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Sample records for bacteriochlorophyll synthesis genes

  1. Molecular genetic and molecular evolutionary studies on the bacteriochlorophyll synthesis genes of Rhodobacter capsulatus

    Energy Technology Data Exchange (ETDEWEB)

    Burke-Agueero, D.H.

    1992-08-01

    Rhodobacter capsulatus, purple bacterium capable of either aerobic or photosynthetic growth, has proven to be very useful in genetic studies of photosynthesis. Forty-four genes clustered together within a 46 kilobase region are required to establish photosynthetic ability in R. capsulatus. Approximately twenty of these genes are involved in bacteriochlorophyll synthesis of which eight bch'' genes are the subject of this thesis. Six of these genes were found to code for the two ring reductases. The first converts protochlorophyllide (PChlide) into a chlorin, the immediate precursor to chlorophyll a, and then into a bacteriochlorin. Each reductase is shown to be made up of three subunits. PChlide reductase is coded by the genes bchN, bchB, and bchL. Proteins with amino acid sequences markedly similar to those of bchN and bchL have been shown in other organisms to be required for chlorophyll synthesis; hence, their designation as chlN and chlB. A third chloroplast-encoded gene of heretofore unknown function shares amino acid identities with bchB and is probably the third subunit of the plant PChlide reductase. The bchA locus, which encodes the chlorin reductase, is found to be made up of three separate, translationally coupled genes, referred to as bchX, bchY, and bchZ. Amino acid similarities between bchX, bchL, and the nitrogenase reductase protein nifH suggest that all three classes of proteins share certain three-dimensional structural features, including elements that are central to the enzymatic mechanism of nifH. PChlide reductase and chlorin reductase are clearly derived from a common ancestor. Several lines of analysis suggests the ancestor of both enzyme systems reduced PChlide twice to produce bacteriochlorophyll supporting the concept bacteriochlorophyll as the ancestral reaction center pigment.

  2. Molecular genetic and molecular evolutionary studies on the bacteriochlorophyll synthesis genes of Rhodobacter capsulatus

    Energy Technology Data Exchange (ETDEWEB)

    Burke-Agueero, D.H.

    1992-08-01

    Rhodobacter capsulatus, purple bacterium capable of either aerobic or photosynthetic growth, has proven to be very useful in genetic studies of photosynthesis. Forty-four genes clustered together within a 46 kilobase region are required to establish photosynthetic ability in R. capsulatus. Approximately twenty of these genes are involved in bacteriochlorophyll synthesis of which eight ``bch`` genes are the subject of this thesis. Six of these genes were found to code for the two ring reductases. The first converts protochlorophyllide (PChlide) into a chlorin, the immediate precursor to chlorophyll a, and then into a bacteriochlorin. Each reductase is shown to be made up of three subunits. PChlide reductase is coded by the genes bchN, bchB, and bchL. Proteins with amino acid sequences markedly similar to those of bchN and bchL have been shown in other organisms to be required for chlorophyll synthesis; hence, their designation as chlN and chlB. A third chloroplast-encoded gene of heretofore unknown function shares amino acid identities with bchB and is probably the third subunit of the plant PChlide reductase. The bchA locus, which encodes the chlorin reductase, is found to be made up of three separate, translationally coupled genes, referred to as bchX, bchY, and bchZ. Amino acid similarities between bchX, bchL, and the nitrogenase reductase protein nifH suggest that all three classes of proteins share certain three-dimensional structural features, including elements that are central to the enzymatic mechanism of nifH. PChlide reductase and chlorin reductase are clearly derived from a common ancestor. Several lines of analysis suggests the ancestor of both enzyme systems reduced PChlide twice to produce bacteriochlorophyll supporting the concept bacteriochlorophyll as the ancestral reaction center pigment.

