biofix the carbon dioxide and can be used as an additional food ingredient. The variations of flow rate and carbon dioxide concentration in the process resulting different value of biomass production and carbon dioxide biofixation. Biomass production at 40% carbon dioxide concentration obtained 5.685 gr/dm3 at 10% carbon dioxide concentration obtained 4.892 gr/dm3. The greatest value of carbon dioxide absorption occurs at a 40% concentration amounting to 12.09%. The rate of growth and productivity of microalgae tend to rise in 10% and 20% (%v carbon dioxide concentration, but began started a constant at 30% and 40% (%v carbon dioxide concentration. Biomass production tends to increase in light conditions while a constant in dark conditions. This study used Chlamydomonas sp. as media culture and performed on bubble column and tubular reactor with 6 litres of culture medium at a temperature of 28oC and atmospheric pressure.

  12. Optical and electrical measurement of energy transfer between nanocrystalline quantum dots and photosystem I.

    Science.gov (United States)

    Jung, Hyeson; Gulis, Galina; Gupta, Subhadra; Redding, Kevin; Gosztola, David J; Wiederrecht, Gary P; Stroscio, Michael A; Dutta, Mitra

    2010-11-18

    In the natural photosynthesis process, light harvesting complexes (LHCs) absorb light and pass excitation energy to photosystem I (PSI) and photosystem II (PSII). In this study, we have used nanocrystalline quantum dots (NQDs) as an artificial LHC by integrating them with PSI to extend their spectral range. We have performed photoluminescence (PL) and ultrafast time-resolved absorption measurements to investigate this process. Our PL experiments showed that emission from the NQDs is quenched, and the fluorescence from PSI is enhanced. Transient absorption and bleaching results can be explained by fluorescence resonance energy transfer (FRET) from the NQDs to the PSI. This nonradiative energy transfer occurs in ∼6 ps. Current-voltage (I-V) measurements on the composite NQD-PSI samples demonstrate a clear photoresponse.

  13. Optical and Electrical Measurement of Energy Transfer between Nanocrystalline Quantum Dots and Photosystem I

    Energy Technology Data Exchange (ETDEWEB)

    Jung, Hyeson [Univ. of Illinois, Chicago, IL (United States); Gulis, G. [Univ. of Alabama, Tuscaloosa, AL (United States); Gupta, S. [Univ. of Alabama, Tuscaloosa, AL (United States); Redding, K. [Arizona State Univ., Mesa, AZ (United States); Gosztola, D. J. [Argonne National Lab. (ANL), Argonne, IL (United States); Wiederrecht, Gary P [Argonne National Lab. (ANL), Argonne, IL (United States); Stroscio, M. A. [Univ. of Illinois, Chicago, IL (United States); Dutta, M. [Univ. of Illinois, Chicago, IL (United States)

    2010-08-31

    In the natural photosynthesis process, light harvesting complexes (LHCs) absorb light and pass excitation energy to photosystem I (PSI) and photosystem II (PSII). In this study, we have used nanocrystalline quantum dots (NQDs) as an artificial LHC by integrating them with PSI to extend their spectral range. We have performed photoluminescence (PL) and ultrafast time-resolved absorption measurements to investigate this process. Our PL experiments showed that emission from the NQDs is quenched, and the fluorescence from PSI is enhanced. Transient absorption and bleaching results can be explained by fluorescence resonance energy transfer (FRET) from the NQDs to the PSI. This nonradiative energy transfer occurs in ~6 ps. Current-voltage (I-V) measurements on the composite NQD-PSI samples demonstrate a clear photoresponse.

  14. Excitation energy distribution between two photosystems in Porphyra yezoensis and its sig-nificance in photosynthesisevolution

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    Comparative investigation on energy distribution between twophotosystems were carried out in the sporo- phytes and gametophytes of Porphyra yezoensis. By perfor- ming 77 K fluorescence spectra, we suggested that there probably existed a pathway for energy transfer from PSⅡ to PSⅠ to redistribute the absorbed energy in gametophytes, while no such a way or at minor level in sporophytes. Electron transfer inhibitor DCMU blocked the energy transfer from PSⅡ to PSⅠ in gametophytes, but no obvious effects on sporophytes. These indicated that excitation energy dis-tribution between two photosystems in gametophytes was more cooperative than that in sporophytes. These data in ontogenesis reflected the evolution process of photosynthetic organisms and supported the hypothesis of independent evolution of each photosystem.

  15. Concentric-flow electrokinetic injector enables serial crystallography of ribosome and photosystem II.

    Science.gov (United States)

    Sierra, Raymond G; Gati, Cornelius; Laksmono, Hartawan; Dao, E Han; Gul, Sheraz; Fuller, Franklin; Kern, Jan; Chatterjee, Ruchira; Ibrahim, Mohamed; Brewster, Aaron S; Young, Iris D; Michels-Clark, Tara; Aquila, Andrew; Liang, Mengning; Hunter, Mark S; Koglin, Jason E; Boutet, Sébastien; Junco, Elia A; Hayes, Brandon; Bogan, Michael J; Hampton, Christina Y; Puglisi, Elisabetta V; Sauter, Nicholas K; Stan, Claudiu A; Zouni, Athina; Yano, Junko; Yachandra, Vittal K; Soltis, S Michael; Puglisi, Joseph D; DeMirci, Hasan

    2016-01-01

    We describe a concentric-flow electrokinetic injector for efficiently delivering microcrystals for serial femtosecond X-ray crystallography analysis that enables studies of challenging biological systems in their unadulterated mother liquor. We used the injector to analyze microcrystals of Geobacillus stearothermophilus thermolysin (2.2-Å structure), Thermosynechococcus elongatus photosystem II (<3-Å diffraction) and Thermus thermophilus small ribosomal subunit bound to the antibiotic paromomycin at ambient temperature (3.4-Å structure).

  16. Light-harvesting features revealed by the structure of plant Photosystem I

    CERN Document Server

    Ben-Shem, A; Nelson, N; 10.1023/B:PRES.0000036881.23512.42

    2004-01-01

    Oxygenic photosynthesis is driven by two multi-subunit membrane protein complexes, Photosystem I and Photosystem II. In plants and green algae, both complexes are composed of two moieties: a reaction center (RC), where light-induced charge translocation occurs, and a peripheral antenna that absorbs light and funnels its energy to the reaction center. The peripheral antenna of PS I (LHC I) is composed of four gene products (Lhca 1-4) that are unique among the chlorophyll a/b binding proteins in their pronounced long-wavelength absorbance and in their assembly into dimers. The recently determined structure of plant Photosystem I provides the first relatively high- resolution structural model of a super-complex containing a reaction center and its peripheral antenna. We describe some of the structural features responsible for the unique properties of LHC I and discuss the advantages of the particular LHC I dimerization mode over monomeric or trimeric forms. In addition, we delineate some of the interactions betw...

  17. Differential responses of photosystems I and II to seasonal drought in two Ficus species

    Science.gov (United States)

    Zhang, Shubin; Huang, Wei; Zhang, Jiaolin; Cao, Kunfang

    2016-05-01

    Hemiepiphytic Ficus species exhibit more conservative water use strategy and are more drought-tolerant compared with their non-hemiepiphytic congeners, but a difference in the response of photosystem I (PSI) and photosystem II (PSII) to drought stress has not been documented to date. The enhancement of non-photochemical quenching (NPQ) and cyclic electron flow (CEF) have been identified as important mechanisms that protect the photosystems under drought conditions. Using the hemiepiphytic Ficus tinctoria and the non-hemiepiphytic Ficus racemosa, we studied the water status and the electron fluxes through PSI and PSII under seasonal water stress. Our results clearly indicated that the decline in the leaf predawn water potential (ψpd), the maximum photosynthetic rate (Amax) and the predawn maximum quantum yield of PSII (Fv/Fm) were more pronounced in F. racemosa than in F. tinctoria at peak drought. The Fv/Fm of F. racemosa was reduced to 0.69, indicating net photoinhibition of PSII. Concomitantly, the maximal photo-oxidizable P700 (Pm) decreased significantly in F. racemosa but remained stable in F. tinctoria. The fraction of non-photochemical quenching [Y(NPQ)] and the ratio of effective quantum yield of PSI to PSII [Y(I)/Y(II)] increased for both Ficus species at peak drought, with a stronger increase in F. racemosa. These results indicated that the enhancement of NPQ and the activation of CEF contributed to the photoprotection of PSI and PSII for both Ficus species under seasonal drought, particularly for F. racemosa.

  18. Characteristics of Photosystem II Behavior in Cotton (Gossypium hirsutum L.) Bract and Capsule Wall

    Institute of Scientific and Technical Information of China (English)

    ZHANG Ya-li; LUO Hong-hai; HU Yuan-yuan; Reto J. Strasser; ZHANG Wang-feng

    2013-01-01

    Though bract and capsule wall of boll in cotton (Gossypium hirsutum L.) have different photosynthetic capacities, the features of photosystem II (PS II) in these organs are scarce. In this paper, chlorophyll a lfuorescence emission was measured to investigate the difference in the photosynthetic apparatus of dark-acclimated (JIP-test) and light-acclimated (light-saturation pulse method) bract and capsule wall. Compared with leaves, the oxygen evolving system of non-foliar organs had lower efifciency. The pool size of PS II electron acceptor of non-foliar organs was small, and the photochemical activity of leaves was higher than that of the bract and capsule wall. In regard to the photosystem I (PS I) electron acceptor side, the pool size of end electron acceptors of leaves was larger, and the quantum yield of electron transport from QA (PS II primary plastoquinone acceptor) further than the PS I electron acceptors of leaves was higher than that of bract and capsule wall. In all green organs, the actual quantum yield of photochemistry decreased with light. The thermal dissipation fraction of light absorbed by the PS II antennae was the highest in bract and the lowest in capsule wall relative to leaves. Compared with leaves, capsule wall was characterized by less constitutive thermal dissipation and via dissipation as lfuorescence emission. These results suggested that lower PS II photochemical activity in non-foliar organs may be result from limitations at the donor side of PS II and the acceptor sides of both photosystems.

  19. Cryo-imaging of photosystems and phycobilisomes in Anabaena sp. PCC 7120 cells.

    Science.gov (United States)

    Steinbach, Gábor; Schubert, Félix; Kaňa, Radek

    2015-11-01

    Primary photosynthetic reactions take place inside thylakoid membrane where light-to-chemical energy conversion is catalyzed by two pigment-protein complexes, photosystem I (PSI) and photosystem II (PSII). Light absorption in cyanobacteria is increased by pigment-protein supercomplexes--phycobilisomes (PBSs) situated on thylakoid membrane surfaces that transfer excitation energy into both photosystems. We have explored the localization of PSI, PSII and PBSs in thylakoid membrane of native cyanobacteria cell Anabaena sp. 7120 by means of cryogenic confocal microscopy. We have adapted a conventional temperature controlling stage to an Olympus FV1000 confocal microscope. The presence of red shifted emission of chlorophylls from PSI has been confirmed by spectral measurements. Confocal fluorescence images of PSI (in a spectral range 710-750 nm), PSII (in a spectral range 690-705 nm) and PBSs (in a spectral range 650-680 nm) were recorded at low temperature. Co-localization of images showed spatial heterogeneity of PSI, PSII and PBSs over the thylakoid membrane, and three dominant areas were identified: PSI-PSII-PBS supercomplex area, PSII-PBS supercomplex area and PSI area. The observed results were discussed with regard to light-harvesting regulation in cyanobacteria.

  20. Probing the connection of PBSs to the photosystems in Spirulina platensis by artificially induced fluorescence fluctuations

    Energy Technology Data Exchange (ETDEWEB)

    Li Heng [Key Laboratory of Photochemistry, Center for Molecular Science, Institute of Chemistry, Chinese Academy of Sciences, Beijing 100080 (China); Yang Shuzhen [Key Laboratory of Photochemistry, Center for Molecular Science, Institute of Chemistry, Chinese Academy of Sciences, Beijing 100080 (China); Xie Jie [Key Laboratory of Photochemistry, Center for Molecular Science, Institute of Chemistry, Chinese Academy of Sciences, Beijing 100080 (China); Zhao Jingquan [Key Laboratory of Photochemistry, Center for Molecular Science, Institute of Chemistry, Chinese Academy of Sciences, Beijing 100080 (China)]. E-mail: zhaojq@iccas.ac.cn

    2007-01-15

    The molecular architecture and the structural connections of phycobilisomes (PBSs) to the photosystems in the intact cells of Spirulina platensis were studied by taking advantage of glycerol- and betaine-induced fluorescence fluctuations. Generally, with a selective excitation of C-phycocyanin (C-PC), glycerol could induce not only decoupling of PBSs from the photosystems but also of C-PC rods from allophycocyanin (APC) cores, while betaine could strengthen the connection of PBSs to the thylakoid membrane but induce a partial dissociation of PBS. On the other hand, glycerol did not exert an influence on the fluorescence spectra of the photosystems in isolated thylakoid membrane. Therefore, it was deduced that glycerol could provide a molecular environment to weaken the hydrophobic interactions of not only the L{sub CM} with the membrane but also the linker polypeptides to the water-soluble phycobiliproteins (C-PC and APC), while the betaine could strengthen the hydrophobic interaction of L{sub CM} with the membrane but weaken the electrostatic interaction of linker polypeptides to C-PC and/or APC.

  1. The experimental evolution of herbicide resistance in Chlamydomonas reinhardtii results in a positive correlation between fitness in the presence and absence of herbicides.

    Science.gov (United States)

    Vogwill, T; Lagator, M; Colegrave, N; Neve, P

    2012-10-01

    Pleiotropic fitness trade-offs will be key determinants of the evolutionary dynamics of selection for pesticide resistance. However, for herbicide resistance, empirical support for a fitness cost of resistance is mixed, and it is therefore also questionable what further ecological trade-offs can be assumed to apply to herbicide resistance. Here, we test the existence of trade-offs by experimentally evolving herbicide resistance in Chlamydomonas reinhardtii. Although fitness costs are detected for all herbicides, we find that, counterintuitively, the most resistant populations also have the lowest fitness costs as measured by growth rate in the ancestral environment. Furthermore, after controlling for differences in the evolutionary dynamics of resistance to different herbicides, we also detect significant positive correlations between resistance, fitness in the ancestral environment and cross-resistance to other herbicides. We attribute this to the highest levels of nontarget-site resistance being achieved by fixing mutations that more broadly affect cellular physiology, which results in both more cross-resistance and less overall antagonistic pleiotropy on maximum growth rate. Consequently, the lack of classical ecological trade-offs could present a major challenge for herbicide resistance management.

  2. Ultrafast Primary Reactions in the Photosystems of Oxygen-Evolving Organisms

    Science.gov (United States)

    Holzwarth, A. R.

    In oxygen-evolving photosynthetic organisms (plants, green algae, cyanobacteria), the primary steps of photosynthesis occur in two membrane-bound protein supercomplexes, Photosystem I (PS I) and Photosystem II (PS II), located in the thylakoid membrane (c.f. Fig. 7.1) along with two other important protein complexes, the cytochrome b6/f complex and the ATP-synthase [1]. Each of the photosystems consists of a reaction center (RC) where the photoinduced early electron transfer processes occur, of a so-called core antenna consisting of chlorophyll (Chl) protein complexes responsible for light absorption and ultrafast energy transfer to the RC pigments, and additional peripheral antenna complexes of various kinds that increase the absorption cross-section. The peripheral complexes are Chl a/b-protein complexes in higher plants and green algae (LHC I or LHC II for PS I or PS II, respectively) and so-called phycobilisomes in cyanobacteria and red algae [2-4]. The structures and light-harvesting functions of these antenna systems have been extensively reviewed [2, 5-9]. Recently, X-ray structures of both PS I and PS II antenna/RC complexes have been determined, some to atomic resolution. Although many details of the pigment content and organization of the RCs and antenna systems of PS I and PS II have been known before, the high resolution structures of the integral complexes allow us for the first time to try to understand structure/function relationships in detail. This article covers our present understanding of the ultrafast energy transfer and early electron transfer processes occurring in the photosystems of oxygen-evolving organisms. The main emphasis will be on the electron transfer processes. However, in both photosystems the kinetics of the energy transfer processes in the core antennae is intimately interwoven with the kinetics of the electron transfer steps. Since both types of processes occur on a similar time scale, their kinetics cannot be considered

  3. Deletion of PsbM in tobacco alters the QB site properties and the electron flow within photosystem II.

    Science.gov (United States)

    Umate, Pavan; Schwenkert, Serena; Karbat, Izhar; Dal Bosco, Cristina; Mlcòchová, Lada; Volz, Stefanie; Zer, Hagit; Herrmann, Reinhold G; Ohad, Itzhak; Meurer, Jörg

    2007-03-30

    Photosystem II, the oxygen-evolving complex of photosynthetic organisms, includes an intriguingly large number of low molecular weight polypeptides, including PsbM. Here we describe the first knock-out of psbM using a transplastomic, reverse genetics approach in a higher plant. Homoplastomic Delta psbM plants exhibit photoautotrophic growth. Biochemical, biophysical, and immunological analyses demonstrate that PsbM is not required for biogenesis of higher order photosystem II complexes. However, photosystem II is highly light-sensitive, and its activity is significantly decreased in Delta psbM, whereas kinetics of plastid protein synthesis, reassembly of photosystem II, and recovery of its activity are comparable with the wild type. Unlike wild type, phosphorylation of the reaction center proteins D1 and D2 is severely reduced, whereas the redox-controlled phosphorylation of photosystem II light-harvesting complex is reversely regulated in Delta psbM plants because of accumulation of reduced plastoquinone in the dark and a limited photosystem II-mediated electron transport in the light. Charge recombination in Delta psbM measured by thermoluminescence oscillations significantly differs from the 2/6 patterns in the wild type. A simulation program of thermoluminescence oscillations indicates a higher Q(B)/Q(-)(B) ratio in dark-adapted mutant thylakoids relative to the wild type. The interaction of the Q(A)/Q(B) sites estimated by shifts in the maximal thermoluminescence emission temperature of the Q band, induced by binding of different herbicides to the Q(B) site, is changed indicating alteration of the activation energy for back electron flow. We conclude that PsbM is primarily involved in the interaction of the redox components important for the electron flow within, outward, and backward to photosystem II.

  4. Sulphate, more than a nutrient, protects the microalga Chlamydomonas moewusii from cadmium toxicity

    Energy Technology Data Exchange (ETDEWEB)

    Mera, Roi; Torres, Enrique, E-mail: torres@udc.es; Abalde, Julio

    2014-03-01

    Highlights: • Sulphate effect on cadmium toxicity in the microalga Chlamydomonas moewusii Gerloff. • Cadmium increases the sulphur requirements in Chlamydomonas moewusii. • Kinetic coefficients for sulphate utilization and cadmium effect on them. • Sulphate and cadmium influence on the biosynthesis of low-molecular mass thiols. • Cadmium toxicity reduction by sulphate due to higher biosynthesis of thiols. - Abstract: Sulphur is an essential macroelement that plays important roles in living organisms. The thiol rich sulphur compounds, such as cysteine, γ-Glu–Cys, glutathione and phytochelatins participate in the tolerance mechanisms against cadmium toxicity. Plants, algae, yeasts and most prokaryotes cover their demand for reduced sulphur by reduction of inorganic sulphate. The aim of this study was to investigate, using a bifactorial experimental design, the effect of different sulphate concentrations in the nutrient solution on cadmium toxicity in the freshwater microalga Chlamydomonas moewusii. Cell growth, kinetic parameters of sulphate utilization and intracellular concentrations of low-molecular mass thiol compounds were determined. A mathematical model to describe the growth of this microalga based on the effects of sulphate and cadmium was obtained. An ANOVA revealed an interaction between them, 16% of the effect sizes was explained by this interaction. A higher amount of sulphate in the culture medium allowed a higher cadmium tolerance due to an increase in the thiol compound biosynthesis. The amount of low-molecular mass thiol compounds, mainly phytochelatins, synthesized by this microalga was significantly dependent on the sulphate and cadmium concentrations; the higher phytochelatin content was obtained in cultures with 4 mg Cd/L and 1 mM sulphate. The maximum EC{sub 50} value (based on nominal cadmium concentration) reached for this microalga was 4.46 ± 0.42 mg Cd/L when the sulphate concentration added to the culture medium was also 1 m

  5. Supercomplexes of plant photosystem I with cytochrome b6f, light-harvesting complex II and NDH.

    Science.gov (United States)

    Yadav, K N Sathish; Semchonok, Dmitry A; Nosek, Lukáš; Kouřil, Roman; Fucile, Geoffrey; Boekema, Egbert J; Eichacker, Lutz A

    2017-01-01

    Photosystem I (PSI) is a pigment-protein complex required for the light-dependent reactions of photosynthesis and participates in light-harvesting and redox-driven chloroplast metabolism. Assembly of PSI into supercomplexes with light harvesting complex (LHC) II, cytochrome b6f (Cytb6f) or NAD(P)H dehydrogenase complex (NDH) has been proposed as a means for regulating photosynthesis. However, structural details about the binding positions in plant PSI are lacking. We analyzed large data sets of electron microscopy single particle projections of supercomplexes obtained from the stroma membrane of Arabidopsis thaliana. By single particle analysis, we established the binding position of Cytb6f at the antenna side of PSI. The rectangular-shaped Cytb6f dimer binds at the side where Lhca1 is located. The complex binds with its short side rather than its long side to PSI, which may explain why these supercomplexes are difficult to purify and easily disrupted. Refined analysis of the interaction between PSI and the NDH complex indicates that in total up to 6 copies of PSI can arrange with one NDH complex. Most PSI-NDH supercomplexes appeared to have 1-3 PSI copies associated. Finally, the PSI-LHCII supercomplex was found to bind an additional LHCII trimer at two positions on the LHCI side in Arabidopsis. The organization of PSI, either in a complex with NDH or with Cytb6f, may improve regulation of electron transport by the control of binding partners and distances in small domains.

  6. Dynamic curvature regulation accounts for the symmetric and asymmetric beats of Chlamydomonas flagella

    CERN Document Server

    Sartori, Pablo; Scholich, Andre; Jülicher, Frank; Howard, Jonathon

    2015-01-01

    Axonemal dyneins are the molecular motors responsible for the beating of cilia and flagella. These motors generate sliding forces between adjacent microtubule doublets within the axoneme, the motile cytoskeletal structure inside the flagellum. To create regular, oscillatory beating patterns, the activities of the axonemal dyneins must be coordinated both spatially and temporally. It is thought that coordination is mediated by stresses or strains that build up within the moving axoneme, but it is not known which components of stress or strain are involved, nor how they feed back on the dyneins. To answer this question, we used isolated, reactivate axonemes of the unicellular alga Chlamydomonas as a model system. We derived a theory for beat regulation in a two-dimensional model of the axoneme. We then tested the theory by measuring the beat waveforms of wild type axonemes, which have asymmetric beats, and mutant axonemes, in which the beat is nearly symmetric, using high-precision spatial and temporal imaging....

  7. Kinesin-II is not essential for mitosis and cell growth in Chlamydomonas.

    Science.gov (United States)

    Matsuura, Kumi; Lefebvre, Paul A; Kamiya, Ritsu; Hirono, Masafumi

    2002-08-01

    The FLA10 gene product (Fla10p) in Chlamydomonas, a heterotrimeric kinesin-II, plays a crucial role in flagellar assembly as a motor protein driving intraflagellar transport. This protein has also been suggested to play a role in mitosis based on its localization to mitotic spindle. A role for Fla10p in mitosis has been difficult to test because to date only conditional (temperature-sensitive) mutant alleles were available, and it is not known whether these retain residual function for mitosis at the non-permissive temperature. In this report, we describe a null allele of fla10 produced by insertional mutagenesis. This mutant does not assemble flagella, but proliferates at a rate identical to that of wild type cells. Observation of microtubule organization in the cell body revealed that normal mitotic spindles are formed in dividing mutant cells. Thus, we conclude that FLA10 kinesin plays no significant roles in mitosis.

  8. Inhomogeneous distribution of Chlamydomonas in a cylindrical container with a bubble plume.

    Science.gov (United States)

    Nonaka, Yuki; Kikuchi, Kenji; Numayama-Tsuruta, Keiko; Kage, Azusa; Ueno, Hironori; Ishikawa, Takuji

    2016-01-19

    Swimming microalgae show various taxes, such as phototaxis and gravitaxis, which sometimes result in the formation of a cell-rich layer or a patch in a suspension. Despite intensive studies on the effects of shear flow and turbulence on the inhomogeneous distribution of microalgae, the effect of a bubble plume has remained unclear. In this study, we used Chlamydomonas as model microalgae, and investigated the spatial distribution of cells in a cylindrical container with a bubble plume. The results illustrate that cells become inhomogeneously distributed in the suspension due to their motility and photo-responses. A vortical ring distribution was observed below the free surface when the bubble flow rate was sufficiently small. We performed a scaling analysis on the length scale of the vortical ring, which captured the main features of the experimental results. These findings are important in understanding transport phenomena in a microalgae suspension with a bubble plume.

  9. Trichloroacetate affects the EPR SignalⅡslow and SignalⅠin the thylakoid of Chlamydomonas reinhardtii

    Institute of Scientific and Technical Information of China (English)

    2000-01-01

    One electron paramagnetic resonance (EPR) signal, named SignalⅡslow, originates from the oxidized Tyrosine 160 (YDo) of D2 polypeptide of photosystemⅡ reaction center. After adding high concentration trichloroacetate (TCA) to the Chlamydomonas reinhardtii thylakoid suspension, this signal was abolished in a minute. Treatment of TCA also removes a few of polypeptides, including three extrinsic polypeptides of oxygen-evolving complex, from the thylakoid membrane. Based upon the analysis of the microenvironment around YD with a three-dimensional model, it is indicated that relatively high hydrophobicity of this microenvironment may be the essential prerequisite for TCA to affect YD. It has been observed that TCA treatment also retards the decay of the SignalⅠ, produced by the oxidized reaction center chlorophyll dimer (P700+) of photosys- temⅠ.

  10. A chloroplast pathway for the de novo biosynthesis of triacylglycerol in Chlamydomonas reinhardtii

    Energy Technology Data Exchange (ETDEWEB)

    Fan, J.; Xu, C.; Andre, C.

    2011-06-23

    Neutral lipid metabolism has been extensively studied in yeast, plants and mammals. In contrast, little information is available regarding the biochemical pathway, enzymes and regulatory factors involved in the biosynthesis of triacylglycerol (TAG) in microalgae. In the conventional TAG biosynthetic pathway widely accepted for yeast, plants and mammals, TAG is assembled in the endoplasmic reticulum (ER) from its immediate precursor diacylglycerol (DAG) made by ER-specific acyltransferases, and is deposited exclusively in lipid droplets in the cytosol. Here, we demonstrated that the unicellular microalga Chlamydomonas reinhardtii employs a distinct pathway that uses DAG derived almost exclusively from the chloroplast to produce TAG. This unique TAG biosynthesis pathway is largely dependent on de novo fatty acid synthesis, and the TAG formed in this pathway is stored in lipid droplets in both the chloroplast and the cytosol. These findings have wide implications for understanding TAG biosynthesis and storage and other areas of lipid metabolism in microalgae and other organisms.

  11. Efficient phototrophic production of a high-value sesquiterpenoid from the eukaryotic microalga Chlamydomonas reinhardtii.

    Science.gov (United States)

    Lauersen, Kyle J; Baier, Thomas; Wichmann, Julian; Wördenweber, Robin; Mussgnug, Jan H; Hübner, Wolfgang; Huser, Thomas; Kruse, Olaf

    2016-11-01

    The heterologous expression of terpene synthases in microbial hosts has opened numerous possibilities for bioproduction of desirable metabolites. Photosynthetic microbial hosts present a sustainable alternative to traditional fermentative systems, using freely available (sun)light and carbon dioxide as inputs for bio-production. Here, we report the expression of a patchoulol synthase from Pogostemon cablin Benth in the model green microalga Chlamydomonas reinhardtii. The sesquiterpenoid patchoulol was produced from the alga and was used as a marker of sesquiterpenoid production capacity. A novel strategy for gene loading was employed and patchoulol was produced up to 922±242µgg(-1) CDW in six days. We additionally investigated the effect of carbon source on sesquiterpenoid productivity from C. reinhardtii in scale-up batch cultivations. It was determined that up to 1.03mgL(-1) sesquiterpenoid products could be produced in completely photoautotrophic conditions and that the alga exhibited altered sesquiterpenoid production metabolism related to carbon source.

  12. Quantification of phytochelatins in Chlamydomonas reinhardtii using ferrocene-based derivatization.

    Science.gov (United States)

    Bräutigam, Anja; Bomke, Susanne; Pfeifer, Thorben; Karst, Uwe; Krauss, Gerd-Joachim; Wesenberg, Dirk

    2010-08-01

    A method for the identification and quantification of canonic and isoforms of phytochelatins (PCs) from Chlamydomonas reinhardtii was developed. After disulfide reduction with tris(2-carboxyethyl)phosphine (TCEP) PCs were derivatized with ferrocenecarboxylic acid (2-maleimidoyl)ethylamide (FMEA) in order to avoid oxidation of the free thiol functions during analysis. Liquid chromatography (LC) coupled to electrospray mass spectrometry (ESI-MS) and inductively coupled plasma-mass spectrometry (ICP-MS) was used for rapid and quantitative analysis of the precolumn derivatized PCs. PC(2-4), CysGSH, CysPC(2-4), CysPC(2)desGly, CysPC(2)Glu and CysPC(2)Ala were determined in the algal samples depending on the exposure of the cells to cadmium ions.

  13. Investigating the link between fermentative metabolism and hydrogen production in the unicellular green alga Chlamydomonas reinhardtii

    Energy Technology Data Exchange (ETDEWEB)

    Burgess, S.J.; Nixon, P.J. [Imperial College London (United Kingdom)

    2010-07-01

    In the model green alga Chlamydomonas reinhardtii, the electrons required for hydrogen production can come from both the biophotolysis of water and from the fermentation of carbohydrate reserves. Anoxia leads to the activation of several fermentative pathways, which produce a number of end products including formic, malic and acetic acid along with ethanol, carbon dioxide and hydrogen. It has been proposed that by switching off competing fermentative pathways hydrogen production can be increased. Therefore the aim of this study was to devise an experimental strategy to down-regulate the expression of enzymes thought to control C. reinhardtii's fermentative metabolism. We demonstrate here that it is possible to use artificial microRNA (amiRNA) technology to generate knock-down mutants with reduced expression of pyruvate formate lyase (PFL1), a key fermentative enzyme in C. reinhardtii. This work opens up new possibilities to improve hydrogen yields through metabolic engineering. (orig.)

  14. Cell types and their status in Chlamydomonas-like algae (Chlorophyceae on agar medium culture

    Directory of Open Access Journals (Sweden)

    M.М. Pavlovska

    2014-04-01

    Full Text Available The classification of cell types under agar culture was proposed. Six cell morphotypes were allocated. The statuses were identified depending on the reduction of monade attributes of cells. The variants of transition from one cell morphotype to another under dissolving mucilage were shown. The monade, cocciod, palmeloid and gloeocysta morphotypes approximately equally represented in all clades. The asterococcus and mucogleocysta morphotypes presented only in Reinhardtinia аnd Oogamochlamydinia clades. Any morphotype isn’t typical for all clades of Chlamydomonas-like algae at once. The most of morphotypes numbers (5 from 6 are presented in Reinhardtinia clade. This demonstrates the diversity of the Reinhardtinia clade species. There are only one morphotype presented in Polytominia and Monadinia clades. There are four morphotypes presented in Oogamochlamydinia clade, three – in Moewusinia, two morphotypes – in Chloromonadinia.

  15. Stable expression of a bifunctional diterpene synthase in the chloroplast of Chlamydomonas reinhardtii

    DEFF Research Database (Denmark)

    Zedler, Julie A Z; Gangl, Doris; Hamberger, Björn Robert;

    2015-01-01

    Chlamydomonas reinhardtii has been shown to hold significant promise as a production platform for recombinant proteins, but transformation of the nuclear genome is still a non-trivial process due to random gene insertion and frequent silencing. Insertion of transgenes into the chloroplasts...... is an alternative strategy, and we report here the stable expression of a large (91 kDa) protein in the chloroplast using a recently developed low-cost transformation protocol. Moreover, selection of transformants is based on restoration of prototrophy using an endogenous gene (psbH) as the marker, thereby allowing...... the generation of transgenic lines without the use of antibiotic-resistance genes. Here, we have expressed a bifunctional diterpene synthase in C. reinhardtii chloroplasts. Homoplasmic transformants were obtained with the expressed enzyme accounting for 3.7 % of total soluble protein. The enzyme was purified...

  16. DNA-free two-gene knockout in Chlamydomonas reinhardtii via CRISPR-Cas9 ribonucleoproteins.

    Science.gov (United States)

    Baek, Kwangryul; Kim, Duk Hyoung; Jeong, Jooyeon; Sim, Sang Jun; Melis, Anastasios; Kim, Jin-Soo; Jin, EonSeon; Bae, Sangsu

    2016-07-28

    Microalgae are versatile organisms capable of converting CO2, H2O, and sunlight into fuel and chemicals for domestic and industrial consumption. Thus, genetic modifications of microalgae for enhancing photosynthetic productivity, and biomass and bio-products generation are crucial for both academic and industrial applications. However, targeted mutagenesis in microalgae with CRISPR-Cas9 is limited. Here we report, a one-step transformation of Chlamydomonas reinhardtii by the DNA-free CRISPR-Cas9 method rather than plasmids that encode Cas9 and guide RNAs. Outcome was the sequential CpFTSY and ZEP two-gene knockout and the generation of a strain constitutively producing zeaxanthin and showing improved photosynthetic productivity.

  17. Zinc Deficiency Impacts CO2 Assimilation and Disrupts Copper Homeostasis in Chlamydomonas reinhardtii*

    Science.gov (United States)

    Malasarn, Davin; Kropat, Janette; Hsieh, Scott I.; Finazzi, Giovanni; Casero, David; Loo, Joseph A.; Pellegrini, Matteo; Wollman, Francis-André; Merchant, Sabeeha S.

    2013-01-01

    Zinc is an essential nutrient because of its role in catalysis and in protein stabilization, but excess zinc is deleterious. We distinguished four nutritional zinc states in the alga Chlamydomonas reinhardtii: toxic, replete, deficient, and limited. Growth is inhibited in zinc-limited and zinc-toxic cells relative to zinc-replete cells, whereas zinc deficiency is visually asymptomatic but distinguished by the accumulation of transcripts encoding ZIP family transporters. To identify targets of zinc deficiency and mechanisms of zinc acclimation, we used RNA-seq to probe zinc nutrition-responsive changes in gene expression. We identified genes encoding zinc-handling components, including ZIP family transporters and candidate chaperones. Additionally, we noted an impact on two other regulatory pathways, the carbon-concentrating mechanism (CCM) and the nutritional copper regulon. Targets of transcription factor Ccm1 and various CAH genes are up-regulated in zinc deficiency, probably due to reduced carbonic anhydrase activity, validated by quantitative proteomics and immunoblot analysis of Cah1, Cah3, and Cah4. Chlamydomonas is therefore not able to grow photoautotrophically in zinc-limiting conditions, but supplementation with 1% CO2 restores growth to wild-type rates, suggesting that the inability to maintain CCM is a major consequence of zinc limitation. The Crr1 regulon responds to copper limitation and is turned on in zinc deficiency, and Crr1 is required for growth in zinc-limiting conditions. Zinc-deficient cells are functionally copper-deficient, although they hyperaccumulate copper up to 50-fold over normal levels. We suggest that zinc-deficient cells sequester copper in a biounavailable form, perhaps to prevent mismetallation of critical zinc sites. PMID:23439652

  18. A revised mineral nutrient supplement increases biomass and growth rate in Chlamydomonas reinhardtii

    Science.gov (United States)

    Kropat, Janette; Hong-Hermesdorf, Anne; Casero, David; Ent, Petr; Castruita, Madeli; Pellegrini, Matteo; Merchant, Sabeeha S.; Malasarn, Davin

    2011-01-01

    Summary Interest in exploiting algae as a biofuel source and the role of inorganic nutrient deficiency in inducing triacylglyceride (TAG) accumulation in cells necessitates a strategy to efficiently formulate species-specific culture media that can easily be manipulated. Using the reference organism Chlamydomonas reinhardtii, we tested the hypothesis that modeling trace element supplements after the cellular ionome would result in optimized cell growth. We determined the trace metal content of several commonly used Chlamydomonas strains in various culture conditions and developed a revised trace element solution to parallel these measurements. Comparison of cells growing in the revised supplement versus a traditional trace element solution revealed faster growth rates and higher maximum cell densities with the revised recipe. RNA-seq analysis of cultures growing in the traditional versus revised medium suggest that the variation in transcriptomes was smaller than that found between different wild-type strains grown in traditional Hutner’s supplement. Visual observation did not reveal defects in cell motility or mating efficiency in the new supplement. Ni2+-inducible expression from the CYC6 promoter remained a useful tool, albeit with an increased requirement for Ni2+ because of the introduction of an EDTA buffer system in the revised medium. Other advantages include more facile preparation of trace element stock solutions, a reduction in total chemical use, a more consistent batch-to-batch formulation, and long-term stability (tested up to 5 years). Under the new growth regime, we analyzed cells growing under different macro- and micronutrient-deficiencies. TAG accumulation in N deficiency is comparable in the new medium. Fe and Zn deficiency also induced TAG accumulation, as suggested by Nile Red staining. This approach can be used to efficiently optimize culture conditions for other algal species to improve growth and to assay cell physiology. PMID:21309872

  19. CO2 acquisition in Chlamydomonas acidophila is influenced mainly by CO2, not phosphorus, availability.

    Science.gov (United States)

    Spijkerman, Elly; Stojkovic, Slobodanka; Beardall, John

    2014-09-01

    The extremophilic green microalga Chlamydomonas acidophila grows in very acidic waters (pH 2.3-3.4), where CO2 is the sole inorganic carbon source. Previous work has revealed that the species can accumulate inorganic carbon (Ci) and exhibits high affinity CO2 utilization under low-CO2 (air-equilibrium) conditions, similar to organisms with an active CO2 concentrating mechanism (CCM), whereas both processes are down-regulated under high CO2 (4.5 % CO2) conditions. Responses of this species to phosphorus (Pi)-limited conditions suggested a contrasting regulation of the CCM characteristics. Therefore, we measured external carbonic anhydrase (CAext) activities and protein expression (CAH1), the internal pH, Ci accumulation, and CO2-utilization in cells adapted to high or low CO2 under Pi-replete and Pi-limited conditions. Results reveal that C. acidophila expressed CAext activity and expressed a protein cross-reacting with CAH1 (the CAext from Chlamydomonas reinhardtii). Although the function of this CA remains unclear, CAext activity and high affinity CO2 utilization were the highest under low CO2 conditions. C. acidophila accumulated Ci and expressed the CAH1 protein under all conditions tested, and C. reinhardtii also contained substantial amounts of CAH1 protein under Pi-limitation. In conclusion, Ci utilization is optimized in C. acidophila under ecologically relevant conditions, which may enable optimal survival in its extreme Ci- and Pi-limited habitat. The exact physiological and biochemical acclimation remains to be further studied.

  20. Survival and proliferation characteristics of the microalga Chlamydomonas sp. ICE-L after hypergravitational stress pretreatment

    Science.gov (United States)

    Gao, Zhengquan; Li, Demao; Meng, Chunxiao; Xu, Dong; Zhang, Xiaowen; Ye, Naihao

    2013-09-01

    Seeking extraterrestrial life, transferring between planets, even migrating to other planets attracts more and more attention of public and scientists. However, to make it clear for the ability to survive the forces studies is prerequisite to enable the speculations by natural means. Gravity is a critical force involved in all the life on Earth and, possibly, others planets. Organisms have been grown in microgravity habitats and in centrifuges to characterize the biological response to a range of gravitational forces and radiation levels in space and on Earth. However, little is known about the profiles of eukaryotic life under conditions of hyperacceleration attributable to extreme gravities. In this study, a eukaryotic extremophile, the Antarctic green microalga Chlamydomonas sp. ICE-L, showed amazing proliferation capacity during and after hypergravitational stress for 30 min to 48 h at 110,200, 423,400, and 670,800g. These extreme gravities also had profound effects on viability, reproduction rate, photosynthesis efficiency, and gene transcriptional expression of this microalga. Most notably, all three supergravities efficiently stimulated algal cell division, but the greater the centrifugal force and the longer the duration of treatment, the lower the viable rate and breeding potential of samples in the following incubation. These results illustrated Chlamydomonas sp. ICE-L is a useful eukaryotic model system candidate for space research. Further studies could provide new insight into the physical limits of life and its evolution and enhance the possibility for interstellar space travel and the quest for extraterrestrial life according to panspermia theory. Also, it indicated that life come from the outer space is not always prokaryotes but may be eukaryotes.

  1. A Chlamydomonas-derived Human Papillomavirus 16 E7 vaccine induces specific tumor protection.

    Directory of Open Access Journals (Sweden)

    Olivia C Demurtas

    Full Text Available BACKGROUND: The E7 protein of the Human Papillomavirus (HPV type 16, being involved in malignant cellular transformation, represents a key antigen for developing therapeutic vaccines against HPV-related lesions and cancers. Recombinant production of this vaccine antigen in an active form and in compliance with good manufacturing practices (GMP plays a crucial role for developing effective vaccines. E7-based therapeutic vaccines produced in plants have been shown to be active in tumor regression and protection in pre-clinical models. However, some drawbacks of in whole-plant vaccine production encouraged us to explore the production of the E7-based therapeutic vaccine in Chlamydomonas reinhardtii, an organism easy to grow and transform and fully amenable to GMP guidelines. METHODOLOGY/PRINCIPAL FINDINGS: An expression cassette encoding E7GGG, a mutated, attenuated form of the E7 oncoprotein, alone or as a fusion with affinity tags (His6 or FLAG, under the control of the C. reinhardtii chloroplast psbD 5' UTR and the psbA 3' UTR, was introduced into the C. reinhardtii chloroplast genome by homologous recombination. The protein was mostly soluble and reached 0.12% of total soluble proteins. Affinity purification was optimized and performed for both tagged forms. Induction of specific anti-E7 IgGs and E7-specific T-cell proliferation were detected in C57BL/6 mice vaccinated with total Chlamydomonas extract and with affinity-purified protein. High levels of tumor protection were achieved after challenge with a tumor cell line expressing the E7 protein. CONCLUSIONS: The C. reinhardtii chloroplast is a suitable expression system for the production of the E7GGG protein, in a soluble, immunogenic form. The production in contained and sterile conditions highlights the potential of microalgae as alternative platforms for the production of vaccines for human uses.

  2. Photosystem I shows a higher tolerance to sorbitol-induced osmotic stress than photosystem II in the intertidal macro-algae Ulva prolifera (Chlorophyta).

    Science.gov (United States)

    Gao, Shan; Zheng, Zhenbing; Gu, Wenhui; Xie, Xiujun; Huan, Li; Pan, Guanghua; Wang, Guangce

    2014-10-01

    The photosynthetic performance of the desiccation-tolerant, intertidal macro-algae Ulva prolifera was significantly affected by sorbitol-induced osmotic stress. Our results showed that photosynthetic activity decreased significantly with increases in sorbitol concentration. Although the partial activity of both photosystem I (PS I) and photosystem II (PS II) was able to recover after 30 min of rehydration, the activity of PS II decreased more rapidly than PS I. At 4 M sorbitol concentration, the activity of PS II was almost 0 while that of PS I was still at about one third of normal levels. Following prolonged treatment with 1 and 2 M sorbitol, the activity of PS I and PS II decreased slowly, suggesting that the effects of moderate concentrations of sorbitol on PS I and PS II were gradual. Interestingly, an increase in non-photochemical quenching occurred under these conditions in response to moderate osmotic stress, whereas it declined significantly under severe osmotic stress. These results suggest that photoprotection in U. prolifera could also be induced by moderate osmotic stress. In addition, the oxidation of PS I was significantly affected by osmotic stress. P700(+) in the thalli treated with high concentrations of sorbitol could still be reduced, as PS II was inhibited by 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU), but it could not be fully oxidized. This observation may be caused by the higher quantum yield of non-photochemical energy dissipation in PS I due to acceptor-side limitation (Y(NA)) during rehydration in seawater containing DCMU.

  3. D1-arginine257 mutants (R257E, K, and Q) of Chlamydomonas reinhardtii have a lowered QB redox potential: analysis of thermoluminescence and fluorescence measurements.

    Science.gov (United States)

    Rose, Stuart; Minagawa, Jun; Seufferheld, Manfredo; Padden, Sean; Svensson, Bengt; Kolling, Derrick R J; Crofts, Antony R; Govindjee

    2008-01-01

    Arginine257 (R257), in the de-helix that caps the Q(B) site of the D1 protein, has been shown by mutational studies to play a key role in the sensitivity of Photosystem II (PS II) to bicarbonate-reversible binding of the formate anion. In this article, the role of this residue has been further investigated through D1 mutations (R257E, R257Q, and R257K) in Chlamydomonas reinhardtii. We have investigated the activity of the Q(B) site by studying differences from wild type on the steady-state turnover of PS II, as assayed through chlorophyll (Chl) a fluorescence yield decay after flash excitation. The effects of p-benzoquinone (BQ, which oxidizes reduced Q(B), Q(B)(-) ) and 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU, which blocks electron flow from Q(A)(-) to Q(B)) were measured. The equilibrium constants of the two-electron gate were obtained through thermoluminescence measurements. The thermoluminescence properties were changed in the mutants, especially when observed after pretreatment with 100 microM BQ. A theoretical analysis of the thermoluminescence data, based mainly on the recombination pathways model of Rappaport et al. (2005), led to the conclusion that the free-energy difference for the recombination of Q(B)(-) with S(2) was reduced by 20-40 mV in the three mutants (D1-R257K, D1-R257Q, and D1-R257E); this was interpreted to be due to a lowering of the redox potential of Q(B)/Q(B)(-). Further, since the recombination of Q(A)(-) with S(2) was unaffected, we suggest that no significant change in redox potential of Q(A)/Q(A)(-) occurred in these three mutants. The maximum variable Chl a fluorescence yield is lowered in the mutants, in the order R257K > R257Q > R257E, compared to wild type. Our analysis of the binary oscillations in Chl a fluorescence following pretreatment of cells with BQ showed that turnover of the Q(B) site was relatively unaffected in the three mutants. The mutant D1-R257E had the lowest growth rate and steady-state activity and showed

  4. Variety in excitation energy transfer processes from phycobilisomes to photosystems I and II.

    Science.gov (United States)

    Ueno, Yoshifumi; Aikawa, Shimpei; Niwa, Kyosuke; Abe, Tomoko; Murakami, Akio; Kondo, Akihiko; Akimoto, Seiji

    2017-02-09

    The light-harvesting antennas of oxygenic photosynthetic organisms capture light energy and transfer it to the reaction centers of their photosystems. The light-harvesting antennas of cyanobacteria and red algae, called phycobilisomes (PBSs), supply light energy to both photosystem I (PSI) and photosystem II (PSII). However, the excitation energy transfer processes from PBS to PSI and PSII are not understood in detail. In the present study, the energy transfer processes from PBS to PSs in various cyanobacteria and red algae were examined in vivo by selectively exciting their PSs or PBSs, and measuring the resulting picosecond to nanosecond time-resolved fluorescences. By observing the delayed fluorescence spectrum of PBS-selective excitation in Arthrospira platensis, we demonstrated that energy transfer from PBS to PSI via PSII (PBS→PSII→PSI transfer) occurs even for PSI trimers. The contribution of PBS→PSII→PSI transfer was species dependent, being largest in the wild-type of red alga Pyropia yezoensis (formerly Porphyra yezoensis) and smallest in Synechococcus sp. PCC 7002. Comparing the time-resolved fluorescence after PSs- and PBS-selective excitation, we revealed that light energy flows from CP43 to CP47 by energy transfer between the neighboring PSII monomers in PBS-PSII supercomplexes. We also suggest two pathways of energy transfer: direct energy transfer from PBS to PSI (PBS→PSI transfer) and indirect transfer through PSII (PBS→PSII→PSI transfer). We also infer that PBS→PSI transfer conveys light energy to a lower-energy red chlorophyll than PBS→PSII→PSI transfer.