  3. IMPACT OF LIGHT DARK REGIMEN ON GROWTH-RATE, BIOMASS FORMATION AND BACTERIOCHLOROPHYLL SYNTHESIS IN ERYTHROMICROBIUM-HYDROLYTICUM

    NARCIS (Netherlands)

    YURKOV, VV; VANGEMERDEN, H

    1993-01-01

    The impact of illumination on specific growth rate, biomass formation, and synthesis of photopigment was studied in Erythromicrobium hydrolyticum, an obligately aerobic heterotrophic bacterium having the ability to synthesize bacteriochlorophyll a. In dark-grown continuous cultures the concentration

  4. Chlorobium tepidum mutant lacking bacteriochlorophyll c made by inactivation of the bchK gene, encoding bacteriochlorophyll c synthase

    DEFF Research Database (Denmark)

    Frigaard, Niels-Ulrik; Voigt, Ginny D; Bryant, Donald A

    2002-01-01

    The gene encoding bacteriochlorophyll (BChl) c synthase was identified by insertional inactivation in the photosynthetic green sulfur bacterium Chlorobium tepidum and was named bchK. The bchK mutant of C. tepidum was rusty-orange in color and completely lacked BChl c. Because of the absence of the...... found in the wild type, the bchK mutant should prove valuable for future analyses of the photosynthetic reaction center and of the roles of BChl a in photosynthesis in green bacteria. An evolutionary implication of our findings is that the photosynthetic ancestor of green sulfur bacteria could have...

  5. Mimics of the self-assembling chlorosomal bacteriochlorophylls: regio- and stereoselective synthesis and stereoanalysis of acyl(1-hydroxyalkyl)porphyrins.

    Science.gov (United States)

    Balaban, Teodor Silviu; Bhise, Anil Dnyanoba; Bringmann, Gerhard; Bürck, Jochen; Chappaz-Gillot, Cyril; Eichhöfer, Andreas; Fenske, Dieter; Götz, Daniel C G; Knauer, Michael; Mizoguchi, Tadashi; Mössinger, Dennis; Rösner, Harald; Roussel, Christian; Schraut, Michaela; Tamiaki, Hitoshi; Vanthuyne, Nicolas

    2009-10-14

    Diacylation of copper 10,20-bis(3,5-di-tert-butylphenylporphyrin) using Friedel-Crafts conditions at short reaction times, high concentrations of catalyst, and 0-4 degrees C affords only the 3,17-diacyl-substituted porphyrins, out of the 12 possible regioisomers. At longer reaction times and higher temperatures, the 3,13-diacyl compounds are also formed, and the two isomers can be conveniently separated by normal chromatographic techniques. Monoreduction of these diketones affords in good yields the corresponding acyl(1-hydroxyalkyl)porphyrins, which after zinc metalation are mimics of the natural chlorosomal bacteriochlorophyll (BChl) d. Racemate resolution by HPLC on a variety of chiral columns was achieved and further optimized, thus permitting easy access to enantiopure porphyrins. Enantioselective reductions proved to be less effective in this respect, giving moderate yields and only 79% ee in the best case. The absolute configuration of the 3(1)-stereocenter was assigned by independent chemical and spectroscopic methods. Self-assembly of a variety of these zinc BChl d mimics proves that a collinear arrangement of the hydroxyalkyl substituent with the zinc atom and the carbonyl substituent is not a stringent requirement, since both the 3,13 and the 3,17 regioisomers self-assemble readily as the racemates. Interestingly, the separated enantiomers self-assemble less readily, as judged by absorption, fluorescence, and transmission electron microscopy studies. Circular dichroism spectra of the self-assemblies show intense Cotton effects, which are mirror-images for the two 3(1)-enantiomers, proving that the supramolecular chirality is dependent on the configuration at the 3(1)-stereocenter. Upon disruption of these self-assemblies with methanol, which competes with zinc ligation, only very weak monomeric Cotton effects are present. The favored heterochiral self-assembly process may also be encountered for the natural BChls. This touches upon the long

  6. Bacteriochlorophyll dimers in photosynthesis

    International Nuclear Information System (INIS)

    The X-ray crysallagraphic study of reaction center (RC) single crystals of the photosynthetic bacteria Rps.Viridis and Rb. sphaeroides confirms the existence of bacteriochlorophyll (BChl) dimers which were postulated earlier from EPR and ENDOR studies at low temperature to be the primary electron donors P960 and P870. Apart from the spatial structure of these dimers a knowlegde of the electron density distribution in various electronic stated in indispensable for an understanding of their functional properties. For P870+ and P960+ under physiological ocnditions the electron spin density distrubutions were obtained by ENDOR-in-solution via the hyperifne couplings. The comparison between the EPR/ENDOR data of P870+ and P960+ in RC's and of onomeric BChl a. and BChl b. shows that the primary donors are pie conjugated supermolecules with more or less asymmetric spin dnesity distrubutions over the dimer halves. Theoretical spian and charge densities were calculated by an all-valence electron SCF MO method, RHF-INDO/SP, using coordinates from refined X-ray data. These calculations yield asymmetry ratios similar to those observed.Consequences of the asymmetries in the charge distribution with respect to the observed unidirectionality of the electron transfer are discussed. (author). 29 refs.; 4 figs