  5. Responsibility of regulatory gene expression and repressed protein synthesis for triacylglycerol accumulation on sulfur-starvation in Chlamydomonas reinhardtii

    OpenAIRE

    2014-01-01

    Triacylglycerol (TG) synthesis is induced for energy and carbon storage in algal cells under nitrogen(N)-starved conditions, and helps prevent reactive oxygen species (ROS) production through fatty acid synthesis that consumes excessive reducing power. Here, the regulatory mechanism for the TG content in sulfur(S)-starved cells of Chlamydomonas reinhardtii was examined, in comparison to that in N- or phosphorus(P)-starved cells. S- and N- starved cells exhibited markedly increased TG contents...

  6. Electron transport, Photosystem-2 reaction centers and chlorophyll-protein complexes of thylakoids of drought resistant and sensitive Lupin piants.

    Science.gov (United States)

    Meyer, S; de Kouchkovsky, Y

    1993-07-01

    Two genotypes ofLupinus albus L., resistant and susceptible to drought, were subjected to water deficiency for up to two weeks. Such treatment progressively lowered the leaf water content from about 85% to about 60% (water potential from -0.8 to -4.3 MPa). Light-saturation curves of the uncoupled electron transport were analyzed according to a simple kinetic model of separated or connected reversible photoreactions. It gives an extrapolated maximum rate (Vmax) and the efficiency for capturing light (Im, which is the light intensity at Vmax/2). For Photosystem 2, Vmax and, less markedly, Im, declined with increasing severity of drought treatment; the artificial donor, diphenylcarbazide, could not restore the activity. One cause of this Photosystem 2 inhibition could be the loss of active Photosystem 2 centers. Indeed, their concentration relative to chlorophyll, estimated by flash-induced reduction of dimethylquinone, was halved by a medium stress. To the extent that it was still not restored by diphenylcarbazide, the site of Photosystem 2 inactivation must have been close to the photochemical trap, after water oxidation and before or at plastoquinone pool. By relating electron transport rate to active centers instead of chlorophyll, no inhibition by drought was detected. Therefore, water stress inactivates specifically Photosystem 2, without impairing a downhill thermal step of electron transport. On the other hand, the decrease of Im suggests that antennae connected to inactive centers may transfer their excitation energy to active neighbors, which implies that antenna network remains essentially intact. Gel electrophoresis confirmed that the apoproteins of the pigment complexes were well conserved. In conclusion, the inactivation of Photosystem 2 may not be a physical loss of its centers and core antennae but probably reflects protein alterations or conformational changes. These may result from the massive decrease of lipids induced by drought (Meyer et al. 1992

  7. Light and Heat Induced Denaturation of Photosystem Ⅱ Core Antenna Complex CP47

    Institute of Scientific and Technical Information of China (English)

    1999-01-01

    Light and heat induced denaturation of CP47, the core antenna complex of photosystem Ⅱ purified from spinach, were investigated using absorption and circular dichroism spectra.Light caused the destruction of chlorophyll a and excitonic interaction of chlorophyll a in CP47, while the protein secondary structure was not apparently changed.Heat induced the destruction of protein secondary structure and excitonic interaction of chlorophyll a, but the chlorophyll a molecule was not damaged.The results suggest that both the chlorophyll a molecular structure and the protein native conformation are necessary for excitonic interaction of chlorophyll a and the energy transfer function of the chlorophyll a binding protein.

  8. Isolation of monomeric photosystem II that retains the subunit PsbS.

    Science.gov (United States)

    Haniewicz, Patrycja; De Sanctis, Daniele; Büchel, Claudia; Schröder, Wolfgang P; Loi, Maria Cecilia; Kieselbach, Thomas; Bochtler, Matthias; Piano, Dario

    2013-12-01

    Photosystem II has been purified from a transplastomic strain of Nicotiana tabacum according to two different protocols. Using the procedure described in Piano et al. (Photosynth Res 106:221-226, 2010) it was possible to isolate highly active PSII composed of monomers and dimers but depleted in their PsbS protein content. A "milder" procedure than the protocol reported by Fey et al. (Biochim Biophys Acta 1777:1501-1509, 2008) led to almost exclusively monomeric PSII complexes which in part still bind the PsbS protein. This finding might support a role for PSII monomers in higher plants.

  9. Molecular dynamics studies of pathways of water movement in cyanobacterial photosystem II

    Energy Technology Data Exchange (ETDEWEB)

    Gabdulkhakov, A. G., E-mail: azat@vega.protres.ru; Kljashtorny, V. G.; Dontsova, M. V. [Russian Academy of Sciences, Institute of Protein Research (Russian Federation)

    2015-01-15

    Photosystem II (PSII) catalyzes the light-induced generation of oxygen from water. The oxygen-evolving complex is buried deep in the protein on the lumenal side of PSII, and water molecules need to pass through protein subunits to reach the active site—the manganese cluster. Previous studies on the elucidation of water channels in PSII were based on an analysis of the cavities in the static PSII structure determined by X-ray diffraction. In the present study, we perform molecular dynamics simulations of the water movement in the transport system of PSII.

  10. Effect of salt stress on photosystem Ⅱ heterogeneity in wheat leaves

    Institute of Scientific and Technical Information of China (English)

    裴真明; 傅伟; 郭连旺; 汤章城

    1996-01-01

    In order to study the effects of salt stress on photosystem Ⅱ (PSⅡ) heterogeneity, signal from the fluorometer was digitized via a microcomputer interface to record PSⅡ fluorescence induction kinetics. Changes of parameters (Fm, Fp1 and F0) from the fast phase of fluorescence induction curve showed that the percentage of QB-nonreducing PSⅡ reaction centers dropped at first, and then rose with the increase of stress intensity and time. This indicated that the electron transfer from QA to QB is one of the sites where the photochemical and photophysical processes of PSⅡ are damaged by the salt stress.

  11. Hydrogen ions directly regulating the oligomerization state of Photosystem I in intact Spirulina platensis cells

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    H+ concentration induced-monomerization or trimerization of photosystem I (PSI) in cyanobacteria has never been directly observed. In this work, taking characteristic spectra for the trimers and monomers as the indicators, it was experimentally demonstrated that H+ could induce the oligomeric changes of PSI reaction centers in the intact Spirulina ,platensis cells and also in the isolated thylakoid membrane complexes. Especially, the higher concentration of H+ would induce the monomerization while the lower the trimerization, suggesting the electrostatic interaction should be mainly responsible forchanges in the oligomeric state of PSI in Spirulina platensis.

  12. The ferredoxin-thioredoxin system of a green alga, Chlamydomonas reinhardtii: identification and characterization of thioredoxins and ferredoxin-thioredoxin reductase components

    Science.gov (United States)

    Huppe, H. C.; de Lamotte-Guery, F.; Buchanan, B. B.

    1990-01-01

    The components of the ferredoxin-thioredoxin (FT) system of Chlamydomonas reinhardtii have been purified and characterized. The system resembled that of higher plants in consisting of a ferredoxin-thioredoxin reductase (FTR) and two types of thioredoxin, a single f and two m species, m1 and m2. The Chlamydomonas m and f thioredoxins were antigenically similar to their higher-plant counterparts, but not to one another. The m thioredoxins were recognized by antibodies to both higher plant m and bacterial thioredoxins, whereas the thioredoxin f was not. Chlamydomonas thioredoxin f reacted, although weakly, with the antibody to spinach thioredoxin f. The algal thioredoxin f differed from thioredoxins studied previously in behaving as a basic protein on ion-exchange columns. Purification revealed that the algal thioredoxins had molecular masses (Mrs) typical of thioredoxins from other sources, m1 and m2 being 10700 and f 11500. Chlamydomonas FTR had two dissimilar subunits, a feature common to all FTRs studied thus far. One, the 13-kDa ("similar") subunit, resembled its counterpart from other sources in both size and antigenicity. The other, 10-kDa ("variable") subunit was not recognized by antibodies to any FTR tested. When combined with spinach, (Spinacia oleracea L.) thylakoid membranes, the components of the FT system functioned in the light activation of the standard target enzymes from chloroplasts, corn (Zea mays L.) NADP-malate dehydrogenase (EC 1.1.1.82) and spinach fructose 1,6-bisphosphatase (EC 3.1.3.11) as well as the chloroplast-type fructose 1,6-bisphosphatase from Chlamydomonas. Activity was greatest if ferredoxin and other components of the FT system were from Chlamydomonas. The capacity of the Chlamydomonas FT system to activate autologous FBPase indicates that light regulates the photosynthetic carbon metabolism of green algae as in other oxygenic photosynthetic organisms.

  13. Whole-Genome Resequencing Reveals Extensive Natural Variation in the Model Green Alga Chlamydomonas reinhardtii[OPEN

    Science.gov (United States)

    Hazzouri, Khaled M.; Rosas, Ulises; Bahmani, Tayebeh; Nelson, David R.; Abdrabu, Rasha; Harris, Elizabeth H.; Salehi-Ashtiani, Kourosh; Purugganan, Michael D.

    2015-01-01

    We performed whole-genome resequencing of 12 field isolates and eight commonly studied laboratory strains of the model organism Chlamydomonas reinhardtii to characterize genomic diversity and provide a resource for studies of natural variation. Our data support previous observations that Chlamydomonas is among the most diverse eukaryotic species. Nucleotide diversity is ∼3% and is geographically structured in North America with some evidence of admixture among sampling locales. Examination of predicted loss-of-function mutations in field isolates indicates conservation of genes associated with core cellular functions, while genes in large gene families and poorly characterized genes show a greater incidence of major effect mutations. De novo assembly of unmapped reads recovered genes in the field isolates that are absent from the CC-503 assembly. The laboratory reference strains show a genomic pattern of polymorphism consistent with their origin as the recombinant progeny of a diploid zygospore. Large duplications or amplifications are a prominent feature of laboratory strains and appear to have originated under laboratory culture. Extensive natural variation offers a new source of genetic diversity for studies of Chlamydomonas, including naturally occurring alleles that may prove useful in studies of gene function and the dissection of quantitative genetic traits. PMID:26392080

  14. Characterization of a Mutant Deficient for Ammonium and Nitric Oxide Signalling in the Model System Chlamydomonas reinhardtii.

    Directory of Open Access Journals (Sweden)

    Emanuel Sanz-Luque

    Full Text Available The ubiquitous signalling molecule Nitric Oxide (NO is characterized not only by the variety of organisms in which it has been described, but also by the wealth of biological processes that it regulates. In contrast to the expanding repertoire of functions assigned to NO, however, the mechanisms of NO action usually remain unresolved, and genes that work within NO signalling cascades are seldom identified. A recent addition to the list of known NO functions is the regulation of the nitrogen assimilation pathway in the unicellular alga Chlamydomonas reinhardtii, a well-established model organism for genetic and molecular studies that offers new possibilities in the search for mediators of NO signalling. By further exploiting a collection of Chlamydomonas insertional mutant strains originally isolated for their insensitivity to the ammonium (NH4+ nitrogen source, we found a mutant which, in addition to its ammonium insensitive (AI phenotype, was not capable of correctly sensing the NO signal. Similarly to what had previously been described in the AI strain cyg56, the expression of nitrogen assimilation genes in the mutant did not properly respond to treatments with various NO donors. Complementation experiments showed that NON1 (NO Nitrate 1, a gene that encodes a protein containing no known functional domain, was the gene underlying the mutant phenotype. Beyond the identification of NON1, our findings broadly demonstrate the potential for Chlamydomonas reinhardtii to be used as a model system in the search for novel components of gene networks that mediate physiological responses to NO.

  15. Enzymatic modification by point mutation and functional analysis of an omega-6 fatty acid desaturase from Arctic Chlamydomonas sp.

    Science.gov (United States)

    Jung, Woongsic; Kim, Eun Jae; Han, Se Jong; Kang, Sung-Ho; Choi, Han-Gu; Kim, Sanghee

    2017-02-07

    Arctic Chlamydomonas sp. is a dominant microalgal strain in cold or frozen freshwater in the Arctic region. The full-length open reading frame of the omega-6 fatty acid desaturase gene (AChFAD6) was obtained from the transcriptomic database of Arctic Chlamydomonas sp. from the KOPRI culture collection of polar micro-organisms. Amino acid sequence analysis indicated the presence of three conserved histidine-rich segments as unique characteristics of omega-6 fatty acid desaturases, and three transmembrane regions transported to plastidic membranes by chloroplast transit peptides in the N-terminal region. The AChFAD6 desaturase activity was examined by expressing wild-type and V254A mutant (Mut-AChFAD6) heterologous recombinant proteins. Quantitative gas chromatography indicated that the concentration of linoleic acids in AChFAD6-transformed cells increased more than 3-fold [6.73 ± 0.13 mg g(-1) dry cell weight (DCW)] compared with cells transformed with vector alone. In contrast, transformation with Mut-AChFAD6 increased the concentration of oleic acid to 9.23 ± 0.18 mg g(-1) DCW, indicating a change in enzymatic activity to mimic that of stearoyl-CoA desaturase. These results demonstrate that AChFAD6 of Arctic Chlamydomonas sp. increases membrane fluidity by enhancing denaturation of C18 fatty acids and facilitates production of large quantities of linoleic fatty acids in prokaryotic expression systems.

  16. Oil accumulation in the model green alga Chlamydomonas reinhardtii: characterization, variability between common laboratory strains and relationship with starch reserves

    Directory of Open Access Journals (Sweden)

    Carrier Patrick

    2011-01-01

    Full Text Available Abstract Background When cultivated under stress conditions, many microalgae species accumulate both starch and oil (triacylglycerols. The model green microalga Chlamydomonas reinhardtii has recently emerged as a model to test genetic engineering or cultivation strategies aiming at increasing lipid yields for biodiesel production. Blocking starch synthesis has been suggested as a way to boost oil accumulation. Here, we characterize the triacylglycerol (TAG accumulation process in Chlamydomonas and quantify TAGs in various wild-type and starchless strains. Results In response to nitrogen deficiency, Chlamydomonas reinhardtii produced TAGs enriched in palmitic, oleic and linoleic acids that accumulated in oil-bodies. Oil synthesis was maximal between 2 and 3 days following nitrogen depletion and reached a plateau around day 5. In the first 48 hours of oil deposition, a ~80% reduction in the major plastidial membrane lipids occurred. Upon nitrogen re-supply, mobilization of TAGs started after starch degradation but was completed within 24 hours. Comparison of oil content in five common laboratory strains (CC124, CC125, cw15, CC1690 and 11-32A revealed a high variability, from 2 μg TAG per million cell in CC124 to 11 μg in 11-32A. Quantification of TAGs on a cell basis in three mutants affected in starch synthesis (cw15sta1-2, cw15sta6 and cw15sta7-1 showed that blocking starch synthesis did not result in TAG over-accumulation compared to their direct progenitor, the arginine auxotroph strain 330. Moreover, no significant correlation was found between cellular oil and starch levels among the twenty wild-type, mutants and complemented strains tested. By contrast, cellular oil content was found to increase steeply with salt concentration in the growth medium. At 100 mM NaCl, oil level similar to nitrogen depletion conditions could be reached in CC124 strain. Conclusion A reference basis for future genetic studies of oil metabolism in Chlamydomonas

  17. Isolated photosystem I reaction centers on a functionalized gated high electron mobility transistor.

    Science.gov (United States)

    Eliza, Sazia A; Lee, Ida; Tulip, Fahmida S; Mostafa, Salwa; Greenbaum, Elias; Ericson, M Nance; Islam, Syed K

    2011-09-01

    In oxygenic plants, photons are captured with high quantum efficiency by two specialized reaction centers (RC) called Photosystem I (PS I) and Photosystem II (PS II). The captured photon triggers rapid charge separation and the photon energy is converted into an electrostatic potential across the nanometer-scale (~6 nm) reaction centers. The exogenous photovoltages from a single PS I RC have been previously measured using the technique of Kelvin force probe microscopy (KFM). However, biomolecular photovoltaic applications require two-terminal devices. This paper presents for the first time, a micro-device for detection and characterization of isolated PS I RCs. The device is based on an AlGaN/GaN high electron mobility transistor (HEMT) structure. AlGaN/GaN HEMTs show high current throughputs and greater sensitivity to surface charges compared to other field-effect devices. PS I complexes immobilized on the floating gate of AlGaN/GaN HEMTs resulted in significant changes in the device characteristics under illumination. An analytical model has been developed to estimate the RCs of a major orientation on the functionalized gate surface of the HEMTs.

  18. A protein dynamics study of photosystem II: the effects of protein conformation on reaction center function.

    Science.gov (United States)

    Vasil'ev, Sergej; Bruce, Doug

    2006-05-01

    Molecular dynamics simulations have been performed to study photosystem II structure and function. Structural information obtained from simulations was combined with ab initio computations of chromophore excited states. In contrast to calculations based on the x-ray structure, the molecular-dynamics-based calculations accurately predicted the experimental absorbance spectrum. In addition, our calculations correctly assigned the energy levels of reaction-center (RC) chromophores, as well as the lowest-energy antenna chlorophyll. The primary and secondary quinone electron acceptors, Q(A) and Q(B), exhibited independent changes in position over the duration of the simulation. Q(B) fluctuated between two binding sites similar to the proximal and distal sites previously observed in light- and dark-adapted RC from purple bacteria. Kinetic models were used to characterize the relative influence of chromophore geometry, site energies, and electron transport rates on RC efficiency. The fluctuating energy levels of antenna chromophores had a larger impact on quantum yield than did their relative positions. Variations in electron transport rates had the most significant effect and were sufficient to explain the experimentally observed multi-component decay of excitation in photosystem II. The implications of our results are discussed in the context of competing evolutionary selection pressures for RC structure and function.

  19. Detection of herbicide subclasses by an optical multibiosensor based on an array of photosystem II mutants.

    Science.gov (United States)

    Giardi, Maria Teresa; Guzzella, Licia; Euzet, Pierre; Rouillon, Regis; Esposito, Dania

    2005-07-15

    Massive use of herbicides in agriculture over the last few decades has become a serious environmental problem. The residual concentration of these compounds frequently exceeds the maximum admissible concentration in drinking water for human consumption and is a real environmental risk for the aquatic ecosystem. Herbicides inhibiting photosynthesis via targeting photosystem II function still represent the basic means of weed control. A multibiosensor was constructed for detecting herbicides using as biosensing elements photosynthetic preparations coupled to an optical fluorescence transduction system (Giardi et al. EU patent EP1134585, 01830148.1-2204); this paper is about its application in the detection of herbicide subclasses in river water. Photosynthetic material was immobilized on a silicio septum inside a series of flow cells, close to diodes so as to activate photosystem II (PSII) fluorescence. The principle of the detection was based on the factthat herbicides selectively modify PSII fluorescence activity. The multibiosensor has the original feature of being able to distinguish the subclasses of the photosynthetic herbicides by using specific immobilized biomediators isolated from mutated organisms. This setup resulted in a reusable, portable multibiosensor for the detection of herbicide subclasses with a half-life of 54 h for spinach thylakoids and limit of detection of 3 x 10(-9) M for herbicides present in river water.

  20. Protein delivery of a Ni catalyst to photosystem I for light-driven hydrogen production.

    Science.gov (United States)

    Silver, Sunshine C; Niklas, Jens; Du, Pingwu; Poluektov, Oleg G; Tiede, David M; Utschig, Lisa M

    2013-09-11

    The direct conversion of sunlight into fuel is a promising means for the production of storable renewable energy. Herein, we use Nature's specialized photosynthetic machinery found in the Photosystem I (PSI) protein to drive solar fuel production from a nickel diphosphine molecular catalyst. Upon exposure to visible light, a self-assembled PSI-[Ni(P2(Ph)N2(Ph))2](BF4)2 hybrid generates H2 at a rate 2 orders of magnitude greater than rates reported for photosensitizer/[Ni(P2(Ph)N2(Ph))2](BF4)2 systems. The protein environment enables photocatalysis at pH 6.3 in completely aqueous conditions. In addition, we have developed a strategy for incorporating the Ni molecular catalyst with the native acceptor protein of PSI, flavodoxin. Photocatalysis experiments with this modified flavodoxin demonstrate a new mechanism for biohybrid creation that involves protein-directed delivery of a molecular catalyst to the reducing side of Photosystem I for light-driven catalysis. This work further establishes strategies for constructing functional, inexpensive, earth-abundant solar fuel-producing PSI hybrids that use light to rapidly produce hydrogen directly from water.

  1. Comparison of the light-harvesting networks of plant and cyanobacterial photosystem I.

    Science.gov (United States)

    Sener, Melih K; Jolley, Craig; Ben-Shem, Adam; Fromme, Petra; Nelson, Nathan; Croce, Roberta; Schulten, Klaus

    2005-09-01

    With the availability of structural models for photosystem I (PSI) in cyanobacteria and plants it is possible to compare the excitation transfer networks in this ubiquitous photosystem from two domains of life separated by over one billion years of divergent evolution, thus providing an insight into the physical constraints that shape the networks' evolution. Structure-based modeling methods are used to examine the excitation transfer kinetics of the plant PSI-LHCI supercomplex. For this purpose an effective Hamiltonian is constructed that combines an existing cyanobacterial model for structurally conserved chlorophylls with spectral information for chlorophylls in the Lhca subunits. The plant PSI excitation migration network thus characterized is compared to its cyanobacterial counterpart investigated earlier. In agreement with observations, an average excitation transfer lifetime of approximately 49 ps is computed for the plant PSI-LHCI supercomplex with a corresponding quantum yield of 95%. The sensitivity of the results to chlorophyll site energy assignments is discussed. Lhca subunits are efficiently coupled to the PSI core via gap chlorophylls. In contrast to the chlorophylls in the vicinity of the reaction center, previously shown to optimize the quantum yield of the excitation transfer process, the orientational ordering of peripheral chlorophylls does not show such optimality. The finding suggests that after close packing of chlorophylls was achieved, constraints other than efficiency of the overall excitation transfer process precluded further evolution of pigment ordering.

  2. Light driven CO2 fixation by using cyanobacterial photosystem I and NADPH-dependent formate dehydrogenase.

    Directory of Open Access Journals (Sweden)

    Masaki Ihara

    Full Text Available The ultimate goal of this research is to construct a new direct CO2 fixation system using photosystems in living algae. Here, we report light-driven formate production from CO2 by using cyanobacterial photosystem I (PS I. Formate, a chemical hydrogen carrier and important industrial material, can be produced from CO2 by using the reducing power and the catalytic function of formate dehydrogenase (FDH. We created a bacterial FDH mutant that experimentally switched the cofactor specificity from NADH to NADPH, and combined it with an in vitro-reconstituted cyanobacterial light-driven NADPH production system consisting of PS I, ferredoxin (Fd, and ferredoxin-NADP(+-reductase (FNR. Consequently, light-dependent formate production under a CO2 atmosphere was successfully achieved. In addition, we introduced the NADPH-dependent FDH mutant into heterocysts of the cyanobacterium Anabaena sp. PCC 7120 and demonstrated an increased formate concentration in the cells. These results provide a new possibility for photo-biological CO2 fixation.

  3. Excitation energy transfer between Light-harvesting complex II and Photosystem I in reconstituted membranes.

    Science.gov (United States)

    Akhtar, Parveen; Lingvay, Mónika; Kiss, Teréz; Deák, Róbert; Bóta, Attila; Ughy, Bettina; Garab, Győző; Lambrev, Petar H

    2016-04-01

    Light-harvesting complex II (LHCII), the major peripheral antenna of Photosystem II in plants, participates in several concerted mechanisms for regulation of the excitation energy and electron fluxes in thylakoid membranes. In part, these include interaction of LHCII with Photosystem I (PSI) enhancing the latter's absorption cross-section - for example in the well-known state 1 - state 2 transitions or as a long-term acclimation to high light. In this work we examined the capability of LHCII to deliver excitations to PSI in reconstituted membranes in vitro. Proteoliposomes with native plant thylakoid membrane lipids and different stoichiometric ratios of LHCII:PSI were reconstituted and studied by steady-state and time-resolved fluorescence spectroscopy. Fluorescence emission from LHCII was strongly decreased in PSI-LHCII membranes due to trapping of excitations by PSI. Kinetic modelling of the time-resolved fluorescence data revealed the existence of separate pools of LHCII distinguished by the time scale of energy transfer. A strongly coupled pool, equivalent to one LHCII trimer per PSI, transferred excitations to PSI with near-unity efficiency on a time scale of less than 10ps but extra LHCIIs also contributed significantly to the effective antenna size of PSI, which could be increased by up to 47% in membranes containing 3 LHCII trimers per PSI. The results demonstrate a remarkable competence of LHCII to increase the absorption cross-section of PSI, given the opportunity that the two types of complexes interact in the membrane.

  4. Quality control of Photosystem II: the mechanisms for avoidance and tolerance of light and heat stresses are closely linked to membrane fluidity of the thylakoids

    Directory of Open Access Journals (Sweden)

    Yasusi Yamamoto

    2016-08-01

    Full Text Available When oxygenic photosynthetic organisms are exposed to excessive light and/or heat, Photosystem II is damaged and electron transport is blocked. In these events, reactive oxygen species, endogenous radicals and lipid peroxidation products generated by photochemical reaction and/or heat cause the damage. Regarding light stress, plants first dissipate excessive light energy captured by light-harvesting chlorophyll protein complexes as heat to avoid the hazards, but once light stress is unavoidable, they tolerate the stress by concentrating damage in a particular protein in photosystem II, i.e. the reaction-center binding D1 protein of Photosystem II. The damaged D1 is removed by specific proteases and replaced with a new copy produced through de novo synthesis (reversible photoinhibition. When light intensity becomes extremely high, irreversible aggregation of D1 occurs and thereby D1 turnover is prevented. Once the aggregated products accumulate in Photosystem II complexes, removal of them by proteases is difficult, and irreversible inhibition of Photosystem II takes place (irreversible photoinhibition. Important is that various aspects of both the reversible and irreversible photoinhibition are highly dependent on the membrane fluidity of the thylakoids. Heat stress-induced inactivation of photosystem II is an irreversible process, which may be also affected by the fluidity of the thylakoid membranes. Here I describe why the membrane fluidity is a key to regulate the avoidance and tolerance of Photosystem II on environmental stresses.

  5. Hydrogen Photoevolution Indicates an Increase in the Antenna Size of Photosystem I in Chlamydobotrys stellata during Transition from Autotrophic to Photoheterotrophic Nutrition.

    Science.gov (United States)

    Boichenko, V A; Wiessner, W; Klimov, V V; Mende, D; Demeter, S

    1992-09-01

    The changes in the light-harvesting antenna size of photosystem I were investigated in the green alga Chlamydobotrys stellata during transition from autotrophic to photoheterotrophic nutrition by measuring the light-saturation behavior of hydrogen evolution following single turnover flashes. It was found that during autotrophic-to-photoheterotrophic transition the antenna size of photosystem I increased from 180 to 250 chlorophyll. The chlorophyll (a + b)/P700 ratio decreased from 800 to 550. The electron transport of photosystem I measured from reduced 2,6-dichloro-phenolindophenol to methylviologen was accelerated 1.4 times. In the 77K fluorescence spectra, the photosystem II fluorescence yield was considerably lowered relative to the photosystem I fluorescence yield. It is suggested that the increased light-harvesting capacity and redistribution of absorbed excitation energy in favor of photosystem I is a response of photoheterotrophic algae to meet the ATP demand for acetate metabolism by efficient photosystem I cyclic electron transport when the noncyclic photophosphorylation is inhibited by CO(2) deficiency.

  6. The interaction of quinones, herbicides and bicarbonate with their binding environment and the acceptor side of photosystem II in photosynthesis

    NARCIS (Netherlands)

    Vermaas, W.

    1984-01-01

    In this thesis experiments are described which are directed towards a further characterization of the interaction of the native bound plastoquinone Q B , artificial quinones, herbicides and bicarbonate with their binding environment at the acceptor side of Photosystem II in the thylakoid

  7. Cyanobacterial photosystem II at 2.9-A resolution and the role of quinones, lipids, channels and chloride

    NARCIS (Netherlands)

    Guskov, Albert; Kern, Jan; Gabdulkhakov, Azat; Broser, Matthias; Zouni, Athina; Saenger, Wolfram

    2009-01-01

    Photosystem II (PSII) is a large homodimeric protein-cofactor complex located in the photosynthetic thylakoid membrane that acts as light-driven water:plastoquinone oxidoreductase. The crystal structure of PSII from Thermosynechococcus elongatus at 2.9-A resolution allowed the unambiguous assignment

  8. Determination of the excitation migration time in Photosystem II consequences for the membrane organization and charge separation parameters

    NARCIS (Netherlands)

    Broess, K.; Trinkunas, G.; Hoek, van A.; Croce, R.; Amerongen, van H.

    2008-01-01

    The fluorescence decay kinetics of Photosystem II (PSII) membranes from spinach with open reaction centers (RCs), were compared after exciting at 420 and 484 nm. These wavelengths lead to preferential excitation of chlorophyll (Chl) a and Chl b, respectively, which causes different initial excited-s

  9. Determination of the excitation migration time in Photosystem II - Consequences for the membrane organization and charge separation parameters

    NARCIS (Netherlands)

    Broess, Koen; Trinkunas, Gediminas; van Hoek, Arie; Croce, Roberta; van Amerongen, Herbert

    2008-01-01

    The fluorescence decay kinetics of Photosystem II (PSII) membranes from spinach with open reaction centers (RCs), were compared after exciting at 420 and 484 nm. These wavelengths lead to preferential excitation of chlorophyll (Chl) a and Chl b, respectively, which causes different initial excited-s

  10. Photoprotection in higher plants : The putative quenching site is conserved in all outer light-harvesting complexes of Photosystem II

    NARCIS (Netherlands)

    Mozzo, Milena; Passarini, Francesca; Bassi, Roberto; van Amerongen, Herbert; Croce, Roberta

    2008-01-01

    In bright sunlight, the amount of energy harvested by plants exceeds the electron transport capacity of Photosystem II in the chloroplasts. The excess energy can lead to severe damage of the photosynthetic apparatus and to avoid this, part of the energy is thermally dissipated via a mechanism called

  11. The cytoplasmic domain of the gamete membrane fusion protein HAP2 targets the protein to the fusion site in Chlamydomonas and regulates the fusion reaction

    Science.gov (United States)

    Liu, Yanjie; Pei, Jimin; Grishin, Nick; Snell, William J.

    2015-01-01

    Cell-cell fusion between gametes is a defining step during development of eukaryotes, yet we know little about the cellular and molecular mechanisms of the gamete membrane fusion reaction. HAP2 is the sole gamete-specific protein in any system that is broadly conserved and shown by gene disruption to be essential for gamete fusion. The wide evolutionary distribution of HAP2 (also known as GCS1) indicates it was present in the last eukaryotic common ancestor and, therefore, dissecting its molecular properties should provide new insights into fundamental features of fertilization. HAP2 acts at a step after membrane adhesion, presumably directly in the merger of the lipid bilayers. Here, we use the unicellular alga Chlamydomonas to characterize contributions of key regions of HAP2 to protein location and function. We report that mutation of three strongly conserved residues in the ectodomain has no effect on targeting or fusion, although short deletions that include those residues block surface expression and fusion. Furthermore, HAP2 lacking a 237-residue segment of the cytoplasmic region is expressed at the cell surface, but fails to localize at the apical membrane patch specialized for fusion and fails to rescue fusion. Finally, we provide evidence that the ancient HAP2 contained a juxta-membrane, multi-cysteine motif in its cytoplasmic region, and that mutation of a cysteine dyad in this motif preserves protein localization, but substantially impairs HAP2 fusion activity. Thus, the ectodomain of HAP2 is essential for its surface expression, and the cytoplasmic region targets HAP2 to the site of fusion and regulates the fusion reaction. PMID:25655701

  12. Lack of Set Theory Relevant Prerequisite Knowledge

    Science.gov (United States)

    Dogan-Dunlap, Hamide

    2006-01-01

    Many students struggle with college mathematics topics due to a lack of mastery of prerequisite knowledge. Set theory language is one such prerequisite for linear algebra courses. Many students' mistakes on linear algebra questions reveal a lack of mastery of set theory knowledge. This paper reports the findings of a qualitative analysis of a…

  13. Nitrogen-Sparing Mechanisms in Chlamydomonas Affect the Transcriptome, the Proteome, and Photosynthetic Metabolism.

    Science.gov (United States)

    Schmollinger, Stefan; Mühlhaus, Timo; Boyle, Nanette R; Blaby, Ian K; Casero, David; Mettler, Tabea; Moseley, Jeffrey L; Kropat, Janette; Sommer, Frederik; Strenkert, Daniela; Hemme, Dorothea; Pellegrini, Matteo; Grossman, Arthur R; Stitt, Mark; Schroda, Michael; Merchant, Sabeeha S

    2014-04-01

    Nitrogen (N) is a key nutrient that limits global primary productivity; hence, N-use efficiency is of compelling interest in agriculture and aquaculture. We used Chlamydomonas reinhardtii as a reference organism for a multicomponent analysis of the N starvation response. In the presence of acetate, respiratory metabolism is prioritized over photosynthesis; consequently, the N-sparing response targets proteins, pigments, and RNAs involved in photosynthesis and chloroplast function over those involved in respiration. Transcripts and proteins of the Calvin-Benson cycle are reduced in N-deficient cells, resulting in the accumulation of cycle metabolic intermediates. Both cytosolic and chloroplast ribosomes are reduced, but via different mechanisms, reflected by rapid changes in abundance of RNAs encoding chloroplast ribosomal proteins but not cytosolic ones. RNAs encoding transporters and enzymes for metabolizing alternative N sources increase in abundance, as is appropriate for the soil environmental niche of C. reinhardtii. Comparison of the N-replete versus N-deplete proteome indicated that abundant proteins with a high N content are reduced in N-starved cells, while the proteins that are increased have lower than average N contents. This sparing mechanism contributes to a lower cellular N/C ratio and suggests an approach for engineering increased N-use efficiency.

  14. Nitrogen-Sparing Mechanisms in Chlamydomonas Affect the Transcriptome, the Proteome, and Photosynthetic Metabolism[W

    Science.gov (United States)

    Schmollinger, Stefan; Mühlhaus, Timo; Boyle, Nanette R.; Blaby, Ian K.; Casero, David; Mettler, Tabea; Moseley, Jeffrey L.; Kropat, Janette; Sommer, Frederik; Strenkert, Daniela; Hemme, Dorothea; Pellegrini, Matteo; Grossman, Arthur R.; Stitt, Mark; Schroda, Michael; Merchant, Sabeeha S.

    2014-01-01

    Nitrogen (N) is a key nutrient that limits global primary productivity; hence, N-use efficiency is of compelling interest in agriculture and aquaculture. We used Chlamydomonas reinhardtii as a reference organism for a multicomponent analysis of the N starvation response. In the presence of acetate, respiratory metabolism is prioritized over photosynthesis; consequently, the N-sparing response targets proteins, pigments, and RNAs involved in photosynthesis and chloroplast function over those involved in respiration. Transcripts and proteins of the Calvin-Benson cycle are reduced in N-deficient cells, resulting in the accumulation of cycle metabolic intermediates. Both cytosolic and chloroplast ribosomes are reduced, but via different mechanisms, reflected by rapid changes in abundance of RNAs encoding chloroplast ribosomal proteins but not cytosolic ones. RNAs encoding transporters and enzymes for metabolizing alternative N sources increase in abundance, as is appropriate for the soil environmental niche of C. reinhardtii. Comparison of the N-replete versus N-deplete proteome indicated that abundant proteins with a high N content are reduced in N-starved cells, while the proteins that are increased have lower than average N contents. This sparing mechanism contributes to a lower cellular N/C ratio and suggests an approach for engineering increased N-use efficiency. PMID:24748044

  15. The Proteome of Copper, Iron, Zinc, and Manganese Micronutrient Deficiency in Chlamydomonas reinhardtii*

    Science.gov (United States)

    Hsieh, Scott I.; Castruita, Madeli; Malasarn, Davin; Urzica, Eugen; Erde, Jonathan; Page, M. Dudley; Yamasaki, Hiroaki; Casero, David; Pellegrini, Matteo; Merchant, Sabeeha S.; Loo, Joseph A.

    2013-01-01

    Trace metals such as copper, iron, zinc, and manganese play important roles in several biochemical processes, including respiration and photosynthesis. Using a label-free, quantitative proteomics strategy (MSE), we examined the effect of deficiencies in these micronutrients on the soluble proteome of Chlamydomonas reinhardtii. We quantified >103 proteins with abundances within a dynamic range of 3 to 4 orders of magnitude and demonstrated statistically significant changes in ∼200 proteins in each metal-deficient growth condition relative to nutrient-replete media. Through analysis of Pearson's coefficient, we also examined the correlation between protein abundance and transcript abundance (as determined via RNA-Seq analysis) and found moderate correlations under all nutritional states. Interestingly, in a subset of transcripts known to significantly change in abundance in metal-replete and metal-deficient conditions, the correlation to protein abundance is much stronger. Examples of new discoveries highlighted in this work include the accumulation of O2 labile, anaerobiosis-related enzymes (Hyd1, Pfr1, and Hcp2) in copper-deficient cells; co-variation of Cgl78/Ycf54 and coprogen oxidase; the loss of various stromal and lumenal photosynthesis-related proteins, including plastocyanin, in iron-limited cells; a large accumulation (from undetectable amounts to over 1,000 zmol/cell) of two COG0523 domain-containing proteins in zinc-deficient cells; and the preservation of photosynthesis proteins in manganese-deficient cells despite known losses in photosynthetic function in this condition. PMID:23065468

  16. Copper response regulator1-dependent and -independent responses of the Chlamydomonas reinhardtii transcriptome to dark anoxia.

    Science.gov (United States)

    Hemschemeier, Anja; Casero, David; Liu, Bensheng; Benning, Christoph; Pellegrini, Matteo; Happe, Thomas; Merchant, Sabeeha S

    2013-09-01

    Anaerobiosis is a stress condition for aerobic organisms and requires extensive acclimation responses. We used RNA-Seq for a whole-genome view of the acclimation of Chlamydomonas reinhardtii to anoxic conditions imposed simultaneously with transfer to the dark. Nearly 1.4 × 10(3) genes were affected by hypoxia. Comparing transcript profiles from early (hypoxic) with those from late (anoxic) time points indicated that cells activate oxidative energy generation pathways before employing fermentation. Probable substrates include amino acids and fatty acids (FAs). Lipid profiling of the C. reinhardtii cells revealed that they degraded FAs but also accumulated triacylglycerols (TAGs). In contrast with N-deprived cells, the TAGs in hypoxic cells were enriched in desaturated FAs, suggesting a distinct pathway for TAG accumulation. To distinguish transcriptional responses dependent on copper response regulator1 (CRR1), which is also involved in hypoxic gene regulation, we compared the transcriptomes of crr1 mutants and complemented strains. In crr1 mutants, ~40 genes were aberrantly regulated, reaffirming the importance of CRR1 for the hypoxic response, but indicating also the contribution of additional signaling strategies to account for the remaining differentially regulated transcripts. Based on transcript patterns and previous results, we conclude that nitric oxide-dependent signaling cascades operate in anoxic C. reinhardtii cells.

  17. Transcriptome Analysis of Manganese-deficient Chlamydomonas reinhardtii Provides Insight on the Chlorophyll Biosynthesis Pathway

    Energy Technology Data Exchange (ETDEWEB)

    Lockhart, Ainsley; Zvenigorodsky, Natasha; Pedraza, Mary Ann; Lindquist, Erika

    2011-08-11

    The biosynthesis of chlorophyll and other tetrapyrroles is a vital but poorly understood process. Recent genomic advances with the unicellular green algae Chlamydomonas reinhardtii have created opportunity to more closely examine the mechanisms of the chlorophyll biosynthesis pathway via transcriptome analysis. Manganese is a nutrient of interest for complex reactions because of its multiple stable oxidation states and role in molecular oxygen coordination. C. reinhardtii was cultured in Manganese-deplete Tris-acetate-phosphate (TAP) media for 24 hours and used to create cDNA libraries for sequencing using Illumina TruSeq technology. Transcriptome analysis provided intriguing insight on possible regulatory mechanisms in the pathway. Evidence supports similarities of GTR (Glutamyl-tRNA synthase) to its Chlorella vulgaris homolog in terms of Mn requirements. Data was also suggestive of Mn-related compensatory up-regulation for pathway proteins CHLH1 (Manganese Chelatase), GUN4 (Magnesium chelatase activating protein), and POR1 (Light-dependent protochlorophyllide reductase). Intriguingly, data suggests possible reciprocal expression of oxygen dependent CPX1 (coproporphyrinogen III oxidase) and oxygen independent CPX2. Further analysis using RT-PCR could provide compelling evidence for several novel regulatory mechanisms in the chlorophyll biosynthesis pathway.

  18. Antagonistic and synergistic effects of light irradiation on the effects of copper on Chlamydomonas reinhardtii

    Energy Technology Data Exchange (ETDEWEB)

    Cheloni, Giulia; Cosio, Claudia; Slaveykova, Vera I., E-mail: vera.slaveykova@unige.ch

    2014-10-15

    Highlights: • Light intensity and spectral composition affect Cu uptake and effects to C. reinhardtii. • High light (HL) reduced Cu effect on growth inhibition, oxidative stress and damage. • HL in combination with Cu up-regulated genes involved in the antioxidant responses. • HL with increased UVB radiation exacerbated Cu uptake and Cu-induced toxic effects. - Abstract: The present study showed the important role of light intensity and spectral composition on Cu uptake and effects on green alga Chlamydomonas reinhardtii. High-intenisty light (HL) increased cellular Cu concentrations, but mitigated the Cu-induced decrease in chlorophyll fluorescence, oxidative stress and lipid peroxidation at high Cu concentrations, indicating that Cu and HL interact in an antagonistic manner. HL up-regulated the transcription of genes involved in the antioxidant response in C. reinhardtii and thus reduced the oxidative stress upon exposure to Cu and HL. Combined exposure to Cu and UVBR resulted in an increase of cellular Cu contents and caused severe oxidative damage to the cells. The observed effects were higher than the sum of the effects corresponding to exposure to UVBR or Cu alone suggesting a synergistic interaction.

  19. Relief of arsenate toxicity by Cd-stimulated phytochelatin synthesis in the green alga Chlamydomonas reinhardtii.

    Science.gov (United States)

    Kobayashi, Isao; Fujiwara, Shoko; Saegusa, Hirotaka; Inouhe, Masahiro; Matsumoto, Hiroko; Tsuzuki, Mikio

    2006-01-01

    In most photosynthetic organisms, inorganic arsenic taken up into the cells inhibits photosynthesis and cellular growth. In a green alga, Chlamydomonas reinhardtii, 0.5 mM arsenate inhibited photosynthesis almost completely within 30 min. However, in cells acclimated with a sublethal concentration (0.05 to 0.1 mM) of Cd, the inhibition of photosynthesis at 30 min after the addition of arsenate was relieved by more than 50%. The concentrations of arsenic incorporated into the cells were not significantly different between the Cd-acclimated and the non-acclimated cells. The Cd-acclimated cells accumulated Cd and synthesized phytochelatin (PC) peptides, which are known to play an important role in detoxification of heavy metals in plants. By the addition of an inhibitor of glutathione (an intermediate in the PC biosynthetic pathway) biosynthesis, buthionine sulfoximine, cells lost not only Cd tolerance but also arsenate tolerance. These results suggest that glutathione and/or PCs synthesized in Cd-acclimated cells are involved in mechanisms of arsenate tolerance.

  20. Integrated quantitative analysis of nitrogen stress response in Chlamydomonas reinhardtii using metabolite and protein profiling.

    Science.gov (United States)

    Wase, Nishikant; Black, Paul N; Stanley, Bruce A; DiRusso, Concetta C

    2014-03-01

    Nitrogen starvation induces a global stress response in microalgae that results in the accumulation of lipids as a potential source of biofuel. Using GC-MS-based metabolite and iTRAQ-labeled protein profiling, we examined and correlated the metabolic and proteomic response of Chlamydomonas reinhardtii under nitrogen stress. Key amino acids and metabolites involved in nitrogen sparing pathways, methyl group transfer reactions, and energy production were decreased in abundance, whereas certain fatty acids, citric acid, methionine, citramalic acid, triethanolamine, nicotianamine, trehalose, and sorbitol were increased in abundance. Proteins involved in nitrogen assimilation, amino acid metabolism, oxidative phosphorylation, glycolysis, TCA cycle, starch, and lipid metabolism were elevated compared with nonstressed cultures. In contrast, the enzymes of the glyoxylate cycle, one carbon metabolism, pentose phosphate pathway, the Calvin cycle, photosynthetic and light harvesting complex, and ribosomes were reduced. A noteworthy observation was that citrate accumulated during nitrogen stress coordinate with alterations in the enzymes that produce or utilize this metabolite, demonstrating the value of comparing protein and metabolite profiles to understand complex patterns of metabolic flow. Thus, the current study provides unique insight into the global metabolic adjustments leading to lipid storage during N starvation for application toward advanced biofuel production technologies.

  1. Chlamydomonas as a model for biofuels and bio-products production.

    Science.gov (United States)

    Scranton, Melissa A; Ostrand, Joseph T; Fields, Francis J; Mayfield, Stephen P

    2015-05-01

    Developing renewable energy sources is critical to maintaining the economic growth of the planet while protecting the environment. First generation biofuels focused on food crops like corn and sugarcane for ethanol production, and soybean and palm for biodiesel production. Second generation biofuels based on cellulosic ethanol produced from terrestrial plants, has received extensive funding and recently pilot facilities have been commissioned, but to date output of fuels from these sources has fallen well short of what is needed. Recent research and pilot demonstrations have highlighted the potential of algae as one of the most promising sources of sustainable liquid transportation fuels. Algae have also been established as unique biofactories for industrial, therapeutic, and nutraceutical co-products. Chlamydomonas reinhardtii's long established role in the field of basic research in green algae has paved the way for understanding algal metabolism and developing genetic engineering protocols. These tools are now being utilized in C. reinhardtii and in other algal species for the development of strains to maximize biofuels and bio-products yields from the lab to the field.

  2. Recombination and Heterologous Expression of Allophycocyanin Gene in the Chloroplast of Chlamydomonas reinhardtii

    Institute of Scientific and Technical Information of China (English)

    Zhong-Liang SU; Kai-Xian QIAN; Cong-Ping TAN; Chun-Xiao MENG; Song QIN

    2005-01-01

    Heterogeneous expression of multiple genes in the nucleus of transgenic plants requires the introduction of an individual gene and the subsequent backcross to reconstitute multi-subunit proteins or metabolic pathways. In order to accomplish the expression of multiple genes in a single transformation event, we inserted both large and small subunits of allophycocyanin gene (apcA and apcB) into Chlamydomonas reinhardtii chloroplast expression vector, resulting in papc-S. The constructed vector was then introduced into the chloroplast of C. reinhardtii by micro-particle bombardment. Polymerase chain reaction and Southern blot analysis revealed that the two genes had integrated into the chloroplast genome. Western blot and enzyme-linked immunosorbent assay showed that the two genes from the prokaryotic cyanobacteria could be correctly expressed in the chloroplasts of C. reinhardtii. The expressed foreign protein in transformants accounted for about 2%-3% of total soluble proteins. These findings pave the way to the reconstitution of multi-subunit proteins or metabolic pathways in transgenic C. reinhardtii chloroplasts in a single transformation event.

  3. Linear systems analysis of the ciliary steering behavior associated with negative-phototaxis in Chlamydomonas reinhardtii.

    Science.gov (United States)

    Josef, Keith; Saranak, Jureepan; Foster, Kenneth W

    2006-12-01

    In response to light stimulation Chlamydomonas reinhardtii changes the beating frequency, beating pattern, and beating synchrony of the trans and cis cilia to steer the freely-swimming cell relative to light sources. To understand the cell steering behavior the impulse responses of the beating frequency and stroke velocity of each cilium have been obtained with high temporal resolution on cells held with a micropipette. Interestingly the response of each cilium is quite different. The trans cilium responds with less delay than the cis cilium for both beating frequency and stroke velocity. For light stimulation at 2 Hz, the critical cell-rotation frequency, both responses of the trans and cis cilia are about 180 degrees out of phase. The trans-cilium beating frequency response peaks at a stimulus frequency of 5-6 Hz, higher than the cis at 1-2 Hz. The stroke velocities of the trans and cis cilia have the same stimulus-frequency response (2 Hz), but the trans cilium has a shorter delay than the cis. The times to maximum response are much shorter than the time for a rotation of the cell. The use of two different approaches that enable the trans cilium to respond ahead of the cis for both the beating frequency and stroke velocity responses suggests the importance of both responses to phototaxis. Internal cell processing responsible for the time course of the responses is proposed.

  4. Enhancement of lipid production and fatty acid profiling in Chlamydomonas reinhardtii, CC1010 for biodiesel production.

    Science.gov (United States)

    Karpagam, R; Preeti, R; Ashokkumar, B; Varalakshmi, P

    2015-11-01

    Lipid from microalgae is one of the putative oil resources to facilitate the biodiesel production during this era of energy dissipation and environmental pollution. In this study, the key parameters such as biomass productivity, lipid productivity and lipid content were evaluated at the early stationary phase of Chlamydomonas reinhardtii, CC1010 cultivated in nutrient starved (nitrogen, phosphorous), glucose (0.05%, 0.1%, 0.15% and 0.2%) and vitamin B12 supplementation (0.001%, 0.002% and 0.003%) in Tris-Acetate-Phosphate (TAP) medium. The lipid content in nitrogen starved media was 61% which is 2.34 folds higher than nutrient sufficient TAP medium. Glucose supplementation has lead to proportional increase in biomass productivity with the increasing concentration of glucose whereas vitamin B12 supplementations had not shown any influence in lipid and biomass production. Further, fatty acid methyl ester (FAME) profiling of C. reinhardtii, CC 1010 has revealed more than 80% of total SFA (saturated fatty acid) and MUFA (mono unsaturated fatty acid) content. Quality checking parameters of biodiesel like cetane number, saponification value, iodine number and degree of unsaturation were analyzed and the biodiesel fuel properties were found to be appropriate as per the international standards, EN 14214 and ASTM D6751. Conclusively, among all the treatments, nitrogen starvation with 0.1% glucose supplementation had yielded high lipid content in C. reinhardtii, CC 1010.

  5. Ciprofloxacin toxicity and its co-metabolic removal by a freshwater microalga Chlamydomonas mexicana.

    Science.gov (United States)

    Xiong, Jiu-Qiang; Kurade, Mayur B; Kim, Jung Rae; Roh, Hyun-Seog; Jeon, Byong-Hun

    2017-02-05

    This study evaluated the toxicity and cellular stresses of ciprofloxacin (CIP) and its co-metabolic removal in a freshwater microalga Chlamydomonas mexicana. The toxicological effects of CIP on C. mexicana were assessed by studying the growth and biochemical characteristics of the microalga including total chlorophyll, carotenoid content, malondialdehyde (MDA) and superoxide dismutase (SOD) activity. The calculated effective concentration (EC50) of CIP on C. mexicana was 65±4mgL(-1) at 96h. The growth of C. mexicana was significantly inhibited at increased concentrations of CIP, showing 36±1, 75±3. and 88±3% inhibition at 40, 60 and 100mgL(-1) CIP, respectively, compared to the control after 11days of cultivation. The total chlorophyll, carotenoid, MDA and SOD activity were significantly increased as a result of relatively high concentrations of CIP stress. C. mexicana showed 13±1% removal of CIP (2mgL(-1)) after 11days of cultivation; however, the addition of an electron donor (sodium acetate, 4gL(-1)) highly enhanced the removal of CIP (2mgL(-1)) by>3-fold after 11days. Kinetic studies showed that removal of CIP followed a first-order model (R(2) 0.94-0.97) with the apparent rate constants (k) ranging from 0.0121 to 0.079 d(-1).

  6. Influence of sulphate on the reduction of cadmium toxicity in the microalga Chlamydomonas moewusii.

    Science.gov (United States)

    Mera, Roi; Torres, Enrique; Abalde, Julio

    2016-06-01

    Cadmium is considered as one of the most hazardous metals for living organism and ecosystems. Environmental factors play an important role since they alter the toxicity of metals by varying the bioavailability of these elements for the organisms. The aim of the present study was to investigate, using the freshwater microalga Chlamydomonas moewusii, the existence of an interaction between cadmium and sulphate as a factor that varied the toxicity of this metal. Different cell parameters such as cell growth, content of chlorophylls and biosynthesis of phytochelatins (PCs) were determined. A two-way ANOVA showed that the interaction had a significant effect size of 21% (pmicroalga and around of a 6% on the content of chlorophylls/cell. The effect of this inhibition was that when the concentration of sulphate increased, a lower toxic effect of cadmium on the growth and on the content of chlorophylls was observed. In addition, the increase of sulphate concentration allowed the biosynthesis of a higher amount of PCs and/or PCs with higher chain length. This higher biosynthesis was responsible for the reduction of the toxic effect of cadmium and explained the interaction.

  7. Toxicity of atrazine and its bioaccumulation and biodegradation in a green microalga, Chlamydomonas mexicana.

    Science.gov (United States)

    Kabra, Akhil N; Ji, Min-Kyu; Choi, Jaewon; Kim, Jung Rae; Govindwar, Sanjay P; Jeon, Byong-Hun

    2014-11-01

    This study evaluated the toxicity of herbicide atrazine, along with its bioaccumulation and biodegradation in the green microalga Chlamydomonas mexicana. At low concentration (10 μg L(-1)), atrazine had no profound effect on the microalga, while higher concentrations (25, 50, and 100 μg L(-1)) imposed toxicity, leading to inhibition of cell growth and chlorophyll a accumulation by 22 %, 33 %, and 36 %, and 13 %, 24 %, and 27 %, respectively. Atrazine 96-h EC50 for C. mexicana was estimated to be 33 μg L(-1). Microalga showed a capability to accumulate atrazine in the cell and to biodegrade the cell-accumulated atrazine resulting in 14-36 % atrazine degradation at 10-100 μg L(-1). Increasing atrazine concentration decreased the total fatty acids (from 102 to 75 mg g(-1)) and increased the unsaturated fatty acid content in the microalga. Carbohydrate content increased gradually with the increase in atrazine concentration up to 15 %. This study shows that C. mexicana has the capability to degrade atrazine and can be employed for the remediation of atrazine-contaminated streams.

  8. An organelle K+ channel is required for osmoregulation in Chlamydomonas reinhardtii.

    Science.gov (United States)

    Xu, Feifei; Wu, Xiaoan; Jiang, Lin-Hua; Zhao, Hucheng; Pan, Junmin

    2016-08-01

    Fresh water protozoa and algae face hypotonic challenges in their living environment. Many of them employ a contractile vacuole system to uptake excessive water from the cytoplasm and expel it to the environment to achieve cellular homeostasis. K(+), a major osmolyte in contractile vacuole, is predicted to create higher osmolarity for water influx. Molecular mechanisms for K(+) permeation through the plasma membrane have been well studied. However, how K(+) permeates organelles such as the contractile vacuole is not clear. Here, we show that the six-transmembrane K(+) channel KCN11 in Chlamydomonas is exclusively localized to contractile vacuole. Ectopic expression of KCN11 in HEK293T cells results in voltage-gated K(+) channel activity. Disruption of the gene or mutation of key residues for K(+) permeability of the channel leads to dysfunction of cell osmoregulation in very hypotonic conditions. The contractile cycle is inhibited in the mutant cells with a slower rate of contractile vacuole swelling, leading to cell death. These data demonstrate a new role for six-transmembrane K(+) channels in contractile vacuole functioning and provide further insights into osmoregulation mediated by the contractile vacuole.

  9. The Unicellular Green Alga Chlamydomonas reinhardtii as an Experimental System to Study Chloroplast RNA Metabolism

    Science.gov (United States)

    Nickelsen, J.; Kück, U.

    Chloroplasts are typical organelles of photoautotrophic eukaryotic cells which drive a variety of functions, including photosynthesis. For many years the unicellular green alga Chlamydomonas reinhardtii has served as an experimental organism for studying photosynthetic processes. The recent development of molecular tools for this organism together with efficient methods of genetic analysis and the availability of many photosynthesis mutants has now made this alga a powerful model system for the analysis of chloroplast biogenesis. For example, techniques have been developed to transfer recombinant DNA into both the nuclear and the chloroplast genome. This allows both complementation tests and analyses of gene functions in vivo. Moreover, site-specific DNA recombinations in the chloroplast allow targeted gene disruption experiments which enable a "reverse genetics" to be performed. The potential of the algal system for the study of chloroplast biogenesis is illustrated in this review by the description of regulatory systems of gene expression involved in organelle biogenesis. One example concerns the regulation of trans-splicing of chloroplast mRNAs, a process which is controlled by both multiple nuclear- and chloroplast-encoded factors. The second example involves the stabilization of chloroplast mRNAs. The available data lead us predict distinct RNA elements, which interact with trans-acting factors to protect the RNA against nucleolytic attacks.

  10. Phosphoribulokinase from Chlamydomonas reinhardtii: a Benson-Calvin cycle enzyme enslaved to its cysteine residues.

    Science.gov (United States)

    Thieulin-Pardo, Gabriel; Remy, Thérèse; Lignon, Sabrina; Lebrun, Régine; Gontero, Brigitte

    2015-04-01

    Phosphoribulokinase (PRK) in the green alga Chlamydomonas reinhardtii is a finely regulated and well-studied enzyme of the Benson-Calvin cycle. PRK can form a complex with glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and the small chloroplast protein CP12. This study aimed to determine the molecular determinants on PRK involved in the complex and the mechanism of action of a recently described novel regulation of PRK that involves glutathionylation. A combination of mass spectrometry, mutagenesis and activity analyses showed that Cys16, besides its role as the binding site of ATP, was also the site for S-glutathionylation. Previous kinetic analysis of the C55S mutant showed that in the oxidized inactive form of PRK, this residue formed a disulfide bridge with the Cys16 residue. This is the only bridge reported for PRK in the literature. Our data show for the first time that a disulfide bridge between Cys243 and Cys249 on PRK is required to form the PRK-GAPDH-CP12 complex. These results uncover a new mechanism for the PRK-GAPDH-CP12 formation involving a thiol disulfide exchange reaction with CP12 and identify Cys16 of PRK as a target of glutathionylation acting against oxidative stress. Although Cys16 is the key residue involved in binding ATP and acting as a defense against oxidative damage, the formation of the algal ternary complex requires the formation of another disulfide bridge on PRK involving Cys243 and Cys249.

  11. Towards elucidation of the toxic mechanism of copper on the model green alga Chlamydomonas reinhardtii.

    Science.gov (United States)

    Jiang, Yongguang; Zhu, Yanli; Hu, Zhangli; Lei, Anping; Wang, Jiangxin

    2016-09-01

    Toxic effects of copper on aquatic organisms in polluted water bodies have garnered particular attention in recent years. Microalgae play an important role in aquatic ecosystems, and they are sensitive to heavy metal pollution. Thus, it is important to clarify the mechanism of copper toxicity first for ecotoxicology studies. In this study, the physiological, biochemical and gene expression characteristics of a model green microalga, Chlamydomonas reinhardtii, with 0, 50, 150 and 250 μM copper treatments were investigated. The response of C. reinhardtii to copper stress was significantly shown at a dose dependent manner. Inhibition of cell growth and variation of total chlorophyll content were observed with copper treatments. The maximum photochemical efficiency of PSII, actual photochemical efficiency of PSII and photochemical quenching value decreased in the 250 μM copper treatment with minimum values equal to 28, 24 and 60 % of the control values respectively. The content of lipid peroxidation biomarker malondialdehyde with copper treatments increased with a maximum value sevenfold higher than the control value. Inhibition of cell growth and photosynthesis was ascribed to peroxidation of membrane lipids. The glutathione content and activities of antioxidant enzymes, glutathione S-transferase, glutathione peroxidase, superoxide dismutase and peroxidase were induced by copper. Interestingly, the expression of antioxidant genes and the photosynthetic gene decreased in most copper treatments. In conclusion, oxidative stress caused by production of excess reactive oxidative species might be the major mechanism of copper toxicity on C. reinhardtii.

  12. Phytotoxicity of 15 common pharmaceuticals on the germination of Lactuca sativa and photosynthesis of Chlamydomonas reinhardtii.

    Science.gov (United States)

    Pino, Ma Rosa; Muñiz, Selene; Val, Jonatan; Navarro, Enrique

    2016-11-01

    Pharmaceuticals reach terrestrial environments through the application of treated wastewaters and biosolids to agricultural soils. We have investigated the toxicity of 15 common pharmaceuticals, classified as nonsteroidal anti-inflammatory drugs (NSAIDs), blood lipid-lowering agents, β-blockers and antibiotics, in two photosynthetic organisms. Twelve pharmaceuticals caused inhibitory effects on the radicle and hypocotyl elongation of Lactuca sativa seeds. The EC50 values obtained were in the range of 170-5656 mg L(-1) in the case of the radicle and 188-4558 mg L(-1) for the hypocotyl. Propranolol was the most toxic drug for both root and hypocotyl elongation, followed by the NSAIDs, then gemfibrozil and tetracycline. Other effects, such as root necrosis, inhibition of root growth and curly hairs, were detected. However, even at the highest concentrations tested (3000 mg L(-1)), seed germination was not affected. NSAIDs decreased the photosynthetic yield of Chlamydomonas reinhardtii, but only salicylic acid showed EC50 values below 1000 mg L(-1). The first effects detected at low concentrations, together with the concentrations found in environmental samples, indicate that the use of biosolids and wastewaters containing pharmaceuticals should be regulated and their compositions assessed in order to prevent medium- and long-term impacts on agricultural soils and crops.

  13. Light/electricity conversion by defined cocultures of Chlamydomonas and Geobacter.

    Science.gov (United States)

    Nishio, Koichi; Hashimoto, Kazuhito; Watanabe, Kazuya

    2013-04-01

    Biological energy-conversion systems are attractive in terms of their self-organizing and self-sustaining properties and are expected to be applied towards environmentally friendly bioenergy processes. Recent studies have demonstrated that sustainable light/electricity-conversion systems, termed microbial solar cells (MSCs), can be constructed using naturally occurring microbial communities. To better understand the energy-conversion mechanisms in microbial communities, the present study attempted to construct model MSCs comprised of defined cocultures of a green alga, Chlamydomonas reinhardtii, and an iron-reducing bacterium, Geobacter sulfurreducens, and examined their metabolism and interactions in MSCs. When MSC bioreactors were inoculated with these microbes and irradiated on a 12-h light/dark cycle, periodic current was generated in the dark with energy-conversion efficiencies of 0.1%. Metabolite analyses revealed that G. sulfurreducens generated current by oxidizing formate that was produced by C. reinhardtii in the dark. These results demonstrate that the light/electricity conversion occurs via syntrophic interactions between phototrophs and electricity-generating bacteria. Based on the results and data in literatures, it is estimated that the excretion of organics by the phototroph was the bottleneck step in the syntrophic light/electricity conversion. We also discuss differences between natural-community and defined-coculture MSCs.

  14. Genetic Analysis of the Chlamydomonas Reinhardtii I-Crei Mobile Intron Homing System in Escherichia Coli

    Science.gov (United States)

    Seligman, L. M.; Stephens, K. M.; Savage, J. H.; Monnat-Jr., R. J.

    1997-01-01

    We have developed and used a genetic selection system in Escherichia coli to study functional requirements for homing site recognition and cleavage by a representative eukaryotic mobile intron endonuclease. The homing endonuclease, I-CreI, was originally isolated from the chloroplast of the unicellular green alga Chlamydomonas reinhardtii. I-CreI homing site mutants contained base pair substitutions or single base deletions that altered the rate of homing site cleavage and/or product release. I-CreI endonuclease mutants fell into six phenotypic classes that differed in in vivo activity, toxicity or genetic dominance. Inactivating mutations clustered in the N-terminal 60% of the I-CreI amino acid sequence, and two frameshift mutations were isolated that resulted in premature translation termination though retained partial activity. These mutations indicate that the N-terminal two-thirds of the I-CreI endonuclease is sufficient for homing site recognition and cleavage. Substitution mutations altered in four potential active site residues were examined: D20N, Q47H or R70A substitutions inactivated endonuclease activity, whereas S22A did not. The genetic approach we have taken complements phylogenetic and structural studies of mobile intron endonucleases and has provided new information on the mechanistic basis of I-CreI homing site recognition and cleavage. PMID:9409828

  15. An omics based assessment of cadmium toxicity in the green alga Chlamydomonas reinhardtii

    Energy Technology Data Exchange (ETDEWEB)

    Jamers, An; Blust, Ronny; De Coen, Wim [Laboratory for Ecophysiology, Biochemistry and Toxicology, Department of Biology, University of Antwerp, Groenenborgerlaan 171, 2020 Antwerp (Belgium); Griffin, Julian L. [Department of Biochemistry, University of Cambridge, 80 Tennis Court Road, Cambridge CB2 2QA (United Kingdom); Jones, Oliver A.H., E-mail: oliver.jones@rmit.edu.au [School of Applied Sciences, RMIT University, GPO Box 2476, Melbourne, VIC 3001 (Australia)

    2013-01-15

    The effects of cadmium were assessed in the freshwater alga Chlamydomonas reinhardtii. Algae were exposed to concentrations of 0, 8.1 or 114.8 {mu}M of cadmium and growth rates, gene transcription and metabolite profiles were examined after 48 and 72 h of exposure. In algae exposed to 8.1 {mu}M Cd, several genes were differentially transcribed after 48 h but no adverse growth related effects were detected. A transient effect on both gene transcription patterns and metabolite profiles could be discerned after 48 h of exposure but the majority of these changes disappeared after 72 h. In contrast, all effects were more pronounced at the 114.8 {mu}M cadmium exposure. Here growth was clearly reduced and transcription of a large number of genes involved in oxidative stress defense mechanisms was differentially increased. Metabolites involved in the glutathione synthesis pathway (an important antioxidant defense) were also affected but the effects of cadmium were found to be more pronounced at the transcript level than in the metabolome, suggesting that the former exhibits greater sensitivity toward cadmium exposure.

  16. A simple and non-invasive method for nuclear transformation of intact-walled Chlamydomonas reinhardtii.

    Directory of Open Access Journals (Sweden)

    Sora Kim

    Full Text Available Genetic engineering in microalgae is gaining attraction but nuclear transformation methods available so far are either inefficient or require special equipment. In this study, we employ positively charged nanoparticles, 3-aminopropyl-functionalized magnesium phyllosilicate (aminoclay, approximate unit cell composition of [H2N(CH23]8Si8Mg6O12(OH4, for nuclear transformation into eukaryotic microalgae. TEM and EDX analysis of the process of transformation reveals that aminoclay coats negatively-charged DNA biomolecules and forms a self-assembled hybrid nanostructure. Subsequently, when this nanostructure is mixed with microalgal cells and plated onto selective agar plates with high friction force, cell wall is disrupted facilitating delivery of plasmid DNA into the cell and ultimately to the nucleus. This method is not only simple, inexpensive, and non-toxic to cells but also provides efficient transformation (5.03×10(2 transformants/µg DNA, second only to electroporation which needs advanced instrumentation. We present optimized parameters for efficient transformation including pre-treatment, friction force, concentration of foreign DNA/aminoclay, and plasticity of agar plates. It is also confirmed the successful integration and stable expression of foreign gene in Chlamydomonas reinhardtii through molecular methods.

  17. Effects of chromium on photosynthetic and photoreceptive apparatus of the alga Chlamydomonas reinhardtii.

    Science.gov (United States)

    Rodríguez, M Cecilia; Barsanti, Laura; Passarelli, Vincenzo; Evangelista, Valter; Conforti, Visitacion; Gualtieri, Paolo

    2007-10-01

    Chromium is a highly toxic non-essential metal for microorganisms and plants. Due to its widespread industrial use, chromium (Cr) has become a serious pollutant in diverse environmental settings. The presence of Cr leads to the selection of algal populations able to tolerate high levels of Cr compounds. The diverse Cr-resistance mechanisms displayed by microorganisms include biosorption, diminished accumulation, precipitation, reduction of Cr(6+) to Cr(3+), and chromate efflux. In this paper we describe the effects of Cr(6+) (the more toxic species) on the photosynthetic and photoreceptive apparatus of the fresh water unicellular alga Chlamydomonas reinhardtii. We measured the effect of the heavy metal by means of in vivo absorption microspectroscopy of both the thylakoid compartments and the eyespot. The decomposition of the overall absorption spectra in pigment constituents indicates that Cr(6+) induced a complete pheophinitization of the chrorophylls and a modification of the carotenoids present in the eyespot only when its concentration is equal or greater than 10 microM. Due to this low tolerance level, C. reinhardtii could be used as indicator of Cr pollution, but it is not feasible for bioremediation purposes.

  18. Characterization of a Native Algae Species Chlamydomonas debaryana: Strain Selection, Bioremediation Ability, and Lipid Characterization

    Directory of Open Access Journals (Sweden)

    Bo Zhang

    2014-08-01

    Full Text Available Native microalgal species may offer a favorable combination of both wastewater treatment and biofuel production. In this research, a green microalgae, Chlamydomonas debaryana, was isolated from a local lagoon, screened for its lipid content using flow cytometry, and further identified with microscopic observations and DNA sequence analysis. When using swine wastewater as a medium, the biomass yields were between 0.6 and 1.62 g/L, giving a median value of 1.11 g/L. By increasing mass transfer rates and providing sufficient light intensity, the microalgal growth was intrinsically enhanced. The growth of C. debaryana reduced most nutritional contents of the wastewater except iron. When combining the microalgal growth and nutrient removal, C. debaryana was able to utilize 1.3 to 1.6×103 mg COD (chemical oxygen demand/g biomass, 55 to 90 ppm ammonia/g biomass, and 48 to 89 ppm phosphorous/g biomass, The lipid content of C. debaryana was 19.9 ± 4.3% of cell dry weight. The transesterified microalgal oil mostly consisted of 14 kinds of fatty acids, ranging from C5 to C22, which can be refined into renewable jet fuel or used as sources of omega-3 and omega-6 fatty acids.

  19. Phosphoregulation of an Inner Dynein Arm Complex in Chlamydomonas reinhardtii Is Altered in Phototactic Mutant Strains

    Science.gov (United States)

    King, Stephen J.; Dutcher, Susan K.

    1997-01-01

    To gain a further understanding of axonemal dynein regulation, mutant strains of Chlamydomonas reinhardtii that had defects in both phototactic behavior and flagellar motility were identified and characterized. ptm1, ptm2, and ptm3 mutant strains exhibited motility phenotypes that resembled those of known inner dynein arm region mutant strains, but did not have biochemical or genetic phenotypes characteristic of other inner dynein arm mutations. Three other mutant strains had defects in the f class of inner dynein arms. Dynein extracts from the pf9-4 strain were missing the entire f complex. Strains with mutations in pf9/ida1, ida2, or ida3 failed to assemble the f dynein complex and did not exhibit phototactic behavior. Fractionated dynein from mia1-1 and mia2-1 axonemes exhibited a novel f class inner dynein arm biochemical phenotype; the 138-kD f intermediate chain was present in altered phosphorylation forms. In vitro axonemal dynein activity was reduced by the mia1-1 and mia2-1 mutations. The addition of kinase inhibitor restored axonemal dynein activity concomitant with the dephosphorylation of the 138-kD f intermediate chain. Dynein extracts from uni1-1 axonemes, which specifically assemble only one of the two flagella, contained relatively high levels of the altered phosphorylation forms of the 138-kD intermediate chain. We suggest that the f dynein complex may be phosphoregulated asymmetrically between the two flagella to achieve phototactic turning. PMID:9008712

  20. The Chlamydomonas reinhardtii alternative oxidase 1 is regulated by heat stress.

    Science.gov (United States)

    Zalutskaya, Zhanneta; Lapina, Tatiana; Ermilova, Elena

    2015-12-01

    The alternative oxidase (AOX) is a non-energy conserving terminal oxidase that has emerged as an important mitochondrial component of the cell stress responses. Although the most studied abiotic condition in relation to Chlamydomonas reinhardtii is high temperature, changes in AOX capacity of the alga were studied only under oxidative stress and cold. To examine whether elevated temperatures affected AOX1 expression, we applied quantitative real-time PCR and pharmaceutical approaches. In this work, we demonstrated a sharp increase in AOX1 transcript and protein abundance under heat stress. Furthermore, C. reinhardtii cells displayed a large increase in alternative respiration in response to high temperature. Feeding with the protein kinase inhibitor staurosporine strongly retarded the AOX1 transcription. Finally, the addition of the calcium chelator EGTA prevented heat-induced AOX1 expression. Together, our results imply that heat-inducible Ca(2+) influx and protein kinase(s) may mediate AOX1 expression at elevated temperatures. Characterization of heat-induced AOX1 regulation in the green alga C. reinhardtii provides a framework for a more complete understanding of the function of this conserved protein.

  1. Trophic transfer of gold nanoparticles from Euglena gracilis or Chlamydomonas reinhardtii to Daphnia magna.

    Science.gov (United States)

    Lee, Woo-Mi; Yoon, Sung-Ji; Shin, Yu-Jin; An, Youn-Joo

    2015-06-01

    Understanding the trophic transfer of nanoparticles (NPs) is important because NPs are small enough to easily penetrate into organisms. In this study, we evaluated the trophic transfer of gold NPs (AuNPs) within the aquatic food chain. We observed AuNPs transfer from 2 species of primary producers (Chlamydomonas reinhardtii or Euglena gracilis) to the primary consumer (Daphnia magna). Also, bioaccumulation of AuNPs in E. gracilis was higher than that in C. reinhardtii. The reasons for the difference in Au accumulation may be the physical structure of these organisms, and the surface area that is available for interaction with NPs. C. reinhardtii has a cell wall that may act as a barrier to the penetration of NPs. The size of E. gracilis is larger than that of C. reinhardtii. This study demonstrates the trophic transfer of AuNPs from a general producer to a consumer in an aquatic environment.

  2. A small multifunctional pentatricopeptide repeat protein in the chloroplast of Chlamydomonas reinhardtii.

    Science.gov (United States)

    Jalal, Abdullah; Schwarz, Christian; Schmitz-Linneweber, Christian; Vallon, Olivier; Nickelsen, Jörg; Bohne, Alexandra-Viola

    2015-03-01

    Organellar biogenesis is mainly regulated by nucleus-encoded factors, which act on various steps of gene expression including RNA editing, processing, splicing, stabilization, and translation initiation. Among these regulatory factors, pentatricopeptide repeat (PPR) proteins form the largest family of RNA binding proteins, with hundreds of members in flowering plants. In striking contrast, the genome of the unicellular green alga Chlamydomonas reinhardtii encodes only 14 such proteins. In this study, we analyzed PPR7, the smallest and most highly expressed PPR protein in C. reinhardtii. Green fluorescent protein-based localization and gel-filtration analysis revealed that PPR7 forms a part of a high-molecular-weight ribonucleoprotein complex in the chloroplast stroma. RIP-chip analysis of PPR7-bound RNAs demonstrated that the protein associates with a diverse set of chloroplast transcripts in vivo, i.e. rrnS, psbH, rpoC2, rbcL, atpA, cemA-atpH, tscA, and atpI-psaJ. Furthermore, the investigation of PPR7 RNAi strains revealed that depletion of PPR7 results in a light-sensitive phenotype, accompanied by altered levels of its target RNAs that are compatible with the defects in their maturation or stabilization. PPR7 is thus an unusual type of small multifunctional PPR protein, which interacts, probably in conjunction with other RNA binding proteins, with numerous target RNAs to promote a variety of post-transcriptional events.

  3. Identification of regulatory network hubs that control lipid metabolism in Chlamydomonas reinhardtii.

    Science.gov (United States)

    Gargouri, Mahmoud; Park, Jeong-Jin; Holguin, F Omar; Kim, Min-Jeong; Wang, Hongxia; Deshpande, Rahul R; Shachar-Hill, Yair; Hicks, Leslie M; Gang, David R

    2015-08-01

    Microalgae-based biofuels are promising sources of alternative energy, but improvements throughout the production process are required to establish them as economically feasible. One of the most influential improvements would be a significant increase in lipid yields, which could be achieved by altering the regulation of lipid biosynthesis and accumulation. Chlamydomonas reinhardtii accumulates oil (triacylglycerols, TAG) in response to nitrogen (N) deprivation. Although a few important regulatory genes have been identified that are involved in controlling this process, a global understanding of the larger regulatory network has not been developed. In order to uncover this network in this species, a combined omics (transcriptomic, proteomic and metabolomic) analysis was applied to cells grown in a time course experiment after a shift from N-replete to N-depleted conditions. Changes in transcript and protein levels of 414 predicted transcription factors (TFs) and transcriptional regulators (TRs) were monitored relative to other genes. The TF and TR genes were thus classified by two separate measures: up-regulated versus down-regulated and early response versus late response relative to two phases of polar lipid synthesis (before and after TAG biosynthesis initiation). Lipidomic and primary metabolite profiling generated compound accumulation levels that were integrated with the transcript dataset and TF profiling to produce a transcriptional regulatory network. Evaluation of this proposed regulatory network led to the identification of several regulatory hubs that control many aspects of cellular metabolism, from N assimilation and metabolism, to central metabolism, photosynthesis and lipid metabolism.

  4. The response of Chlamydomonas reinhardtii to nitrogen deprivation: a systems biology analysis.

    Science.gov (United States)

    Park, Jeong-Jin; Wang, Hongxia; Gargouri, Mahmoud; Deshpande, Rahul R; Skepper, Jeremy N; Holguin, F Omar; Juergens, Matthew T; Shachar-Hill, Yair; Hicks, Leslie M; Gang, David R

    2015-02-01

    Drastic alteration in macronutrients causes large changes in gene expression in the photosynthetic unicellular alga Chlamydomonas reinhardtii. Preliminary data suggested that cells follow a biphasic response to this change hinging on the initiation of lipid accumulation, and we hypothesized that drastic repatterning of metabolism also followed this biphasic modality. To test this hypothesis, transcriptomic, proteomic, and metabolite changes that occur under nitrogen (N) deprivation were analyzed. Eight sampling times were selected covering the progressive slowing of growth and induction of oil synthesis between 4 and 6 h after N deprivation. Results of the combined, systems-level investigation indicated that C. reinhardtii cells sense and respond on a large scale within 30 min to a switch to N-deprived conditions turning on a largely gluconeogenic metabolic state, which then transitions to a glycolytic stage between 4 and 6 h after N depletion. This nitrogen-sensing system is transduced to carbon- and nitrogen-responsive pathways, leading to down-regulation of carbon assimilation and chlorophyll biosynthesis, and an increase in nitrogen metabolism and lipid biosynthesis. For example, the expression of nearly all the enzymes for assimilating nitrogen from ammonium, nitrate, nitrite, urea, formamide/acetamide, purines, pyrimidines, polyamines, amino acids and proteins increased significantly. Although arginine biosynthesis enzymes were also rapidly up-regulated, arginine pool size changes and isotopic labeling results indicated no increased flux through this pathway.

  5. Coordinated Regulation of Gene Expression for Carotenoid Metabolism in Chlamydomonas reinhardtii

    Institute of Scientific and Technical Information of China (English)

    Tian-Hu Sun; Cheng-Qian Liu; Yuan-Yuan Hui; Wen-Kai Wu; Zhi-Gang Zhou; Shan Lu

    2010-01-01

    Carotenoids are important plant pigments for both light harvesting and photooxidation protection.Using the model system of the unicellular green alga Chlamydomonas reinhardtii,we characterized the regulation of gene expression for carotenoid metabolism by quantifying changes in the transcript abundance of dxs,dxr and ipi in the plastidic methylerythritol phosphate pathway and of ggps,psy,pds,lcyb and bchy,directly involved in carotenoid metabolism,under different photoperiod,light and metabolite treatments.The expression of these genes fluctuated with light/dark shifting.Light treatment also promoted the accumulation of transcripts of all these genes.Of the genes studied,dxs,ggps and lcyb displayed the typical circadian pattern by retaining a rhythmic fluctuation of transcript abundance under both constant light and constant dark entrainments.The expression of these genes could also be regulated by metabolic intermediates.For example,ggps was significantly suppressed by a geranylgeranyl pyrophosphate supplement and ipi was upregulated by isopentenyl pyrophosphate.Furthermore,CrOr,a C.reinhardtii homolog of the recently characterized Or gene that accounts for carotenoid accumulation,also showed co-expression with carotenoid biosynthetic genes such as pds and lcyb.Our data suggest a coordinated regulation on carotenoid metabolism in C.reinhardtii at the transcriptional level.

  6. Alleviation of copper-induced oxidative damage in Chlamydomonas reinhardtii by carbon monoxide.

    Science.gov (United States)

    Zheng, Qi; Meng, Qian; Wei, Yuan Yuan; Yang, Zhi Min

    2011-08-01

    Carbon monoxide (CO) is an endogenous gaseous molecule in plants and animals. Recent studies have shown that it is one of the most essential cellular components regulating many aspects of plant growth and development. However, whether CO regulates the green algae adaptive response to heavy metal toxicity is unknown. The present study investigated the role of CO in regulating Cu-induced oxidative stress in eukaryotic algae Chlamydomonas reinhardtii. Cells pretreated with 5 μM CO for 30 min and followed by exposure to 5 μM Cu(II) for 4 days showed attenuated toxicity. The CO-improved growth of algae was correlated with reduced lipid peroxidation and increased chlorophyll accumulation. The beneficial effect of CO was confirmed by histochemical staining with reactive oxygen species. Further, treatment with 5 μM CO increased the activity of catalase with Cu. However, a reduced superoxide dismutase activity was observed in the CO + Cu-treated algae compared to the control (activity of Cu treatment alone). Under the same condition, the activity of ascorbate peroxidase was not significantly changed. These results suggest that CO can play an important role in regulating the response of algae to Cu stress.

  7. Investigation of the calcium-binding site of the oxygen evolving complex of photosystem II using 87Sr ESEEM spectroscopy.

    Science.gov (United States)

    Kim, Sun Hee; Gregor, Wolfgang; Peloquin, Jeffrey M; Brynda, Marcin; Britt, R David

    2004-06-16

    The proximity of the calcium/strontium binding site of the oxygen evolving complex (OEC) of photosystem II (PSII) to the paramagnetic Mn cluster is explored with (87)Sr three-pulse electron spin-echo envelope modulation (ESEEM) spectroscopy. CW-EPR spectra of Sr(2+)-substituted Ca(2+)-depleted PSII membranes show the modified g = 2 multiline EPR signal as previously reported. We performed three-pulse ESEEM on this modified multiline signal of the Mn cluster using natural abundance Sr and (87)Sr, respectively. Three-pulse ESEEM of the natural abundance Sr sample exhibits no detectable modulation by the 7% abundance (87)Sr. On the other hand, that of the (87)Sr enriched (93%) sample clearly reveals modulation arising from the I = (9)/(2) (87)Sr nucleus weakly magnetically coupled to the Mn cluster. Using a simple point dipole approximation for the electron spin, analysis of the (87)Sr ESEEM modulation depth via an analytic expression suggests a Mn-Ca (Sr) distance of 4.5 A. Simulation of three-pulse ESEEM with a numerical matrix diagonalization procedure gave good agreement with this analytical result. A more appropriate tetranuclear magnetic/structural model for the Mn cluster converts the 4.5 A point dipole distance to a 3.8-5.0 A range of distances. DFT calculations of (43)Ca and (87)Sr quadrupolar interactions on Ca (and Sr substituted) binding sites in various proteins suggest that the lack of the nuclear quadrupole induced splitting in the ESEEM spectrum of (87)Sr enriched PSII samples is related to a very high degree of symmetry of the ligands surrounding the Sr(2+) ion in the substituted Ca site. Numerical simulations show that moderate (87)Sr quadrupolar couplings decrease the envelope modulation relative to the zero quadrupole case, and therefore we consider that the 3.8-5.0 A range obtained without quadrupolar coupling included in the simulation represents an upper limit to the actual manganese-calcium distance. This (87)Sr pulsed EPR spectroscopy provides

  8. Cyclic electron flow around photosystem I via chloroplast NAD(P)H dehydrogenase (NDH) complex performs a significant physiological role during photosynthesis and plant growth at low temperature in rice.

    Science.gov (United States)

    Yamori, Wataru; Sakata, Naoki; Suzuki, Yuji; Shikanai, Toshiharu; Makino, Amane

    2011-12-01

    The role of NAD(P)H dehydrogenase (NDH)-dependent cyclic electron flow around photosystem I in photosynthetic regulation and plant growth at several temperatures was examined in rice (Oryza sativa) that is defective in CHLORORESPIRATORY REDUCTION 6 (CRR6), which is required for accumulation of sub-complex A of the chloroplast NDH complex (crr6). NdhK was not detected by Western blot analysis in crr6 mutants, resulting in lack of a transient post-illumination increase in chlorophyll fluorescence, and confirming that crr6 mutants lack NDH activity. When plants were grown at 28 or 35°C, all examined photosynthetic parameters, including the CO(2) assimilation rate and the electron transport rate around photosystems I and II, at each growth temperature at light intensities above growth light (i.e. 800 μmol photons m(-2) sec(-1)), were similar between crr6 mutants and control plants. However, when plants were grown at 20°C, all the examined photosynthetic parameters were significantly lower in crr6 mutants than control plants, and this effect on photosynthesis caused a corresponding reduction in plant biomass. The F(v)/F(m) ratio was only slightly lower in crr6 mutants than in control plants after short-term strong light treatment at 20°C. However, after long-term acclimation to the low temperature, impairment of cyclic electron flow suppressed non-photochemical quenching and promoted reduction of the plastoquinone pool in crr6 mutants. Taken together, our experiments show that NDH-dependent cyclic electron flow plays a significant physiological role in rice during photosynthesis and plant growth at low temperature.

  9. Effects of Exogenous Spermidine on Photosystem Ⅱ of Wheat Seedlings Under Water Stress

    Institute of Scientific and Technical Information of China (English)

    Hui-Guo Duan; Shu Yuan; Wen-Juan Liu; De-Hui Xi; Dong-Hong Qing; Hou-Guo Liang; Hong-Hui Lin

    2006-01-01

    The effects of exogenous spermidine (Spd) on lipid peroxidation, relative plasma membrane permeability,photosystem Ⅱ (PSⅡ) gene expression and PSⅡ photochemical activity in water-stressed wheat seedlings were investigated. The decrease in relative water content (RWC), Chi content, and 2,6-dichlorophenol indophenol (DCIP) photoreduction of PSⅡ, and increases in electrolyte leakage of plasma membranes and malonyldialdehyde (MDA) in water-stressed leaves was alleviated by Spd pretreatment. Furthermore, Western and Northern blot analysis showed that decreases in the PSⅡ major proteins D1, D2 and LHCⅡ and the transcripts of corresponding genes psbA, psbD and cab were also alleviated by Spd pretreatment under water stress. These results suggest that the application of exogenous Spd protects PSⅡ against water stress at both the transcriptional level and the translational level, and allows PSⅡ to retain a higher activity level during water stress. The protective role of Spd in the photosynthetic apparatus also is discussed.

  10. Methodology of Pulsed Photoacoustics and Its Application to Probe Photosystems and Receptors

    Directory of Open Access Journals (Sweden)

    Harvey J.M. Hou

    2010-06-01

    Full Text Available We review recent advances in the methodology of pulsed time-resolved photoacoustics and its application to studies of photosynthetic reaction centers and membrane receptors such as the G protein-coupled receptor rhodopsin. The experimental parameters accessible to photoacoustics include molecular volume change and photoreaction enthalpy change. Light-driven volume change secondary to protein conformational changes or electrostriction is directly related to the photoreaction and thus can be a useful measurement of activity and function. The enthalpy changes of the photochemical reactions observed can be measured directly by photoacoustics. With the measurement of enthalpy change, the reaction entropy can also be calculated when free energy is known. Dissecting the free energy of a photoreaction into enthalpic and entropic components may provide critical information about photoactivation mechanisms of photosystems and photoreceptors. The potential limitations and future applications of time-resolved photoacoustics are also discussed.

  11. Effect of Phosphatidylcholine on the Steady State Fluorescence of Chlorophyll in Photosystem Ⅱ Particles

    Institute of Scientific and Technical Information of China (English)

    1999-01-01

    Phosphatidylcholine (PC) accounts for less than 1% of the total lipids in plant photosystem II (PSII) particles.In this experiment, PSII particles were reconstituted with PC to construct PSII-PC vesicles.The effect of PC on the steady state fluorescence of chlorophyll (Chl) in PSII particles was studied.The results show that PC significantly affected the fluorescence intensity, but did not obviously affect the fluorescence emission band peak position.PC also did not obviously affect the absorbance at 436 nm or the amide I band peak position in FT-IR spectroscopy of PSII particles.The results suggest that PC may affect the light energy transfer from the antenna chlorophyll molecules to the reaction center chlorophyll molecule (P680).

  12. Photosystem I in Langmuir-Blodgett and Langmuir-Schaefer monolayers.

    Science.gov (United States)

    Yan, Xun; Faulkner, Christopher J; Jennings, G Kane; Cliffel, David E

    2012-10-23

    Photosystem I (PSI) is a membrane protein complex that generates photoinduced electrons and transfers them across the thylakoid membrane during photosynthesis. The PSI complex, separated from spinach leaves, was spread onto the air-water interface as a monolayer and transferred onto a gold electrode surface that was precoated with a self-assembled monolayer (SAM). The electrochemical properties of the transferred PSI monolayer, including cyclic voltammetry and photoinduced chronoamperometry, were measured. The results showed that PSI retained its bioactivity after the manipulation. Its capability of converting photoenergy into electrical potential was demonstrated by its reducing an electron acceptor, dichloroindophenol (DCIP), and by oxidizing an electron donor, sodium ascorbate (ASC). We have shown that the protein has two possible orientations at the water interface. The orientation distribution was determined by comparing the controlled reductive and oxidative photocurrents generated from Langmuir-Blodgett and Langmuir-Schaefer monolayers.

  13. Simultaneous measurements of photocurrents and H2O2 evolution from solvent exposed photosystem 2 complexes.

    Science.gov (United States)

    Vöpel, Tobias; Ning Saw, En; Hartmann, Volker; Williams, Rhodri; Müller, Frank; Schuhmann, Wolfgang; Plumeré, Nicolas; Nowaczyk, Marc; Ebbinghaus, Simon; Rögner, Matthias

    2015-03-22

    In plants, algae, and cyanobacteria, photosystem 2 (PS2) catalyzes the light driven oxidation of water. The main products of this reaction are protons and molecular oxygen. In vitro, however, it was demonstrated that reactive oxygen species like hydrogen peroxide are obtained as partially reduced side products. The transition from oxygen to hydrogen peroxide evolution might be induced by light triggered degradation of PS2's active center. Herein, the authors propose an analytical approach to investigate light induced bioelectrocatalytic processes such as PS2 catalyzed water splitting. By combining chronoamperometry and fluorescence microscopy, the authors can simultaneously monitor the photocurrent and the hydrogen peroxide evolution of light activated, solvent exposed PS2 complexes, which have been immobilized on a functionalized gold electrode. The authors show that under limited electron mediation PS2 displays a lower photostability that correlates with an enhanced H2O2 generation as a side product of the light induced water oxidation.

  14. Effect of phosphatidylglycerol on conformation and microenvironment of tyrosyl residue in photosystem II

    Institute of Scientific and Technical Information of China (English)

    阳振乐; 李良璧; 许亦农; 匡廷云

    2000-01-01

    The structural aspects in the interaction of phosphatidylglycerol (PG) with photosystem II (PSIl), mainly the effect of PQ on conformation and microenvironment of tyrosine residues of PSIl proteins were studied by Fourier transform infrared (FTIR) spectroscopy. It was found that the binding of PG to PSIl particle induces changes in the conformation and micropolarity of phenol ring in the tyrosine residues. In other words, the PG effect on the PSIl results in blue shift of the stretch vibrational band in the phenol ring from 1620 to 1500 cm-1 with the enhancement of the absorb-ance intensity. Additionally, a new spectrum of hydrogen bond was also observed. The results imply that the hydrogen-bond formation between the OH group of phenol and one of PG might cause changes in the structures of tyrosine residues in PSIl proteins.

  15. Effects of Irregular Bimetallic Nanostructures on the Optical Properties of Photosystem I from Thermosynechococcus elongatus

    Directory of Open Access Journals (Sweden)

    Imran Ashraf

    2015-07-01

    Full Text Available The fluorescence of photosystem I (PSI trimers in proximity to bimetallic plasmonic nanostructures have been explored by single-molecule spectroscopy (SMS at cryogenic temperature (1.6 K. PSI serves as a model for biological multichromophore-coupled systems with high potential for biotechnological applications. Plasmonic nanostructures are fabricated by thermal annealing of thin metallic films. The fluorescence of PSI has been intensified due to the coupling with plasmonic nanostructures. Enhancement factors up to 22.9 and 5.1 are observed for individual PSI complexes coupled to Au/Au and Ag/Au samples, respectively. Additionally, a wavelength dependence of fluorescence enhancement is observed, which can be explained by the multichromophoric composition of PSI.

  16. Solar Photo Catalytic Hydrogen Production from water using a dual bed photosystem

    Energy Technology Data Exchange (ETDEWEB)

    Florida Solar Energy Center

    2003-03-30

    A body of work was performed in which the feasibility of photocatalytically decomposing water into its constituent elements using a dual bed, or modular photosystem, under solar radiation was investigated. The system envisioned consists of two modules, each consisting of a shallow, flat, sealed container, in which microscopic photocatalytic particles are immobilized. The photocatalysts absorb light, generating free electrons and lattice vacancy holes, which are capable of performing reductive and oxidative chemistry, respectively. The photocatalysts would be chosen as to whether they specifically promote H{sub 2} or O{sub 2} evolution in their respective containers. An aqueous solution containing a redox mediator is pumped between the two chambers in order to transfer electron equivalents from one reaction to the other.

  17. Effect of phosphatidylglycerol on conformation and micro-environment of tyrosyl residue in photosystem

    Institute of Scientific and Technical Information of China (English)

    2000-01-01

    The structural aspects in the interaction of phosphatidylglycerol (PG) with photosystem II (PSII),mainly the effect of PG on conformation and microenvironment of tyrosine residues of PSII proteins were studied by Fourier transform infrared (FTIR) spectroscopy.It was found that the binding of PG to PSII particle induces changes in the conformation and micropolarity of phenol ring in the tyrosine residues.In other words,the PG effect on the PSII results in blue shift of the stretch vibrational band in the phenol ring from 1620 to 1500 cm-1 with the enhancement of the absorbance intensity.Additionally,a new spectrum of hydrogen bond was also observed.The results imply that the hydrogen-bond formation between the OH group of phenol and one of PG might cause changes in the structures of tyrosine residues in PSII proteins.