  7. Novel algorithms for in vitro gene synthesis

    OpenAIRE

    Thachuk, Chris

    2007-01-01

    Methods for reliable synthesis of long genes offer great promise for novel protein synthesis via expression of synthetic genes. Current technologies use computational methods for design of short oligos, which can then be reliably synthesized and assembled into the desired target gene. A precursor to this process is optimization of the gene sequence for improved protein expression. In this thesis, we provide the first results on the computational complexity of oligo design for gene synthesis. ...

  8. Mutational analysis of three bchH paralogs in (bacterio-)chlorophyll biosynthesis in Chlorobaculum tepidum

    DEFF Research Database (Denmark)

    Gomez Maqueo Chew, Aline; Frigaard, Niels-Ulrik; Bryant, Donald A

    2009-01-01

    The first committed step in the biosynthesis of (bacterio-)chlorophyll is the insertion of Mg2+ into protoporphyrin IX by Mg-chelatase. In all known (B)Chl-synthesizing organisms, Mg-chelatase is encoded by three genes that are homologous to bchH, bchD, and bchI of Rhodobacter spp. The genomes of...

  9. Homeobox genes and melatonin synthesis

    DEFF Research Database (Denmark)

    Rohde, Kristian; Møller, Morten; Rath, Martin Fredensborg

    2014-01-01

    Nocturnal synthesis of melatonin in the pineal gland is controlled by a circadian rhythm in arylalkylamine N-acetyltransferase (AANAT) enzyme activity. In the rodent, Aanat gene expression displays a marked circadian rhythm; release of norepinephrine in the gland at night causes a c......AMP-based induction of Aanat transcription. However, additional transcriptional control mechanisms exist. Homeobox genes, which are generally known to encode transcription factors controlling developmental processes, are also expressed in the mature rodent pineal gland. Among these, the cone-rod homeobox (CRX......) transcription factor is believed to control pineal-specific Aanat expression. Based on recent advances in our understanding of Crx in the rodent pineal gland, we here suggest that homeobox genes play a role in adult pineal physiology both by ensuring pineal-specific Aanat expression and by facilitating c...

  10. The bchU gene of Chlorobium tepidum encodes the C-20 methyltransferase in bacteriochlorophyll c biosynthesis

    DEFF Research Database (Denmark)

    Maresca, Julia A; Gomez Maqueo Chew, Aline; Ponsatí, Marta Ros;

    2004-01-01

    that restores the correct reading frame in bchU. The bchU gene was inactivated in C. tepidum, a BChl c-producing species, and the resulting mutant produced only BChl d. Growth rate measurements showed that BChl c- and d-producing strains of the same organism (C. tepidum or C. vibrioforme) have similar...... c rather than BChl d confers a significant competitive advantage to green sulfur bacteria living at limiting red and near-infrared light intensities....

  11. Benchmarking calculations of excitonic couplings between bacteriochlorophylls

    OpenAIRE

    Kenny, Elise P.; Kassal, Ivan

    2015-01-01

    Excitonic couplings between (bacterio)chlorophyll molecules are necessary for simulating energy transport in photosynthetic complexes. Many techniques for calculating the couplings are in use, from the simple (but inaccurate) point-dipole approximation to fully quantum-chemical methods. We compared several approximations to determine their range of applicability, noting that the propagation of experimental uncertainties poses a fundamental limit on the achievable accuracy. In particular, the ...

  12. Benchmarking calculations of excitonic couplings between bacteriochlorophylls

    CERN Document Server

    Kenny, Elise P

    2015-01-01

    Excitonic couplings between (bacterio)chlorophyll molecules are necessary for simulating energy transport in photosynthetic complexes. Many techniques for calculating the couplings are in use, from the simple (but inaccurate) point-dipole approximation to fully quantum-chemical methods. We compared several approximations to determine their range of applicability, noting that the propagation of experimental uncertainties poses a fundamental limit on the achievable accuracy. In particular, the uncertainty in crystallographic coordinates yields an uncertainty of about 20% in the calculated couplings. Because quantum-chemical corrections are smaller than 20% in most biologically relevant cases, their considerable computational cost is rarely justified. We therefore recommend the electrostatic TrEsp method across the entire range of molecular separations and orientations because its cost is minimal and it generally agrees with quantum-chemical calculations to better than the geometric uncertainty. We also caution ...

  13. Benchmarking Calculations of Excitonic Couplings between Bacteriochlorophylls.

    Science.gov (United States)

    Kenny, Elise P; Kassal, Ivan

    2016-01-14

    Excitonic couplings between (bacterio)chlorophyll molecules are necessary for simulating energy transport in photosynthetic complexes. Many techniques for calculating the couplings are in use, from the simple (but inaccurate) point-dipole approximation to fully quantum-chemical methods. We compared several approximations to determine their range of applicability, noting that the propagation of experimental uncertainties poses a fundamental limit on the achievable accuracy. In particular, the uncertainty in crystallographic coordinates yields an uncertainty of about 20% in the calculated couplings. Because quantum-chemical corrections are smaller than 20% in most biologically relevant cases, their considerable computational cost is rarely justified. We therefore recommend the electrostatic TrEsp method across the entire range of molecular separations and orientations because its cost is minimal and it generally agrees with quantum-chemical calculations to better than the geometric uncertainty. Understanding these uncertainties can guard against striving for unrealistic precision; at the same time, detailed benchmarks can allow important qualitative questions-which do not depend on the precise values of the simulation parameters-to be addressed with greater confidence about the conclusions. PMID:26651217

  14. From chlorophyll a towards bacteriochlorophyll a: Excited-state processes of modified pigments

    OpenAIRE

    Teuchner, K.; Stiel, H.; Leupold, D; Katheder, Ingrid; Scheer, Hugo

    1994-01-01

    By means of fluorescence spectroscopy and nonlinear absorption experiments, excited-state processes of the modified pigments [3-acetyl]-chlorophyll a, [31-OH]-bacteriochlorophyll a and [3-vinyl]-bacteriochlorophyll a were investigated and compared with those of chlorophyll a and bacteriochlorophyll a.

  15. Bacteriochlorophyll f: properties of chlorosomes containing the "forbidden chlorophyll"

    Directory of Open Access Journals (Sweden)

    Kajetan Vogl

    2012-08-01

    Full Text Available The chlorosomes of green sulfur bacteria are mainly assembled from one of three types of bacteriochlorophylls, BChls c, d, and e. By analogy to the relationship between BChl c and BChl d (20-desmethyl-BChl c, a fourth type of BChl, BChl f (20-desmethyl-BChl e, should exist but has not yet been observed in nature. The bchU gene (bacteriochlorophyllide C-20 methyltransferase of the brown-colored green sulfur bacterium Chlorobaculum limnaeum was inactivated by conjugative transfer from Eshcerichia coli and homologous recombination of a suicide plasmid carrying a portion of the bchU. The resulting bchU mutant was greenish brown in color and synthesized BChl fF. The chlorosomes of the bchU mutant had similar size and polypeptide composition as those of the wild type (WT, but the Qy absorption band of the BChl f aggregates was blue-shifted 16 nm (705 nm vs. 721 nm for the WT. Fluorescence spectroscopy showed that energy transfer to the baseplate was much less efficient in chlorosomes containing BChl f than in WT chlorosomes containing BChl e. When cells were grown at high irradiance with tungsten or fluorescent light, the WT and bchU mutant had identical growth rates. However, the WT grew about 50% faster than the bchU mutant at low irradiance (10 µmol photons m-2 s-1. Less efficient energy transfer from BChl f aggregates to BChl a in the baseplate, the much slower growth of the strain producing BChl f relative to the WT, and competition from other phototrophs, may explain why BChl f is not observed naturally.