  18. Effect of Mn cluster on the formation of superoxide radicals in photoinhibition of photosystem

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    To further realize the action of superoxide radicals (O-2) in photoinhibition of photosystem Ⅱ (PS Ⅱ),we employed 5,5-dimethyl-1-pyrroline-N-oxide (DMPO) as a spin trap, associated with EPR spectroscopy, to study the effect of illumination time on O-*2 formation during high light photoinhibition in PS Ⅱ membranes and Mn-depleted PS Ⅱ membranes. Results indicated that the removal of Mn cluster from PSⅡmembranes has a strong influence on the dynamnics of superoxide formation.The relative mechanism was also discussed.These novel findings may further promote the studies of the structure and function of PSⅡand the mechanism of photoinhibition.

  19. Effects of CeCl3 on Energy Transfer and Oxygen Evolution in Spinach Photosystem

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    Due to 4f electron characteristics and alternation valence, cerium involved in an oxidation-reduction reaction in plant, closely relating to photosynthesis. Our studies proved that cerium could promote photosynthesis and greatly improve spinach growth. However, the mechanism of promoting energy transfer and conversion by cerium remains unclear. Here we reported that the effects of Ce3+ on energy transfer and oxygen evolution in photosystem Ⅱ(PSⅡ) isolated from spinach, which was related to 4f electron characteristics and alternation valence in Ce molecule. The methods of absorption spectrum, fluorescence spectrum were used in the research. Results showed that Ce3+ treatment at low concentration could suitably change PSⅡ microenvironment, increase the absorbance of visible light, improve the energy transfer among amino acids within PSⅡ protein-pigment complex, and accelerate energy transport from tyrosine residue to chlorophyll a. In summary, the photochemical activity of PSⅡ(fluorescence quantum yield) and its oxygen evolving rate were enhanced by Ce3+.

  20. Filling the Green Gap of a Megadalton Photosystem I Complex by Conjugation of Organic Dyes.

    Science.gov (United States)

    Gordiichuk, Pavlo I; Rimmerman, Dolev; Paul, Avishek; Gautier, Daniel A; Gruszka, Agnieszka; Saller, Manfred; de Vries, Jan Willem; Wetzelaer, Gert-Jan A H; Manca, Marianna; Gomulya, Widianta; Matmor, Maayan; Gloukhikh, Ekaterina; Loznik, Mark; Ashkenasy, Nurit; Blom, Paul W M; Rögner, Matthias; Loi, Maria Antonietta; Richter, Shachar; Herrmann, Andreas

    2016-01-20

    Photosynthesis is Nature's major process for converting solar into chemical energy. One of the key players in this process is the multiprotein complex photosystem I (PSI) that through absorption of incident photons enables electron transfer, which makes this protein attractive for applications in bioinspired photoactive hybrid materials. However, the efficiency of PSI is still limited by its poor absorption in the green part of the solar spectrum. Inspired by the existence of natural phycobilisome light-harvesting antennae, we have widened the absorption spectrum of PSI by covalent attachment of synthetic dyes to the protein backbone. Steady-state and time-resolved photoluminescence reveal that energy transfer occurs from these dyes to PSI. It is shown by oxygen-consumption measurements that subsequent charge generation is substantially enhanced under broad and narrow band excitation. Ultimately, surface photovoltage (SPV) experiments prove the enhanced activity of dye-modified PSI even in the solid state.

  1. Lack of Exercise Might Invite Dementia

    Science.gov (United States)

    ... fullstory_163264.html Lack of Exercise Might Invite Dementia Study found being sedentary may make you as ... TV may make you as likely to develop dementia as people genetically predisposed to the condition, a ...

  2. What computational chemistry and magnetic resonance reveal concerning the oxygen evolving centre in Photosystem II.

    Science.gov (United States)

    Terrett, Richard; Petrie, Simon; Stranger, Rob; Pace, Ron J

    2016-09-01

    Density Functional Theory (DFT) computational studies of the Mn4/Ca Oxygen Evolving Complex (OEC) region of Photosystem II in the paramagnetic S2 and S3 states of the water oxdizing catalytic cycle are described. These build upon recent advances in computationally understanding the detailed S1 state OEC geometries, revealed by the recent high resolution Photosystem II crystal structures of Shen et al., at 1.90Å and 1.95Å (Petrie et al., 2015, Angew. Chem. Int. Ed., 54, 7120). The models feature a 'Low Oxidation Paradigm' assumption for the mean Mn oxidation states in the functional enzyme, with the mean oxidation levels being 3.0, 3.25 and 3.5 in S1, S2 and S3, respectively. These calculations are used to infer magnetic exchange interactions within the coupled OEC cluster, particularly in the Electron Paramagnetic Resonance (EPR)-visible S2 and S3 states. Detailed computational estimates of the intrinsic magnitudes and molecular orientations of the (55)Mn hyperfine tensors in the S2 state are presented. These parameters, together with the resultant spin projected hyperfine values are compared with recent appropriate experimental EPR data (Continuous Wave (CW), Electron-Nuclear Double Resonance (ENDOR) and ELDOR (Electron-Electron Double Resonance)-Detected Nuclear Magnetic Resonance (EDNMR)) from the OEC. It is found that an effective Coupled Dimer magnetic organization of the four Mn in the OEC cluster in the S2 and S3 states is able to quantitatively rationalize the observed (55)Mn hyperfine data. This is consistent with structures we propose to represent the likely state of the OEC in the catalytically active form of the enzyme.

  3. A miniature bioassay for testing the acute phytotoxicity of photosystem II herbicides on seagrass.

    Science.gov (United States)

    Wilkinson, Adam D; Collier, Catherine J; Flores, Florita; Mercurio, Phil; O'Brien, Jake; Ralph, Peter J; Negri, Andrew P

    2015-01-01

    Photosystem II (PSII) herbicides have been detected in nearshore tropical waters such as those of the Great Barrier Reef and may add to the pressure posed by runoff containing sediments and nutrients to threatened seagrass habitats. There is a growing number of studies into the potential effects of herbicides on seagrass, generally using large experimental setups with potted plants. Here we describe the successful development of an acute 12-well plate phytotoxicity assay for the PSII herbicide Diuron using isolated Halophila ovalis leaves. Fluorescence images demonstrated Diuron affected the entire leaf surface evenly and responses were not influenced by isolating leaves from the plant. The optimum exposure duration was 24 h, by which time the inhibition of effective quantum yield of PSII (∆F/F(m)') was highest and no deterioration of photosystems was evident in control leaves. The inhibition of ∆F/F(m)' by Diuron in isolated H. ovalis leaves was identical to both potted and hydroponically grown plants (with leaves remaining attached to rhizomes), indicating similar reductions in photosynthetic activity in these acute well-plate assays. The sensitivity of the assay was not influenced by irradiance (range tested 40 to 400 μmol photons m(-2) s(-1)). High irradiance, however, caused photo-oxidative stress in H. ovalis and this generally impacted in an additive or sub-additive way with Diuron to damage PSII. The bioassay using isolated leaves is more rapid, uses far less biological material and does not rely on specialised aquarium facilities in comparison with assays using potted plants. The development and validation of this sensitive bioassay will be useful to reliably screen and monitor the phytotoxicity of existing and emerging PSII herbicides and contribute to risk assessments and water quality guideline development in the future.

  4. A miniature bioassay for testing the acute phytotoxicity of photosystem II herbicides on seagrass.

    Directory of Open Access Journals (Sweden)

    Adam D Wilkinson

    Full Text Available Photosystem II (PSII herbicides have been detected in nearshore tropical waters such as those of the Great Barrier Reef and may add to the pressure posed by runoff containing sediments and nutrients to threatened seagrass habitats. There is a growing number of studies into the potential effects of herbicides on seagrass, generally using large experimental setups with potted plants. Here we describe the successful development of an acute 12-well plate phytotoxicity assay for the PSII herbicide Diuron using isolated Halophila ovalis leaves. Fluorescence images demonstrated Diuron affected the entire leaf surface evenly and responses were not influenced by isolating leaves from the plant. The optimum exposure duration was 24 h, by which time the inhibition of effective quantum yield of PSII (∆F/F(m' was highest and no deterioration of photosystems was evident in control leaves. The inhibition of ∆F/F(m' by Diuron in isolated H. ovalis leaves was identical to both potted and hydroponically grown plants (with leaves remaining attached to rhizomes, indicating similar reductions in photosynthetic activity in these acute well-plate assays. The sensitivity of the assay was not influenced by irradiance (range tested 40 to 400 μmol photons m(-2 s(-1. High irradiance, however, caused photo-oxidative stress in H. ovalis and this generally impacted in an additive or sub-additive way with Diuron to damage PSII. The bioassay using isolated leaves is more rapid, uses far less biological material and does not rely on specialised aquarium facilities in comparison with assays using potted plants. The development and validation of this sensitive bioassay will be useful to reliably screen and monitor the phytotoxicity of existing and emerging PSII herbicides and contribute to risk assessments and water quality guideline development in the future.

  5. Application of peptide gemini surfactants as novel solubilization surfactants for photosystems I and II of cyanobacteria.

    Science.gov (United States)

    Koeda, Shuhei; Umezaki, Katsunari; Noji, Tomoyasu; Ikeda, Atsushi; Kawakami, Keisuke; Kondo, Masaharu; Yamamoto, Yasushi; Shen, Jian-Ren; Taga, Keijiro; Dewa, Takehisa; Ito, Shigeru; Nango, Mamoru; Tanaka, Toshiki; Mizuno, Toshihisa

    2013-09-17

    We designed novel peptide gemini surfactants (PG-surfactants), DKDKC12K and DKDKC12D, which can solubilize Photosystem I (PSI) of Thermosynecoccus elongatus and Photosystem II (PSII) of Thermosynecoccus vulcanus in an aqueous buffer solution. To assess the detailed effects of PG-surfactants on the original supramolecular membrane protein complexes and functions of PSI and PSII, we applied the surfactant exchange method to the isolated PSI and PSII. Spectroscopic properties, light-induced electron transfer activity, and dynamic light scattering measurements showed that PSI and PSII could be solubilized not only with retention of the original supramolecular protein complexes and functions but also without forming aggregates. Furthermore, measurement of the lifetime of light-induced charge-separation state in PSI revealed that both surfactants, especially DKDKC12D, displayed slight improvement against thermal denaturation below 60 °C compared with that using β-DDM. This degree of improvement in thermal resistance still seems low, implying that the peptide moieties did not interact directly with membrane protein surfaces. By conjugating an electron mediator such as methyl viologen (MV(2+)) to DKDKC12K (denoted MV-DKDKC12K), we obtained derivatives that can trap the generated reductive electrons from the light-irradiated PSI. After immobilization onto an indium tin oxide electrode, a cathodic photocurrent from the electrode to the PSI/MV-DKDKC12K conjugate was observed in response to the interval of light irradiation. These findings indicate that the PG-surfactants DKDKC12K and DKDKC12D provide not only a new class of solubilization surfactants but also insights into designing other derivatives that confer new functions on PSI and PSII.

  6. Excess manganese differentially inhibits photosystem I versus II in Arabidopsis thaliana.

    Science.gov (United States)

    Millaleo, R; Reyes-Díaz, M; Alberdi, M; Ivanov, A G; Krol, M; Hüner, N P A

    2013-01-01

    The effects of exposure to increasing manganese concentrations (50-1500 µM) from the start of the experiment on the functional performance of photosystem II (PSII) and photosystem I (PSI) and photosynthetic apparatus composition of Arabidopsis thaliana were compared. In agreement with earlier studies, excess Mn caused minimal changes in the PSII photochemical efficiency measured as F(v)/F(m), although the characteristic peak temperature of the S(2/3)Q(B) (-) charge recombinations was shifted to lower temperatures at the highest Mn concentration. SDS-PAGE and immunoblot analyses also did not exhibit any significant change in the relative abundance of PSII-associated polypeptides: PSII reaction centre protein D1, Lhcb1 (major light-harvesting protein of LHCII complex), and PsbO (OEC33, a 33 kDa protein of the oxygen-evolving complex). In addition, the abundance of Rubisco also did not change with Mn treatments. However, plants grown under excess Mn exhibited increased susceptibility to PSII photoinhibition. In contrast, in vivo measurements of the redox transients of PSI reaction centre (P700) showed a considerable gradual decrease in the extent of P700 photooxidation (P700(+)) under increased Mn concentrations compared to control. This was accompanied by a slower rate of P700(+) re-reduction indicating a downregulation of the PSI-dependent cyclic electron flow. The abundance of PSI reaction centre polypeptides (PsaA and PsaB) in plants under the highest Mn concentration was also significantly lower compared to the control. The results demonstrate for the first time that PSI is the major target of Mn toxicity within the photosynthetic apparatus of Arabidopsis plants. The possible involvement mechanisms of Mn toxicity targeting specifically PSI are discussed.

  7. Photoinhibition of photosystem I in a pea mutant with altered LHCII organization.

    Science.gov (United States)

    Ivanov, A G; Morgan-Kiss, R M; Krol, M; Allakhverdiev, S I; Zanev, Yu; Sane, P V; Huner, N P A

    2015-11-01

    Comparative analysis of in vivo chlorophyll fluorescence imaging revealed that photosystem II (PSII) photochemical efficiency (Fv/Fm) of leaves of the Costata 2/133 pea mutant with altered pigment composition and decreased level of oligomerization of the light harvesting chlorophyll a/b-protein complexes (LHCII) of PSII (Dobrikova et al., 2000; Ivanov et al., 2005) did not differ from that of WT. In contrast, photosystem I (PSI) activity of the Costata 2/133 mutant measured by the far-red (FR) light inducible P700 (P700(+)) signal exhibited 39% lower steady state level of P700(+), a 2.2-fold higher intersystem electron pool size (e(-)/P700) and higher rate of P700(+) re-reduction, which indicate an increased capacity for PSI cyclic electron transfer (CET) in the Costata 2/133 mutant than WT. The mutant also exhibited a limited capacity for state transitions. The lower level of oxidizable P700 (P700(+)) is consistent with a lower amount of PSI related chlorophyll protein complexes and lower abundance of the PsaA/PsaB heterodimer, PsaD and Lhca1 polypeptides in Costata 2/133 mutant. Exposure of WT and the Costata 2/133 mutant to high light stress resulted in a comparable photoinhibition of PSII measured in vivo, although the decrease of Fv/Fm was modestly higher in the mutant plants. However, under the same photoinhibitory conditions PSI photochemistry (P700(+)) measured as ΔA820-860 was inhibited to a greater extent (50%) in the Costata 2/133 mutant than in the WT (22%). This was accompanied by a 50% faster re-reduction rate of P700(+) in the dark indicating a higher capacity for CET around PSI in high light treated mutant leaves. The role of chloroplast thylakoid organization on the stability of the PSI complex and its susceptibility to high light stress is discussed.

  8. Comparison of excitation energy transfer in cyanobacterial photosystem I in solution and immobilized on conducting glass.

    Science.gov (United States)

    Szewczyk, Sebastian; Giera, Wojciech; D'Haene, Sandrine; van Grondelle, Rienk; Gibasiewicz, Krzysztof

    2017-05-01

    Excitation energy transfer in monomeric and trimeric forms of photosystem I (PSI) from the cyanobacterium Synechocystis sp. PCC 6803 in solution or immobilized on FTO conducting glass was compared using time-resolved fluorescence. Deposition of PSI on glass preserves bi-exponential excitation decay of ~4-7 and ~21-25 ps lifetimes characteristic of PSI in solution. The faster phase was assigned in part to photochemical quenching (charge separation) of excited bulk chlorophylls and in part to energy transfer from bulk to low-energy (red) chlorophylls. The slower phase was assigned to photochemical quenching of the excitation equilibrated over bulk and red chlorophylls. The main differences between dissolved and immobilized PSI (iPSI) are: (1) the average excitation decay in iPSI is about 11 ps, which is faster by a few ps than for PSI in solution due to significantly faster excitation quenching of bulk chlorophylls by charge separation (~10 ps instead of ~15 ps) accompanied by slightly weaker coupling of bulk and red chlorophylls; (2) the number of red chlorophylls in monomeric PSI increases twice-from 3 in solution to 6 after immobilization-as a result of interaction with neighboring monomers and conducting glass; despite the increased number of red chlorophylls, the excitation decay accelerates in iPSI; (3) the number of red chlorophylls in trimeric PSI is 4 (per monomer) and remains unchanged after immobilization; (4) in all the samples under study, the free energy gap between mean red (emission at ~710 nm) and mean bulk (emission at ~686 nm) emitting states of chlorophylls was estimated at a similar level of 17-27 meV. All these observations indicate that despite slight modifications, dried PSI complexes adsorbed on the FTO surface remain fully functional in terms of excitation energy transfer and primary charge separation that is particularly important in the view of photovoltaic applications of this photosystem.

  9. 衣藻有性生殖的分子机制%Molecular Mechanism of Chlamydomonas Mating

    Institute of Scientific and Technical Information of China (English)

    李修岭; 李夜光

    2009-01-01

    衣藻作为分子生物学研究的模式材料,被广泛用于植物光合作用、鞭毛组装与功能、细胞周期与节律、细胞信号传导与光感受、细胞识别等重要生物学过程的研究,而且衣藻有性生殖的分子机制与人类某些疾病的发生机制存在联系.该文对国内外近年来有关莱茵衣藻在有性生殖过程中凝集素的动态分布,包括鞭毛粘连、补充、传递、脱粘连、凝集素合成的正调节,以及与性凝集素行为有关的基因研究进展进行综述,以阐明衣藻有性生殖的分子机制,为人类的疾病研究提供参考.%As a model organism for studying photosynthesis of plant,flagella assembly and function,cell cycle and circadian rhythms,signal transduction,light perception and cell recognition,Chlamydomonas has been investigated widely.This review reported the advances in molecular mechanism of Chlamydomonas reinhardtii mating.We focused on adhension,tipping,disadhension and regulation of synthesis of sex agglutinins as well as some genes related with adhension of agglutinins.The molecular mechanism of Chlamydomonas mating might provide reference to the study of some human diseases.

  10. LACK - a VoIP Steganographic Method

    CERN Document Server

    Mazurczyk, Wojciech

    2008-01-01

    The paper presents a new steganographic method called LACK (Lost Audio PaCKets Steganography) which is intended mainly for VoIP. The method is presented in a broader context of network steganography and of VoIP steganography in particular. The analytical results presented in the paper concern the influence of LACK's hidden data insertion procedure on the method's impact on quality of voice transmission and its resistance to steganalysis.

  11. Filling Knowledge Gaps in Biological Networks: integrating global approaches to understand H2 metabolism in Chlamydomonas reinhardtii - Final Report

    Energy Technology Data Exchange (ETDEWEB)

    Posewitz, Matthew C

    2011-06-30

    The green alga Chlamydomonas reinhardtii (Chlamydomonas) has numerous genes encoding enzymes that function in fermentative pathways. Among these genes, are the [FeFe]-hydrogenases, pyruvate formate lyase, pyruvate ferredoxin oxidoreductase, acetate kinase, and phosphotransacetylase. We have systematically undertaken a series of targeted mutagenesis approaches to disrupt each of these key genes and omics techniques to characterize alterations in metabolic flux. Funds from DE-FG02-07ER64423 were specifically leveraged to generate mutants with disruptions in the genes encoding the [FeFe]-hydrogenases HYDA1 and HYDA2, pyruvate formate lyase (PFL1), and in bifunctional alcohol/aldehyde alcohol dehydrogenase (ADH1). Additionally funds were used to conduct global transcript profiling experiments of wildtype Chlamydomonas cells, as well as of the hydEF-1 mutant, which is unable to make H2 due to a lesion in the [FeFe]-hydrogenase biosynthetic pathway. In the wildtype cells, formate, acetate and ethanol are the dominant fermentation products with traces of CO2 and H2 also being produced. In the hydEF-1 mutant, succinate production is increased to offset the loss of protons as a terminal electron acceptor. In the pfl-1 mutant, lactate offsets the loss of formate production, and in the adh1-1 mutant glycerol is made instead of ethanol. To further probe the system, we generated a double mutant (pfl1-1 adh1) that is unable to synthesize both formate and ethanol. This strain, like the pfl1 mutants, secreted lactate, but also exhibited a significant increase in the levels of extracellular glycerol, acetate, and intracellular reduced sugars, and a decline in dark, fermentative H2 production. Whereas wild-type Chlamydomonas fermentation primarily produces formate and ethanol, the double mutant performs a complete rerouting of the glycolytic carbon to lactate and glycerol. Lastly, transcriptome data have been analysed for both the wildtype and hydEF-1, that correlate with our

  12. The path to triacylglyceride obesity in the sta6 strain of Chlamydomonas reinhardtii.

    Science.gov (United States)

    Goodenough, Ursula; Blaby, Ian; Casero, David; Gallaher, Sean D; Goodson, Carrie; Johnson, Shannon; Lee, Jae-Hyeok; Merchant, Sabeeha S; Pellegrini, Matteo; Roth, Robyn; Rusch, Jannette; Singh, Manmilan; Umen, James G; Weiss, Taylor L; Wulan, Tuya

    2014-05-01

    When the sta6 (starch-null) strain of the green microalga Chlamydomonas reinhardtii is nitrogen starved in acetate and then "boosted" after 2 days with additional acetate, the cells become "obese" after 8 days, with triacylglyceride (TAG)-filled lipid bodies filling their cytoplasm and chloroplasts. To assess the transcriptional correlates of this response, the sta6 strain and the starch-forming cw15 strain were subjected to RNA-Seq analysis during the 2 days prior and 2 days after the boost, and the data were compared with published reports using other strains and growth conditions. During the 2 h after the boost, ∼425 genes are upregulated ≥2-fold and ∼875 genes are downregulated ≥2-fold in each strain. Expression of a small subset of "sensitive" genes, encoding enzymes involved in the glyoxylate and Calvin-Benson cycles, gluconeogenesis, and the pentose phosphate pathway, is responsive to culture conditions and genetic background as well as to boosting. Four genes-encoding a diacylglycerol acyltransferase (DGTT2), a glycerol-3-P dehydrogenase (GPD3), and two candidate lipases (Cre03.g155250 and Cre17.g735600)-are selectively upregulated in the sta6 strain. Although the bulk rate of acetate depletion from the medium is not boost enhanced, three candidate acetate permease-encoding genes in the GPR1/FUN34/YaaH superfamily are boost upregulated, and 13 of the "sensitive" genes are strongly responsive to the cell's acetate status. A cohort of 64 autophagy-related genes is downregulated by the boost. Our results indicate that the boost serves both to avert an autophagy program and to prolong the operation of key pathways that shuttle carbon from acetate into storage lipid, the combined outcome being enhanced TAG accumulation, notably in the sta6 strain.

  13. The Path to Triacylglyceride Obesity in the sta6 Strain of Chlamydomonas reinhardtii

    Science.gov (United States)

    Blaby, Ian; Casero, David; Gallaher, Sean D.; Goodson, Carrie; Johnson, Shannon; Lee, Jae-Hyeok; Merchant, Sabeeha S.; Pellegrini, Matteo; Roth, Robyn; Rusch, Jannette; Singh, Manmilan; Umen, James G.; Weiss, Taylor L.; Wulan, Tuya

    2014-01-01

    When the sta6 (starch-null) strain of the green microalga Chlamydomonas reinhardtii is nitrogen starved in acetate and then “boosted” after 2 days with additional acetate, the cells become “obese” after 8 days, with triacylglyceride (TAG)-filled lipid bodies filling their cytoplasm and chloroplasts. To assess the transcriptional correlates of this response, the sta6 strain and the starch-forming cw15 strain were subjected to RNA-Seq analysis during the 2 days prior and 2 days after the boost, and the data were compared with published reports using other strains and growth conditions. During the 2 h after the boost, ∼425 genes are upregulated ≥2-fold and ∼875 genes are downregulated ≥2-fold in each strain. Expression of a small subset of “sensitive” genes, encoding enzymes involved in the glyoxylate and Calvin-Benson cycles, gluconeogenesis, and the pentose phosphate pathway, is responsive to culture conditions and genetic background as well as to boosting. Four genes—encoding a diacylglycerol acyltransferase (DGTT2), a glycerol-3-P dehydrogenase (GPD3), and two candidate lipases (Cre03.g155250 and Cre17.g735600)—are selectively upregulated in the sta6 strain. Although the bulk rate of acetate depletion from the medium is not boost enhanced, three candidate acetate permease-encoding genes in the GPR1/FUN34/YaaH superfamily are boost upregulated, and 13 of the “sensitive” genes are strongly responsive to the cell's acetate status. A cohort of 64 autophagy-related genes is downregulated by the boost. Our results indicate that the boost serves both to avert an autophagy program and to prolong the operation of key pathways that shuttle carbon from acetate into storage lipid, the combined outcome being enhanced TAG accumulation, notably in the sta6 strain. PMID:24585881

  14. Experimental Definition and Validation of Protein Coding Transcripts in Chlamydomonas reinhardtii

    Energy Technology Data Exchange (ETDEWEB)

    Kourosh Salehi-Ashtiani; Jason A. Papin

    2012-01-13

    Algal fuel sources promise unsurpassed yields in a carbon neutral manner that minimizes resource competition between agriculture and fuel crops. Many challenges must be addressed before algal biofuels can be accepted as a component of the fossil fuel replacement strategy. One significant challenge is that the cost of algal fuel production must become competitive with existing fuel alternatives. Algal biofuel production presents the opportunity to fine-tune microbial metabolic machinery for an optimal blend of biomass constituents and desired fuel molecules. Genome-scale model-driven algal metabolic design promises to facilitate both goals by directing the utilization of metabolites in the complex, interconnected metabolic networks to optimize production of the compounds of interest. Using Chlamydomonas reinhardtii as a model, we developed a systems-level methodology bridging metabolic network reconstruction with annotation and experimental verification of enzyme encoding open reading frames. We reconstructed a genome-scale metabolic network for this alga and devised a novel light-modeling approach that enables quantitative growth prediction for a given light source, resolving wavelength and photon flux. We experimentally verified transcripts accounted for in the network and physiologically validated model function through simulation and generation of new experimental growth data, providing high confidence in network contents and predictive applications. The network offers insight into algal metabolism and potential for genetic engineering and efficient light source design, a pioneering resource for studying light-driven metabolism and quantitative systems biology. Our approach to generate a predictive metabolic model integrated with cloned open reading frames, provides a cost-effective platform to generate metabolic engineering resources. While the generated resources are specific to algal systems, the approach that we have developed is not specific to algae and

  15. Cellular oxido-reductive proteins of Chlamydomonas reinhardtii control the biosynthesis of silver nanoparticles

    Directory of Open Access Journals (Sweden)

    Barwal Indu

    2011-12-01

    Full Text Available Abstract Background Elucidation of molecular mechanism of silver nanoparticles (SNPs biosynthesis is important to control its size, shape and monodispersity. The evaluation of molecular mechanism of biosynthesis of SNPs is of prime importance for the commercialization and methodology development for controlling the shape and size (uniform distribution of SNPs. The unicellular algae Chlamydomonas reinhardtii was exploited as a model system to elucidate the role of cellular proteins in SNPs biosynthesis. Results The C. reinhardtii cell free extract (in vitro and in vivo cells mediated synthesis of silver nanoparticles reveals SNPs of size range 5 ± 1 to 15 ± 2 nm and 5 ± 1 to 35 ± 5 nm respectively. In vivo biosynthesized SNPs were localized in the peripheral cytoplasm and at one side of flagella root, the site of pathway of ATP transport and its synthesis related enzymes. This provides an evidence for the involvement of oxidoreductive proteins in biosynthesis and stabilization of SNPs. Alteration in size distribution and decrease of synthesis rate of SNPs in protein-depleted fractions confirmed the involvement of cellular proteins in SNPs biosynthesis. Spectroscopic and SDS-PAGE analysis indicate the association of various proteins on C. reinhardtii mediated in vivo and in vitro biosynthesized SNPs. We have identified various cellular proteins associated with biosynthesized (in vivo and in vitro SNPs by using MALDI-MS-MS, like ATP synthase, superoxide dismutase, carbonic anhydrase, ferredoxin-NADP+ reductase, histone etc. However, these proteins were not associated on the incubation of pre-synthesized silver nanoparticles in vitro. Conclusion Present study provides the indication of involvement of molecular machinery and various cellular proteins in the biosynthesis of silver nanoparticles. In this report, the study is mainly focused towards understanding the role of diverse cellular protein in the synthesis and capping of silver

  16. A novel screening protocol for the isolation of hydrogen producing Chlamydomonas reinhardtii strains

    Directory of Open Access Journals (Sweden)

    Happe Thomas

    2008-10-01

    Full Text Available Abstract Background Sealed Chlamydomonas reinhardtii cultures evolve significant amounts of hydrogen gas under conditions of sulfur depletion. However, the eukaryotic green alga goes through drastic metabolic changes during this nutritional stress resulting in cell growth inhibition and eventually cell death. This study aimed at isolating C. reinhardtii transformants which produce hydrogen under normal growth conditions to allow a continuous hydrogen metabolism without the stressful impact of nutrient deprivation. Results To achieve a steady photobiological hydrogen production, a screening protocol was designed to identify C. reinhardtii DNA insertional mutagenesis transformants with an attenuated photosynthesis to respiration capacity ratio (P/R ratio. The screening protocol entails a new and fast method for mutant strain selection altered in their oxygen production/consumption balance. Out of 9000 transformants, four strains with P/R ratios varying from virtually zero to three were isolated. Strain apr1 was found to have a slightly higher respiration rate and a significantly lower photosynthesis rate than the wild type. Sealed cultures of apr1 became anaerobic in normal growth medium (TAP under moderate light conditions and induced [FeFe]-hydrogenase activity, yet without significant hydrogen gas evolution. However, Calvin-Benson cycle inactivation of anaerobically adapted apr1 cells in the light led to a 2-3-fold higher in vivo hydrogen production than previously reported for the sulfur-deprived C. reinhardtii wild type. Conclusion Attenuated P/R capacity ratio in microalgal mutants constitutes a platform for achieving steady state photobiological hydrogen production. Using this platform, algal hydrogen metabolism can be analyzed without applying nutritional stress. Furthermore, these strains promise to be useful for biotechnological hydrogen generation, since high in vivo hydrogen production rates are achievable under normal growth

  17. Inorganic polyphosphate occurs in the cell wall of Chlamydomonas reinhardtii and accumulates during cytokinesis

    Directory of Open Access Journals (Sweden)

    Freimoser Florian M

    2007-09-01

    Full Text Available Abstract Background Inorganic polyphosphate (poly P, linear chains of phosphate residues linked by energy rich phosphoanhydride bonds, is found in every cell and organelle and is abundant in algae. Depending on its localization and concentration, poly P is involved in various biological functions. It serves, for example, as a phosphate store and buffer against alkali, is involved in energy metabolism and regulates the activity of enzymes. Bacteria defective in poly P synthesis are impaired in biofilm development, motility and pathogenicity. PolyP has also been found in fungal cell walls and bacterial envelopes, but has so far not been measured directly or stained specifically in the cell wall of any plant or alga. Results Here, we demonstrate the presence of poly P in the cell wall of Chlamydomonas reinhardtii by staining with specific poly P binding proteins. The specificity of the poly P signal was verified by various competition experiments, by staining with different poly P binding proteins and by correlation with biochemical quantification. Microscopical investigation at different time-points during growth revealed fluctuations of the poly P signal synchronous with the cell cycle: The poly P staining peaked during late cytokinesis and was independent of the high intracellular poly P content, which fluctuated only slightly during the cell cycle. Conclusion The presented staining method provides a specific and sensitive tool for the study of poly P in the extracellular matrices of algae and could be used to describe the dynamic behaviour of cell wall poly P during the cell cycle. We assume that cell wall poly P and intracellular poly P are regulated by distinct mechanisms and it is suggested that cell wall bound poly P might have important protective functions against toxic compounds or pathogens during cytokinesis, when cells are more vulnerable.

  18. L,L-diaminopimelate aminotransferase from Chlamydomonas reinhardtii: a target for algaecide development.

    Directory of Open Access Journals (Sweden)

    Renwick C J Dobson

    Full Text Available In some bacterial species and photosynthetic cohorts, including algae, the enzyme L,L-diaminopimelate aminotransferase (DapL (E.C. 2.6.1.83 is involved in the anabolism of the essential amino acid L-lysine. DapL catalyzes the conversion of tetrahydrodipicolinate (THDPA to L,L-diaminopimelate (L,L-DAP, in one step bypassing the DapD, DapC and DapE enzymatic reactions present in the acyl DAP pathways. Here we present an in vivo and in vitro characterization of the DapL ortholog from the alga Chlamydomonas reinhardtii (Cr-DapL. The in vivo analysis illustrated that the enzyme is able to functionally complement the E. coli dap auxotrophs and was essential for plant development in Arabidopsis. In vitro, the enzyme was able to inter-convert THDPA and L,L-DAP, showing strong substrate specificity. Cr-DapL was dimeric in both solution and when crystallized. The structure of Cr-DapL was solved in its apo form, showing an overall architecture of a α/β protein with each monomer in the dimer adopting a pyridoxal phosphate-dependent transferase-like fold in a V-shaped conformation. The active site comprises residues from both monomers in the dimer and shows some rearrangement when compared to the apo-DapL structure from Arabidopsis. Since animals do not possess the enzymatic machinery necessary for the de novo synthesis of the amino acid L-lysine, enzymes involved in this pathway are attractive targets for the development of antibiotics, herbicides and algaecides.

  19. Preventive effect of the microalga Chlamydomonas debaryana on the acute phase of experimental colitis in rats.

    Science.gov (United States)

    Avila-Román, Javier; Talero, Elena; Alcaide, Antonio; Reyes, Carolina de Los; Zubía, Eva; García-Mauriño, Sofía; Motilva, Virginia

    2014-10-14

    Inflammatory bowel diseases (IBD) are characterised by chronic uncontrolled inflammation of intestinal mucosa. Diet and nutritional factors have emerged as possible interventions for IBD. Microalgae are rich sources of n-3 PUFA and derived oxylipins. Oxylipins are lipid mediators involved in the resolution of many inflammatory disorders. The aim of the present study was to investigate the effects of the oxylipin-containing biomass of the microalga Chlamydomonas debaryana and its major oxylipin constituent, (9Z,11E,13S,15Z)-13-hydroxyoctadeca-9,11,15-trienoic acid ((13S)-HOTE), on acute 2,4,6-trinitrobenzenesulfonic acid (TNBS)-induced colitis in rats. Lyophilised microalgal biomass and (13S)-HOTE were administered by oral route 48, 24 and 1 h before the induction of colitis and 24 h later, and the rats were killed after 48 h. The treatment with the lyophilised microalga and (13S)-HOTE improved body-weight loss and colon shortening, as well as attenuated the extent of colonic damage and increased mucus production. Cellular neutrophil infiltration, with the subsequent increase in myeloperoxidase levels induced by TNBS, were also reduced after the administration of the lyophilised microalga or (13S)-HOTE. The anti-inflammatory effects of these treatments were confirmed by the inhibition of colonic TNF-α production. Moreover, lyophilised microalga or (13S)-HOTE down-regulated cyclo-oxygenase-2 and inducible nitric oxide synthase expression. The present study was the first to show the prophylactic effects of a lyophilised biomass sample of the microalga C. debaryana and the oxylipin (13S)-HOTE on TNBS-induced acute colitis in rats. Our findings suggest that the microalga C. debaryana or derived oxylipins could be used as nutraceuticals in the treatment of the active phase of IBD.

  20. Sulphate, more than a nutrient, protects the microalga Chlamydomonas moewusii from cadmium toxicity.

    Science.gov (United States)

    Mera, Roi; Torres, Enrique; Abalde, Julio

    2014-03-01

    Sulphur is an essential macroelement that plays important roles in living organisms. The thiol rich sulphur compounds, such as cysteine, γ-Glu-Cys, glutathione and phytochelatins participate in the tolerance mechanisms against cadmium toxicity. Plants, algae, yeasts and most prokaryotes cover their demand for reduced sulphur by reduction of inorganic sulphate. The aim of this study was to investigate, using a bifactorial experimental design, the effect of different sulphate concentrations in the nutrient solution on cadmium toxicity in the freshwater microalga Chlamydomonas moewusii. Cell growth, kinetic parameters of sulphate utilization and intracellular concentrations of low-molecular mass thiol compounds were determined. A mathematical model to describe the growth of this microalga based on the effects of sulphate and cadmium was obtained. An ANOVA revealed an interaction between them, 16% of the effect sizes was explained by this interaction. A higher amount of sulphate in the culture medium allowed a higher cadmium tolerance due to an increase in the thiol compound biosynthesis. The amount of low-molecular mass thiol compounds, mainly phytochelatins, synthesized by this microalga was significantly dependent on the sulphate and cadmium concentrations; the higher phytochelatin content was obtained in cultures with 4 mg Cd/L and 1mM sulphate. The maximum EC50 value (based on nominal cadmium concentration) reached for this microalga was 4.46 ± 0.42 mg Cd/L when the sulphate concentration added to the culture medium was also 1mM. An increase in the sulphate concentration, in deficient environments, could alleviate the toxic effect of this metal; however, a relative excess is also negative. The results obtained showed a substrate inhibition for this nutrient. An uncompetitive model for sulphate was chosen to establish the mathematical model that links both factors.

  1. High yield secretion of recombinant proteins from the microalga Chlamydomonas reinhardtii.

    Science.gov (United States)

    Ramos-Martinez, E M; Fimognari, L; Sakuragi, Y

    2017-02-16

    Microalga-based biomanufacturing of recombinant proteins is attracting growing attention due to its advantages in safety, metabolic diversity, scalability, and sustainability. Secretion of recombinant proteins can accelerate the use of microalgal platforms by allowing post-translational modifications and easy recovery of products from the culture media. However, currently, the yields of secreted recombinant proteins are low, which hampers the commercial application of this strategy. This study aimed at expanding the genetic tools for enhancing secretion of recombinant proteins in Chlamydomonas reinhardtii, a widely used green microalga as a model organism and a potential industrial biotechnology platform. We demonstrated that the putative signal sequence from C. reinhardtii gametolysin can assist the secretion of the yellow fluorescent protein Venus into the culture media. In order to increase the secretion yields, Venus was C-terminally fused with synthetic glycomodules comprised of tandem serine (Ser) and proline (Pro) repeats of 10 and 20 units [hereafter (SP)n, wherein n=10 or 20]. The yields of the (SP)n-fused Venus were higher than Venus without the glycomodule by up to 12 folds, with the maximum yield of 15 mg L(-1) . Moreover, the presence of the glycomodules confererred an enhanced proteolytic protein stability. The Venus-(SP)n proteins were shown to be glycosylated, and a treatment of the cells with Brefeldin A led to a suggestion that glycosylation of the (SP)n glycomodules starts in the endoplasmic reticulum (ER). Taken together, the results demonstrate the utility of the gametolysin signal sequence and (SP)n glycomodule to promote a more efficient biomanufacturing of microalgae-based recombinant proteins. This article is protected by copyright. All rights reserved.

  2. Characterization of lead induced metal-phytochelatin complexes in Chlamydomonas reinhardtii.

    Science.gov (United States)

    Scheidegger, Christian; Sigg, Laura; Behra, Renata

    2011-11-01

    Accumulation of Pb and induction of phytochelatin synthesis were observed in Chlamydomonas reinhardtii upon Pb(II) exposure. Our aim was to examine whether Pb(II) is bound by phytochelatins (PCs) in C. reinhardtii and to examine formed complexes for their stoichiometry and composition. Metal-phytochelatin (Me-PC) complexes induced by Pb were isolated by size-exclusion chromatography in 13 collected fractions, which were analyzed for their PC and metal content by high-performance liquid chromatography and inductively coupled plasma mass spectrometry. A recovery of more than 90% of Pb from standard Pb-PC₂ complexes within the total volume of the size-exclusion column indicated the adequacy of the method for Pb-PC(n) complex separation and characterization. Phytochelatins were detected mainly in a molecular weight ranging from 1,000 to 5,300 daltons (Da), indicating the formation of complexes with various stoichiometries. Approximately 72% of total PC₂ eluted in the range from 1,000 to 1,600 Da, and 80% of total PC₃ eluted in the molecular weight range from 1,600 to 2,300 Da. The distribution of Cu, Zn, and Pb showed that more than 70% of these metals were associated with the high-molecular-weight fractions. Copper, zinc, and lead were also observed in PC-containing fractions, suggesting the formation of various Me-PC complexes. The results of the present study indicate that the role of PCs in Pb detoxification is minor, because only 13% of total Pb was associated with PCs.

  3. Saturating Light Induces Sustained Accumulation of Oil in Plastidal Lipid Droplets in Chlamydomonas reinhardtii.

    Science.gov (United States)

    Goold, Hugh Douglas; Cuiné, Stéphan; Légeret, Bertrand; Liang, Yuanxue; Brugière, Sabine; Auroy, Pascaline; Javot, Hélène; Tardif, Marianne; Jones, Brian; Beisson, Fred; Peltier, Gilles; Li-Beisson, Yonghua

    2016-08-01

    Enriching algal biomass in energy density is an important goal in algal biotechnology. Nitrogen (N) starvation is considered the most potent trigger of oil accumulation in microalgae and has been thoroughly investigated. However, N starvation causes the slow down and eventually the arrest of biomass growth. In this study, we show that exposing a Chlamydomonas reinhardtii culture to saturating light (SL) under a nonlimiting CO2 concentration in turbidostatic photobioreactors induces a sustained accumulation of lipid droplets (LDs) without compromising growth, which results in much higher oil productivity than N starvation. We also show that the polar membrane lipid fraction of SL-induced LDs is rich in plastidial lipids (approximately 70%), in contrast to N starvation-induced LDs, which contain approximately 60% lipids of endoplasmic reticulum origin. Proteomic analysis of LDs isolated from SL-exposed cells identified more than 200 proteins, including known proteins of lipid metabolism, as well as 74 proteins uniquely present in SL-induced LDs. LDs induced by SL and N depletion thus differ in protein and lipid contents. Taken together, lipidomic and proteomic data thus show that a large part of the sustained oil accumulation occurring under SL is likely due to the formation of plastidial LDs. We discuss our data in relation to the different metabolic routes used by microalgae to accumulate oil reserves depending on cultivation conditions. Finally, we propose a model in which oil accumulation is governed by an imbalance between photosynthesis and growth, which can be achieved by impairing growth or by boosting photosynthetic carbon fixation, with the latter resulting in higher oil productivity.

  4. Tying Down Loose Ends in the Chlamydomonas Genome: Functional Significance of Abundant Upstream Open Reading Frames

    Directory of Open Access Journals (Sweden)

    Frederick R. Cross

    2016-02-01

    Full Text Available The Chlamydomonas genome has been sequenced, assembled, and annotated to produce a rich resource for genetics and molecular biology in this well-studied model organism. The annotated genome is very rich in open reading frames upstream of the annotated coding sequence (‘uORFs’: almost three quarters of the assigned transcripts have at least one uORF, and frequently more than one. This is problematic with respect to the standard ‘scanning’ model for eukaryotic translation initiation. These uORFs can be grouped into three classes: class 1, initiating in-frame with the coding sequence (CDS (thus providing a potential in-frame N-terminal extension; class 2, initiating in the 5′ untranslated sequences (5UT and terminating out-of-frame in the CDS; and class 3, initiating and terminating within the 5UT. Multiple bioinformatics criteria (including analysis of Kozak consensus sequence agreement and BLASTP comparisons to the closely related Volvox genome, and statistical comparison to cds and to random sequence controls indicate that of ∼4000 class 1 uORFs, approximately half are likely in vivo translation initiation sites. The proposed resulting N-terminal extensions in many cases will sharply alter the predicted biochemical properties of the encoded proteins. These results suggest significant modifications in ∼2000 of the ∼20,000 transcript models with respect to translation initiation and encoded peptides. In contrast, class 2 uORFs may be subject to purifying selection, and the existent ones (surviving selection are likely inefficiently translated. Class 3 uORFs are found in more than half of transcripts, frequently multiple times per transcript; however, they are remarkably similar to random sequence expectations with respect to size, number, and composition, and therefore may in most cases be selectively neutral.

  5. Cyclic AMP functions as a primary sexual signal in gametes of Chlamydomonas reinhardtii.

    Science.gov (United States)

    Pasquale, S M; Goodenough, U W

    1987-11-01

    When Chlamydomonas reinhardtii gametes of opposite mating type are mixed together, they adhere by a flagella-mediated agglutination that triggers three rapid mating responses: flagellar tip activation, cell wall loss, and mating structure activation accompanied by actin polymerization. Here we show that a transient 10-fold elevation of intracellular cAMP levels is also triggered by sexual agglutination. We further show that gametes of a single mating type can be induced to undergo all three mating responses when presented with exogenous dibutyryl-cAMP (db-cAMP). These events are also induced by cyclic nucleotide phosphodiesterase inhibitors, which elevate endogenous cAMP levels and act synergistically with db-cAMP. Non-agglutinating mutants of opposite mating type will fuse efficiently in the presence of db-cAMP. No activation of mating events is induced by calcium plus ionophores, 8-bromo-cGMP, dibutyryl-cGMP, nigericin at alkaline pH, phorbol esters, or forskolin. H-8, an inhibitor of cyclic nucleotide-dependent protein kinase, inhibits mating events in agglutinating cells and antagonizes the effects of cAMP on non-agglutinating cells. Adenylate cyclase activity was detected in both the gamete cell body and flagella, with the highest specific activity displayed in flagellar membrane fractions. The flagellar membrane adenylate cyclase is preferentially stimulated by Mn++, unresponsive to NaF, GTP, GTP gamma S, AlF4-, and forskolin, and is inhibited by trifluoperazine. Cyclic nucleotide phosphodiesterase activity is also present in flagella. Our observations indicate that cAMP is a sufficient initial signal for all of the known mating reaction events in C. reinhardtii, and suggest that the flagellar cyclase and/or phosphodiesterase may be important loci of control for the agglutination-stimulated production of this signal.

  6. Use of microalgae Chlamydomonas reinhardtii for production of double-stranded RNA against shrimp virus

    Directory of Open Access Journals (Sweden)

    Parinyachat Somchai

    2016-05-01

    Full Text Available RNA interference has been proposed to be a promising tool for combating shrimp viruses. Antiviral double-stranded (dsRNA has been mostly produced in Escherichia coli-expression system because of its high efficiency and inexpensive operations. However, overusing the bacteria may raise concerns regarding public health and environmental contamination, and seeking for a new dsRNA production platform would be alternative for future molecular farming. In this study, we exploited the green microalgae Chlamydomonas reinhardtii to produce dsRNA targeting the lethal shrimp yellow head virus (YHV. The expression plasmid pSL18 for C. reinhardtii was constructed to contain YHV-specific hairpin RNA expression cassette, and the successful assembly of pSL18-YHV was confirmed by PCR and enzymatic digestions. Glass bead method was employed for transformation of C. reinhardtii nuclear genome with pSL18-YHV. Microalgal expression of dsRNA-YHV, approximately 45 ng from 100-mL culture, was detected by qRT-PCR. Oral feeding experiment on postlarval shrimp revealed that the formulated feed with C. reinhardtii expressing dsRNA-YHV, at the ratio of 1 × 108 transformants per gram feed, improved 22% survival rate after YHV challenge. The present study suggests that C. reinhardtii can be bioengineered to produce viral-specific dsRNA for shrimp viral disease control, and the developed qRT-PCR could detect microalgal dsRNA with detection limit of subpicogram.

  7. Structure, circadian regulation and bioinformatic analysis of the unique sigma factor gene in Chlamydomonas reinhardtii.

    Science.gov (United States)

    Carter, Matthew L; Smith, Annette C; Kobayashi, Hirokazu; Purton, Saul; Herrin, David L

    2004-01-01

    In higher plants, the transcription of plastid genes is mediated by at least two types of RNA polymerase (RNAP); a plastid-encoded bacterial RNAP in which promoter specificity is conferred by nuclear-encoded sigma factors, and a nuclear-encoded phage-like RNAP. Green algae, however, appear to possess only the bacterial enzyme. Since transcription of much, if not most, of the chloroplast genome in Chlamydomonas reinhardtii is regulated by the circadian clock and the nucleus, we sought to identify sigma factor genes that might be responsible for this regulation. We describe a nuclear gene (RPOD) that is predicted to encode an 80 kDa protein that, in addition to a predicted chloroplast transit peptide at the N-terminus, has the conserved motifs (2.1- 4.2) diagnostic of bacterial sigma-70 factors. We also identified two motifs not previously recognized for sigma factors, adjacent PEST sequences and a leucine zipper, both suggested to be involved in protein-protein interactions. PEST sequences were also found in approximately 40% of sigma factors examined, indicating they may be of general significance. Southern blot hybridization and BLAST searches of the genome and EST databases suggest that RPODmay be the only sigma factor gene in C. reinhardtii. The levels of RPODmRNA increased 2- 3-fold in the mid-to-late dark period of light-dark cycling cells, just prior to, or coincident with, the peak in chloroplast transcription. Also, the dark-period peak in RPOD mRNA persisted in cells shifted to continuous light or continuous dark for at least one cycle, indicating that RPODis under circadian clock control. These results suggest that regulation of RPODexpression contributes to the circadian clock's control of chloroplast transcription.

  8. Age dependent alterations in photosystem II acceptor side in Cucumis sativus cotyledonary leaf thylakoids: analysis of binding characteristics of herbicide [14C]-atrazine.

    Science.gov (United States)

    Prakash, J S; Baig, M A; Mohanty, P

    1999-02-01

    Senescence induced temporal changes in photosystems can be conveniently studied in cotyledonary leaves. We monitored the protein, chlorophyll and electron transport activities in Cucumis sativus cv Poinsette cotyledonary leaves and observed that by 20th day, there was a 50%, 41% and 30-33% decline in the chlorophyll, protein and photosystem II activity respectively when compared to 6th day cotyledonary leaves taken as control. We investigated the changes in photosystem II activity (O2 evolution) as a function of light intensity. The photosystem II functional antenna decreased by 27% and the functional photosystem II units decreased by 30% in 20-day old cotyledonary leaf thylakoids. The herbicide [14C]-atrazine binding assay to monitor specific binding of the herbicide to the acceptor side of photosystem II reaction centre protein, D1, showed an increase in the affinity for atrazine towards D1 protein and decrease in the QB binding sites in 20th day leaf thylakoids when compared to 6th day leaf thylakoids. The western blot analysis also suggested a decrease in steady state levels of D1 protein in 20th day cotyledonary leaf thylakoids as compared to 6th day sample which is in agreement with [14C]-atrazine binding assay and light saturation kinetics.

  9. High-and low-intensity photosystems in Phycomyces phototropism: Effects of mutations in genes madA, madB, and madC.

    Science.gov (United States)

    Palit, A; Galland, P; Lipson, E D

    1989-04-01

    Sporangiophores of Phycomyces blakesleeanus Burgeff that have been grown in darkness and are then suddenly exposed to unilateral light show a two-step bending response rather than a smooth, monotonic response found in light-adapted specimens (Galland and Lipson, 1987, Proc. Natl. Acad. Sci. USA 84, 104-108). The stepwise bending is controlled by two photosystems optimized for the low-and high-intensity ranges. These two photosystems have now been studied in phototropism mutants with defects in genes madA, madB, and madC. All three mutations raise the threshold of the low-intensity (low-fluence) photosystem by about 10(6)-fold and that of the high-intensity (high-fluence) system by about 10(3)-fold. Estimates for the light-adaptation time constants of the low-and high-intensity photosystems show that the mutants are affected in adaptation. In the mutants, the light-adaptation kinetics are only slightly affected in the low-intensity photosystem but, for the high-intensity photosystem, the kinetics are considerably slower than in the wild type.

  10. Bioinformatic Analysis of the Nitrate Reductase Gene in Antartic Ice Algae Chlamydomonas sp. ICE-L%南极衣藻Chlamydomonas sp.ICE-L硝酸还原酶基因的生物信息学分析

    Institute of Scientific and Technical Information of China (English)

    林敏卓; 刘晨临; 黄晓航; 杨平平

    2012-01-01

    Nitrate reductase (NR) plays an important role in the abiotic stress adaptation in plants by regulating nitrogen metabolism. A nitrate reductase (NR) gene of Antarctic ice algae, Chlamydomonas sp. ICE-L, was identified from the cDNA library and sequenced. The encoded protein sequence of NR gene was investigated by bioinformatic analysis. Through sequence alignment the active sites of ICE-L NR protein sequence which may related to stress acclimation was identified. In addition, the tertiary structure of ICE- L NR protein sequence was predicted. The full-length of Chlamydomonas ICE-L NR gene contained an open reading frame of 2,589 bp encoding a nitrate reductase of 863 amino acids. Phylogenetic analysis showed that the gene was homologous to known green algae NRs with identity of 63%, 61%, 60% and 54% to Volvox carteri, Chlamydomonas reinhardtii, Dunaliella tertiolecta and Chlorella vulgaris respectively. The functional prediction analysis revealed that NR gene sequence has 3 different functional domains which was similar to higher plant. This bioinformatic analysis about NR gene of ICE- L will help us further understand and deeply expand the recearch on the acclimatizing mechanism of Antarctic ice alga Chlamydomonas in the extreme environment from the angle of NR gene.%硝酸还原酶(NR)除调节植物的氮代谢外,在植物的各种非生物胁迫的适应过程中也发挥着重要的作用.从南极冰藻Chlamydomonas sp.ICE-L的cDNA文库中筛选到了硝酸还原酶的全长基因,对其进行测序并对其编码的蛋白序列进行了生物信息学分析,构建了NR的系统进化树,通过多序列比对探讨了可能与该酶逆境适应性相关的活性位点,并对该蛋白进行了三级结构预测分析.结果显示,NR基因的编码区长2 589 bp,编码863个氨基酸.在以氨基酸序列构建的系统进化树中,南极衣藻的NR序列和其他绿藻类的聚在一起,与团藻、莱茵衣藻、杜氏盐藻和小球藻

  11. Comparative genomics in Chlamydomonas and Plasmodium identifies an ancient nuclear envelope protein family essential for sexual reproduction in protists, fungi, plants, and vertebrates.

    Science.gov (United States)

    Ning, Jue; Otto, Thomas D; Pfander, Claudia; Schwach, Frank; Brochet, Mathieu; Bushell, Ellen; Goulding, David; Sanders, Mandy; Lefebvre, Paul A; Pei, Jimin; Grishin, Nick V; Vanderlaan, Gary; Billker, Oliver; Snell, William J

    2013-05-15

    Fertilization is a crucial yet poorly characterized event in eukaryotes. Our previous discovery that the broadly conserved protein HAP2 (GCS1) functioned in gamete membrane fusion in the unicellular green alga Chlamydomonas and the malaria pathogen Plasmodium led us to exploit the rare biological phenomenon of isogamy in Chlamydomonas in a comparative transcriptomics strategy to uncover additional conserved sexual reproduction genes. All previously identified Chlamydomonas fertilization-essential genes fell into related clusters based on their expression patterns. Out of several conserved genes in a minus gamete cluster, we focused on Cre06.g280600, an ortholog of the fertilization-related Arabidopsis GEX1. Gene disruption, cell biological, and immunolocalization studies show that CrGEX1 functions in nuclear fusion in Chlamydomonas. Moreover, CrGEX1 and its Plasmodium ortholog, PBANKA_113980, are essential for production of viable meiotic progeny in both organisms and thus for mosquito transmission of malaria. Remarkably, we discovered that the genes are members of a large, previously unrecognized family whose first-characterized member, KAR5, is essential for nuclear fusion during yeast sexual reproduction. Our comparative transcriptomics approach provides a new resource for studying sexual development and demonstrates that exploiting the data can lead to the discovery of novel biology that is conserved across distant taxa.

  12. Efficient expression of nuclear transgenes in the green alga Chlamydomonas: synthesis of an HIV antigen and development of a new selectable marker.

    Science.gov (United States)

    Barahimipour, Rouhollah; Neupert, Juliane; Bock, Ralph

    2016-03-01

    The unicellular green alga Chlamydomonas reinhardtii has become an invaluable model system in plant biology. There is also considerable interest in developing this microalga into an efficient production platform for biofuels, pharmaceuticals, green chemicals and industrial enzymes. However, the production of foreign proteins in the nucleocytosolic compartment of Chlamydomonas is greatly hampered by the inefficiency of transgene expression from the nuclear genome. We have recently addressed this limitation by isolating mutant algal strains that permit high-level transgene expression and by determining the contributions of GC content and codon usage to gene expression efficiency. Here we have applied these new tools and explored the potential of Chlamydomonas to produce a recombinant biopharmaceutical, the HIV antigen P24. We show that a codon-optimized P24 gene variant introduced into our algal expression strains give rise to recombinant protein accumulation levels of up to 0.25% of the total cellular protein. Moreover, in combination with an expression strain, a resynthesized nptII gene becomes a highly efficient selectable marker gene that facilitates the selection of transgenic algal clones at high frequency. By establishing simple principles of successful transgene expression, our data open up new possibilities for biotechnological research in Chlamydomonas.

  13. Acetate versus sulfur deprivation role in creating anaerobiosis in light for hydrogen production by Chlamydomonas reinhardtii and Spirulina platensis: two different organisms and two different mechanisms.

    Science.gov (United States)

    Morsy, Fatthy Mohamed

    2011-01-01

    This work was devoted to separate acetate role in creating anaerobiosis from that of sulfur deprivation. Chlamydomonas reinhardtii grown in TAP (Tris-acetate-phosphate) medium was resuspended in sulfur-replete or -deprived medium in sealed or nonsealed cultures. Sulfur deprivation was substantial for starch accumulation and hydrogen evolution; however, acetate induced anaerobiosis in the presence or absence of sulfur in only sealed cultures. In nonsealed cultures, Chlamydomonas did not lose its photosynthetic activity; however, it was arrested in anoxia with no photosynthetic activity as long as the culture was sealed. The sealed cultures resumed photosynthesis upon unsealing overnight unless the cells died by anoxia at late stage of the experiment. These results indicate that the enhanced oxygen consumption for the enormous acetate respiration and inhibition of the external oxygen supply in sealed cultures of Chlamydomonas are the main reasons for the steady anaerobic conditions. Although acetate was substantial for creating anaerobiosis in Chlamydomonas, sulfur deprivation alone could create anaerobiosis in Spirulina platensis grown autotrophically. Hydrogen evolution and glycogen accumulation were induced under such conditions. Severely reduced phycocyanin, chlorophyll and photosynthesis, while respiration had increased, induced anaerobiosis in Spirulina. This study reports for the first time anaerobiosis under autotrophic conditions in a cyanobacterium.

  14. Overexpressing Ferredoxins in Chlamydomonas reinhardtii Increase Starch and Oil Yields and Enhance Electric Power Production in a Photo Microbial Fuel Cell

    Directory of Open Access Journals (Sweden)

    Li-Fen Huang

    2015-08-01

    Full Text Available Ferredoxins (FDX are final electron carrier proteins in the plant photosynthetic pathway, and function as major electron donors in diverse redox-driven metabolic pathways. We previously showed that overexpression of a major constitutively expressed ferredoxin gene PETF in Chlamydomonas decreased the reactive oxygen species (ROS level and enhanced tolerance to heat stress. In addition to PETF, an endogenous anaerobic induced FDX5 was overexpressed in transgenic Chlamydomonas lines here to address the possible functions of FDX5. All the independent FDX transgenic lines showed decreased cellular ROS levels and enhanced tolerance to heat and salt stresses. The transgenic Chlamydomonas lines accumulated more starch than the wild-type line and this effect increased almost three-fold in conditions of nitrogen depletion. Furthermore, the lipid content was higher in the transgenic lines than in the wild-type line, both with and without nitrogen depletion. Two FDX-overexpressing Chlamydomonas lines were assessed in a photo microbial fuel cell (PMFC; power density production by the transgenic lines was higher than that of the wild-type cells. These findings suggest that overexpression of either PETF or FDX5 can confer tolerance against heat and salt stresses, increase starch and oil production, and raise electric power density in a PMFC.

  15. Taxonomic revision of Chlamydomonas subg. Amphichloris (Volvocales, Chlorophyceae), with resurrection of the genus Dangeardinia and descriptions of Ixipapillifera gen. nov. and Rhysamphichloris gen. nov.

    Science.gov (United States)

    Nakada, Takashi; Tomita, Masaru; Wu, Jiunn-Tzong; Nozaki, Hisayoshi

    2016-04-01

    Chlamydomonas (Cd.) is one of the largest but most polyphyletic genera of freshwater unicellular green algae. It consists of 400-600 morphological species and requires taxonomic revision. Toward reclassification, each morphologically defined classical subgenus (or subgroup) should be examined using culture strains. Chlamydomonas subg. Amphichloris is characterized by a central nucleus between two axial pyrenoids, however, the phylogenetic structure of this subgenus has yet to be examined using molecular data. Here, we examined 12 strains including six newly isolated strains, morphologically identified as Chlamydomonas subg. Amphichloris, using 18S rRNA gene phylogeny, light microscopy, and mitochondria fluorescent microscopy. Molecular phylogenetic analyses revealed three independent lineages of the subgenus, separated from the type species of Chlamydomonas, Cd. reinhardtii. These three lineages were further distinguished from each other by light and fluorescent microscopy-in particular by the morphology of the papillae, chloroplast surface, stigmata, and mitochondria-and are here assigned to three genera: Dangeardinia emend., Ixipapillifera gen. nov., and Rhysamphichloris gen. nov. Based on the molecular and morphological data, two to three species were recognized in each genus, including one new species, I. pauromitos. In addition, Cd. deasonii, which was previously assigned to subgroup "Pleiochloris," was included in the genus Ixipapillifera as I. deasonii comb. nov.

  16. Productivity and selective accumulation of carotenoids of the novel extremophile microalga Chlamydomonas acidophila grown with different carbon sources in batch systems

    NARCIS (Netherlands)

    Cuaresma, M.; Casal, C.; Forján, E.; Vílchez, C.

    2011-01-01

    Cultivation of extremophile microorganisms has attracted interest due to their ability to accumulate highvalue compounds. Chlamydomonas acidophila is an acidophile green microalga isolated by our group from Tinto River, an acidic river that flows down from the mining area in Huelva, Spain. This micr

  17. Physico-chemical property of rare earths-effects on the energy regulation of photosystem II in Arabidopsis thaliana.

    Science.gov (United States)

    Xiaoqing, Liu; Hao, Huang; Chao, Liu; Min, Zhou; Fashui, Hong

    2009-08-01

    Photosystem II (PSII) from Arabidopsis thaliana treated by lanthanum (La(3+)), cerium (Ce(3+)), and neodymium (Nd(3+)) were isolated to investigate the effects of 4f electron characteristics and alternation valence of rare earth elements (REEs) on PSII function regulation comparatively. Results showed that REE treatment could induce the generous expression of LhcII b in A. thaliana and increase the content of light-harvesting complex II and its trimer on the thylakoid membrane significantly. Meanwhile, the light absorption in the red and blue region and fluorescence quantum yield near 683 nm were obviously increased; oxygen evolution rate was greatly improved too, suggesting that REEs could enhance the efficiency of light absorption, regulate excitation energy distribution from photosystem I (PSI) to PSII, and thus increase the activity of photochemical reaction and oxygen evolution accordingly. The efficiency order of the four treatments was Ce(3+) > Nd(3+) > La(3+) > control.

  18. Systems Biology Approach in Chlamydomonas Reveals Connections between Copper Nutrition and Multiple Metabolic Steps[C][W][OA

    Science.gov (United States)

    Castruita, Madeli; Casero, David; Karpowicz, Steven J.; Kropat, Janette; Vieler, Astrid; Hsieh, Scott I.; Yan, Weihong; Cokus, Shawn; Loo, Joseph A.; Benning, Christoph; Pellegrini, Matteo; Merchant, Sabeeha S.

    2011-01-01

    In this work, we query the Chlamydomonas reinhardtii copper regulon at a whole-genome level. Our RNA-Seq data simulation and analysis pipeline validated a 2-fold cutoff and 10 RPKM (reads per kilobase of mappable length per million mapped reads) (~1 mRNA per cell) to reveal 63 CRR1 targets plus another 86 copper-responsive genes. Proteomic and immunoblot analyses captured 25% of the corresponding proteins, whose abundance was also dependent on copper nutrition, validating transcriptional regulation as a major control mechanism for copper signaling in Chlamydomonas. The impact of copper deficiency on the expression of several O2-dependent enzymes included steps in lipid modification pathways. Quantitative lipid profiles indicated increased polyunsaturation of fatty acids on thylakoid membrane digalactosyldiglycerides, indicating a global impact of copper deficiency on the photosynthetic apparatus. Discovery of a putative plastid copper chaperone and a membrane protease in the thylakoid suggest a mechanism for blocking copper utilization in the chloroplast. We also found an example of copper sparing in the N assimilation pathway: the replacement of copper amine oxidase by a flavin-dependent backup enzyme. Forty percent of the targets are previously uncharacterized proteins, indicating considerable potential for new discovery in the biology of copper. PMID:21498682

  19. Construction of modular tandem expression vectors for the green alga Chlamydomonas reinhardtii using the Cre/lox-system.

    Science.gov (United States)

    Heitzer, Markus; Zschoernig, Barbara

    2007-09-01

    The successful expression of foreign genes mainly depends on both a reliable method for transformation and a suitable promoter sequence. We created a series of modular plasmids that facilitate the rapid construction of large tandem vectors for transgene expression under the control of different promoter sequences in Chlamydomonas reinhardtii. Tandem vectors carrying expression cassettes for Renilla luciferase and a metabolic selection marker (ARG7) were manufactured by fusing two plasmids in vitro using Cre/lox site-specific recombination. Supercoiled and linear plasmids were used to transform an arginine auxotrophic Chlamydomonas strain, and rates of co-expression as well as levels of luciferase activity were monitored for frequently used promoters (HSP70A, LHCB1, PSAD, and the chimeric HSP70A/RBCS2). Linearized tandem vectors generally increased the co-expression frequency (up to 77%) compared with standard cotransformation protocols. Most transformants showed a single and complete integration event confirming the close linkage of active selectable marker and reporter gene within the nuclear genome. The analysis of luciferase activity showed expression levels within three orders of magnitude for the promoters used, with the artificial HSP70A/RRBCS2 being the most active. For 69% of all luminescent transformants carrying the HSP70A promoter luciferase expression was enhanced by heatshock, indicating physiological promoter function in a transgenic context.

  20. Effect of Cd on GSH and GSH-related enzymes of Chlamydomonas sp. ICE-L existing in Antarctic ice

    Institute of Scientific and Technical Information of China (English)

    DING Yu; MIAO Jin-lai; LI Guang-you; WANG Quan-fu; KAN Guang-feng; WANG Guo-dong

    2005-01-01

    Glutathione(GSH) and GSH-related enzymes play a great role in protecting organisms from oxidative damage. The GSH level and GSH-related enzymes activities were investigated as well as the growth yield and malonyldialdehyde(MDA) content in the Antarctic ice microalga Chlamydomonas sp. ICE-L exposure to the different cadmium concentration in this paper. The results showed that the higher concentration Cd inhibited the growth of ICE-L significantly and Cd would induce formation of MDA. At the same time, it is clear that GSH level, glutathione peroxidases(GPx) activity and glutathione S-transferases(GST), activity were higher in ICE-L exposed to Cd than the control. But GR activity dropped notably when ICE-L were cultured in the medium containing Cd. Increase of GSH level, GPx and GST activities acclimate to oxidative stress induced by Cd and protect Antarctic ice microalga Chlamydomonas sp. ICE-L from toxicity caused by Cd exposure. These parameters may be used to assess the biological impact of Cd in the Antarctic pole region environment.

  1. Ecophysiology, secondary pigments and ultrastructure of Chlainomonas sp. (Chlorophyta) from the European Alps compared with Chlamydomonas nivalis forming red snow.

    Science.gov (United States)

    Remias, Daniel; Pichrtová, Martina; Pangratz, Marion; Lütz, Cornelius; Holzinger, Andreas

    2016-04-01

    Red snow is a well-known phenomenon caused by microalgae thriving in alpine and polar regions during the melting season. The ecology and biodiversity of these organisms, which are adapted to low temperatures, high irradiance and freeze-thaw events, are still poorly understood. We compared two different snow habitats containing two different green algal genera in the European Alps, namely algae blooming in seasonal rock-based snowfields (Chlamydomonas nivalis) and algae dominating waterlogged snow bedded over ice (Chlainomonassp.). Despite the morphological similarity of the red spores found at the snow surface, we found differences in intracellular organization investigated by light and transmission electron microscopy and in secondary pigments investigated by chromatographic analysis in combination with mass spectrometry. Spores ofChlainomonassp. show clear differences fromChlamydomonas nivalisin cell wall arrangement and plastid organization. Active photosynthesis at ambient temperatures indicates a high physiological activity, despite no cell division being present. Lipid bodies containing the carotenoid astaxanthin, which produces the red color, dominate cells of both species, but are modified differently. While inChlainomonassp. astaxanthin is mainly esterified with two fatty acids and is more apolar, inChamydomonas nivalis, in contrast, less apolar monoesters prevail.

  2. Interaction of Herbicides and Quinone with the QB-Protein of the Diuron-Resistant Chlamydomonas reinhardtii Mutant Dr2

    Science.gov (United States)

    Haworth, Philip; Steinback, Katherine E.

    1987-01-01

    We have used the diuron-resistant Dr2 mutant of Chlamydomonas reinhardtii which is altered in the 32 kilodalton QB-protein at amino acid 219 (valine to isoleucine), to investigate the interactions of herbicides and plastoquinone with the 32 kilodalton QB-protein. The data contained in this report demonstrate that the effects of this mutation are different from those of the more completely characterized mutant which confers extreme resistance to triazines in higher plants. The mutation in C. reinhardtii Dr2 confers only slight resistance to a number of inhibitors of photosynthetic electron transport. Extreme triazine resistance results from an increase in the binding constant of the herbicide with the 32 kilodalton QB-protein, in contrast the diuron binding constant for chloroplasts isolated from wild-type (sensitive) Chlamydomonas and the resistant Dr2 are indistinguishable. We conclude that the altered structure in the 32 kilodalton QB-protein of Dr2 does not directly affect the diuron binding site. This mutation appears to alter the steric properties of the binding protein in such a way that diuron and plastoquinone do not directly compete for binding. This steric perturbation confers mild resistance to other herbicidal inhibitors of photosynthesis and alters the kinetics of QA to QB electron transfer. PMID:16665318

  3. Synergism between Inositol Polyphosphates and TOR Kinase Signaling in Nutrient Sensing, Growth Control, and Lipid Metabolism in Chlamydomonas[OPEN

    Science.gov (United States)

    Evans, Bradley S.; Li, Jia; Liu, Yu; Diamond, Spencer

    2016-01-01

    The networks that govern carbon metabolism and control intracellular carbon partitioning in photosynthetic cells are poorly understood. Target of Rapamycin (TOR) kinase is a conserved growth regulator that integrates nutrient signals and modulates cell growth in eukaryotes, though the TOR signaling pathway in plants and algae has yet to be completely elucidated. We screened the unicellular green alga Chlamydomonas reinhardtii using insertional mutagenesis to find mutants that conferred hypersensitivity to the TOR inhibitor rapamycin. We characterized one mutant, vip1-1, that is predicted to encode a conserved inositol hexakisphosphate kinase from the VIP family that pyrophosphorylates phytic acid (InsP6) to produce the low abundance signaling molecules InsP7 and InsP8. Unexpectedly, the rapamycin hypersensitive growth arrest of vip1-1 cells was dependent on the presence of external acetate, which normally has a growth-stimulatory effect on Chlamydomonas. vip1-1 mutants also constitutively overaccumulated triacylglycerols (TAGs) in a manner that was synergistic with other TAG inducing stimuli such as starvation. vip1-1 cells had reduced InsP7 and InsP8, both of which are dynamically modulated in wild-type cells by TOR kinase activity and the presence of acetate. Our data uncover an interaction between the TOR kinase and inositol polyphosphate signaling systems that we propose governs carbon metabolism and intracellular pathways that lead to storage lipid accumulation. PMID:27600537

  4. Changes of proteins in the Antarctic ice microalga Chlamydomonas sp. cultured under UV-B radiation stress

    Institute of Scientific and Technical Information of China (English)

    KAN Guangfeng; MIAO Jinlai; SHI Cuijuan; LI Guangyou

    2006-01-01

    Antarctic ice microalga Chlamydomonas sp. can thrive undisturbed under high UV radiation in the Antarctic ice layer. However, it is unknown that the initial adaptation mechanisms in protein level occurring in response to high UV radiation. Global-expression profiling of proteins in response to stress was analyzed by two-dimensional electrophoresis (2-DE) and image analysis. In the 2-DE analysis,protein preparation is the key step. Three different protein extract methods were compared, and the results showed that the trichloroacetic acid (TCA)-acetone fractional precipitation method was the fittest one. At the same time, the proteins in Chlamydomonas sp. were compared in 2-DE way, and the synthesis of seven protein spots was found disappeared and 18 decreased after exposed to UV-B radiation. In addition, 14 protein spots were enhanced or induced, among which two new peptides (20 and 21 kDa) appeared whose isoelectric point (pI) was 7.05 and 4.60 respectively. These changed proteins might act as key role in the acclimation of Antarctic ice microalga to UV-B radiation

  5. Using single cell cultivation system for on-chip monitoring of the interdivision timer in Chlamydomonas reinhardtii cell cycle

    Directory of Open Access Journals (Sweden)

    Soloviev Mikhail

    2010-09-01

    Full Text Available Abstract Regulation of cell cycle progression in changing environments is vital for cell survival and maintenance, and different regulation mechanisms based on cell size and cell cycle time have been proposed. To determine the mechanism of cell cycle regulation in the unicellular green algae Chlamydomonas reinhardtii, we developed an on-chip single-cell cultivation system that allows for the strict control of the extracellular environment. We divided the Chlamydomonas cell cycle into interdivision and division phases on the basis of changes in cell size and found that, regardless of the amount of photosynthetically active radiation (PAR and the extent of illumination, the length of the interdivision phase was inversely proportional to the rate of increase of cell volume. Their product remains constant indicating the existence of an 'interdivision timer'. The length of the division phase, in contrast, remained nearly constant. Cells cultivated under light-dark-light conditions did not divide unless they had grown to twice their initial volume during the first light period. This indicates the existence of a 'commitment sizer'. The ratio of the cell volume at the beginning of the division phase to the initial cell volume determined the number of daughter cells, indicating the existence of a 'mitotic sizer'.

  6. Photoinduced hydrogen evolution in an artificial system containing photosystem I,hydrogenase,methyl viologen and mercaptoacetic acid

    Institute of Scientific and Technical Information of China (English)

    Dong Jin Qian; Ai Rong Liu; Chikashi Nakamura; Stephan Olav Wenk; Jun Miyake

    2008-01-01

    Hydrogen evolution was detected in an artificial system composed of light-harvesting unit of purified photosystem Ⅰ, catalyst ofhydrogenase, methyl viologen and electron donor under radiation. Absorption spectral features confirmed that electron transferfrom electron donors to proton was via a photoinduced reductive process of methyl viologen.2008 Dong Jin Qian. Published by Elsevier B.V. on behalf of Chinese Chemical Society. All rights reserved.

  7. EPR characterisation of the triplet state in photosystem II reaction centers with singly reduced primary acceptor Q(A).

    Science.gov (United States)

    Feikema, W Onno; Gast, Peter; Klenina, Irina B; Proskuryakov, Ivan I

    2005-09-05

    The triplet states of photosystem II core particles from spinach were studied using time-resolved cw EPR technique at different reduction states of the iron--quinone complex of the reaction center primary electron acceptor. With doubly reduced primary acceptor, the well-known photosystem II triplet state characterised by zero-field splitting parameters |D|=0.0286 cm(-1), |E|=0.0044 cm(-1) was detected. When the primary acceptor was singly reduced either chemically or photochemically, a triplet state of a different spectral shape was observed, bearing the same D and E values and characteristic spin polarization pattern arising from RC radical pair recombination. The latter triplet state was strongly temperature dependent disappearing at T=100 K, and had a much faster decay than the former one. Based on its properties, this triplet state was also ascribed to the photosystem II reaction center. A sequence of electron-transfer events in the reaction centers is proposed that explains the dependence of the triplet state properties on the reduction state of the iron--quinone primary acceptor complex.

  8. Primary light harvesting system: the relationship of phycobilisomes to photosystem I and II. Progress report, September 1983-March 1984

    Energy Technology Data Exchange (ETDEWEB)

    Gantt, E.

    1984-01-01

    It has long been known that energy absorbed by phycobiliproteins is transferred to photosystem II and is then distributed to I. Consequently, it has been predicted that phycobilisomes would be in close association with photosystem II. Our laboratory has succeeded in isolating phycobilisomes which have photosystem II activity, as measured by oxygen evolution, and reduction of dichlorophenol indophenol. In a special medium (0.5 M phosphate buffer, 0.5 M sucrose, 0.3 M citrate, and 10 mM magnesium chloride) the phycobilisomes remain intact, and the PS II activity is preserved. These preparations are deficient in the PS I reaction center pigment P700, as well as in the PS I polypeptide CP 1. Preparations of this system are being further characterized to determine the P680 content, to ascertain the DCMU-binding protein, cytochromes, and other polypeptides which are specific for PS II, and also for PS I. In addition, the sidedness of thylakoid membranes of red algae and cyanobacteria are also being explored by immunocytochemistry (EM) and fractionation on acrylamide gels. Attachment of the phycobilisomes, and the location of the terminal emitter of phycobilisomes, are an integral part of these studies. Results from these studies will aid in our understanding of the energy transfer in the photosynthetic apparatus of these groups of organisms which appear to have a considerable convergence from green plants.

  9. Photobiological hydrogen production with the unicellular green alga Chlamydomonas reinhardtii under process engineering aspects; Photobiologische Wasserstoffproduktion mit der einzelligen Gruenalge Chlamydomonas reinhardtii unter verfahrenstechnischen Aspekten

    Energy Technology Data Exchange (ETDEWEB)

    Geier, Stephanie

    2011-07-01

    Hydrogen is of high interest as a clean and environmentally friendly energy source as its combustion only emits water and energy. However, currently hydrogen is produced in energy demanding processes by the consumption of fossil fuels. An alternative way of sustainable and non-polluting hydrogen production could be provided by use of photosynthetic active microalgae. Within this work, the photobiological hydrogen production with the unicellular green algae Chlamydomonas reinhardtii is investigated under the aspects of bioprocess-engineering and economics. Objectives are, besides the increase of the photochemical efficiency, the cultivation of the algae and subsequent hydrogen production under cost-free sunlight. It could be demonstrated that outdoor cultivation of C. reinhardtii is possible in Central Europe throughout the year by using e.g. waste heat. Similar cell numbers in the range from 1,2.10{sup 7} cells ml{sup -1} to 1,7.10{sup 7} cells ml{sup -1} could be achieved in closed photobioreactors of the type Photobioreactor Screening Module under controlled laboratory conditions and both continuous illumination (200 {mu}mol.m{sup -2}.s{sup -1}) and simulated outdoor conditions according to the light intensity of idealized summer day as well as in outdoor experiments (up to 2000 {mu}mol.m{sup -2}.s{sup -1}).The use of 10 % CO{sub 2} corresponding to the CO{sub 2} content in flue gas led to a doubling of cell numbers under continuous illumination to 4,2.10{sup 7} cells ml{sup -1}, compared to the reference culture bubbled with 3 % CO{sub 2}. A significant increase of cell numbers under the light profiles of an idealized summer day could not be achieved. The cultivation under the light profile of a winter day at 25 C reduced cell growth to 54 %, compared to the summer simulation. In open 30 L outdoor ponds, only 0,26.10{sup 7} cells ml{sup -1} could be achieved under photoheterotrophic conditions during the summer months, which corresponds to 20 % of the cell

  10. Photobiological hydrogen production with the unicellular green alga Chlamydomonas reinhardtii under process engineering aspects; Photobiologische Wasserstoffproduktion mit der einzelligen Gruenalge Chlamydomonas reinhardtii unter verfahrenstechnischen Aspekten

    Energy Technology Data Exchange (ETDEWEB)

    Geier, Stephanie

    2011-07-01

    Hydrogen is of high interest as a clean and environmentally friendly energy source as its combustion only emits water and energy. However, currently hydrogen is produced in energy demanding processes by the consumption of fossil fuels. An alternative way of sustainable and non-polluting hydrogen production could be provided by use of photosynthetic active microalgae. Within this work, the photobiological hydrogen production with the unicellular green algae Chlamydomonas reinhardtii is investigated under the aspects of bioprocess-engineering and economics. Objectives are, besides the increase of the photochemical efficiency, the cultivation of the algae and subsequent hydrogen production under cost-free sunlight. It could be demonstrated that outdoor cultivation of C. reinhardtii is possible in Central Europe throughout the year by using e.g. waste heat. Similar cell numbers in the range from 1,2.10{sup 7} cells ml{sup -1} to 1,7.10{sup 7} cells ml{sup -1} could be achieved in closed photobioreactors of the type Photobioreactor Screening Module under controlled laboratory conditions and both continuous illumination (200 {mu}mol.m{sup -2}.s{sup -1}) and simulated outdoor conditions according to the light intensity of idealized summer day as well as in outdoor experiments (up to 2000 {mu}mol.m{sup -2}.s{sup -1}).The use of 10 % CO{sub 2} corresponding to the CO{sub 2} content in flue gas led to a doubling of cell numbers under continuous illumination to 4,2.10{sup 7} cells ml{sup -1}, compared to the reference culture bubbled with 3 % CO{sub 2}. A significant increase of cell numbers under the light profiles of an idealized summer day could not be achieved. The cultivation under the light profile of a winter day at 25 C reduced cell growth to 54 %, compared to the summer simulation. In open 30 L outdoor ponds, only 0,26.10{sup 7} cells ml{sup -1} could be achieved under photoheterotrophic conditions during the summer months, which corresponds to 20 % of the cell

  11. Prevalence and evolution of core photosystem II genes in marine cyanobacterial viruses and their hosts.

    Directory of Open Access Journals (Sweden)

    Matthew B Sullivan

    2006-07-01

    Full Text Available Cyanophages (cyanobacterial viruses are important agents of horizontal gene transfer among marine cyanobacteria, the numerically dominant photosynthetic organisms in the oceans. Some cyanophage genomes carry and express host-like photosynthesis genes, presumably to augment the host photosynthetic machinery during infection. To study the prevalence and evolutionary dynamics of this phenomenon, 33 cultured cyanophages of known family and host range and viral DNA from field samples were screened for the presence of two core photosystem reaction center genes, psbA and psbD. Combining this expanded dataset with published data for nine other cyanophages, we found that 88% of the phage genomes contain psbA, and 50% contain both psbA and psbD. The psbA gene was found in all myoviruses and Prochlorococcus podoviruses, but could not be amplified from Prochlorococcus siphoviruses or Synechococcus podoviruses. Nearly all of the phages that encoded both psbA and psbD had broad host ranges. We speculate that the presence or absence of psbA in a phage genome may be determined by the length of the latent period of infection. Whether it also carries psbD may reflect constraints on coupling of viral- and host-encoded PsbA-PsbD in the photosynthetic reaction center across divergent hosts. Phylogenetic clustering patterns of these genes from cultured phages suggest that whole genes have been transferred from host to phage in a discrete number of events over the course of evolution (four for psbA, and two for psbD, followed by horizontal and vertical transfer between cyanophages. Clustering patterns of psbA and psbD from Synechococcus cells were inconsistent with other molecular phylogenetic markers, suggesting genetic exchanges involving Synechococcus lineages. Signatures of intragenic recombination, detected within the cyanophage gene pool as well as between hosts and phages in both directions, support this hypothesis. The analysis of cyanophage psbA and psb

  12. Effects of ethylene on photosystem II and antioxidant enzyme activity in Bermuda grass under low temperature.

    Science.gov (United States)

    Hu, Zhengrong; Fan, Jibiao; Chen, Ke; Amombo, Erick; Chen, Liang; Fu, Jinmin

    2016-04-01

    The phytohormone ethylene has been reported to mediate plant response to cold stress. However, it is still debated whether the effect of ethylene on plant response to cold stress is negative or positive. The objective of the present study was to explore the role of ethylene in the cold resistance of Bermuda grass (Cynodon dactylon (L).Pers.). Under control (warm) condition, there was no obvious effect of the ethylene precursor 1-aminocyclopropane-1-carboxylic acid (ACC) or the antagonist Ag(+) of ethylene signaling on electrolyte leakage (EL) and malondialdehyde (MDA) content. Under cold stress conditions, ACC-treated plant leaves had a greater level of EL and MDA than the untreated leaves. However, the EL and MDA values were lower in the Ag(+) regime versus the untreated. In addition, after 3 days of cold treatment, ACC remarkably reduced the content of soluble protein and also altered antioxidant enzyme activity. Under control (warm) condition, there was no significant effect of ACC on the performance of photosystem II (PS II) as monitored by chlorophyll α fluorescence transients. However, under cold stress, ACC inhibited the performance of PS II. Under cold condition, ACC remarkably reduced the performance index for energy conservation from excitation to the reduction of intersystem electron acceptors (PI(ABS)), the maximum quantum yield of primary photochemistry (φP0), the quantum yield of electron transport flux from Q(A) to Q(B) (φE0), and the efficiency/probability of electron transport (ΨE0). Simultaneously, ACC increased the values of specific energy fluxes for absorption (ABS/RC) and dissipation (DI0/RC) after 3 days of cold treatment. Additionally, under cold condition, exogenous ACC altered the expressions of several related genes implicated in the induction of cold tolerance (LEA, SOD, POD-1 and CBF1, EIN3-1, and EIN3-2). The present study thus suggests that ethylene affects the cold tolerance of Bermuda grass by impacting the antioxidant system

  13. Modulating the Redox Potential of the Stable Electron Acceptor, QB, in Mutagenized Photosystem II Reaction Centers.

    Energy Technology Data Exchange (ETDEWEB)

    Perrine, Zoee [The Ohio State Univ., Columbus, OH (United States); The Donald Danforth Plant Science Center, St. Louis, MO (United States); Sayre, Richard [The Donald Danforth Plant Science Center, St. Louis, MO (United States)

    2011-02-10

    One of the unique features of electron transfer processes in photosystem II (PSII) reaction centers (RC) is the exclusive transfer of electrons down only one of the two parallel cofactor branches. In contrast to the RC core polypeptides (psaA and psaB) of photosystem I (PSI), where electron transfer occurs down both parallel redox-active cofactor branches, there is greater protein-cofactor asymmetry between the PSII RC core polypeptides (D1 and D2). We have focused on the identification of protein-cofactor relationships that determine the branch along which primary charge separation occurs (P680+/pheophytin-(Pheo)). We have previously shown that mutagenesis of the strong hydrogen-bonding residue, D1-E130, to less polar residues (D1-E130Q,H,L) shifted the midpoint potential of the PheoD1/PheoD1- couple to more negative values, reducing the quantum yield of primary charge separation. We did not observe, however, electron transfer down the inactive branch in D1-E130 mutants. The protein residue corresponding to D1-E130 on the inactive branch is D2-Q129 which presumably has a reduced hydrogen-bonding interaction with PheoD2 relative to the D1-E130 residue with PheoD1. Analysis of the recent 2.9 Å cyanobacterial PSII crystal structure indicated, however, that the D2-Q129 residue was too distant from the PheoD2 headgroup to serve as a possible hydrogen bond donor and directly impact its midpoint potential as well as potentially determine the directionality of electron transfer. Our objective was to characterize the function of this highly conserved inactive branch residue by replacing it with a nonconservative leucine or a conservative histidine residue. Measurements of Chl fluorescence decay kinetics and thermoluminescence studies indicate that the mutagenesis of D2-Q129 decreases the redox gap between QA and QB due to a lowering of the redox potential of QB. The

  14. Semiquinone-iron complex of photosystem II: EPR signals assigned to the low-field edge of the ground state doublet of QA•-Fe2+ and QB•-Fe2+.

    Science.gov (United States)

    Sedoud, Arezki; Cox, Nicholas; Sugiura, Miwa; Lubitz, Wolfgang; Boussac, Alain; Rutherford, A William

    2011-07-12

    The quinone-iron complex of the electron acceptor complex of Photosystem II was studied by EPR spectroscopy in Thermosynechococcus elongatus. New g ∼ 2 features belonging to the EPR signal of the semiquinone forms of the primary and secondary quinone, i.e., Q(A)(•-)Fe(2+) and Q(B)(•-)Fe(2+), respectively, are reported. In previous studies, these signals were missed because they were obscured by the EPR signal arising from the stable tyrosyl radical, TyrD(•). When the TyrD(•) signal was removed, either by chemical reduction or by the use of a mutant lacking TyrD, the new signals dominated the spectrum. For Q(A)(•-)Fe(2+), the signal was formed by illumination at 77 K or by sodium dithionite reduction in the dark. For Q(B)(•-)Fe(2+), the signal showed the characteristic period-of-two variations in its intensity when generated by a series of laser flashes. The new features showed relaxation characteristics comparable to those of the well-known features of the semiquinone-iron complexes and showed a temperature dependence consistent with an assignment to the low-field edge of the ground state doublet of the spin system. Spectral simulations are consistent with this assignment and with the current model of the spin system. The signal was also present in Q(B)(•-)Fe(2+) in plant Photosystem II, but in plants, the signal was not detected in the Q(A)(•-)Fe(2+) state.

  15. Generation of a phage-display library of single-domain camelid VH H antibodies directed against Chlamydomonas reinhardtii antigens, and characterization of VH Hs binding cell-surface antigens.

    Science.gov (United States)

    Jiang, Wenzhi; Rosenberg, Julian N; Wauchope, Akelia D; Tremblay, Jacqueline M; Shoemaker, Charles B; Weeks, Donald P; Oyler, George A

    2013-11-01

    Single-domain antibodies (sdAbs) are powerful tools for the detection, quantification, purification and subcellular localization of proteins of interest in biological research. We have generated camelid (Lama pacos) heavy chain-only variable VH domain (VH H) libraries against antigens in total cell lysates from Chlamydomonas reinhardtii. The sdAbs in the sera from immunized animals and VH H antibody domains isolated from the library show specificity to C. reinhardtii and lack of reactivity to antigens from four other algae: Chlorella variabilis, Coccomyxa subellipsoidea, Nannochloropsis oceanica and Thalassiosira pseudonana. Antibodies were produced against a diverse representation of antigens as evidenced by sera ELISA and protein-blot analyses. A phage-display library consisting of the VH H region contained at least 10(6) individual transformants, and thus should represent a wide range of C. reinhardtii antigens. The utility of the phage library was demonstrated by using live C. reinhardtii cells to pan for VH H clones with specific recognition of cell-surface epitopes. The lead candidate VH H clones (designated B11 and H10) bound to C. reinhardtii with EC50 values ≤ 0.5 nm. Treatment of cells with VH H B11 fused to the mCherry or green fluorescent proteins allowed brilliant and specific staining of the C. reinhardtii cell wall and analysis of cell-wall genesis during cell division. Such high-complexity VH H antibody libraries for algae will be valuable tools for algal researchers and biotechnologists.

  16. Conceptualising the lack of health insurance coverage.

    Science.gov (United States)

    Davis, J B

    2000-01-01

    This paper examines the lack of health insurance coverage in the US as a public policy issue. It first compares the problem of health insurance coverage to the problem of unemployment to show that in terms of the numbers of individuals affected lack of health insurance is a problem comparable in importance to the problem of unemployment. Secondly, the paper discusses the methodology involved in measuring health insurance coverage, and argues that the current method of estimation of the uninsured underestimates the extent that individuals go without health insurance. Third, the paper briefly introduces Amartya Sen's functioning and capabilities framework to suggest a way of representing the extent to which individuals are uninsured. Fourth, the paper sketches a means of operationalizing the Sen representation of the uninsured in terms of the disability-adjusted life year (DALY) measure.

  17. Enhanced Ascorbate Regeneration Via Dehydroascorbate Reductase Confers Tolerance to Photo-Oxidative Stress in Chlamydomonas reinhardtii.

    Science.gov (United States)

    Lin, Shu-Tseng; Chiou, Chih-Wen; Chu, Yen-Lin; Hsiao, Yu; Tseng, Yu-Fei; Chen, Yi-Chun; Chen, Hsien-Jung; Chang, Hsin-Yang; Lee, Tse-Min

    2016-10-01

    The role of ascorbate (AsA) recycling via dehydroascorbate reductase (DHAR) in the tolerance of Chlamydomonas reinhardtii to photo-oxidative stress was examined. The activity of DHAR and the abundance of the CrDHAR1 (Cre10.g456750) transcript increased after moderate light (ML; 750 µmol m(-2) s(-1)) or high light (HL; 1,800 µmol m(-2) s(-1)) illumination, accompanied by dehydroascorbate (DHA) accumulation, decreased AsA redox state, photo-inhibition, lipid peroxidation, H2O2 overaccumulation, growth inhibition and cell death. It suggests that DHAR and AsA recycling is limiting under high-intensity light stress. The CrDHAR1 gene was cloned and its recombinant CrDHAR1 protein was a monomer (25 kDa) detected by Western blot that exhibits an enzymatic activity of 965 µmol min(-1)( )mg(-1) protein. CrDHAR1 was overexpressed driven by a HSP70A:RBCS2 fusion promoter or down-regulated by artificial microRNA (amiRNA) to examine whether DHAR-mediated AsA recycling is critical for the tolerance of C. reinahartii cells to photo-oxidative stress. The overexpression of CrDHAR1 increased DHAR protein abundance and enzyme activity, AsA pool size, AsA:DHA ratio and the tolerance to ML-, HL-, methyl viologen- or H2O2-induced oxidative stress. The CrDHAR1-knockdown amiRNA lines that have lower DHAR expression and AsA recycling ability were sensitive to high-intensity illumination and oxidative stress. The glutathione pool size, glutathione:oxidized glutathione ratio and glutathione reductase and ascorbate peroxidase activities were increased in CrDHAR1-overexpressing cells and showed a further increase after high-intensity illumination but decreased in wild-type cells after light stress. The present results suggest that increasing AsA regeneration via enhanced DHAR activity modulates the ascorbate-glutathione cycle activity in C. reinhardtii against photo-oxidative stress.