  16. Integrating Gene Synthesis & Microfluidic Protein Analysis for Rapid Protein Engineering

    OpenAIRE

    Matthew Blackburn

    2015-01-01

    The ability to rapidly design proteins with novel functions will have a significant impact on medicine, biotechnology, and synthetic biology. While synthetic genes are becoming a commodity, integrated approaches that take full advantage of gene synthesis have yet to be produced. We developed a bench-top, solid-phase gene synthesis method based on asymmetric primer extension (APE) and demonstrated that linear templates from this technique can be used directly for high-throughput, on-chip prote...

  17. Direct observation of monomer film structure of bacteriochlorophyll c

    Institute of Scientific and Technical Information of China (English)

    JIANG Yaxin; XU Qingmin; WAN Lijun; WANG Chen; FANG Xiaohong; BAI Chunli; WANG Zhengyu; Nozawa Tsunenori

    2003-01-01

    The adsorption and assembling of bacteriochlorophyll c (BChl c) on a highly oriented pyrolytic graphite (HOPG) surface have been investigated by the scanning tunneling microscopy (STM). BChl c molecules are found to self-organize a well-defined adlayer on HOPG surface with a monomeric structure different fromthe dimer association seen on adlayers of Chls a, c and BChlide c, d. A structural model is proposed for the adlayer.

  18. Computational study of short-range interactions in bacteriochlorophyll aggregates

    Czech Academy of Sciences Publication Activity Database

    Alster, J.; Kabeláč, Martin; Tůma, R.; Pšenčík, J.; Burda, J. V.

    2012-01-01

    Roč. 998, SI (2012), s. 87-97. ISSN 2210-271X R&D Projects: GA AV ČR IAA400550808 Grant ostatní: GA ČR(CZ) GA206/09/0375; GA MŠk(CZ) ME10149 Institutional support: RVO:61388963 Keywords : chlorosome * bacteriochlorophyll * molecular simulations Subject RIV: CF - Physical ; Theoretical Chemistry Impact factor: 1.139, year: 2012

  19. Origin of Bacteriochlorophyll a and the Early Diversification of Photosynthesis.

    Directory of Open Access Journals (Sweden)

    Tanai Cardona

    Full Text Available Photosynthesis originated in the domain Bacteria billions of years ago; however, the identity of the last common ancestor to all phototrophic bacteria remains undetermined and speculative. Here I present the evolution of BchF or 3-vinyl-bacteriochlorophyll hydratase, an enzyme exclusively found in bacteria capable of synthetizing bacteriochlorophyll a. I show that BchF exists in two forms originating from an early divergence, one found in the phylum Chlorobi, including its paralogue BchV, and a second form that was ancestral to the enzyme found in the remaining anoxygenic phototrophic bacteria. The phylogeny of BchF is consistent with bacteriochlorophyll a evolving in an ancestral phototrophic bacterium that lived before the radiation event that gave rise to the phylum Chloroflexi, Chlorobi, Acidobacteria, Proteobacteria, and Gemmatimonadetes, but only after the divergence of Type I and Type II reaction centers. Consequently, it is suggested that the lack of phototrophy in many groups of extant bacteria is a derived trait.

  20. Enzyme-catalyzed organic syntheses: transesterification reactions of chlorophyl a, bacteriochlorophyll a, and derivatives with chlorophyllase

    Energy Technology Data Exchange (ETDEWEB)

    Michalski, T.J.; Hunt, J.E.; Bradshaw, C.; Wagner, A.M.; Norris, J.R.; Katz, J.J.

    1988-08-17

    The green plant enzyme chlorophyllase (EC 3.1.1.14, chlorophyll chlorophyllido-hydroase) has been used for the synthesis of a variety of primary alcohol and diol esters of chlorophyll a, bacteriochlorophyll a, and pyrobacteriochlorophyll a. Green plant chlorophyllase accepts a much larger range of alcohol and chlorophyll substrates than had previously been realized. Thus, chlorophyllide and bacteriochlorophyllide esters of primary alcohols such as retinol and the detergent Triton X-100 and of dihydric alcohols such as ethylene glycol, butanediol, or 2-hydroxyethyl disulfide can readily be obtained by enzyme-assisted transesterification. The diol chlorophyllide esters are valuable intermediates for the synthesis of reaction center special pair models. Chlorophyllase-assisted reactions can be carried out in media containing up to 95% of organic solvents without the concomitant side reactions that important chlorophyll functional groups readily undergo even under mild conditions in conventional chemical synthetic procedures. In competitive chlorophyllase-catalyzed transesterification reactions, long-chain alcohols such as farnesol and retinol vs simple aliphatic alcohols and diols, the enzyme shows a definite preference for the long-chain alcohol. 37 references, 1 figure, 2 tables.