  18. Interactive effects of copper oxide nanoparticles and light to green alga Chlamydomonas reinhardtii

    Energy Technology Data Exchange (ETDEWEB)

    Cheloni, Giulia; Marti, Elodie; Slaveykova, Vera I., E-mail: vera.slaveykova@unige.ch

    2016-01-15

    Highlights: • Comparable stability of CuO-NP suspensions under different light conditions. • UVR* inhibits growth, bleaches chlorophyll fluorescence and damages membrane. • Below 1 mg L{sup −1} CuO-NPs do not attenuate light in algal suspension. • SNL enhances significantly the effect of 0.8 mg L{sup −1} CuO-NPs on microalgae. • Synergistic interactions between UVR* and CuO-NPs. - Abstract: The present study explores the effect of light with different spectral composition on the stability of CuO-nanoparticle (CuO-NP) dispersions and their effects to green alga Chlamydomonas reinhardtii. The results showed that simulated natural light (SNL) and light with enhanced UVB radiation (UVR*) do not affect the dissolution of CuO-NPs as compared to light irradiation conditions typically used in laboratory incubator (INC). Comparable values of ζ-potential and hydrodynamic size during 24 h were found under all studied conditions. Concentrations of CuO-NPs below 1 mg L{sup −1} do not attenuate the light penetration in the algal suspensions in comparison with NP-free system. Exposure to a combination of 8 μg L{sup −1} or 0.8 mg L{sup −1} CuO-NPs and INC or SNL has no significant effect on the algal growth inhibition, algal fluorescence and membrane integrity under short-term exposure. However, an enhancement of the percentage of cells experiencing oxidative stress was observed upon exposure to 0.8 mg L{sup −1} CuO-NPs and SNL for 4 and 8 h. Combination of UVR* and 0.8 mg L{sup −1} CuO-NPs resulted in synergistic effects for all biological endpoints. Despite the photocatalytic properties of CuO-NPs no significant increase in abiotic reactive oxygen species (ROS) production under simulated solar radiation was observed suggesting that the synergistic effect observed might be correlated to other factors than CuO-NP-mediated ROS photoproduction. Tests performed with CuSO{sub 4} confirmed the important role of dissolution as toxicity driving force for lower

  19. Harvesting microalgae cultures with superabsorbent polymers: desulfurization of Chlamydomonas reinhardtii for hydrogen production.

    Science.gov (United States)

    Martín del Campo, Julia S; Patiño, Rodrigo

    2013-12-01

    It is presented in this work a new methodology to harvest fresh water microalgae cultures by extracting the culture medium with superabsorbent polymers (SAPs). The microalgae Chlamydomonas reinhardtii were grown in the Sueoka culture medium, harvested with polyacrylic SAPs and re-suspended in the culture medium tris-acetate-potassium without sulfur (TAP-S) to generate hydrogen (H2 ) under anoxic conditions. The H2 production as an alternative fuel is relevant since this gas has high-energy recovery without involving carbon. Before microalgae harvesting, a number of range diameters (1-7 mm) for SAPs spherical particles were tested, and the initial rate (V0 ) and the maximal capacity (Qmax ) were determined for the Sueoka medium absorption. The SAP particles with the diameter range 2.0-2.5 mm performed the best and these were employed for the rest of the experiments. The Sueoka medium has a high salt content and the effect of the ionic strength was also studied for different medium concentrations (0-400%). The SAPs were reused in consecutive absorption/desorption cycles, maintaining their absorption capacity. Although the Sueoka medium reduces the SAPs absorption capacity to 40% compared with deionized water, the use of SAPs was very significant for the desulfurization process of C. reihardtii. The presence of C. reinhardtii at different concentrations does not affect the absorption capacity of the Sueoka culture medium by the SAPs. In order to reduce the time of the process, an increase of the SAPs concentration was tested, being 20 g of SAP per liter of medium, a condition to harvest the microalgae culture in 4 h. There were no evident cell ruptures during the harvesting process and the cells remained alive. Finally, the harvested biomass was re-suspended in TAP-S medium and kept under anaerobic conditions and illumination to produce H2 that was monitored by a PEM fuel cell. The use of SAPs for microalgae harvesting is a feasible non-invasive procedure to obtain

  20. Reduced alcohol consumption in mice lacking preprodynorphin.

    OpenAIRE

    Blednov, Yuri A.; Walker, Danielle; Martinez, Marni; Harris, R. Adron

    2006-01-01

    Many studies suggest a role for endogenous opioid peptides and their receptors in regulation of ethanol intake. It is commonly accepted that the κ-opioid receptors and their endogenous ligands, dynorphins, produce a dysphoric state and therefore may be responsible for avoidance of alcohol. We used mutant mice lacking preprodynorphin in a variety of behavioral tests of alcohol actions. Null mutant female, but not male, mice showed significantly lower preference for alcohol and consumed lower a...

  1. The plastoquinol-plastoquinone exchange mechanism in photosystem II: insight from molecular dynamics simulations.

    Science.gov (United States)

    Zobnina, Veranika; Lambreva, Maya D; Rea, Giuseppina; Campi, Gaetano; Antonacci, Amina; Scognamiglio, Viviana; Giardi, Maria Teresa; Polticelli, Fabio

    2017-01-01

    In the photosystem II (PSII) of oxygenic photosynthetic organisms, the reaction center (RC) core mediates the light-induced electron transfer leading to water splitting and production of reduced plastoquinone molecules. The reduction of plastoquinone to plastoquinol lowers PSII affinity for the latter and leads to its release. However, little is known about the role of protein dynamics in this process. Here, molecular dynamics simulations of the complete PSII complex embedded in a lipid bilayer have been used to investigate the plastoquinol release mechanism. A distinct dynamic behavior of PSII in the presence of plastoquinol is observed which, coupled to changes in charge distribution and electrostatic interactions, causes disruption of the interactions seen in the PSII-plastoquinone complex and leads to the "squeezing out" of plastoquinol from the binding pocket. Displacement of plastoquinol closes the second water channel, recently described in a 2.9 Å resolution PSII structure (Guskov et al. in Nat Struct Mol Biol 16:334-342, 2009), allowing to rule out the proposed "alternating" mechanism of plastoquinol-plastoquinone exchange, while giving support to the "single-channel" one. The performed simulations indicated a pivotal role of D1-Ser264 in modulating the dynamics of the plastoquinone binding pocket and plastoquinol-plastoquinone exchange via its interaction with D1-His252 residue. The effects of the disruption of this hydrogen bond network on the PSII redox reactions were experimentally assessed in the D1 site-directed mutant Ser264Lys.

  2. Two-Dimensional Electronic Spectroscopies for Probing Electronic Structure and Charge Transfer: Applications to Photosystem II

    Energy Technology Data Exchange (ETDEWEB)

    Ogilvie, Jennifer P. [Univ. of Michigan, Ann Arbor, MI (United States). Dept. of Physics

    2016-11-22

    Photosystem II (PSII) is the only known natural enzyme that uses solar energy to split water, making the elucidation of its design principles critical for our fundamental understanding of photosynthesis and for our ability to mimic PSII’s remarkable properties. This report discusses progress towards addressing key open questions about the PSII RC. It describes new spectroscopic methods that were developed to answer these questions, and summarizes the outcomes of applying these methods to study the PSII RC. Using 2D electronic spectroscopy and 2D electronic Stark spectroscopy, models for the PSII RC were tested and refined. Work is ongoing to use the collected data to elucidate the charge separation mechanism in the PSII RC. Coherent dynamics were also observed in the PSII RC for the first time. Through extensive characterization and modeling we have assigned these coherences as vibronic in nature, and believe that they reflect resonances between key vibrational pigment modes and electronic energy gaps that may facilitate charge separation. Work is ongoing to definitively test the functional relevance of electronic-vibrational resonances.

  3. Interaction of polyamines with proteins of photosystem II: Cation binding and photosynthetic oxygen evolution

    Science.gov (United States)

    Beauchemin, R.; Harnois, J.; Rouillon, R.; Tajmir-Riahi, H. A.; Carpentier, R.

    2007-05-01

    Polyamines are organic cations that function in diverse physiological processes that share as a common thread a close relationship to cell proliferation and growth. Polyamines also affect photosynthetic oxygen evolution and therefore, this study was designed to investigate the interaction of 1,3-diaminopropane, 1,4-diaminobutane (putrescine), and 1,5-diaminopentane (cadaverine) cations with proteins of photosystem II (PSII) using PSII-enriched submembrane fractions with diamine concentrations between 0.01 and 20 mM. Fourier transformed infrared (FTIR) difference spectroscopy with its self-deconvolution and second derivative resolution enhancement, as well as curve-fitting procedures were applied in order to determine the diamine binding mode, the protein conformational changes, and the structural properties of diamine-protein complexes. Spectroscopic evidence showed that diamines interact with proteins (H-bonding) through polypeptide C dbnd O groups with no major perturbations of protein secondary structure. At very low diamine concentration (0.01 mM), no inhibition of oxygen-evolution occurred, while at higher diamine content (5-10 mM), 100% inhibition was observed. Chorophyll fluorescence measurements demonstrated that the inhibition mainly affects the oxygen evolving complex of PSII. Comparisons of the effects of these dipositive organic cations with divalent metal cations on one hand and with polyvalent spermine and spermidine on the other hand, show major alterations of the protein secondary structure as positive charge increases.

  4. Long-wavelength chlorophylls in photosystem I of cyanobacteria: origin, localization, and functions.

    Science.gov (United States)

    Karapetyan, N V; Bolychevtseva, Yu V; Yurina, N P; Terekhova, I V; Shubin, V V; Brecht, M

    2014-03-01

    The structural organization of photosystem I (PSI) complexes in cyanobacteria and the origin of the PSI antenna long-wavelength chlorophylls and their role in energy migration, charge separation, and dissipation of excess absorbed energy are discussed. The PSI complex in cyanobacterial membranes is organized preferentially as a trimer with the core antenna enriched with long-wavelength chlorophylls. The contents of long-wavelength chlorophylls and their spectral characteristics in PSI trimers and monomers are species-specific. Chlorophyll aggregates in PSI antenna are potential candidates for the role of the long-wavelength chlorophylls. The red-most chlorophylls in PSI trimers of the cyanobacteria Arthrospira platensis and Thermosynechococcus elongatus can be formed as a result of interaction of pigments peripherally localized on different monomeric complexes within the PSI trimers. Long-wavelength chlorophylls affect weakly energy equilibration within the heterogeneous PSI antenna, but they significantly delay energy trapping by P700. When the reaction center is open, energy absorbed by long-wavelength chlorophylls migrates to P700 at physiological temperatures, causing its oxidation. When the PSI reaction center is closed, the P700 cation radical or P700 triplet state (depending on the P700 redox state and the PSI acceptor side cofactors) efficiently quench the fluorescence of the long-wavelength chlorophylls of PSI and thus protect the complex against photodestruction.

  5. Limitations to photosynthesis by proton motive force-induced photosystem II photodamage

    Science.gov (United States)

    Davis, Geoffry A; Kanazawa, Atsuko; Schöttler, Mark Aurel; Kohzuma, Kaori; Froehlich, John E; Rutherford, A William; Satoh-Cruz, Mio; Minhas, Deepika; Tietz, Stefanie; Dhingra, Amit; Kramer, David M

    2016-01-01

    The thylakoid proton motive force (pmf) generated during photosynthesis is the essential driving force for ATP production; it is also a central regulator of light capture and electron transfer. We investigated the effects of elevated pmf on photosynthesis in a library of Arabidopsis thaliana mutants with altered rates of thylakoid lumen proton efflux, leading to a range of steady-state pmf extents. We observed the expected pmf-dependent alterations in photosynthetic regulation, but also strong effects on the rate of photosystem II (PSII) photodamage. Detailed analyses indicate this effect is related to an elevated electric field (Δψ) component of the pmf, rather than lumen acidification, which in vivo increased PSII charge recombination rates, producing singlet oxygen and subsequent photodamage. The effects are seen even in wild type plants, especially under fluctuating illumination, suggesting that Δψ-induced photodamage represents a previously unrecognized limiting factor for plant productivity under dynamic environmental conditions seen in the field. DOI: http://dx.doi.org/10.7554/eLife.16921.001 PMID:27697149

  6. Triplet state in photosystem II reaction centers as studied by 130 GHz EPR

    Energy Technology Data Exchange (ETDEWEB)

    Pashenko, S.V.; Proskuryakov, I.I.; Germano, M.; Gorkom, H.J. van; Gast, P

    2003-11-01

    The triplet state in the reaction centers of photosystem II was studied by high-field/high-frequency (130 GHz) EPR in the temperature range 50-90 K. At 50 K, the zero-field splitting parameters of the EPR spectrum correspond well to those of a chlorophyll monomer, in agreement with earlier studies. In the high magnetic field of 4.6 T employed in this study, the g-anisotropy of the triplet state becomes apparent and leads to a shift of the canonical positions of the triplet EPR spectrum. Assuming that triplet g- and zero-field tensors are coaxial, the principal values of the triplet g-tensor are determined to be 2.00324, 2.00306 and 2.00231 with an error of {+-}0.00004. Lifting this assumption results in higher g-anisotropy. At higher temperatures, the shape of the spectra changes significantly. Triplet excitation hopping involving the accessory chlorophyll B{sub A} and P{sub A} or P{sub B} (equivalents of the special pair bacteriochlorophylls of the bacterial reaction centers) can partially explain those changes, but the most prominent features indicate that also the electron acceptor I{sub A} (a pheophytin molecule) must be involved.

  7. Effects of Iron Limitation on Photosystem II Composition and Light Utilization in Dunaliella tertiolecta.

    Science.gov (United States)

    Vassiliev, I. R.; Kolber, Z.; Wyman, K. D.; Mauzerall, D.; Shukla, V. K.; Falkowski, P. G.

    1995-11-01

    The effects of iron limitation on photosystem II (PSII) composition and photochemical energy conversion efficiency were studied in the unicellular chlorophyte alga Dunaliella tertiolecta. The quantum yield of photochemistry in PSII, inferred from changes in variable fluorescence normalized to the maximum fluorescence yield, was markedly lower in iron-limited cells and increased 3-fold within 20 h following the addition of iron. The decrease in the quantum yield of photochemistry was correlated with increased fluorescence emission from the antenna. In iron-limited cells, flash intensity saturation profiles of variable fluorescence closely followed a cumulative one-hit Poisson model, suggesting that PSII reaction centers are energetically isolated, whereas in iron-replete cells, the slope of the profile was steeper and the calculated probability of energy transfer between reaction centers increased to >0.6. Immunoassays revealed that in iron-limited cells the reaction center proteins, D1, CP43, and CP47, were markedly reduced relative to the peripheral light-harvesting Chl-protein complex of PSII, whereas the [alpha] subunit of cytochrome b559 was about 10-fold higher. Spectroscopic analysis established that the cytochrome b559 peptide did not contain an associated functional heme. We conclude that the photochemical conversion of absorbed excitation energy in iron-limited cells is limited by the number of photochemical traps per unit antenna.

  8. Detection of photosynthetic energy storage in a photosystem I reaction center preparation by photoacoustic spectroscopy.

    Science.gov (United States)

    Owens, T G; Carpentier, R; Leblanc, R M

    1990-06-01

    Thermal emission and photochemical energy storage were examined in photosystem I reaction center/core antenna complexes (about 40 Chl a/P700) using photoacoustic spectroscopy. Satisfactory signals could only be obtained from samples bound to hydroxyapatite and all samples had a low signal-to-noise ratio compared to either PS I or PS II in thylakoid membranes. The energy storage signal was saturated at low intensity (half saturation at 1.5 W m(-2)) and predicted a photochemical quantum yield of >90%. Exogenous donors and acceptors had no effect on the signal amplitudes indicating that energy storage is the result of charge separation between endogenous components. Fe(CN)6 (-3) oxidation of P700 and dithionite-induced reduction of acceptors FA-FB inhibited energy storage. These data are compatible with the hypothesis that energy storage in PS I arises from charge separation between P700 and Fe-S centers FA-FB that is stable on the time scale of the photoacoustic modulation. High intensity background light (160 W m(-2)) caused an irreversible loss of energy storage and correlated with a decrease in oxidizable P700; both are probably the result of high light-induced photoinhibition. By analogy to the low fluorescence yield of PS I, the low signal-to-noise ratio in these preparations is attributed to the short lifetime of Chl singlet excited states in PS I-40 and its indirect effect on the yield of thermal emission.

  9. Reconstitution of Photosystem Ⅱ Reaction Center with Cu-Chlorophyll a

    Institute of Scientific and Technical Information of China (English)

    Shuang Liu; Feng-Qin Dong; Chun-Hong Yang; Chong-Qin Tang; Ting-Yun Kuang

    2006-01-01

    An isolated photosystem (PS) Ⅱ reaction center (RC) with altered pigment content was obtained by chemical exchange of native chlorophyll a (Chl) with externally added Cu-Chl a (Cu-Chl). Pigment composition and spectroscopic properties of the RC exchanged with Cu-Chl were compared with native RC and RC treated with Chl in the same way. High-performance liquid chromatography analysis showed approximately 0.5 Cu-Chl per two pheophytin in the Cu-Chl-reconstituted RC preparation. Insertion of Cu-Chl resulted in a decrease in absorption at 670 nm and an increase at 660 nm, suggesting that the peripheral Chl may have been displaced. Fluorescence emission spectra of the Cu-Chl-reconstituted RC displayed a marked decrease in fluorescence yield and a blue shift of the band maximum, accompanied by the appearance of a broad peak at a shorter wavelength, indicating that energy transfer in the modified RC was disturbed by Cu-Chl, a quencher of the excited state. However, there were few differences in the circular dichroism (CD) spectra,suggesting that the arrangement of pigments and proteins responsible for the CD signal was not significantly affected. In addition, no obvious change in peptide components was found after the exchange procedure.

  10. Protein synthesis is the primary target of reactive oxygen species in the photoinhibition of photosystem II.

    Science.gov (United States)

    Nishiyama, Yoshitaka; Allakhverdiev, Suleyman I; Murata, Norio

    2011-05-01

    Photoinhibition of photosystem II (PSII) occurs when the rate of photodamage to PSII exceeds the rate of the repair of photodamaged PSII. Recent examination of photoinhibition by separate determinations of photodamage and repair has revealed that the rate of photodamage to PSII is directly proportional to the intensity of incident light and that the repair of PSII is particularly sensitive to the inactivation by reactive oxygen species (ROS). The ROS-induced inactivation of repair is attributable to the suppression of the synthesis de novo of proteins, such as the D1 protein, that are required for the repair of PSII at the level of translational elongation. Furthermore, molecular analysis has revealed that the ROS-induced suppression of protein synthesis is associated with the specific inactivation of elongation factor G via the formation of an intramolecular disulfide bond. Impairment of various mechanisms that protect PSII against photoinhibition, including photorespiration, thermal dissipation of excitation energy, and the cyclic transport of electrons, decreases the rate of repair of PSII via the suppression of protein synthesis. In this review, we present a newly established model of the mechanism and the physiological significance of repair in the regulation of the photoinhibition of PSII.

  11. Theoretical study on the role of Ca(2+) at the S2 state in photosystem II.

    Science.gov (United States)

    Yang, Jingxiu; Hatakeyama, Makoto; Ogata, Koji; Nakamura, Shinichiro; Li, Can

    2014-12-11

    In photosynthesis, calcium is crucial for oxygen evolution. In the absence of Ca(2+), the Kok cycle has been proven to stop at S2 with Yz•. To explore the reason, photosystem II (PSII) S2 models (in total 32452 atoms) with different metal ions (Ca(2+), Sr(2+), and K(+)) or without Ca(2+) involved in the oxygen evolution complex (OEC) have been theoretically studied based on the previous dynamic study of PSII. It is found that the portion of the Mn1 d-orbital decreases in the highest occupied molecular orbitals for Ca(2+)-depleted PSII. This feature is unfavorable for the electron transfer from the OEC to the Yz•. Furthermore, the proton donor-acceptor distance was found elongated by the alternation of the binding water in the absence of Ca(2+). The isolated vibrational modes of the key water molecules along the path and their high frequency of the OH stretching modes also suggested the difficulty of the proton transfer from the OEC toward the proton exit channel. This work provides one mechanistic explanation for the inactivity of Ca(2+)-depleted PSII.

  12. Acute and additive toxicity of ten photosystem-II herbicides to seagrass

    Science.gov (United States)

    Wilkinson, Adam D.; Collier, Catherine J.; Flores, Florita; Negri, Andrew P.

    2015-11-01

    Photosystem II herbicides are transported to inshore marine waters, including those of the Great Barrier Reef, and are usually detected in complex mixtures. These herbicides inhibit photosynthesis, which can deplete energy reserves and reduce growth in seagrass, but the toxicity of some of these herbicides to seagrass is unknown and combined effects of multiple herbicides on seagrass has not been tested. Here we assessed the acute phytotoxicity of 10 PSII herbicides to the seagrass Halophila ovalis over 24 and/or 48 h. Individual herbicides exhibited a broad range of toxicities with inhibition of photosynthetic activity (∆F/Fm‧) by 50% at concentrations ranging from 3.5 μg l-1 (ametryn) to 132 μg l-1 (fluometuron). We assessed potential additivity using the Concentration Addition model of joint action for binary mixtures of diuron and atrazine as well as complex mixtures of all 10 herbicides. The effects of both mixture types were largely additive, validating the application of additive effects models for calculating the risk posed by multiple PSII herbicides to seagrasses. This study extends seagrass ecotoxicological data to ametryn, metribuzin, bromacil, prometryn and fluometuron and demonstrates that low concentrations of PSII herbicide mixtures have the potential to impact ecologically relevant endpoints in seagrass, including ∆F/Fm‧.

  13. Structural investigation of PsbO from plant and cyanobacterial photosystem II.

    Science.gov (United States)

    Slowik, Daria; Rossmann, Maxim; Konarev, Petr V; Irrgang, Klaus-D; Saenger, Wolfram

    2011-03-18

    The manganese-stabilizing protein PsbO is associated with the luminal side of thylakoids close to the redox-active Mn(4)Ca cluster at the catalytically active site of photosystem II (PSII). PsbO is believed to increase the efficiency of oxygen evolution and to stabilize the Mn(4)Ca cluster against photoinhibition. Using small-angle X-ray scattering, we investigated the low-resolution structure of wild-type spinach PsbO and that of chimeric spinach PsbO fused with maltose-binding protein. Small-angle X-ray scattering data revealed that both proteins are monomeric in solution, and that plant and cyanobacterial PsbO have similar structures. We show a highly efficient expression system that allows recombinant production of the active wild type and the chimeric PsbO from spinach and cyanobacteria, with yields compatible with biophysical and structural studies. The binding of spinach PsbO fused with maltose-binding protein to PSII depleted of extrinsic subunits (PSII-ΔpsbO,P,Q) was confirmed by matrix-assisted laser desorption ionization time-of-flight mass spectrometry. The reconstituted PSII was shown to have similar oxygen evolution rates as obtained with wild-type spinach PsbO.

  14. Structural basis of cyanobacterial photosystem II Inhibition by the herbicide terbutryn.

    Science.gov (United States)

    Broser, Matthias; Glöckner, Carina; Gabdulkhakov, Azat; Guskov, Albert; Buchta, Joachim; Kern, Jan; Müh, Frank; Dau, Holger; Saenger, Wolfram; Zouni, Athina

    2011-05-01

    Herbicides that target photosystem II (PSII) compete with the native electron acceptor plastoquinone for binding at the Q(B) site in the D1 subunit and thus block the electron transfer from Q(A) to Q(B). Here, we present the first crystal structure of PSII with a bound herbicide at a resolution of 3.2 Å. The crystallized PSII core complexes were isolated from the thermophilic cyanobacterium Thermosynechococcus elongatus. The used herbicide terbutryn is found to bind via at least two hydrogen bonds to the Q(B) site similar to photosynthetic reaction centers in anoxygenic purple bacteria. Herbicide binding to PSII is also discussed regarding the influence on the redox potential of Q(A), which is known to affect photoinhibition. We further identified a second and novel chloride position close to the water-oxidizing complex and in the vicinity of the chloride ion reported earlier (Guskov, A., Kern, J., Gabdulkhakov, A., Broser, M., Zouni, A., and Saenger, W. (2009) Nat. Struct. Mol. Biol. 16, 334-342). This discovery is discussed in the context of proton transfer to the lumen.

  15. Thermal stability of oxygen evolution in photosystem Ⅱ core complex in the presence of digalactosyl diacylglycerol

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    The influence of digalactosyldiacylglycerol (DGDG), one of the photosynthetic membrane lipids, on heat inactivation of the process of oxygen evolution has been studied in vitro in photosystem Ⅱ (PSⅡ) core complex. It was found that the temperature of semi-inactivation of oxygen evolution in the complex increased from 40.0 to about 43.0℃ in the presence of DGDG with 5-min heat treatment in the dark. Furthermore, when PSⅡ core complex was incubated for 5 min at 45.0℃, the oxygen evolution in the complex was completely lost, whilst the DGDG-complexed PSⅡ core complex still retained a 16% of activity (100% for 25.0℃). In addition, a 1-h incubation at 38.0℃ inactivated absolutely the oxygen evolution for the PSⅡ core complex. By contrast, there remained about 20% of activity (zero time for 100%) for the complex in the presence of DGDG under the same condition. These results indicate a new role of DGDG in the protection of PSⅡ core complex against the deleterious effects of temperature. It was most likely that DGDG-mediated stability toward thermal denaturation of oxygen evolution in PSⅡ core complex is due to the protective effect of DGDG on the release of the 33 kD protein from PSⅡ core complex.

  16. Structural organization of an intact phycobilisome and its association with photosystem II.

    Science.gov (United States)

    Chang, Leifu; Liu, Xianwei; Li, Yanbing; Liu, Cui-Cui; Yang, Fan; Zhao, Jindong; Sui, Sen-Fang

    2015-06-01

    Phycobilisomes (PBSs) are light-harvesting antennae that transfer energy to photosynthetic reaction centers in cyanobacteria and red algae. PBSs are supermolecular complexes composed of phycobiliproteins (PBPs) that bear chromophores for energy absorption and linker proteins. Although the structures of some individual components have been determined using crystallography, the three-dimensional structure of an entire PBS complex, which is critical for understanding the energy transfer mechanism, remains unknown. Here, we report the structures of an intact PBS and a PBS in complex with photosystem II (PSII) from Anabaena sp. strain PCC 7120 using single-particle electron microscopy in combination with biochemical and molecular analyses. In the PBS structure, all PBP trimers and the conserved linker protein domains were unambiguously located, and the global distribution of all chromophores was determined. We provide evidence that ApcE and ApcF are critical for the formation of a protrusion at the bottom of PBS, which plays an important role in mediating PBS interaction with PSII. Our results provide insights into the molecular architecture of an intact PBS at different assembly levels and provide the basis for understanding how the light energy absorbed by PBS is transferred to PSII.

  17. Analysis of light-induced transmembrane ion gradients and membrane potential in Photosystem I proteoliposomes.

    Science.gov (United States)

    Pennisi, Cristian Pablo; Greenbaum, Elias; Yoshida, Ken

    2010-01-01

    Photosystem I (PSI) complexes can support a light-driven electrochemical gradient for protons, which is the driving force for energy-conserving reactions across biological membranes. In this work, a computational model that enables a quantitative description of the light-induced proton gradients across the membrane of PSI proteoliposomes is presented. Using a set of electrodiffusion equations, a compartmental model of a vesicle suspended in aqueous medium was studied. The light-mediated proton movement was modeled as a single proton pumping step with backpressure of the electric potential. The model fits determinations of pH obtained from PSI proteoliposomes illuminated in the presence of mediators of cyclic electron transport. The model also allows analysis of the proton gradients in relation to the transmembrane ion fluxes and electric potential. Sensitivity analysis enabled a determination of the parameters that have greater influence on steady-state levels and onset/decay rates of transmembrane pH and electric potential. This model could be used as a tool for optimizing PSI proteoliposomes for photo-electrochemical applications.

  18. Analysis of Light-Induced Transmembrane Ion Gradients and Membrane Potential in Photosystem I Proteoliposomes

    Energy Technology Data Exchange (ETDEWEB)

    Pennisi, Cristian P. [Aalborg University, Aalborg, Denmark; Greenbaum, Elias [ORNL; Yoshida, Ken [Aalborg University, Aalborg, Denmark

    2010-01-01

    Photosystem I (PSI) complexes can support a light-driven electrochemical gradient for protons, which is the driving force for energy-conserving reactions across biological membranes. In this work, a computational model that enables a quantitative description of the light-induced proton gradients across the membrane of PSI proteoliposomes is presented. Using a set of electrodiffusion equations, a compartmental model of a vesicle suspended in aqueous medium was studied. The light-mediated proton movement was modeled as a single proton pumping step with backpressure of the electric potential. The model fits determinations of pH obtained from PSI proteoliposomes illuminated in the presence of mediators of cyclic electron transport. The model also allows analysis of the proton gradients in relation to the transmembrane ion fluxes and electric potential. Sensitivity analysis enabled a determination of the parameters that have greater influence on steady-state levels and onset/decay rates of transmembrane pH and electric potential. This model could be used as a tool for optimizing PSI proteoliposomes for photo-electrochemical applications.

  19. Structural Coupling of Extrinsic Proteins with the Oxygen-Evolving Center in Photosystem II

    Directory of Open Access Journals (Sweden)

    Kentaro eIfuku

    2016-02-01

    Full Text Available Photosystem II (PSII, which catalyzes photosynthetic water oxidation, is composed of more than 20 subunits, including membrane-intrinsic and -extrinsic proteins. The PSII extrinsic proteins shield the catalytic Mn4CaO5 cluster from the outside bulk solution and enhance binding of inorganic cofactors, such as Ca2+ and Cl-, in the oxygen-evolving center (OEC of PSII. Among PSII extrinsic proteins, PsbO is commonly found in all oxygenic organisms, while PsbP and PsbQ are specific to higher plants and green algae, and PsbU, PsbV, CyanoQ, and CyanoP exist in cyanobacteria. In addition, red algae and diatoms have unique PSII extrinsic proteins, such as PsbQ’ and Psb31, suggesting functional divergence during evolution. Recent studies with reconstitution experiments combined with Fourier transform infrared spectroscopy have revealed how the individual PSII extrinsic proteins affect the structure and function of the OEC in different organisms. In this review, we summarize our recent results and discuss changes that have occurred in the structural coupling of extrinsic proteins with the OEC during evolutionary history.

  20. Photosynthetic Properties of Photosystem Ⅱ in Arabidopsis thaliana Ipa1 Mutant

    Institute of Scientific and Technical Information of China (English)

    Lian-Wei Peng; Jin-Kui Guo; Jin-Fang Ma; Wei Chi; Li-Xin Zhang

    2006-01-01

    In a previous study, we characterized a high chlorophyll fluorescence Ipa1 mutant of Arabidopsis thaliana,in which approximately 20% photosystem (PS) Ⅱ protein is accumulated. In the present study, analysis of fluorescence decay kinetics and thermoluminescence profiles demonstrated that the electron transfer reaction on either the donor or acceptor side of PSll remained largely unaffected in the Ipa1 mutant. In the mutant, maximal photochemical efficiency (Fv/Fm, where Fm is the maximum fluorescence yield and Fv is variable fluorescence) decreased with increasing light intensity and remained almost unchanged in wildtype plants under different light conditions. The Fv/Fm values also increased when mutant plants were transferred from standard growth light to low light conditions. Analysis of PSⅡ protein accumulation further confirmed that the amount of PSll reaction center protein is correlated with changes in Fv/Fm in Ipa1 plants.Thus, the assembled PSll in the mutant was functional and also showed increased photosensitivity compared with wild-type plants.

  1. Phosphatidylglycerol effect on oxygen-evolving activity in Ca2+-depleted photosystem

    Institute of Scientific and Technical Information of China (English)

    2000-01-01

    The effect of anionic phosphatidylglycerol (PG) on oxygen evolution in a photosystem Ⅱ (PSⅡ) particle depleted of Ca2+ (designated dCaPSⅡ) has been investigated. The major finding is the observation of a new role of PG in the PSⅡ function. That is, PG restores nearly the lost oxygen evolution in dCaPSⅡ particle owing to Ca2+ depletion to the levels in intact PSⅡ. Furthermore, there is a stimulation of oxygen-evolving activity in the dCaPSⅡ complexed with PG in the presence of exogenous CaCl2, which PG enhances increasingly oxygen evolution with increasing CaCl2 concentration. It is suggested that PG-induced oxygen evolution recovery of dCa PSⅡ particle results from resumption of normal structure in protein by PG effect, whereas the enhancement of oxygen evolution in complex subject to CaCl2 is ascribed to the optimization of such a structure due to coordination complex formation of Ca2+ ions with PG.

  2. A supramolecular ruthenium macrocycle with high catalytic activity for water oxidation that mechanistically mimics photosystem II

    Science.gov (United States)

    Schulze, Marcus; Kunz, Valentin; Frischmann, Peter D.; Würthner, Frank

    2016-06-01

    Mimicking the ingenuity of nature and exploiting the billions of years over which natural selection has developed numerous effective biochemical conversions is one of the most successful strategies in a chemist's toolbox. However, an inability to replicate the elegance and efficiency of the oxygen-evolving complex of photosystem II (OEC-PSII) in its oxidation of water into O2 is a significant bottleneck in the development of a closed-loop sustainable energy cycle. Here, we present an artificial metallosupramolecular macrocycle that gathers three Ru(bda) centres (bda = 2,2‧-bipyridine-6,6‧-dicarboxylic acid) that catalyses water oxidation. The macrocyclic architecture accelerates the rate of water oxidation via a water nucleophilic attack mechanism, similar to the mechanism exhibited by OEC-PSII, and reaches remarkable catalytic turnover frequencies >100 s-1. Photo-driven water oxidation yields outstanding activity, even in the nM concentration regime, with a turnover number of >1,255 and turnover frequency of >13.1 s-1.

  3. Rapid Evolution of the Photosystem II Electronic Structure during Water Splitting

    CERN Document Server

    Davis, Katherine M; Palenik, Mark; Yan, Lifen; Purohit, Vatsal; Robison, Gregory; Kosheleva, Irina; Henning, Robert W; Seidler, Gerald T; Pushkar, Yulia

    2015-01-01

    Photosynthetic water oxidation is a fundamental process that sustains the biosphere. A Mn$_{4}$Ca cluster embedded in the photosystem II protein environment is responsible for the production of atmospheric oxygen. Here, time-resolved x-ray emission spectroscopy (XES) was used to observe the process of oxygen formation in real time. These experiments reveal that the oxygen evolution step, initiated by three sequential laser flashes, is accompanied by rapid (within 50 $\\mu$s) changes to the Mn K$\\beta$ XES spectrum. However, no oxidation of the Mn$_{4}$Ca core above the all Mn$^{\\text{IV}}$ state was detected to precede O-O bond formation. A new mechanism featuring Mn$^{\\text{IV}}$=O formation in the S$_{3}$ state is proposed to explain the spectroscopic results. This chemical formulation is consistent with the unique reactivity of the S$_{3}$ state and explains facilitation of the following S$_{3}$ to S$_{0}$ transition, resolving in part the kinetic limitations associated with O-O bond formation. In the propo...

  4. Endophytic infection alleviates Pb(2+) stress effects on photosystem II functioning of Oryza sativa leaves.

    Science.gov (United States)

    Li, Xuemei; Zhang, Lihong

    2015-09-15

    The aims of this study were to examine the effect of Pb(2+) stress on the primary reaction of photosynthesis and to assess the potential benefits of endophytic infection on the Pb(2+) tolerance of rice seedlings. Rice inoculated with an endophytic fungus (E+) and non-inoculated (E-) were subjected to 0, 50, 100, 150 and 200 μM Pb(2+). The responses to Pb(2+) stress were characterized by the analysis of Chl a fluorescence. A comparison of E- with E+ rice seedlings, as evaluated by their performance index (PI(ABS) and PI(tot)), revealed the inhibitory effects of Pb(2+) on photosystem II (PSII) connectivity, the oxygen evolving complex (OEC), and on the J step of the induction curves, which is associated with an inhibition of electron transport from the quinone acceptor Q(A) to Q(B). Furthermore, the changes of the donor and the acceptor parameters of PSII were greater in E- than in E+ under Pb(2+) stress. These observations suggest that the efficiency and stability of PSII are markedly affected by Pb(2+) stress, and the photosynthetic energy conservation in E+ was more effective than in E-. We showed that endophytic infection plays an important role in enhancing the photosynthetic mechanism of rice seedlings exposed to Pb(2+) stress.

  5. Excitation-energy transfer dynamics of higher plant photosystem I light-harvesting complexes.

    Science.gov (United States)

    Wientjes, Emilie; van Stokkum, Ivo H M; van Amerongen, Herbert; Croce, Roberta

    2011-03-01

    Photosystem I (PSI) plays a major role in the light reactions of photosynthesis. In higher plants, PSI is composed of a core complex and four outer antennas that are assembled as two dimers, Lhca1/4 and Lhca2/3. Time-resolved fluorescence measurements on the isolated dimers show very similar kinetics. The intermonomer transfer processes are resolved using target analysis. They occur at rates similar to those observed in transfer to the PSI core, suggesting competition between the two transfer pathways. It appears that each dimer is adopting various conformations that correspond to different lifetimes and emission spectra. A special feature of the Lhca complexes is the presence of an absorption band at low energy, originating from an excitonic state of a chlorophyll dimer, mixed with a charge-transfer state. These low-energy bands have high oscillator strengths and they are superradiant in both Lhca1/4 and Lhca2/3. This challenges the view that the low-energy charge-transfer state always functions as a quencher in plant Lhc's and it also challenges previous interpretations of PSI kinetics. The very similar properties of the low-energy states of both dimers indicate that the organization of the involved chlorophylls should also be similar, in disagreement with the available structural data.

  6. Solar photocatalytic H{sub 2} production from water using a dual bed photosystem

    Energy Technology Data Exchange (ETDEWEB)

    Linkous, C.A. [Florida Solar Energy Center, Cocoa, FL (United States)

    1996-10-01

    The authors are developing a dual stage, direct photoconversion scheme for water splitting. The overall system consists of an aqueous solution circulated through two modules, or beds, each of which contains a photochemically active surface layer below the solution. The idea is to divide the energy requirement for water decomposition between the two photosystems, so that more abundant, lower energy photons in the solar spectrum can be utilized, and that the H{sub 2} and O{sub 2} products can be evolved separately from each other. Catalyst-modified semiconductor powders, immobilized within a polymer binder are currently being employed as the photoactive layers. TiO{sub 2} and platinized-InP have been employed for the O{sub 2} and H{sub 2} evolution tasks, respectively. A major effort over the last year was identification of a suitable redox mediator that is responsible for transferring electron equivalents from one unit to the other. After testing more than a dozen candidates, spanning a wide range of electropotentials, under a variety of conditions, it was found that the bromide/bromate (Br{sup {minus}}/BrO{sub 3}{sup {minus}}) and iodide/iodate (I{sup {minus}}/IO{sub 3}{sup {minus}}) redox systems could function in both modules, necessary for closed cycle operation.

  7. Molecular functions of PsbP and PsbQ proteins in the photosystem II supercomplex.

    Science.gov (United States)

    Ifuku, Kentaro; Ido, Kunio; Sato, Fumihiko

    2011-01-01

    The PsbP and PsbQ proteins are extrinsic subunits of the photosystem II (PSII) supercomplex, which are found in green plants including higher plants and green algae. These proteins are thought to have evolved from their cyanobacterial homologs; cyanoP and cyanoQ respectively. It has been suggested that the functions of PsbP and PsbQ have largely changed from those of cyanoP and cyanoQ. In addition, multiple isoforms and homologs of PsbP and PsbQ were found in green plants, indicating that the acquisition of PsbP and PsbQ in PSII is not a direct path but a result of intensive functional divergence during evolution from cyanobacterial endosymbiont to chloroplast. In this review, we highlight newly introduced topics related to the functions and structures of both PsbP and PsbQ proteins. The present data suggest that PsbP together with PsbQ have specific and important roles in coordinating the activity of the donor and acceptor sides of PSII and stabilizing the active form of the PSII-light-harvesting complex II (LHCII) supercomplex.

  8. Acute and additive toxicity of ten photosystem-II herbicides to seagrass.

    Science.gov (United States)

    Wilkinson, Adam D; Collier, Catherine J; Flores, Florita; Negri, Andrew P

    2015-11-30

    Photosystem II herbicides are transported to inshore marine waters, including those of the Great Barrier Reef, and are usually detected in complex mixtures. These herbicides inhibit photosynthesis, which can deplete energy reserves and reduce growth in seagrass, but the toxicity of some of these herbicides to seagrass is unknown and combined effects of multiple herbicides on seagrass has not been tested. Here we assessed the acute phytotoxicity of 10 PSII herbicides to the seagrass Halophila ovalis over 24 and/or 48 h. Individual herbicides exhibited a broad range of toxicities with inhibition of photosynthetic activity (∆F/F(m)') by 50% at concentrations ranging from 3.5 μg l(-1) (ametryn) to 132 μg l(-1) (fluometuron). We assessed potential additivity using the Concentration Addition model of joint action for binary mixtures of diuron and atrazine as well as complex mixtures of all 10 herbicides. The effects of both mixture types were largely additive, validating the application of additive effects models for calculating the risk posed by multiple PSII herbicides to seagrasses. This study extends seagrass ecotoxicological data to ametryn, metribuzin, bromacil, prometryn and fluometuron and demonstrates that low concentrations of PSII herbicide mixtures have the potential to impact ecologically relevant endpoints in seagrass, including ∆F/F(m)'.

  9. Unequal allocation of excitation energy between photosystem II and I reduces cyanolichen photosynthesis in blue light.

    Science.gov (United States)

    Solhaug, Knut Asbjørn; Xie, Li; Gauslaa, Yngvar

    2014-08-01

    Photosynthesis was compared in two cyanobacterial lichens (Lobaria hallii and Peltigera praetextata) and two green algal lichens (Lobaria pulmonaria and Peltigera leucophlebia) exposed to red, green or blue light. Cyanolichens had substantially lower photosynthetic CO(2) uptake and O(2) evolution than the green algal lichens in blue light, but slightly higher photosynthesis in red and green light. The effective quantum yield of photosystem (PS) II (Φ(PSII)) decreased with increasing red and green light for all species, but in blue light this response occurred in green algal lichens only. Cyanolichen Φ(PSII) increased with increasing blue light at low irradiances, but decreased at stronger exposures. However, after adding red light the efficiency of blue light for photosynthetic O(2) evolution increased by 2.4 times. Because phycobilisomes associated with PSII have a low blue light absorption, our results are consistent with blue light absorption mainly by Chl in PSI. Thereby, unequal allocation of excitation energy between PSII and PSI results in low cyanolichen photosynthesis under blue light. This is new knowledge in the science of lichenology with important implications for e.g. the reliability of using Chl fluorometers with blue light for cyanolichens.

  10. Nanostructured photosystems for the generation of electricity and fuels from sunlight

    Science.gov (United States)

    Grätzel, Michael

    2013-03-01

    Learning from the concepts used by green plants photosynthesis, we have developed nanostructured systems affording efficient solar light harvesting and conversion to electricity and fuels. Solar cells using dyes or semiconducting nanoparticles as light harvesters supported by mesoscopic oxide films have emerged as credible contenders to conventional p-n junction photovoltaic devices. Separating light absorption from charge carrier transport dye sensitized mesoscopic solar cells (DSCs) were the first to use a three-dimensional nanocrystalline junction for solar electricity production. The standard AM 1.5 solar to electric power conversion efficiency (PCE) has reached 12.9% for laboratory cells and 9.9% for PV modules. Even higher efficiencies are attained under ambient and indoor light conditions. These features along with excellent long-term stability have fostered first commercial applications, the industrial production of DSC's attaining presently the MW/year scale. Very exciting results have recently been obtained with perovskites and quantum dot particles as light harvesters in mesoscopic solar cells. Striking advances in the direct generation of fuels such as hydrogen from water and sunlight have been achieved by the judicious design of photosystems composed of nanostructured Fe2O3 or Cu2O films.

  11. The Psb32 Protein Aids in Repairing Photodamaged Photosystem Ⅱ in the Cyanobacterium Synechocystis 6803

    Institute of Scientific and Technical Information of China (English)

    Kimberly M. Wegener; Stefan Bennewitz; Ralf Oelmüller; Himadri B. Pakrasi

    2011-01-01

    Photosystem Ⅱ (PSⅡ),a membrane protein complex,catalyzes the photochemical oxidation of water to molecular oxygen.This enzyme complex consists of approximately 20 stoichiometric protein components.However,due to the highly energetic reactions it catalyzes as part of its normal activity,PSⅡ is continuously damaged and repaired.With advances in protein detection technologies,an increasing number of sub-stoichiometric PSⅡ proteins have been identified,many of which aid in the biogenesis and assembly of this protein complex.Psb32 (SⅡ1390) has previously been identified as a protein associated with highly active purified PSⅡ preparations from the cyanobacterium Synechocystis sp.PCC 6803.To investigate its function,the subcellular localization of Psb32 and the impact of deletion of the psb32 gene on PSⅡ were analyzed.Here,we show that Psb32 is an integral membrane protein,primarily located in the thylakoid membranes.Although not required for cell viability,Psb32 protects cells from oxidative stress and additionally confers a selective fitness advantage in mixed culture experiments.Specifically,Psb32 protects PSⅡ from photodamage and accelerates its repair.Thus,the data suggest that Psb32 plays an important role in minimizing the effect of photoinhibition on PSⅡ.

  12. Effect of Temperature and light intensity on growth and Photosynthetic Activity of Chlamydomonas reinhard II; Efecto de la temperatura e intensidad luminosa sobre el crecimiento y actividad fotosintetica del alga Chlamydomonas Reinhardt II

    Energy Technology Data Exchange (ETDEWEB)

    Alfonsel Jaen, M.; Fernandez Gonzalez, J.

    1985-07-01

    The effect of five temperatures (15,20,25,30 and 35 degree centigree) and two levels of illumination on growth and photosynthetic activity of Chlamydomonas reinhard II has been studied. The growth of the cultures was evaluated by optical density. Photosynthetic activity has been carried out studying either the assimilation rate of C0{sub 2} labelled with C-14 or the oxygen evolution by means of polarographic measurements. The maximum photosynthetic rate has been obtained at 25 degree centigree for the lower level of illumination (2400 lux) and at 35 degree centigree for the higher one (13200 lux) and at 35 degree centigree for the higher ono (13200 lux). These results suggest an interaction of temperature and illumination on photosynthetic activity. (Author) 37 refs.

  13. Why does Colombia lack agricultural commodity futures?

    Directory of Open Access Journals (Sweden)

    Pablo Moreno-Alemay

    2015-11-01

    Full Text Available This article explores the reasons why futures contracts are not traded as an alternative to price hedging for agricultural goods in Colombia. Based on surveys, interviews and statistical analysis, this study identified that conceptual gaps in contract negotiation, lack of consensus in the agricultural sector regarding the use of financial mechanisms and the sector’s infrequent contact with Colombia’s financial institutions, are the main reasons why a futures contracts market has not emerged.

  14. Influence of Operating Conditions on the Removal Cd Ions from Aqueous Media by Adsorption Using Chlamydomonas Reinhardtii

    Institute of Scientific and Technical Information of China (English)

    Jiang Yongbin; Zhu Yi; Ji Hongbing

    2010-01-01

    Chlamydomonas reinhardtii(C.reinhardtii)was used to study adsorption of cadmium(Cd)from aqueous media within various experimental conditions.Results showed that the adsorption process was very fast,with most of the adsorption occurring within30 min of contact time and the equilibrium state was reached in about 60 min.The adsorption ability of the algae increases with the increasing adsorptions sites on cells.Maximum adsorption was observed at the initial Cd concentration of 100 mg/L and pH 6.0.The adsorption was markedly inhibited in the presence of calcium and magnesium ions at 10 mM and the Cd removal efficiency was reduced by 16.54% and 14.99% respectively.This study would be a finding of note with regard to practical wastewater treatment.

  15. Transcriptomic and Physiological Responses of the Green Microalga Chlamydomonas reinhardtii during Short-Term Exposure to Subnanomolar Methylmercury Concentrations.

    Science.gov (United States)

    Beauvais-Flück, Rebecca; Slaveykova, Vera I; Cosio, Claudia

    2016-07-05

    The effects of short-term exposure to subnanomolar methyl-mercury (MeHg) concentrations, representative of contaminated environments, on the microalga Chlamydomonas reinhardtii were assessed using both physiological end points and gene expression analysis. MeHg bioaccumulated and induced significant increase of the photosynthesis efficiency, while the algal growth, oxidative stress, and chlorophyll fluorescence were unaffected. At the molecular level, MeHg significantly dysregulated the expression of genes involved in motility, energy metabolism, lipid metabolism, metal transport, and antioxidant enzymes. Data suggest that the cells were able to cope with subnanomolar MeHg exposure, but this tolerance resulted in a significant cost to the cell energy and reserve metabolism as well as ample changes in the nutrition and motility of C. reinhardtii. The present results allowed gaining new insights on the effects and uptake mechanisms of MeHg at subnanomolar concentrations in aquatic primary producers.

  16. Protein-Protein Interactions between Intermediate Chains and the Docking Complex of Chlamydomonas Flagellar Outer Arm Dynein

    Science.gov (United States)

    Ide, Takahiro; Owa, Mikito; King, Stephen M.; Kamiya, Ritsu; Wakabayashi, Ken-ichi

    2013-01-01

    Outer arm dynein (OAD) is bound to specific loci on outer-doublet-microtubules by interactions at two sites: via intermediate chain 1 (IC1) and the outer dynein arm docking complex (ODA-DC). Studies using Chlamydomonas mutants have suggested that the individual sites have rather weak affinities for microtubules, and therefore strong OAD attachment to microtubules is achieved by their cooperation. To test this idea, we examined interactions between IC1, IC2 (another intermediate chain) and ODA-DC using recombinant proteins. Recombinant IC1 and IC2 were found to form a 1:1 complex, and this complex associated with ODA-DC in vitro. Binding of IC1 to mutant axonemes revealed that there are specific binding sites for IC1. From these data, we propose a novel model of OAD-outer doublet association. PMID:23747306

  17. Quantum preparation uncertainty and lack of information

    Science.gov (United States)

    Rozpędek, Filip; Kaniewski, Jędrzej; Coles, Patrick J.; Wehner, Stephanie

    2017-02-01

    The quantum uncertainty principle famously predicts that there exist measurements that are inherently incompatible, in the sense that their outcomes cannot be predicted simultaneously. In contrast, no such uncertainty exists in the classical domain, where all uncertainty results from ignorance about the exact state of the physical system. Here, we critically examine the concept of preparation uncertainty and ask whether similarly in the quantum regime, some of the uncertainty that we observe can actually also be understood as a lack of information (LOI), albeit a lack of quantum information. We answer this question affirmatively by showing that for the well known measurements employed in BB84 quantum key distribution (Bennett and Brassard 1984 Int. Conf. on Computer System and Signal Processing), the amount of uncertainty can indeed be related to the amount of available information about additional registers determining the choice of the measurement. We proceed to show that also for other measurements the amount of uncertainty is in part connected to a LOI. Finally, we discuss the conceptual implications of our observation to the security of cryptographic protocols that make use of BB84 states.

  18. A C2H2 zinc finger protein FEMU2 is required for fox1 expression in Chlamydomonas reinhardtii.

    Directory of Open Access Journals (Sweden)

    Xiaodong Deng

    Full Text Available Chlamydomonas reinhardtii fox1 gene encodes a ferroxidase that is involved in cellular Fe uptake and highly induced during Fe deficient conditions. In an effort to identify fox1 promoter regulatory elements, an insertional library was generated in a transgenic Chlamydomonas strain (2A38 harboring an arylsulfatase (ARS reporter gene driven by the fox1 promoter. Mutants with a defective response to low iron conditions were selected for further study. Among these, a strain containing a disrupted femu2 gene was identified. Activation of the fox1 promoter by the femu2 gene product was confirmed by silencing the femu2 gene using RNA interference. In three femu2 RNAi transgenic lines (IR3, IR6, and IR7, ARS reporter gene activities declined by 84.3%, 86.4%, and 88.8%, respectively under Fe deficient conditions. Furthermore, RT-PCR analysis of both the femu2 mutant and the RNAi transgenic lines showed significantly decreased transcript abundance of the endogenous fox1 gene under Fe deficient conditions. Amino acid sequence analysis of the femu2 gene product identified three potential C2H2 zinc finger (ZF motifs and a nuclear localization study suggests that FEMU2 is localized to the nucleus. In addition, a potential FEMU2 binding site ((G/TTTGG(G/T(G/TT was identified using PCR-mediated random binding site selection. Taken together, this evidence suggests that FEMU2 is involved in up-regulation of the fox1 gene in Fe deficient cells.

  19. A fluorescence-activated cell sorting-based strategy for rapid isolation of high-lipid Chlamydomonas mutants.

    Science.gov (United States)

    Terashima, Mia; Freeman, Elizabeth S; Jinkerson, Robert E; Jonikas, Martin C

    2015-01-01

    There is significant interest in farming algae for the direct production of biofuels and valuable lipids. Chlamydomonas reinhardtii is the leading model system for studying lipid metabolism in green algae, but current methods for isolating mutants of this organism with a perturbed lipid content are slow and tedious. Here, we present the Chlamydomonas high-lipid sorting (CHiLiS) strategy, which enables enrichment of high-lipid mutants by fluorescence-activated cell sorting (FACS) of pooled mutants stained with the lipid-sensitive dye Nile Red. This method only takes 5 weeks from mutagenesis to mutant isolation. We developed a staining protocol that allows quantification of lipid content while preserving cell viability. We improved separation of high-lipid mutants from the wild type by using each cell's chlorophyll fluorescence as an internal control. We initially demonstrated 20-fold enrichment of the known high-lipid mutant sta1 from a mixture of sta1 and wild-type cells. We then applied CHiLiS to sort thousands of high-lipid cells from a pool of about 60,000 mutants. Flow cytometry analysis of 24 individual mutants isolated by this approach revealed that about 50% showed a reproducible high-lipid phenotype. We further characterized nine of the mutants with the highest lipid content by flame ionization detection and mass spectrometry lipidomics. All mutants analyzed had a higher triacylglycerol content and perturbed whole-cell fatty acid composition. One arbitrarily chosen mutant was evaluated by microscopy, revealing larger lipid droplets than the wild type. The unprecedented throughput of CHiLiS opens the door to a systems-level understanding of green algal lipid biology by enabling genome-saturating isolation of mutants in key genes.

  20. Nickel and low CO2-controlled motility in Chlamydomonas through complementation of a paralyzed flagella mutant with chemically regulated promoters

    Directory of Open Access Journals (Sweden)

    Rosenbaum Joel L

    2011-01-01

    Full Text Available Abstract Background Chlamydomonas reinhardtii is a model system for the biology of unicellular green algae. Chemically regulated promoters, such as the nickel-inducible CYC6 or the low CO2-inducible CAH1 promoter, may prove useful for expressing, at precise times during its cell cycle, proteins with relevant biological functions, or complementing mutants in genes encoding such proteins. To this date, this has not been reported for the above promoters. Results We fused the CYC6 and CAH1 promoters to an HA-tagged RSP3 gene, encoding a protein of the flagellar radial spoke complex. The constructs were used for chemically regulated complementation of the pf14 mutant, carrying an ochre mutation in the RSP3 gene. 7 to 8% of the transformants showed cells with restored motility after induction with nickel or transfer to low CO2 conditions, but not in non-inducing conditions. Maximum complementation (5% motile cells was reached with very different kinetics (5-6 hours for CAH1, 48 hours for CYC6. The two inducible promoters drive much lower levels of RSP3 protein expression than the constitutive PSAD promoter, which shows almost complete rescue of motility. Conclusions To our knowledge, this is the first example of the use of the CYC6 or CAH1 promoters to perform a chemically regulated complementation of a Chlamydomonas mutant. Based on our data, the CYC6 and CAH1 promoters should be capable of fully complementing mutants in genes whose products exert their biological activity at low concentrations.

  1. Shewanella oneidensis: a new and efficient System for Expression and Maturation of heterologous [Fe-Fe] Hydrogenase from Chlamydomonas reinhardtii

    Directory of Open Access Journals (Sweden)

    Sybirna Kateryna

    2008-09-01

    Full Text Available Abstract Background The eukaryotic green alga, Chlamydomonas reinhardtii, produces H2 under anaerobic conditions, in a reaction catalysed by a [Fe-Fe] hydrogenase HydA1. For further biochemical and biophysical studies a suitable expression system of this enzyme should be found to overcome its weak expression in the host organism. Two heterologous expression systems used up to now have several advantages. However they are not free from some drawbacks. In this work we use bacterium Shewanella oneidensis as a new and efficient system for expression and maturation of HydA1 from Chlamydomonas reinhardtii. Results Based on codon usage bias and hydrogenase maturation ability, the bacterium S. oneidensis, which possesses putative [Fe-Fe] and [Ni-Fe] hydrogenase operons, was selected as the best potential host for C. reinhardtii [Fe-Fe] hydrogenase expression. Hydrogen formation by S. oneidensis strain AS52 (ΔhydAΔhyaB transformed with a plasmid bearing CrHydA1 and grown in the presence of six different substrates for anaerobic respiration was determined. A significant increase in hydrogen evolution was observed for cells grown in the presence of trimethylamine oxide, dimethylsulfoxide and disodium thiosulfate, showing that the system of S. oneidensis is efficient for heterologous expression of algal [Fe-Fe] hydrogenase. Conclusion In the present work a new efficient system for heterologous expression and maturation of C. reinhardtii hydrogenase has been developed. HydA1 of C. reinhardtii was purified and shown to contain 6 Fe atoms/molecule of protein, as expected. Using DMSO, TMAO or thiosulfate as substrates for anaerobic respiration during the cell growth, 0.4 – 0.5 mg l-1(OD600 = 1 of catalytically active HydA1 was obtained with hydrogen evolution rate of ~700 μmol H2 mg-1 min-1.

  2. Acute effects of a prooxidant herbicide on the microalga Chlamydomonas reinhardtii: Screening cytotoxicity and genotoxicity endpoints

    Energy Technology Data Exchange (ETDEWEB)

    Esperanza, Marta; Cid, Ángeles; Herrero, Concepción; Rioboo, Carmen, E-mail: carmen.rioboo@udc.es

    2015-08-15

    Highlights: • Mitochondrial membrane potential constituted the most sensitive parameter assayed. • Several genotoxicity methods were applied for first time in ecotoxicological studies. • Oxidative DNA base damage (8-OHdG) was induced by paraquat exposure. • Cells with DNA strand breakage and subG1-nuclei increased in treated cultures. • Typical apoptosis hallmarks were observed in microalgal cells exposed to paraquat. - Abstract: Since recent evidence has demonstrated that many types of chemicals exhibit oxidative and/or genotoxic potential on living organisms, reactive oxygen species (ROS) formation and DNA damage are currently the best accepted paradigms to assess the potential hazardous biological effects of a wide range of contaminants. The goal of this study was to evaluate the sensitivity of different cytotoxicity and genotoxicity responses on the model microalga Chlamydomonas reinhardtii exposed to the prooxidant herbicide paraquat. In addition to the growth endpoint, cell viability, mitochondrial membrane potential and presence of reactive oxygen species (ROS) were assayed as potential markers of cytotoxicity using flow cytometry (FCM). To study the effects of paraquat on C. reinhardtii DNA, several genotoxicity approaches were implemented for the first time in an ecotoxicological study on microalgae. Oxidative DNA base damage was analysed by measuring the oxidative DNA lesion 8-OHdG by FCM. DNA fragmentation was analysed by different methods: comet assay, and cell cycle analysis by FCM, with a particular focus on the presence of subG1-nuclei. Finally, effects on morphology of nuclei were monitored through DAPI staining. The evaluation of these endpoints showed that several physiological and biochemical parameters reacted to oxidative stress disturbances with greater sensitivity than integrative parameters such as growth rates or cell viability. The experiments revealed concentration-dependent cytotoxicity (ROS formation, depolarization of

  3. Two photon absorption energy transfer in the light-harvesting complex of photosystem II (LHC-II) modified with organic boron dye.

    Science.gov (United States)

    Chen, Li; Liu, Cheng; Hu, Rui; Feng, Jiao; Wang, Shuangqing; Li, Shayu; Yang, Chunhong; Yang, Guoqiang

    2014-07-15

    The plant light-harvesting complexes of photosystem II (LHC-II) play important roles in collecting solar energy and transferring the energy to the reaction centers of photosystems I and II. A two photon absorption compound, 4-(bromomethyl)-N-(4-(dimesitylboryl)phenyl)-N-phenylaniline (DMDP-CH2Br), was synthesized and covalently linked to the LHC-II in formation of a LHC-II-dye complex, which still maintained the biological activity of LHC-II system. Under irradiation with femtosecond laser pulses at 754 nm, the LHC-II-dye complex can absorb two photons of the laser light effectively compared with the wild type LHC-II. The absorbed excitation energy is then transferred to chlorophyll a with an obvious fluorescence enhancement. The results may be interesting and give potentials for developing hybrid photosystems.

  4. Mice lacking neurofibromin develop gastric hyperplasia

    Science.gov (United States)

    Lin, Lu; Chen, Jian; Richardson, James A.

    2009-01-01

    Gastrointestinal (GI) neoplasms are among many manifestations of the genetic disease neurofibromatosis type 1 (NF1). However, the physiological and pathological functions of the Nf1 gene in the GI system have not been fully studied, possibly because of a lack of mouse models. In this study, we generated conditional knockout mice with Nf1 deficiency in the GI tract. These mice develop gastric epithelial hyperplasia and inflammation together with increased cell proliferation and apoptosis. The gastric phenotypes observed in these mutant mice seem to be the consequence of loss of Nf1 in gastric fibroblasts, resulting in paracrine hyperactivation of the ERK pathway in the gastric epithelium. These mice provide a useful model to study the pathogenesis of GI lesions in a subset of patients with NF1 and to investigate the role of the Nf1 gene in the development of GI neoplasms. PMID:19661150

  5. Lack of transplacental transmission of Bartonella bovis.

    Science.gov (United States)

    Chastant-Maillard, S; Boulouis, H-J; Reynaud, K; Thoumire, S; Gandoin, C; Bouillin, C; Cordonnier, N; Maillard, R

    2015-02-01

    Transplacental transmission of Bartonella spp. has been reported for rodents, but not for cats and has never been investigated in cattle. The objective of this study was to assess vertical transmission of Bartonella in cattle. Fifty-six cow-calf pairs were tested before (cows) and after (calves) caesarean section for Bartonella bacteremia and/or serology, and the cotyledons were checked for gross lesions and presence of the bacteria. None of the 29 (52%) bacteremic cows gave birth to bacteremic calves, and all calves were seronegative at birth. Neither placentitis nor vasculitis were observed in all collected cotyledons. Bartonella bovis was not detected in placental cotyledons. Therefore, transplacental transmission of B. bovis and multiplication of the bacteria in the placenta do not seem likely. The lack of transplacental transmission may be associated with the particular structure of the placenta in ruminants or to a poor affinity/agressiveness of B. bovis for this tissue.

  6. Light saturation response of inactive photosystem II reaction centers in spinach.

    Science.gov (United States)

    Chylla, R A; Whitmarsh, J

    1990-07-01

    The effective absorption cross section of inactive photosystem II (PS II) centers, which is the product of the effective antenna size and the quantum yield for photochemistry, was investigated by comparing the light saturation curves of inactive PS II and active reaction centers in intact chloroplasts and thylakoid membranes of spinach (Spinacia oleracea). Inactive PS II centers are defined as the impaired PS II reaction centers that require greater than 50 ms for the reoxidation of QA (-) subsequent to a single turnover flash. Active reaction centers are defined as the rapidly turning over PS II centers (recovery time less than 50 ms) and all of the PS I centers. The electrochromic shift, measured by the flash-induced absorbance increase at 518 nm, was used to probe the activity of the reaction centers. Light saturation curves were generated for inactive PS II centers and active reaction centers by measuring the extent of the absorbance increase at 518 nm induced by red actinic flashes of variable energy. The light saturation curves show that inactive PS II centers required over twice as many photons as active reaction centers to achieve the same yield. The ratio of the flash energy required for 50% saturation for active reaction centers (PS II active + PS I) compared to inactive PS II centers was 0.45±0.04 in intact chloroplasts, and 0.54±0.11 in thylakoid membranes. Analysis of the light saturation curves using a Poisson statistical model in which the ratio of the antenna size of active PS II centers to that of PS I is considered to range from 1 to 1.5, indicates that the effective absorption cross section of inactive PS II centers was 0.54-0.37 times that of active PS II centers. If the quantum yield for photochemistry is assumed to be one, we estimate that the antenna system serving the inactive PS II centers contains approx. 110 chlorophyll molecules.

  7. Fluorescence property of photosystem II protein complexes bound to a gold nanoparticle.

    Science.gov (United States)

    Tahara, Kazuki; Mohamed, Ahmed; Kawahara, Kousuke; Nagao, Ryo; Kato, Yuki; Fukumura, Hiroshi; Shibata, Yutaka; Noguchi, Takumi

    2017-03-08

    Development of an efficient photo-anode system for water oxidation is key to the success of artificial photosynthesis. We previously assembled photosystem II (PSII) proteins, which are an efficient natural photocatalyst for water oxidation, on a gold nanoparticle (GNP) to prepare a PSII-GNP conjugate as an anode system in a light-driven water-splitting nano-device (Noji et al., J. Phys. Chem. Lett., 2011, 2, 2448-2452). In the current study, we characterized the fluorescence property of the PSII-GNP conjugate by static and time-resolved fluorescence measurements, and compared with that of free PSII proteins. It was shown that in a static fluorescence spectrum measured at 77 K, the amplitude of a major peak at 683 nm was significantly reduced and a red shoulder at 693 nm disappeared in PSII-GNP. Time-resolved fluorescence measurements showed that picosecond components at 683 nm decayed faster by factors of 1.4-2.1 in PSII-GNP than in free PSII, explaining the observed quenching of the major fluorescence peak. In addition, a nanosecond-decay component arising from a 'red chlorophyll' at 693 nm was lost in time-resolved fluorescence of PSII-GNP, probably due to a structural perturbation of this chlorophyll by interaction with GNP. Consistently with these fluorescence properties, degradation of PSII during strong-light illumination was two times slower in PSII-GNP than in free PSII. The enhanced durability of PSII is an advantageous property of the PSII-GNP conjugate in the development of an artificial photosynthesis device.

  8. The response of cyclic electron flow around photosystem I to changes in photorespiration and nitrate assimilation.

    Science.gov (United States)

    Walker, Berkley J; Strand, Deserah D; Kramer, David M; Cousins, Asaph B

    2014-05-01

    Photosynthesis captures light energy to produce ATP and NADPH. These molecules are consumed in the conversion of CO2 to sugar, photorespiration, and NO3(-) assimilation. The production and consumption of ATP and NADPH must be balanced to prevent photoinhibition or photodamage. This balancing may occur via cyclic electron flow around photosystem I (CEF), which increases ATP/NADPH production during photosynthetic electron transport; however, it is not clear under what conditions CEF changes with ATP/NADPH demand. Measurements of chlorophyll fluorescence and dark interval relaxation kinetics were used to determine the contribution of CEF in balancing ATP/NADPH in hydroponically grown Arabidopsis (Arabidopsis thaliana) supplied different forms of nitrogen (nitrate versus ammonium) under changes in atmospheric CO2 and oxygen. Measurements of CEF were made under low and high light and compared with ATP/NADPH demand estimated from CO2 gas exchange. Under low light, contributions of CEF did not shift despite an up to 17% change in modeled ATP/NADPH demand. Under high light, CEF increased under photorespiratory conditions (high oxygen and low CO2), consistent with a primary role in energy balancing. However, nitrogen form had little impact on rates of CEF under high or low light. We conclude that, according to modeled ATP/NADPH demand, CEF responded to energy demand under high light but not low light. These findings suggest that other mechanisms, such as the malate valve and the Mehler reaction, were able to maintain energy balance when electron flow was low but that CEF was required under higher flow.

  9. Two roles of thylakoid lipids in modifying the activity of herbicides which inhibit photosystem II

    Energy Technology Data Exchange (ETDEWEB)

    Kupatt, C.C. Jr.

    1985-01-01

    Thylakoid lipids may modify the activity of herbicides which inhibit electron transport at the Q/sub B/ protein of photosystem II in two ways: (1) lipids can act as a hydrophobic barrier to a binding site localized close to the loculus of the membrane, and (2) changes in lipid composition can reduce the ability of inhibitors to block electron transport, possibly due to a change in the conformation of the Q/sub B/ protein. The herbicide binding site was localized close to the locular side of the thylakoid membrane by determining the activity of a number of substituted phenylurea and s-triazine herbicides in inverted and non-inverted thylakoids. Quantitative structure-activity relationship analysis showed that inversion of thylakoids reduced the requirement of molecular lipophilicity deemed necessary for phenylurea activity in non-inverted membranes, whereas s-triazines exhibited no differences in the lipophilicity requirement in thylakoid membranes of either orientation. The binding affinity of /sup 14/C-diuron was reduced in bicarbonate-depleted thylakoids relative to reconstituted or control membranes, as is the case with atrazine binding. These observations support a model of the herbicide binding site containing both common and herbicide family specific binding domains. Thylakoids isolated either from detached lambs quarters (Chenopodium album L.) leaves, treated with SAN 6706, or from soybean (Glycine max L.), with norflurazon or pyrazon applied preemergence, exhibited decreased susceptibility to atrazine. The ability of lipid-modifying treatments to decrease the atrazine susceptibility of field-grown soybeans was also investigated.

  10. The role of calcium in the oxygen evolving center of photosystem II

    Energy Technology Data Exchange (ETDEWEB)

    Latimer, M.J.

    1995-05-01

    The photosynthetic oxygen evolving complex (OEC) contains a cluster of four manganese atoms and requires both Ca and Cl for activity. Ca can be replaced by Sr with retention of activity. The role of Ca in the OEC has been investigated by performing Mn X-ray absorption experiments on Ca-depleted samples of photosystem II (PS II) and on PS II samples depleted of Ca and reconstituted by either Ca or Sr. Mn X-ray K-edge spectra exhibit no significant differences in oxidation state or symmetry between Ca- and Sr-reactivated preparations, but differences are observed in the extended X-ray absorption fine structure (EXAFS). The amplitude of a Fourier transform peak arising from scatterers at distances greater than 3 A is larger for samples reactivated with strontium relative to calcium. Curve-fitting analyses of the EXAFS data using FEFF 5-calculated parameters favor a model where both manganese and calcium (or strontium) scatterers contribute to the {approximately}3 {Angstrom} Fourier peak (Mn-Mn at 3.3{Angstrom} and Mn-Ca(Sr) at 3.4--3.5 {Angstrom}). Possible structural arrangements for a calcium binding site are discussed. Analysis of Mn K-edge spectra from Ca-depleted samples in the S{sub 1}, S{sub 2}, and S{sub 3} states shows an edge shift on the S{sub 1}-S{sub 2} transition, but no edge shift on the S{sub 2}-S{sub 3} transition, supporting a model where the oxidizing equivalent from the S{sub 2} to S{sub 3} transition is stored on a ligand or nearby protein residue rather than on the Mn cluster. Parallels between Ca-depleted and native samples are discussed.

  11. Detergent-protein interactions in aqueous buffer suspensions of Photosystem I (PS I).

    Science.gov (United States)

    Mukherjee, Dibyendu; May, Mark; Khomami, Bamin

    2011-06-15

    Systematic and uniform monolayer formation of Photosystem I (PS I) onto self-assembled monolayer (SAM) substrates to enable unidirectional electron transfer is crucial for its successful use in the fabrication of bio-hybrid solid-state electronic or photovoltaic devices. Yet, our recent studies (Mukherjee et al., 2010) indicate that surface self-assembly of PS I from aqueous buffer suspensions onto alkanethiolate SAM/Au substrates frequently leads to complex columnar structures due to solution phase protein aggregations. We investigate the effect of two prototypical non-ionic detergents, n-Dodecyl-β-D-Maltoside (DM) and Triton X-100 (TX-100), on protein-protein interactions via the protein-detergent interfacial chemistry. Dynamic light scattering (DLS) experiments are used to demonstrate the impact of relative protein/detergent concentrations on aggregation dynamics of PS I suspensions. In turn, the surface attachment characteristics of PS I adsorbed from the aforementioned suspensions onto SAM/Au substrate is examined by atomic force (AFM) microscopy. Our results indicate that relative concentration of PS I and detergents (DM or, TX-100) with respect to their critical micelle concentrations (CMC) determines the extent of self-association between PS I complexes driven by the screening induced by detergent micelles and/or, inter-protein distances. Such interfacial phenomena during the PS I-detergent complexation process drives the colloidal system through various regimes of phase separations, suspension and/or, de-aggregation, wherein individual PS I complexes can exist in a frustrated state that prevents favorable orientations for PS I-PS I interactions. The present study presents a novel strategy, heretofore not considered, for tailoring inter-protein distances and protein-protein interactions in solution phase, thereby allowing a superior control on the surface attachment of PS I onto SAM/Au substrates.

  12. Azide as a competitor of chloride in oxygen evolution by Photosystem II.

    Science.gov (United States)

    Haddy, A; Hatchell, J A; Kimel, R A; Thomas, R

    1999-05-11

    Oxygen evolution by higher plants requires chloride, which binds to a site associated with the oxygen-evolving complex of photosystem II (PSII). In this study, the inhibitory effect of the anion azide was characterized using steady state measurements of oxygen evolution activity in PSII-enriched thylakoid membranes. N3- (7.8 mM) inhibited O2 evolution activity by 50% when a standard buffer containing chloride was used. By considering Cl- as the substrate in O2 evolution assays, we found azide to be primarily competitive with Cl- with an inhibitor dissociation constant Ki of about 0.6 mM. An uncompetitive component with a Ki ' of 11 mM was also found. Removal of the 17 and 23 kDa polypeptides resulted in a decrease in each inhibition constant. A pH dependence study of O2 evolution activity showed that the pH maximum became narrower and shifted to a higher pH in the presence of azide. Analysis of the data indicated that an acidic residue defined the low side of the pH maximum with an apparent pKa of 6.7 in the presence of azide compared with 5.5 for the control. A basic residue was also affected, exhibiting an apparent pKa of 7.1 compared with a value of 7.6 for the control. This result can be explained by a simple model in which azide binding to the chloride site moves negative charge of the anion away from the basic residue and toward the acidic residue relative to chloride. As a competitor of chloride, azide may provide an interesting probe of the oxygen-evolving complex in future studies.

  13. How does the QB site influence propagate to the QA site in photosystem II?

    Science.gov (United States)

    Ishikita, Hiroshi; Hasegawa, Koji; Noguchi, Takumi

    2011-06-21

    The redox potential of the primary quinone Q(A) [E(m)(Q(A))] in photosystem II (PSII) is lowered by replacement of the native plastoquinone (PQ) with bromoxynil (BR) at the secondary quinone Q(B) binding site. Using the BR-bound PSII structure presented in the previous Fourier transform infrared and docking calculation studies, we calculated E(m)(Q(A)) considering both the protein environment in atomic detail and the protonation pattern of the titratable residues. The calculated E(m)(Q(A)) shift in response to the replacement of PQ with deprotonated BR at the Q(B) binding site [ΔE(m)(Q(A))(PQ→BR)] was -55 mV when the three regions, Q(A), the non-heme iron complex, and Q(B) (Q(B) = PQ or BR), were treated as a conjugated supramolecule (Q(A)-Fe-Q(B)). The negative charge of BR apparently contributes to the downshift in ΔE(m)(Q(A))(PQ→BR). This downshift, however, is mostly offset by the influence of the residues near Q(B). The charge delocalization over the Q(A)-Fe-Q(B) complex and the resulting H-bond strength change between Q(A) and D2-His214 are crucial factors that yield a ΔE(m)(Q(A))(PQ→BR) of -55 mV by (i) altering the electrostatic influence of the H-bond donor D2-His214 on E(m)(Q(A)) and (ii) suppressing the proton uptake events of the titratable residues that could otherwise upshift ΔE(m)(Q(A))(PQ→BR) during replacement of PQ with BR at the Q(B) site.

  14. Manganese containing protein complex isolated from Photosystem II preparations of spinach

    Energy Technology Data Exchange (ETDEWEB)

    Frasch, W.D.; Bowlby, N.R.

    1986-05-01

    By using a ligand-receptor crosslinking method the authors have stabilized Mn associated with Photosystem II (PSII) in a protein complex with an apparent molecular weight of 75,000 and with 3-4 Mn per complex. To crosslink the proteins, purified 33 kDa protein (33) was modified to contain about 8 adducts of the heterobifunctional photoaffinity reagent N-succinimidyl-(4-azidophenyl-dithio)-propionate (SADP). The SADP-33 was reconstituted into PSII membranes which had been depleted of 33 by a 1M CaCl/sub 2/ wash and crosslinking was initiated by ultraviolet illumination. The crosslinked membranes were solubilized in lauryl sulfate (SDS) and the constituent proteins were identified by SDS polyacrylamide gel electrophoresis. Evidence which supports the hypothesis that the Mn associated with the crosslinked proteins has been retained at the site of the photosynthetic oxygen evolving system includes: 1, Scatchard analysis of (/sup 125/I)-33 binding to CaCl/sub 2/-washed PSII preparations revealed one tight binding site and several lower affinity sites; 2, reconstitution of O/sub 2/ evolving activity and binding to the tight site had the same concentration dependence on 33; 3, the SADP-33 was able to reconstitute O/sub 2/ evolution in CaCl/sub 2/ washed PSII membranes; 4, the percent of Mn retained by the crosslinked membranes after treatment with edetic acid (EDTA) was approximately equal to the percent reconstitution of O/sub 2/ evolving activity by SADP-33 before photoactivation of SADP.

  15. Self-assembling peptide detergents stabilize isolated photosystem ion a dry surface for an extended time.

    Directory of Open Access Journals (Sweden)

    2005-07-01

    Full Text Available We used a class of designed peptide detergents to stabilize photosystem I (PS-I upon extended drying under N2 on a gold-coated-Ni-NTA glass surface. PS-I is a chlorophyll-containing membrane protein complex that is the primary reducer of ferredoxin and the electron acceptor of plastocyanin. We isolated the complex from the thylakoids of spinach chloroplasts using a chemical detergent. The chlorophyll molecules associated with the PS-I complex provide an intrinsic steady-state emission spectrum between 650 and 800 nm at -196.15 degrees C that reflects the organization of the pigment-protein interactions. In the absence of detergents, a large blue shift of the fluorescence maxima from approximately 735 nm to approximately 685 nm indicates a disruption in light-harvesting subunit organization, thus revealing chlorophyll-protein interactions. The commonly used membrane protein-stabilizing detergents, N-dodecyl-beta-D-maltoside and N-octyl-beta-D-glucoside, only partially stabilized the approximately 735-nm complex with approximately 685-nm spectroscopic shift. However, prior to drying, addition of the peptide detergent acetyl-AAAAAAK at increasing concentration significantly stabilized the PS-I complex. Moreover, in the presence of acetyl-AAAAAAK, the PS-I complex is stable in a dried form at room temperature for at least 3 wk. Another peptide detergent, acetyl-VVVVVVD, also stabilized the complex but to a lesser extent. These observations suggest that the peptide detergents may effectively stabilize membrane proteins in the solid-state. These designed peptide detergents may facilitate the study of diverse types of membrane proteins.

  16. Effects of polyaromatic hydrocarbons on photosystem II activity in pea leaves.

    Science.gov (United States)

    Kreslavski, Vladimir D; Lankin, Anton V; Vasilyeva, Galina K; Luybimov, Valery Yu; Semenova, Galina N; Schmitt, Franz-Josef; Friedrich, Thomas; Allakhverdiev, Suleyman I

    2014-08-01

    The acute effects of three typical polyaromatic hydrocarbons (PAHs): naphthalene (Naph), phenanthrene (Phen) and fluoranthene (Flu) on photochemical activity of photosystem II (PSII) in detached leaves of 3-week-old pea plants were studied. The leaves were exposed in water with PAHs under white light for 0.5-72 h. The activity of PSII was examined by prompt and delayed chlorophyll a (Chl a) fluorescence. The effects of PAHs depended on their concentration and exposure time. This dependency was more significant in the presence of chemical stressors (Triton X-100 or acetone) or under high intensity irradiance. Increased content of PAHs and long-term exposure (24-72 h) led to significant reduction of the maximum photochemical quantum efficiency (Fv/Fm) of PS II, changes in the polyphasic fluorescence induction (OJIP), and to decreasing amplitudes of fast and slow components of delayed Chl a fluorescence. The damage of PSII depended on water solubility of a given type of PAHs, their concentration and exposure time. During short-time exposure the compound with highest water-solubility - naphthalene - revealed the strongest effect. During long-time exposure the compounds with low water-solubility -Phen, Flu-revealed the strongest effect as the corresponding PAH accumulates in the thylakoids especially when the solution is oversaturated containing a solid phase. The reduction of PSII activity at the presence of naphthalene (30 mg L(-1)) was accompanied by transient generation of H2O2 as well as swelling of thylakoids and distortion of cell plasma membranes, which was indicated by electron microscopy images. Distortion of thylakoid membranes due to accumulation of PAHs as well as the development of oxidative stress seems to be the main pathways of PAHs influencing the photochemical activity of PS II.

  17. Directly probing redox-linked quinones in photosystem II membrane fragments via UV resonance Raman scattering.

    Science.gov (United States)

    Chen, Jun; Yao, Mingdong; Pagba, Cynthia V; Zheng, Yang; Fei, Liping; Feng, Zhaochi; Barry, Bridgette A

    2015-01-01

    In photosynthesis, photosystem II (PSII) harvests sunlight with bound pigments to oxidize water and reduce quinone to quinol, which serves as electron and proton mediators for solar-to-chemical energy conversion. At least two types of quinone cofactors in PSII are redox-linked: QA, and QB. Here, we for the first time apply 257-nm ultraviolet resonance Raman (UVRR) spectroscopy to acquire the molecular vibrations of plastoquinone (PQ) in PSII membranes. Owing to the resonance enhancement effect, the vibrational signal of PQ in PSII membranes is prominent. A strong band at 1661 cm(-1) is assigned to ring CC/CO symmetric stretch mode (ν8a mode) of PQ, and a weak band at 469 cm(-1) to ring stretch mode. By using a pump-probe difference UVRR method and a sample jet technique, the signals of QA and QB can be distinguished. A frequency difference of 1.4 cm(-1) in ν8a vibrational mode between QA and QB is observed, corresponding to ~86 mV redox potential difference imposed by their protein environment. In addition, there are other PQs in the PSII membranes. A negligible anharmonicity effect on their combination band at 2130 cm(-1) suggests that the 'other PQs' are situated in a hydrophobic environment. The detection of the 'other PQs' might be consistent with the view that another functional PQ cofactor (not QA or QB) exists in PSII. This UVRR approach will be useful to the study of quinone molecules in photosynthesis or other biological systems.

  18. Kinetic analyses and mathematical modeling of primary photochemical and photoelectrochemical processes in plant photosystems.

    Science.gov (United States)

    Vredenberg, Wim

    2011-02-01

    In this paper the model and simulation of primary photochemical and photo-electrochemical reactions in dark-adapted intact plant leaves is presented. A descriptive algorithm has been derived from analyses of variable chlorophyll a fluorescence and P700 oxidation kinetics upon excitation with multi-turnover pulses (MTFs) of variable intensity and duration. These analyses have led to definition and formulation of rate equations that describe the sequence of primary linear electron transfer (LET) steps in photosystem II (PSII) and of cyclic electron transport (CET) in PSI. The model considers heterogeneity in PSII reaction centers (RCs) associated with the S-states of the OEC and incorporates in a dark-adapted state the presence of a 15-35% fraction of Q(B)-nonreducing RCs that probably is identical with the S₀ fraction. The fluorescence induction algorithm (FIA) in the 10 μs-1s excitation time range considers a photochemical O-J-D, a photo-electrochemical J-I and an I-P phase reflecting the response of the variable fluorescence to the electric trans-thylakoid potential generated by the proton pump fuelled by CET in PSI. The photochemical phase incorporates the kinetics associated with the double reduction of the acceptor pair of pheophytin (Phe) and plastoquinone Q(A) [PheQ(A)] in Q(B) nonreducing RCs and the associated doubling of the variable fluorescence, in agreement with the three-state trapping model (TSTM) of PS II. The decline in fluorescence emission during the so called SMT in the 1-100s excitation time range, known as the Kautsky curve, is shown to be associated with a substantial decrease of CET-powered proton efflux from the stroma into the chloroplast lumen through the ATPsynthase of the photosynthetic machinery.

  19. Novel Features of Eukaryotic Photosystem II Revealed by Its Crystal Structure Analysis from a Red Alga.

    Science.gov (United States)

    Ago, Hideo; Adachi, Hideyuki; Umena, Yasufumi; Tashiro, Takayoshi; Kawakami, Keisuke; Kamiya, Nobuo; Tian, Lirong; Han, Guangye; Kuang, Tingyun; Liu, Zheyi; Wang, Fangjun; Zou, Hanfa; Enami, Isao; Miyano, Masashi; Shen, Jian-Ren

    2016-03-11

    Photosystem II (PSII) catalyzes light-induced water splitting, leading to the evolution of molecular oxygen indispensible for life on the earth. The crystal structure of PSII from cyanobacteria has been solved at an atomic level, but the structure of eukaryotic PSII has not been analyzed. Because eukaryotic PSII possesses additional subunits not found in cyanobacterial PSII, it is important to solve the structure of eukaryotic PSII to elucidate their detailed functions, as well as evolutionary relationships. Here we report the structure of PSII from a red alga Cyanidium caldarium at 2.76 Å resolution, which revealed the structure and interaction sites of PsbQ', a unique, fourth extrinsic protein required for stabilizing the oxygen-evolving complex in the lumenal surface of PSII. The PsbQ' subunit was found to be located underneath CP43 in the vicinity of PsbV, and its structure is characterized by a bundle of four up-down helices arranged in a similar way to those of cyanobacterial and higher plant PsbQ, although helices I and II of PsbQ' were kinked relative to its higher plant counterpart because of its interactions with CP43. Furthermore, two novel transmembrane helices were found in the red algal PSII that are not present in cyanobacterial PSII; one of these helices may correspond to PsbW found only in eukaryotic PSII. The present results represent the first crystal structure of PSII from eukaryotic oxygenic organisms, which were discussed in comparison with the structure of cyanobacterial PSII.

  20. Growing green electricity: progress and strategies for use of photosystem I for sustainable photovoltaic energy conversion.

    Science.gov (United States)

    Nguyen, Khoa; Bruce, Barry D

    2014-09-01

    Oxygenic photosynthesis is driven via sequential action of Photosystem II (PSII) and (PSI)reaction centers via the Z-scheme. Both of these pigment-membrane protein complexes are found in cyanobacteria, algae, and plants. Unlike PSII, PSI is remarkably stable and does not undergo limiting photo-damage. This stability, as well as other fundamental structural differences, makes PSI the most attractive reaction centers for applied photosynthetic applications. These applied applications exploit the efficient light harvesting and high quantum yield of PSI where the isolated PSI particles are redeployed providing electrons directly as a photocurrent or, via a coupled catalyst to yield H₂. Recent advances in molecular genetics, synthetic biology, and nanotechnology have merged to allow PSI to be integrated into a myriad of biohybrid devices. In photocurrent producing devices, PSI has been immobilized onto various electrode substrates with a continuously evolving toolkit of strategies and novel reagents. However, these innovative yet highly variable designs make it difficult to identify the rate-limiting steps and/or components that function as bottlenecks in PSI-biohybrid devices. In this study we aim to highlight these recent advances with a focus on identifying the similarities and differences in electrode surfaces, immobilization/orientation strategies, and artificial redox mediators. Collectively this work has been able to maintain an annual increase in photocurrent density (Acm⁻²) of ~10-fold over the past decade. The potential drawbacks and attractive features of some of these schemes are also discussed with their feasibility on a large-scale. As an environmentally benign and renewable resource, PSI may provide a new sustainable source of bioenergy. This article is part of a special issue entitled: photosynthesis research for sustainability: keys to produce clean energy.

  1. Águas com predominância de Eutreptia lanowi steuer e Chlamydomonas reinhardi dangeard no plancton, na enseada de Inhauma, Baía de Guanabara

    Directory of Open Access Journals (Sweden)

    Lejeune P. H. de Oliveira

    1962-03-01

    Full Text Available In brackish waters of a creek of Guanabara Bay, the author points by the first time the presence of Chlamydomonas reinhardi, Eutreptia lanowi, Oscillatoria putrida, O. limosa, O. chlorina that were unknown in our waters; such biologic indicators proved themselves pollutional conditions, so bad a stark-mesosaprobic regime. Other news are plankton analysis by the Standar methods, of two most expressive samples of water masses;also the mobility of the plankters are measured in micra by second.

  2. Limiting steps of hydrogen production in Chlamydomonas reinhardtii and Synechocystis PCC 6803 as analysed by light-induced gas exchange transients

    Energy Technology Data Exchange (ETDEWEB)

    Cournac, L.; Mus, F.; Bernard, L.; Guedeney, G.; Peltier, G. [CEA Cadarache, Lab. d' Ecophysiologie de la Photosynthese, Saint Paul lez Durance (France); Vignais, P. [CEA Grenoble, Lab. de Biochimie er Biophysique des Systemes Integres (France)

    2002-12-01

    In the cyanobacterium Synechocystis PCC6803 and in the microalga Chlamydomonas reinhardtii, transient hydrogen photo-production is observed when cells are exposed to light in anoxia. We measured changes in H{sub 2}, O{sub 2}, and CO{sub 2} concentrations using time-resolved mass spectrometry in wild-type and mutant strains of Chlamydomonas and Synechocystis. In both organisms, non-photochemical reduction of the plastoquinone pool is shown to contribute to the initial H{sub 2} photo-production. This pathway, which does not produce O{sub 2}, exhibits a low rate in normal conditions. From the effect of the uncoupler FCCP, we conclude that PS II-independent H{sub 2} production in Chlamydomonas is limited by the trans-thylakoidal proton gradient. In Synechocystis, from the study of a mutant deficient in the NDH-1 complex (M55), we conclude that PS II-independent H{sub 2} production is limited by recycling of NAD(P)H through the NDH-1 complex. Based on these conclusions, we propose strategies for optimising H{sub 2} photo-production in these organisms. (Author)

  3. CSL encodes a leucine-rich-repeat protein implicated in red/violet light signaling to the circadian clock in Chlamydomonas

    Science.gov (United States)

    Kinoshita, Ayumi; Niwa, Yoshimi; Onai, Kiyoshi; Fukuzawa, Hideya; Ishiura, Masahiro

    2017-01-01

    The green alga Chlamydomonas reinhardtii shows various light responses in behavior and physiology. One such photoresponse is the circadian clock, which can be reset by external light signals to entrain its oscillation to daily environmental cycles. In a previous report, we suggested that a light-induced degradation of the clock protein ROC15 is a trigger to reset the circadian clock in Chlamydomonas. However, light signaling pathways of this process remained unclear. Here, we screened for mutants that show abnormal ROC15 diurnal rhythms, including the light-induced protein degradation at dawn, using a luciferase fusion reporter. In one mutant, ROC15 degradation and phase resetting of the circadian clock by light were impaired. Interestingly, the impairments were observed in response to red and violet light, but not to blue light. We revealed that an uncharacterized gene encoding a protein similar to RAS-signaling-related leucine-rich repeat (LRR) proteins is responsible for the mutant phenotypes. Our results indicate that a previously uncharacterized red/violet light signaling pathway is involved in the phase resetting of circadian clock in Chlamydomonas. PMID:28333924

  4. Energy brands lack vitality; Energiemerken ontberen vitaliteit

    Energy Technology Data Exchange (ETDEWEB)

    Godri, S.; Wilders, E. [CONSULT Brand Strategy, Y and R Advertising, Amsterdam (Netherlands)

    2004-06-01

    The three Dutch energy companies (Nuon, Essent and Eneco Energie) have relatively little brand strength. The brands are not perceived to be sufficiently different from one another and are not valued by consumers. With liberalisation imminent, this is hardly a strong starting point. How can you win over consumers if it is not clear what is on offer? In the business market, decision-makers are better placed to distinguish between brands. However, the brands lack vitality in this sector of the market too. The only consolation is that the situation is by no means exclusive to the Netherlands. [Dutch] De merksterkte van de drie Nederlandse energiebedrijven Nuon, Essent en Eneco Energie, is gering. De energiemerken onderscheiden zich niet genoeg en missen waardering. Met het oog op de aanstaande liberalisering geen goede uitgangspositie. Hoe kun je consumenten voor je winnen als onduidelijk is waarvoor ze moeten kiezen? Op de zakelijk markt zijn de beslissers beter in staat om de merken te onderscheiden. Maar ook hier missen de energiemerken vitaliteit. Een troost: Nederlandse energiebedrijven zijn niet de enige met dit probleem.

  5. Lack of universal scaling in magnetohydrodynamic turbulence

    Science.gov (United States)

    Pouquet, A.; Brachet, M.; Krstulovic, G.; Lee, E.; Mininni, P.; Rosenberg, D. L.

    2012-12-01

    Universality is often viewed as a hallmark of turbulent flows, with a search for scaling exponents that derive from intrinsic dynamics and do not depend on initial conditions or forcing, the Kolmogorov law for the energy spectrum of an incompressible fluid being the best known case. However, in the presence of waves due to an external agent such as rotation, stratification or a strong large-scale magnetic field B0, different regimes -- such as weak or strong turbulence, may arise and thus, different scaling behavior may arise as well. This is observed for example in the ocean, and it leads to different mixing and transport properties. In this talk, we shall first review, in the context of MHD turbulence, the phenomenological models that can be constructed using the following plausible dimensionless parameters: (i) RT, the ratio of characteristic time scales (here, the wave period Tw=L_0/B_0 and the eddy-turn-over time based on large-scale length and velocity, TNL=L0/U_0; (ii) RE, the ratio of magnetic to kinetic energy EM/E_V; and (iii) RA, the degree of alignment between the velocity and the magnetic field \\cos(v,b), or between the magnetic potential and magnetic induction, \\cos(A,b). Note that these ratios can also be defined at scale ℓ of velocity uℓ (as opposed to L0, U0), and thus one can consider as well the variation of such ratios across scales. We shall then contrast these models with data stemming from (mostly) solar observations that indicate a clear lack of universal scaling behavior. Similarly, a number of direct numerical simulations (DNS) including some at high resolution, in the spin-down of forcing case, in the presence of boundaries or not, and with or without an imposed strong external magnetic field B0, all point-out to different energy spectra, although the attainable Reynolds numbers in present-day DNS are still limited when contrasted with geophysical and astrophysical flows. In particular, we shall show that, when using as initial

  6. Light-driven hydrogen production by a hybrid complex of a [NiFe]-hydrogenase and the cyanobacterial photosystem I.

    Science.gov (United States)

    Ihara, Masaki; Nishihara, Hirofumi; Yoon, Ki-Seok; Lenz, Oliver; Friedrich, Bärbel; Nakamoto, Hitoshi; Kojima, Kouji; Honma, Daisuke; Kamachi, Toshiaki; Okura, Ichiro

    2006-01-01

    In order to generate renewable and clean fuels, increasing efforts are focused on the exploitation of photosynthetic microorganisms for the production of molecular hydrogen from water and light. In this study we engineered a 'hard-wired' protein complex consisting of a hydrogenase and photosystem I (hydrogenase-PSI complex) as a direct light-to-hydrogen conversion system. The key component was an artificial fusion protein composed of the membrane-bound [NiFe] hydrogenase from the beta-proteobacterium Ralstonia eutropha H16 and the peripheral PSI subunit PsaE of the cyanobacterium Thermosynechococcus elongatus. The resulting hydrogenase-PsaE fusion protein associated with PsaE-free PSI spontaneously, thereby forming a hydrogenase-PSI complex as confirmed by sucrose-gradient ultracentrifuge and immunoblot analysis. The hydrogenase-PSI complex displayed light-driven hydrogen production at a rate of 0.58 mumol H(2).mg chlorophyll(-1).h(-1). The complex maintained its accessibility to the native electron acceptor ferredoxin. This study provides the first example of a light-driven enzymatic reaction by an artificial complex between a redox enzyme and photosystem I and represents an important step on the way to design a photosynthetic organism that efficiently converts solar energy and water into hydrogen.

  7. Economic photoprotection in photosystem II that retains a complete light-harvesting system with slow energy traps.

    Science.gov (United States)

    Belgio, Erica; Kapitonova, Ekaterina; Chmeliov, Jevgenij; Duffy, Christopher D P; Ungerer, Petra; Valkunas, Leonas; Ruban, Alexander V

    2014-07-11

    The light-harvesting antenna of higher plant photosystem II has an intrinsic capability for self-defence against intense sunlight. The thermal dissipation of excess energy can be measured as the non-photochemical quenching of chlorophyll fluorescence. It has recently been proposed that the transition between the light-harvesting and self-defensive modes is associated with a reorganization of light-harvesting complexes. Here we show that despite structural changes, the photosystem II cross-section does not decrease. Our study reveals that the efficiency of energy trapping by the non-photochemical quencher(s) is lower than the efficiency of energy capture by the reaction centres. Consequently, the photoprotective mechanism works effectively for closed rather than open centres. This type of defence preserves the exceptional efficiency of electron transport in a broad range of light intensities, simultaneously ensuring high photosynthetic productivity and, under hazardous light conditions, sufficient photoprotection for both the reaction centre and the light-harvesting pigments of the antenna.

  8. O2 evolution and cyclic electron flow around photosystem I in long-term ground batch culture of Euglena gracilis

    Science.gov (United States)

    An, Yanjun; Wang, Suqin; Hao, Zongjie; Zhou, Yiyong; Liu, Yongding

    2014-12-01

    Based on the purpose of better exploring the function of green producers in the closed aquatic biological life support system, the condition of dynamic O2 evolution and performance of cyclic electron flow around photosystem I (CEF-PSI) in long-term ground batch culture of Euglena gracilis were studied, the relationship between linear electron flow (LEF) and CEF-PSI was revealed, the function of CEF-PSI was investigated. Excellent consistency in O2 evolution pattern was observed in cultures grown in both closed and open containers, O2 evolution was strictly suppressed in phase 1, but the rate of it increased significantly in phase 2. CEF-PSI was proposed to be active during the whole course of cultivation, even in the declining phase 3, it still operated at the extent of 47-55%. It is suggested that the relationship between LEF and CEF-PSI is not only competition but also reciprocity. CEF-PSI was proposed to contribute to the considerable growth in phase 1; it was also suggested to play an important protective role against photosystem II (PSII) photoinhibition at the greatly enhanced level (approximately 80-95%) on the 2nd day. Our results in this research suggest that E. gracilis had very particular photosynthetic characteristics, the strict O2 evolution suppression in the initial culture phase might be a special light acclimation behavior, and CEF-PSI could be an important mechanism involved in this kind of adaptation to the changeable light environment.

  9. Economic photoprotection in photosystem II that retains a complete light-harvesting system with slow energy traps

    Science.gov (United States)

    Belgio, Erica; Kapitonova, Ekaterina; Chmeliov, Jevgenij; Duffy, Christopher D. P.; Ungerer, Petra; Valkunas, Leonas; Ruban, Alexander V.

    2014-07-01

    The light-harvesting antenna of higher plant photosystem II has an intrinsic capability for self-defence against intense sunlight. The thermal dissipation of excess energy can be measured as the non-photochemical quenching of chlorophyll fluorescence. It has recently been proposed that the transition between the light-harvesting and self-defensive modes is associated with a reorganization of light-harvesting complexes. Here we show that despite structural changes, the photosystem II cross-section does not decrease. Our study reveals that the efficiency of energy trapping by the non-photochemical quencher(s) is lower than the efficiency of energy capture by the reaction centres. Consequently, the photoprotective mechanism works effectively for closed rather than open centres. This type of defence preserves the exceptional efficiency of electron transport in a broad range of light intensities, simultaneously ensuring high photosynthetic productivity and, under hazardous light conditions, sufficient photoprotection for both the reaction centre and the light-harvesting pigments of the antenna.

  10. Lack of RNase L attenuates macrophage functions.

    Directory of Open Access Journals (Sweden)

    Xin Yi

    Full Text Available Macrophages are one of the major cell types in innate immunity against microbial infection. It is believed that the expression of proinflammatory genes such as tumor necrosis factor-α (TNF-α, interleukin (IL-1β, IL-6, and cyclooxygenase-2 (Cox-2 by macrophages is also crucial for activation of both innate and adaptive immunities. RNase L is an interferon (IFN inducible enzyme which is highly expressed in macrophages. It has been demonstrated that RNase L regulates the expression of certain inflammatory genes. However, its role in macrophage function is largely unknown.Bone marrow-derived macrophages (BMMs were generated from RNase L(+/+and (-/- mice. The migration of BMMs was analyzed by using Transwell migration assays. Endocytosis and phagocytosis of macrophages were assessed by using fluorescein isothiocyanate (FITC-Dextran 40,000 and FITC-E. coli bacteria, respectively. The expression of inflammatory genes was determined by Western Blot and ELISA. The promoter activity of Cox-2 was measured by luciferase reporter assays.Lack of RNase L significantly decreased the migration of BMMs induced by M-CSF, but at a less extent by GM-CSF and chemokine C-C motif ligand-2 (CCL2. Interestingly, RNase L deficient BMMs showed a significant reduction of endocytic activity to FITC-Dextran 40,000, but no any obvious effect on their phagocytic activity to FITC-bacteria under the same condition. RNase L impacts the expression of certain genes related to cell migration and inflammation such as transforming growth factor (TGF-β, IL-1β, IL-10, CCL2 and Cox-2. Furthermore, the functional analysis of the Cox-2 promoter revealed that RNase L regulated the expression of Cox-2 in macrophages at its transcriptional level. Taken together, our findings provide direct evidence showing that RNase L contributes to innate immunity through regulating macrophage functions.

  11. Comparison of the chloroplast peroxidase system in the chlorophyte Chlamydomonas reinhardtii, the bryophyte Physcomitrella patens, the lycophyte Selaginella moellendorffii and the seed plant Arabidopsis thaliana

    Directory of Open Access Journals (Sweden)

    Baier Margarete

    2010-06-01

    Full Text Available Abstract Background Oxygenic photosynthesis is accompanied by the formation of reactive oxygen species (ROS, which damage proteins, lipids, DNA and finally limit plant yield. The enzymes of the chloroplast antioxidant system are exclusively nuclear encoded. During evolution, plastid and mitochondrial genes were post-endosymbiotically transferred to the nucleus, adapted for eukaryotic gene expression and post-translational protein targeting and supplemented with genes of eukaryotic origin. Results Here, the genomes of the green alga Chlamydomonas reinhardtii, the moss Physcomitrella patens, the lycophyte Selaginella moellendorffii and the seed plant Arabidopsis thaliana were screened for ORFs encoding chloroplast peroxidases. The identified genes were compared for their amino acid sequence similarities and gene structures. Stromal and thylakoid-bound ascorbate peroxidases (APx share common splice sites demonstrating that they evolved from a common ancestral gene. In contrast to most cormophytes, our results predict that chloroplast APx activity is restricted to the stroma in Chlamydomonas and to thylakoids in Physcomitrella. The moss gene is of retrotransposonal origin. The exon-intron-structures of 2CP genes differ between chlorophytes and streptophytes indicating an independent evolution. According to amino acid sequence characteristics only the A-isoform of Chlamydomonas 2CP may be functionally equivalent to streptophyte 2CP, while the weakly expressed B- and C-isoforms show chlorophyte specific surfaces and amino acid sequence characteristics. The amino acid sequences of chloroplast PrxII are widely conserved between the investigated species. In the analyzed streptophytes, the genes are unspliced, but accumulated four introns in Chlamydomonas. A conserved splice site indicates also a common origin of chlorobiont PrxQ. The similarity of splice sites also demonstrates that streptophyte glutathione peroxidases (GPx are of common origin. Besides

  12. Protein film voltammetry and co-factor electron transfer dynamics in spinach photosystem II core complex.

    Science.gov (United States)

    Zhang, Yun; Magdaong, Nikki; Frank, Harry A; Rusling, James F

    2014-05-01

    Direct protein film voltammetry (PFV) was used to investigate the redox properties of the photosystem II (PSII) core complex from spinach. The complex was isolated using an improved protocol not used previously for PFV. The PSII core complex had high oxygen-evolving capacity and was incorporated into thin lipid and polyion films. Three well-defined reversible pairs of reduction and oxidation voltammetry peaks were observed at 4 °C in the dark. Results were similar in both types of films, indicating that the environment of the PSII-bound cofactors was not influenced by film type. Based on comparison with various control samples including Mn-depleted PSII, peaks were assigned to chlorophyll a (Chl a) (Em = -0.47 V, all vs. NHE, at pH 6), quinones (-0.12 V), and the manganese (Mn) cluster (Em = 0.18 V). PFV of purified iron heme protein cytochrome b-559 (Cyt b-559), a component of PSII, gave a partly reversible peak pair at 0.004 V that did not have a potential similar to any peaks observed from the intact PSII core complex. The closest peak in PSII to 0.004 V is the 0.18 V peak that was found to be associated with a two-electron process, and thus is inconsistent with iron heme protein voltammetry. The -0.47 V peak had a peak potential and peak potential-pH dependence similar to that found for purified Chl a incorporated into DMPC films. The midpoint potentials reported here may differ to various extents from previously reported redox titration data due to the influence of electrode double-layer effects. Heterogeneous electron transfer (hET) rate constants were estimated by theoretical fitting and digital simulations for the -0.47 and 0.18 V peaks. Data for the Chl a peaks were best fit to a one-electron model, while the peak assigned to the Mn cluster was best fit by a two-electron/one-proton model.

  13. Structural oxidation state studies of the manganese cluster in the oxygen evolving complex of photosystem II

    Energy Technology Data Exchange (ETDEWEB)

    Liang, W.

    1994-11-01

    X-ray absorption spectroscopy (XAS) was performed on Photosystem II (PSII)-enriched membranes prepared from spinach to explore: (1) the correlation between structure and magnetic spin state of the Mn cluster in the oxygen evolving complex (OEC) in the S{sub 2} state; and (2) the oxidation state changes of the Mn cluster in the flash-induced S-states. The structure of the Mn cluster in the S{sub 2} state with the g{approx}4 electron paramagnetic resonance (EPR) signal (S{sub 2}-g4 state) was compared with that in the S{sub 2} state with multiline signal (S{sub 2}-MLS state) and the S{sub 1} state. The S{sub 2}-g4 state has a higher XAS inflection point energy than that of the S{sub 1} state, indicating the oxidation of Mn in the advance from the S{sub 1} to the S{sub 2}-g4 state. Differences in the edge shape and in the extended X-ray absorption fine structure (EXAFS) show that the structure of the Mn cluster in the S{sub 2}-g4 state is different from that in the S{sub 2}-MLS or the S{sub 1} state. In the S{sub 2}-g4 state, the second shell of backscatterers from the Mn absorber contains two Mn-Mn distances of 2.73 {angstrom} and 2.85 {angstrom}. Very little distance disorder exists in the second shell of the S{sub 1} or S{sub 2}-MLS states. The third shell of the S{sub 2}-g4 state at about 3.3 {angstrom} also contains increased heterogeneity relative to that of the S{sub 2}-MLS or the S{sub 1} state. Various S-states were prepared at room-temperature by saturating, single-turnover flashes. The flash-dependent oscillation in the amplitude of the MLS was used to characterize the S-state composition and to construct {open_quotes}pure{close_quotes} S-state Mn K-edge spectra. The edge position shifts to higher energy by 1.8 eV upon the S{sub 1} {yields} S{sub 2} transition.

  14. Growth inhibition and effect on photosystem by three imidazolium chloride ionic liquids in rice seedlings

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Huijun, E-mail: lhj@mail.zjgsu.edu.cn [School of Environmental Science and Engineering, Zhejiang Gongshang University, Hangzhou 310018, Zhejiang Province (China); Zhang, Shuxian [School of Environmental Science and Engineering, Zhejiang Gongshang University, Hangzhou 310018, Zhejiang Province (China); Jiaxing University, Jiaxing 314001, Zhejiang Province (China); Zhang, Xiaoqiang; Chen, Caidong [School of Environmental Science and Engineering, Zhejiang Gongshang University, Hangzhou 310018, Zhejiang Province (China)

    2015-04-09

    Highlights: • The three ILs have phytotoxic on rice growth. • The antioxidant enzyme activities increased first and then declined with ILs concentration increased. • The Hill reaction activity decreased and the PS II of leaves was damaged by ILs. • The toxicity of ILs increased as the alkyl chain length increased as the order: [OMIM]Cl < [DMIM]Cl < [C{sub 12}MIM]Cl. - Abstract: The effects of three imidazolium chloride ionic liquids (ILs) including 1-octyl-3-methylimidazolium chloride ionic liquid ([OMIM]Cl), 1-decyl-3-methylimidazolium chloride ionic liquid ([DMIM]Cl) and 1-dodecyl-3-methylimidazolium chloride ionic liquid ([C{sub 12}MIM]Cl) were studied in hydroponically grown rice seedlings. The growth inhibition rate increased and the Hill reaction activity of isolated rice chloroplasts decreased with increasing ILs concentrations. The IC{sub 50,5d} for stem length was 0.70 mg/L of [OMIM]Cl, 0.15 mg/L of [DMIM]Cl, and 0.055 mg/L of [C{sub 12}MIM]Cl, respectively. The SOD, POD and CAT activities of chloroplast exhibited initial increases followed by decreases in activity with increasing ILs concentrations. Chlorophyll fluorescence parameters such as the maximum effective quantum yield of PSII(F{sub v}/F{sub m}), the potential activity of PSII(F{sub v}/F{sub 0}), the yield of photochemical quantum [Y(II)], the photochemical quenching coefficient (qP), the non-photochemical quenching coefficient (NPQ) and the relative electron transport ratio (rETR) were affected, showing that ILs will damage the PSII. The results demonstrated that imidazolium chloride ILs are phytotoxic to rice growth and their photosystem, the toxicity increased as the alkyl chain length increased with the following order: [OMIM]Cl < [DMIM]Cl < [C{sub 12}MIM]Cl. The results will help to better understand the possible role of the defense mechanism in rice caused by ILs exposure.

  15. Lack of efficacy of ergocalciferol repletion

    Directory of Open Access Journals (Sweden)

    Thomas Wasser

    2012-04-01

    Full Text Available Introduction: Vitamin D has become an area of intensive scrutiny, both in medical and lay literature. However, there are limited data to suggest proper repletion regimens for those patients who have hypovitaminosis D. Consequently, various methods are used in clinical practice. The aim of this study was to assess the efficacy of various treatment strategies for hypovitaminosis D in an ambulatory internal medicine practice. Methods: A retrospective chart review between October 2005 and June 2010 of a suburban internal medicine practice was performed via query of the electronic medical record (Centricity, General Electric Healthcare, UK. Patients with a 25-hydroxyvitamin D concentration less than 32 mg/dl were identified and treated. Treatment success was defined as 25-hydroxyvitamin D concentrations greater than 32 mg/dl. Statistical analysis to assess changes in vitamin D level controlling for season, comorbidities, and demographics were used. Results: A total of 607 treatment episodes were identified, with 395 excluded due to lack of follow-up vitamin D level within 16 weeks, no treatment documented, topical treatment, doxercalciferol treatment, or non-compliance. Of the remaining patients, there were 212 treatment instances on 178 patients. Ergocalciferol 50,000 international units (IU was used most frequently (71.4% of the time.. A higher initial vitamin D level was positively associated with treatment success (adjusted odds ratio = 1.11, p=0.002. Increased doses of ergocalciferol increased the likelihood of treatment success (p=0.0011. Seasonal variation was related to posttreatment 25-hydroxyvitamin D concentration as was body mass index (BMI (p=0.003 and p=0.044. Conclusion: Pretreatment levels of 25-hydroxyvitamin D, BMI, season, and vitamin D dose are predictors of successful hypovitaminosis D treatment. Our data suggest that patients with initial 25-hydroxyvitamin D concentrations of <20 should be treated with a higher total dose of

  16. Probing the accessibility of the Mn(4)Ca cluster in photosystem II : Channels calculation, noble gas derivatization, and cocrystallization with DMSO

    NARCIS (Netherlands)

    Gabdulkhakov, Azat; Guskov, Albert; Broser, Matthias; Kern, Jan; Müh, Frank; Saenger, Wolfram; Zouni, Athina

    2009-01-01

    Using the 2.9 A resolution structure of the membrane-intrinsic protein-cofactor complex photosystem II (PSII) from the cyanobacterium Thermosynechococcus elongatus, we calculated and characterized nine possible substrate/product channels leading to/away from the Mn(4)Ca cluster, where water is oxidi

  17. Time-resolved visible and infrared absorption spectroscopy data obtained using photosystem I particles with non-native quinones incorporated into the A1 binding site.

    Science.gov (United States)

    Makita, Hiroki; Hastings, Gary

    2016-06-01

    Time-resolved visible and infrared absorption difference spectroscopy data at both 298 and 77 K were obtained using cyanobacterial menB (-) mutant photosystem I particles with several non-native quinones incorporated into the A1 binding site. Data was obtained for photosystem I particles with phylloquinone (2-methyl-3-phytyl-1,4-naphthoquinone), 2-bromo-1,4-naphthoquinone, 2-chloro-1,4-naphthoquinone, 2-methyl-1,4-naphthoquinone, 2,3-dibromo-1,4-naphthoquinone, 2,3-dichloro-1,4-naphthoquinone, and 9,10-anthraquinone incorporated. Transient absorption data were obtained at 487 and 703 nm in the visible spectral range, and 1950-1100 cm(-1) in the infrared region. Time constants obtained from fitting the time-resolved infrared and visible data are in good agreement. The measured time constants are crucial for the development of appropriate kinetic models that can describe electron transfer processes in photosystem I, "Modeling Electron Transfer in Photosystem I" Makita and Hastings (2016) [1].

  18. Evidence for the semireduced primary quinone electron acceptor of photosystem II being a photosensitizer for UVB damage to the photosynthetic apparatus

    NARCIS (Netherlands)

    Rodrigues, G.C.; Jansen, M.A.K.; Noort, van den M.E.; Rensen, van J.J.S.

    2006-01-01

    Exposure to ultraviolet-B radiation (UVB) radiation affects plants in multiple ways, including effects on the photosynthetic apparatus. The carbon dioxide reduction reactions are affected as well as the light reactions, especially those of photosystem II. In the literature several UVB chromophores a

  19. Photosystem I light-harvesting complex Lhca4 adopts multiple conformations : Red forms and excited-state quenching are mutually exclusive

    NARCIS (Netherlands)

    Passarini, Francesca; Wientjes, Emilie; van Amerongen, Herbert; Croce, Roberta

    2010-01-01

    In this work we have investigated the origin of the multi-exponential fluorescence decay and of the short excited-state lifetime of Lhca4. Lhca4 is the antenna complex of Photosystem I which accommodates the red-most chlorophyll forms and it has been proposed that these chlorophylls can play a role

  20. Photosystem I light-harvesting complex Lhca4 adopts multiple conformations: red forms and excited-state quenching are mutually exclusive

    NARCIS (Netherlands)

    Passarini, F.; Wientjes, E.; Amerongen, van H.; Croce, R.

    2010-01-01

    In this work we have investigated the origin of the multi-exponential fluorescence decay and of the short excited-state lifetime of Lhca4. Lhca4 is the antenna complex of Photosystem I which accommodates the red-most chlorophyll forms and it has been proposed that these chlorophylls can play a role

  1. Time-resolved visible and infrared absorption spectroscopy data obtained using photosystem I particles with non-native quinones incorporated into the A1 binding site

    Directory of Open Access Journals (Sweden)

    Hiroki Makita

    2016-06-01

    Full Text Available Time-resolved visible and infrared absorption difference spectroscopy data at both 298 and 77 K were obtained using cyanobacterial menB− mutant photosystem I particles with several non-native quinones incorporated into the A1 binding site. Data was obtained for photosystem I particles with phylloquinone (2-methyl-3-phytyl-1,4-naphthoquinone, 2-bromo-1,4-naphthoquinone, 2-chloro-1,4-naphthoquinone, 2-methyl-1,4-naphthoquinone, 2,3-dibromo-1,4-naphthoquinone, 2,3-dichloro-1,4-naphthoquinone, and 9,10-anthraquinone incorporated. Transient absorption data were obtained at 487 and 703 nm in the visible spectral range, and 1950–1100 cm−1 in the infrared region. Time constants obtained from fitting the time-resolved infrared and visible data are in good agreement. The measured time constants are crucial for the development of appropriate kinetic models that can describe electron transfer processes in photosystem I, “Modeling Electron Transfer in Photosystem I” Makita and Hastings (2016 [1].

  2. The light-harvesting complexes of higher-plant Photosystem I : Lhca1/4 and Lhca2/3 form two red-emitting heterodimers

    NARCIS (Netherlands)

    Wientjes, Emilie; Croce, Roberta

    2011-01-01

    The outer antenna of higher-plant PSI (Photosystem I) is composed of four complexes [Lhc (light-harvesting complex) al-Lhca4] belonging to the light-harvesting protein family. Difficulties in their purification have so far prevented the determination of their properties and most of the knowledge abo

  3. Flavodiiron proteins in oxygenic photosynthetic organisms: photoprotection of photosystem II by Flv2 and Flv4 in Synechocystis sp. PCC 6803.

    Directory of Open Access Journals (Sweden)

    Pengpeng Zhang

    Full Text Available BACKGROUND: Flavodiiron proteins (FDPs comprise a group of modular enzymes that function in oxygen and nitric oxide detoxification in Bacteria and Archaea. The FDPs in cyanobacteria have an extra domain as compared to major prokaryotic enzymes. The physiological role of cyanobacteria FDPs is mostly unknown. Of the four putative flavodiiron proteins (Flv1-4 in the cyanobacterium Synechocystis sp. PCC 6803, a physiological function in Mehler reaction has been suggested for Flv1 and Flv3. PRINCIPAL FINDINGS: We demonstrate a novel and crucial function for Flv2 and Flv4 in photoprotection of photosystem II (PSII in Synechocystis. It is shown that the expression of Flv2 and Flv4 is high under air level of CO(2 and negligible at elevated CO(2. Moreover, the rate of accumulation of flv2 and flv4 transcripts upon shift of cells from high to low CO(2 is strongly dependent on light intensity. Characterization of FDP inactivation mutants of Synechocystis revealed a specific decline in PSII centers and impaired translation of the D1 protein in Delta flv2 and Delta flv4 when grown at air level CO(2 whereas at high CO(2 the Flvs were dispensable. Delta flv2 and Delta flv4 were also more susceptible to high light induced inhibition of PSII than WT or Delta flv1 and Delta flv3. SIGNIFICANCE: Analysis of published sequences revealed the presence of cyanobacteria-like FDPs also in some oxygenic photosynthetic eukaryotes like green algae, mosses and lycophytes. Our data provide evidence that Flv2 and Flv4 have an important role in photoprotection of water-splitting PSII against oxidative stress when the cells are acclimated to air level CO(2. It is conceivable that the function of FDPs has changed during evolution from protection against oxygen in anaerobic microbes to protection against reactive oxygen species thus making the sustainable function of oxygen evolving PSII possible. Higher plants lack FDPs and distinctly different mechanisms have evolved for

  4. Whole Genome Re-Sequencing Identifies a Quantitative Trait Locus Repressing Carbon Reserve Accumulation during Optimal Growth in Chlamydomonas reinhardtii.

    Science.gov (United States)

    Goold, Hugh Douglas; Nguyen, Hoa Mai; Kong, Fantao; Beyly-Adriano, Audrey; Légeret, Bertrand; Billon, Emmanuelle; Cuiné, Stéphan; Beisson, Fred; Peltier, Gilles; Li-Beisson, Yonghua

    2016-05-04

    Microalgae have emerged as a promising source for biofuel production. Massive oil and starch accumulation in microalgae is possible, but occurs mostly when biomass growth is impaired. The molecular networks underlying the negative correlation between growth and reserve formation are not known. Thus isolation of strains capable of accumulating carbon reserves during optimal growth would be highly desirable. To this end, we screened an insertional mutant library of Chlamydomonas reinhardtii for alterations in oil content. A mutant accumulating five times more oil and twice more starch than wild-type during optimal growth was isolated and named constitutive oil accumulator 1 (coa1). Growth in photobioreactors under highly controlled conditions revealed that the increase in oil and starch content in coa1 was dependent on light intensity. Genetic analysis and DNA hybridization pointed to a single insertional event responsible for the phenotype. Whole genome re-sequencing identified in coa1 a >200 kb deletion on chromosome 14 containing 41 genes. This study demonstrates that, 1), the generation of algal strains accumulating higher reserve amount without compromising biomass accumulation is feasible; 2), light is an important parameter in phenotypic analysis; and 3), a chromosomal region (Quantitative Trait Locus) acts as suppressor of carbon reserve accumulation during optimal growth.

  5. Changes in Stoichiometry, Cellular RNA, and Alkaline Phosphatase Activity of Chlamydomonas in Response to Temperature and Nutrients

    Science.gov (United States)

    Hessen, Dag O.; Hafslund, Ola T.; Andersen, Tom; Broch, Catharina; Shala, Nita K.; Wojewodzic, Marcin W.

    2017-01-01

    Phytoplankton may respond both to elevated temperatures and reduced nutrients by changing their cellular stoichiometry and cell sizes. Since increased temperatures often cause increased thermal stratification and reduced vertical flux of nutrients into the mixed zone, it is difficult to disentangle these drivers in nature. In this study, we used a factorial design with high and low levels of phosphorus (P) and high and low temperature to assess responses in cellular stoichiometry, levels of RNA, and alkaline phosphatase activity (APA) in the chlorophyte Chlamydomonas reinhardtii. Growth rate, C:P, C:N, N:P, RNA, and APA all responded primarily to P treatment, but except for N:P and APA, also temperature contributed significantly. For RNA, the contribution from temperature was particularly strong with higher cellular levels of RNA at low temperatures, suggesting a compensatory allocation to ribosomes to maintain protein synthesis and growth. These experiments suggest that although P-limitation is the major determinant of growth rate and cellular stoichiometry, there are pronounced effects of temperature also via interaction with P. At the ecosystem level, nutrients and temperature will thus interact, but temperatures would likely exert a stronger impact on these phytoplankton traits indirectly via its force on stratification regimes and vertical nutrient fluxes. PMID:28167934

  6. Flux balance analysis reveals acetate metabolism modulates cyclic electron flow and alternative glycolytic pathways in Chlamydomonas reinhardtii.

    Science.gov (United States)

    Chapman, Stephen P; Paget, Caroline M; Johnson, Giles N; Schwartz, Jean-Marc

    2015-01-01

    Cells of the green alga Chlamydomonas reinhardtii cultured in the presence of acetate perform mixotrophic growth, involving both photosynthesis and organic carbon assimilation. Under such conditions, cells exhibit a reduced capacity for photosynthesis but a higher growth rate, compared to phototrophic cultures. Better understanding of the down regulation of photosynthesis would enable more efficient conversion of carbon into valuable products like biofuels. In this study, Flux Balance Analysis (FBA) and Flux Variability Analysis (FVA) have been used with a genome scale model of C. reinhardtii to examine changes in intracellular flux distribution in order to explain their changing physiology. Additionally, a reaction essentiality analysis was performed to identify which reaction subsets are essential for a given growth condition. Our results suggest that exogenous acetate feeds into a modified tricarboxylic acid (TCA) cycle, which bypasses the CO2 evolution steps, explaining increases in biomass, consistent with experimental data. In addition, reactions of the oxidative pentose phosphate and glycolysis pathways, inactive under phototrophic conditions, show substantial flux under mixotrophic conditions. Importantly, acetate addition leads to an increased flux through cyclic electron flow (CEF), but results in a repression of CO2 fixation via Rubisco, explaining the down regulation of photosynthesis. However, although CEF enhances growth on acetate, it is not essential-impairment of CEF results in alternative metabolic pathways being increased. We have demonstrated how the reactions of photosynthesis interconnect with carbon metabolism on a global scale, and how systems approaches play a viable tool in understanding complex relationships at the scale of the organism.

  7. Molecular toxicity of cerium oxide nanoparticles to the freshwater alga Chlamydomonas reinhardtii is associated with supra-environmental exposure concentrations.

    Science.gov (United States)

    Taylor, Nadine S; Merrifield, Ruth; Williams, Tim D; Chipman, J Kevin; Lead, Jamie R; Viant, Mark R

    2016-01-01

    Ceria nanoparticles (NPs) are widely used as fuel catalysts and consequently are likely to enter the environment. Their potential impacts on. biota at environmentally relevant concentrations, including uptake and toxicity, remain to be elucidated and quantitative data on which to assess risk are sparse. Therefore, a definitive assessment of the molecular and phenotypic effects of ceria NPs was undertaken, using well-characterised mono-dispersed NPs as their toxicity is likely to be higher, enabling a conservative hazard assessment. Unbiased transcriptomics and metabolomics approaches were used to investigate the potential toxicity of tightly constrained 4-5 nm ceria NPs to the unicellular green alga, Chlamydomonas reinhardtii, a sentinel freshwater species. A wide range of exposure concentrations were investigated from predicted environmental levels, to support hazard assessment, to supra-environmental levels to provide insight into molecular toxicity pathways. Ceria NPs were internalised into intracellular vesicles within C. reinhardtii, yet caused no significant effect on algal growth at any exposure concentration. Molecular perturbations were only detected at supra-environmental ceria NP-concentrations, primarily down-regulation of photosynthesis and carbon fixation with associated effects on energy metabolism. For acute exposures to small mono-dispersed particles, it can be concluded there should be little concern regarding their dispersal into the environment for this trophic level.

  8. Evidence for Nuclear Control of the Expression of the atpA and atpB Chloroplast Genes in Chlamydomonas.

    Science.gov (United States)

    Drapier, D.; Girard-Bascou, J.; Wollman, F. A.

    1992-03-01

    We analyzed three nuclear mutants of Chlamydomonas reinhardtii altered in the expression of the chloroplast genes atpA or atpB coding for the [alpha] or [beta] subunit of the chloroplast ATP synthase. These mutants revealed the existence of three nuclear products controlling the expression of the two chloroplast genes: the first one acts on the translation of the atpA transcript, and the two others act specifically on the stability of either the atpB or the atpA mRNAs. The nuclear mutation responsible for the decreased stability of the atpB mRNA prevented translation of the corresponding polypeptide. In contrast, the mutation responsible for the decreased stability of the atpA mRNA had limited effect on the translation of the [alpha] subunit, thereby allowing its accumulation and assembly in an active ATP synthase. Although acting originally on the expression of only one of the two main coupling factor 1 subunits, the three mutations caused a change in the translation rate of the other subunit, as viewed in 5-min pulse labeling experiments. This is indicative of a concerted expression of the [alpha] and [beta] subunits at an early post-translational step, or during translation, that may be critical for the assembly of the chloroplast ATP synthase.

  9. Effects of sodium sulfate on the freshwater microalga Chlamydomonas moewusii: implications for the optimization of algal culture media.

    Science.gov (United States)

    Mera, Roi; Torres, Enrique; Abalde, Julio

    2016-02-01

    The study of the microalgal growth kinetics is an indispensable tool in all fields of phycology. Knowing the optimal nutrient concentration is an important issue that will help to develop efficient growth systems for these microorganisms. Although nitrogen and phosphorus are well studied for this purpose, sulfur seems to be less investigated. Sulfate is a primary sulfur source used by microalgae; moreover, the concentration of this compound is increasing in freshwater systems due to pollution. The aim of this study was to investigate the effects of different sodium sulfate concentrations in the culture medium on growth and growth kinetics of the freshwater microalga Chlamydomonas moewusii. Production of biomass, chl content, kinetic equations, and a mathematical model that describe the microalgal growth in relation with the concentration of sodium sulfate were obtained. The lowest concentration of sodium sulfate allowing optimal growth was 0.1 mM. Concentrations higher than 3 mM generated a toxic effect. This work demonstrates that this toxic effect was not directly due to the excess of sulfate ion but by the elevation of the ionic strength. An inhibition model was successfully used to simulate the relationship between specific growth rate and sodium sulfate in this microalga.

  10. Green algal cytochrome b6-f complexes: isolation and characterization from Dunaliella saline, Chlamydomonas reinhardtii and Scenedesmus obliquus.

    Science.gov (United States)

    Wynn, R M; Bertsch, J; Bruce, B D; Malkin, R

    1988-09-14

    Cytochrome b6-f complexes have been isolated from Chlamydomonas reinhardtii, Dunaliella saline and Scenedesmus obliquus. Each complex is essentially free of chlorophyll and carotenoids and contains cytochrome b6 and cytochrome f hemes in a 2:1 molar ratio. C. reinhardtii and S. obliquus complexes contain the Rieske iron-sulfur protein (present in approx 1:1 molar ratio to cytochrome f) and each catalyzes a DBMIB- and DNP-INT-sensitive electron transfer from duroquinol to spinach plastocyanin. Immunological assays using antibodies to the peptides from the spinach cytochrome complex show varying cross-reactivity patterns except for the complete absence of binding to the Rieske proteins in any of the three complexes, suggesting little structural similarity between the Rieske proteins of algae with those from higher plants. One complex (D. salina) has been uniformly labeled by growth in NaH14CO3 to determine stoichiometries of constituent polypeptide subunits. Results from these studies indicate that all functionally active cytochrome b6-f complexes contain four subunits which occur in equimolar amounts.

  11. A factor related to pseudouridine synthases is required for chloroplast group II intron trans-splicing in Chlamydomonas reinhardtii.

    Science.gov (United States)

    Perron, K; Goldschmidt-Clermont, M; Rochaix, J D

    1999-11-15

    In Chlamydomonas reinhardtii, the psaA mRNA is assembled by a process involving two steps of trans-splicing that remove two group II introns and give rise to the mature mRNA. The products of at least 14 nuclear genes and one chloroplast gene (tscA) are necessary for this process. We have cloned Maa2, one of the nuclear genes involved in trans-splicing of the second intron. Maa2 encodes a protein with similarity to conserved domains of pseudouridine synthases, but mutagenesis of putative catalytic residues showed that this activity may not be required for trans-splicing of psaA RNA. Although it is not clear whether the pseudouridine synthase activity has been maintained in Maa2, it is possible that this enzyme was recruited during evolution as an RNA chaperone for folding or stabilizing the psaA intron. The Maa2 protein appears to be associated through ionic interactions with a low density membrane system in the chloroplast that also contains RNA-binding proteins involved in translation.

  12. Whole Genome Re-Sequencing Identifies a Quantitative Trait Locus Repressing Carbon Reserve Accumulation during Optimal Growth in Chlamydomonas reinhardtii

    Science.gov (United States)

    Goold, Hugh Douglas; Nguyen, Hoa Mai; Kong, Fantao; Beyly-Adriano, Audrey; Légeret, Bertrand; Billon, Emmanuelle; Cuiné, Stéphan; Beisson, Fred; Peltier, Gilles; Li-Beisson, Yonghua

    2016-01-01

    Microalgae have emerged as a promising source for biofuel production. Massive oil and starch accumulation in microalgae is possible, but occurs mostly when biomass growth is impaired. The molecular networks underlying the negative correlation between growth and reserve formation are not known. Thus isolation of strains capable of accumulating carbon reserves during optimal growth would be highly desirable. To this end, we screened an insertional mutant library of Chlamydomonas reinhardtii for alterations in oil content. A mutant accumulating five times more oil and twice more starch than wild-type during optimal growth was isolated and named constitutive oil accumulator 1 (coa1). Growth in photobioreactors under highly controlled conditions revealed that the increase in oil and starch content in coa1 was dependent on light intensity. Genetic analysis and DNA hybridization pointed to a single insertional event responsible for the phenotype. Whole genome re-sequencing identified in coa1 a >200 kb deletion on chromosome 14 containing 41 genes. This study demonstrates that, 1), the generation of algal strains accumulating higher reserve amount without compromising biomass accumulation is feasible; 2), light is an important parameter in phenotypic analysis; and 3), a chromosomal region (Quantitative Trait Locus) acts as suppressor of carbon reserve accumulation during optimal growth. PMID:27141848

  13. A Chlamydomonas Homologue of the Putative Murine t Complex Distorter Tctex-2 Is an Outer Arm Dynein Light Chain

    Science.gov (United States)

    Patel-King, Ramila S.; Benashski, Sharon E.; Harrison, Alistair; King, Stephen M.

    1997-01-01

    Molecular analysis of a 19,000-Mr protein from the Chlamydomonas flagellum reveals that it is homologous to the t complex–encoded protein Tctex-2, which is a candidate for one of the distorter products that cause the extreme transmission ratio distortion (meiotic drive) of the murine t complex. The 19,000-Mr protein is extracted from the axoneme with 0.6 M NaCl and comigrates with the outer dynein arm in sucrose density gradients. This protein also is specifically missing in axonemes prepared from a mutant that does not assemble the outer arm. These data raise the possibility that Tctex-2 is a sperm flagellar dynein component. Combined with the recent identification of Tctex-1 (another distorter candidate) as a light chain of cytoplasmic dynein, these results lead to a biochemical model for how differential defects in spermiogenesis that result in the phenomenon of meiotic drive might be generated in wild-type vs t-bearing sperm. PMID:9166408

  14. Complete elimination of maternal mitochondrial DNA during meiosis resulting in the paternal inheritance of the mitochondrial genome in Chlamydomonas species.

    Science.gov (United States)

    Aoyama, H; Hagiwara, Y; Misumi, O; Kuroiwa, T; Nakamura, S

    2006-09-01

    The non-Mendelian inheritance of organellar DNA is common in most plants and animals. In the isogamous green alga Chlamydomonas species, progeny inherit chloroplast genes from the maternal parent, as paternal chloroplast genes are selectively eliminated in young zygotes. Mitochondrial genes are inherited from the paternal parent. Analogically, maternal mitochondrial DNA (mtDNA) is thought to be selectively eliminated. Nevertheless, it is unclear when this selective elimination occurs. Here, we examined the behaviors of maternal and paternal mtDNAs by various methods during the period between the beginning of zygote formation and zoospore formation. First, we observed the behavior of the organelle nucleoids of living cells by specifically staining DNA with the fluorochrome SYBR Green I and staining mitochondria with 3,3'-dihexyloxacarbocyanine iodide. We also examined the fate of mtDNA of male and female parental origin by real-time PCR, nested PCR with single zygotes, and fluorescence in situ hybridization analysis. The mtDNA of maternal origin was completely eliminated before the first cell nuclear division, probably just before mtDNA synthesis, during meiosis. Therefore, the progeny inherit the remaining paternal mtDNA. We suggest that the complete elimination of maternal mtDNA during meiosis is the primary cause of paternal mitochondrial inheritance.

  15. Application of proton exchange membrane fuel cells for the monitoring and direct usage of biohydrogen produced by Chlamydomonas reinhardtii

    Science.gov (United States)

    Oncel, S.; Vardar-Sukan, F.

    Photo-biologically produced hydrogen by Chlamydomonas reinhardtii is integrated with a proton exchange (PEM) fuel cell for online electricity generation. To investigate the fuel cell efficiency, the effect of hydrogen production on the open circuit fuel cell voltage is monitored during 27 days of batch culture. Values of volumetric hydrogen production, monitored by the help of the calibrated water columns, are related with the open circuit voltage changes of the fuel cell. From the analysis of this relation a dead end configuration is selected to use the fuel cell in its best potential. After the open circuit experiments external loads are tested for their effects on the fuel cell voltage and current generation. According to the results two external loads are selected for the direct usage of the fuel cell incorporating with the photobioreactors (PBR). Experiments with the PEM fuel cell generate a current density of 1.81 mA cm -2 for about 50 h with 10 Ω load and 0.23 mA cm -2 for about 80 h with 100 Ω load.

  16. Application of proton exchange membrane fuel cells for the monitoring and direct usage of biohydrogen produced by Chlamydomonas reinhardtii

    Energy Technology Data Exchange (ETDEWEB)

    Oncel, S.; Vardar-Sukan, F. [Department of Bioengineering, Faculty of Engineering, Ege University, 35100 Bornova, Izmir (Turkey)

    2011-01-01

    Photo-biologically produced hydrogen by Chlamydomonas reinhardtii is integrated with a proton exchange (PEM) fuel cell for online electricity generation. To investigate the fuel cell efficiency, the effect of hydrogen production on the open circuit fuel cell voltage is monitored during 27 days of batch culture. Values of volumetric hydrogen production, monitored by the help of the calibrated water columns, are related with the open circuit voltage changes of the fuel cell. From the analysis of this relation a dead end configuration is selected to use the fuel cell in its best potential. After the open circuit experiments external loads are tested for their effects on the fuel cell voltage and current generation. According to the results two external loads are selected for the direct usage of the fuel cell incorporating with the photobioreactors (PBR). Experiments with the PEM fuel cell generate a current density of 1.81 mA cm{sup -2} for about 50 h with 10 {omega} load and 0.23 mA cm{sup -2} for about 80 h with 100 {omega} load. (author)

  17. Comparison of tubular and panel type photobioreactors for biohydrogen production utilizing Chlamydomonas reinhardtii considering mixing time and light intensity.

    Science.gov (United States)

    Oncel, S; Kose, A

    2014-01-01

    Two different photobioreactor designs; tubular and panel, were investigated for the biohydrogen production utilizing a green microalgae Chlamydomonas reinhardtii strain CC124 following the two stage protocol. Mixing time and light intensity of the systems were adjusted to compare the productivity of both aerobic culture phase and the following anaerobic biohydrogen production phase. The results showed there was an effect on both phases related with the design. During the aerobic phase bigger illumination area serving more energy, tubular photobioreactor reached higher biomass productivity of 31.8±2.1 mg L(-1) h(-1) which was about 11% higher than the panel photobioreactor. On the other hand biohydrogen productivity in the panel photobioreactor reached a value of 1.3±0.05 mL L(-1) h(-1) based on the efficient removal of biohydrogen gas. According to the results it would be a good approach to utilize tubular design for aerobic phase and panel for biohydrogen production phase.

  18. Assessment of a Novel Algal Strain Chlamydomonas debaryana NIREMACC03 for Mass Cultivation, Biofuels Production and Kinetic Studies.

    Science.gov (United States)

    Mishra, Sanjeev; Singh, Neetu; Sarma, Anil Kumar

    2015-08-01

    A novel microalgae strain Chlamydomonas debaryana (KJ210856) was isolated from a freshwater lake of Punjab, India, and cultivated considering climatic sustainability and inherent adaptability concern. C. debaryana was grown in a 30-L indoor photobioreactor to study the mass cultivation prospect and biofuel potential. Physicochemical characterization of biomass and the lipid was performed with effect to nitrogen stress. It showed a higher biomass yield (1.58 ± 0.02 g L(-1), dry weight) and twofold increase in lipid yield (552.78 ± 9 mg L(-1)) with 34.2 ± 0.19 % lipid content under nitrogen deficient condition. Strikingly, increase in triglycerides achieved with nitrogen depletion containing over 96 % of total fatty acids (C 14, C 16, and C 18). Proximate and ultimate analysis suggested the presence of relatively higher volatile matter and carbon-hydrogen ratio. Furthermore, lower moisture and ash content signified C. debaryana biomass has promising features towards biofuel applications. The pyrolytic behavior of the whole biomass was also studied using thermogravimetric analyzer (TGA) and kinetic parameters were estimated using different methods. Promising growth rate and lipid yield leading to feasible biofuel feed stock production in indoor photobioreactor along with autosediment potential of cells validates C. debaryana NIREMACC03, a potential strain for mass cultivation.

  19. Expression of a clostridial [FeFe]-hydrogenase in Chlamydomonas reinhardtii prolongs photo-production of hydrogen from water splitting

    Energy Technology Data Exchange (ETDEWEB)

    Noone, Seth; Ratcliff, Kathleen; Davis, ReAnna; Subramanian, Venkataramanan; Meuser, Jonathan; Posewitz, Matthew C.; King, Paul W.; Ghirardi, Maria L.

    2017-03-01

    The high oxygen (O2) sensitivity of green algal [FeFe]-hydrogenases is a significant limitation for the sustained production of hydrogen gas (H2) from photosynthetic water splitting. To address this limitation we replaced the native [FeFe]-hydrogenases with a more O2-tolerant clostridial [FeFe]-hydrogenase CaI in Chlamydomonas reinhardtii strain D66..delta..HYD (hydA1-hydA2-) that contains insertionally inactivated [FeFe]-hydrogenases genes. Expression and translocation of CaI in D66..delta..HYD led to the recovery of H2 photoproduction at ~ 20% of the rates of the wild-type parent strain D66. We show for the first time that a bacterial [FeFe]-hydrogenase can be expressed, localized and matured to a catalytically active form that couples to photosynthetic electron transport in the green alga C. reinhardtii. The lower rates of O2 inactivation of CaI led to more sustained H2 photoproduction when cultures were challenged with O2 or kept under prolonged illumination at solar intensities. These results provide new insights into the requisites for attaining photobiological H2 production from water splitting using a more O2-tolerant hydrogenase.

  20. Intact anthracene inhibits photosynthesis in algal cells: a fluorescence induction study on Chlamydomonas reinhardtii cw92 strain.

    Science.gov (United States)

    Aksmann, Anna; Tukaj, Zbigniew

    2008-12-01

    Short-term (24h) experiments were performed to examine the effect of anthracene (ANT) on Chlamydomonas reinhardtii cw92 grown in a batch culture system aerated with 2.5% CO(2). At concentrations ranging from 0.7 to 5.6 microM, ANT inhibited the growth of population in a concentration-dependent manner and EC(50) calculated amounted to 1.6 microM. At concentrations from 0.7 to 4.2 microM ANT stimulated respiration and inhibited the intensity of photosynthesis but did not affect chlorophyll content in the cells. ANT influenced chlorophyll a fluorescence parameters, measured by OJIP test (O, J, I and P are the different steps of fluorescence induction curve). ANT diminished the performance index (PI), the yield of primary photochemistry (phi(Po)), the yield of electron transport (phi(Epsilonomicron), the efficiency of moving the electron beyond Qa(-) (Psi(0)) and the fraction of active oxygen evolving complexes (OEC). The fraction of active PS II reaction centres in the treated samples dramatically dropped. The most pronounced changes in ANT-treated cells were observed in the stimulation of energy dissipation parameter (DI(0)/RC). The only OJIP parameter that was not influenced by ANT was energy absorption by photosynthetic antennae (ABS). The results lead to a conclusion that the inhibition of photosynthesis may be a consequence of unspecific ANT-membrane interaction, resulting from hydrophobic character of this hydrocarbon.

  1. How a structured vibrational environment controls the performance of a photosystem II reaction centre-based photocell

    CERN Document Server

    Stones, Richard; van Grondelle, Rienk; Olaya-Castro, Alexandra

    2016-01-01

    Photosynthetic reaction centres are pigment-protein complexes that can transform photo-induced electronic excitations into stable charge separated states with near-unit quantum efficiency. Here we consider a theoretical photovoltaic device that places a single photosystem II reaction centre between electrodes to investigate how the mean photo-current and its fluctuations depend on the vibrational environment that assists energy and electron transfer. Our results indicate that selective coupling to well resolved vibrational modes does not necessarily offer an advantage in terms of power output but does lead to photo-currents with suppressed noise levels. The exciton manifold and the structured vibrations assisting electron transfer can also support the emergence of a phenomenon akin to dynamical channel blockade, whereby excitonic traps can impose competing routes for population transfer under steady state operation. Our results help characterizing the device-like functionality of these complexes for their pot...