  1. Quenching of bacteriochlorophyll fluorescence in chlorosomes from Chloroflexus aurantiacus by exogenous quinones

    DEFF Research Database (Denmark)

    Tokita, S; Frigaard, N-U; Hirota, M;

    2000-01-01

    The quenching of bacteriochlorophyll (BChl) c fluorescence in chlorosomes isolated from Chloroflexus aurantiacus was examined by the addition of various benzoquinones, naphthoquinones (NQ), and anthraquinones (AQ). Many quinones showed strong quenching in the micromolar or submicromolar range. Th...

  2. {beta}-Carotene to bacteriochlorophyll c energy transfer in self-assembled aggregates mimicking chlorosomes

    Energy Technology Data Exchange (ETDEWEB)

    Alster, J. [Faculty of Mathematics and Physics, Charles University, Ke Karlovu 3, 121 16 Praha (Czech Republic); Polivka, T. [Institute of Physical Biology, University of South Bohemia, Zamek 136, 373 33 Nove Hrady (Czech Republic); Biology Centre, Academy of Sciences of the Czech Republic, Branisovska 31, 370 05 Ceske Budejovice (Czech Republic); Arellano, J.B. [Instituto de Recursos Naturales y Agrobiologia de Salamanca (IRNASA-CSIC), Apdo. 257, 37071 Salamanca (Spain); Chabera, P. [Institute of Physical Biology, University of South Bohemia, Zamek 136, 373 33 Nove Hrady (Czech Republic); Vacha, F. [Institute of Physical Biology, University of South Bohemia, Zamek 136, 373 33 Nove Hrady (Czech Republic); Biology Centre, Academy of Sciences of the Czech Republic, Branisovska 31, 370 05 Ceske Budejovice (Czech Republic); Psencik, J., E-mail: psencik@karlov.mff.cuni.cz [Faculty of Mathematics and Physics, Charles University, Ke Karlovu 3, 121 16 Praha (Czech Republic); Institute of Physical Biology, University of South Bohemia, Zamek 136, 373 33 Nove Hrady (Czech Republic)

    2010-07-19

    Carotenoids are together with bacteriochlorophylls important constituents of chlorosomes, the light-harvesting antennae of green photosynthetic bacteria. Majority of bacteriochlorophyll molecules form self-assembling aggregates inside the chlorosomes. Aggregates of bacteriochlorophylls with optical properties similar to those of chlorosomes can also be prepared in non-polar organic solvents or in aqueous environments when a suitable non-polar molecule is added. In this work, the ability of {beta}-carotene to induce aggregation of bacteriochlorophyll c in aqueous buffer was studied. Excitation relaxation and energy transfer in the carotenoid-bacteriochlorophyll assemblies were measured using femtosecond and nanosecond transient absorption spectroscopy. A fast, {approx}100-fs energy transfer from the S{sub 2} state of {beta}-carotene to bacteriochlorophyll c was revealed, while no evidence for significant energy transfer from the S{sub 1} state was found. Picosecond formation of the carotenoid triplet state (T{sub 1}) was observed, which was likely generated by singlet homo-fission from the S{sub 1} state of {beta}-carotene.

  3. De novo gene synthesis design using TmPrime software.

    Science.gov (United States)

    Li, Mo-Huang; Bode, Marcus; Huang, Mo Chao; Cheong, Wai Chye; Lim, Li Shi

    2012-01-01

    This chapter presents TmPrime, a computer program to design oligonucleotide for both ligase chain reaction (LCR)- and polymerase chain reaction (PCR)-based de novo gene synthesis. The program divides a long input DNA sequence based on user-specified melting temperatures and assembly conditions, and dynamically optimizes the length of oligonucleotides to achieve homologous melting temperatures. The output reports the melting temperatures, oligonucleotide sequences, and potential formation of secondary structures in a PDF file, which will be sent to the user via e-mail. The program also provides functions on sequence pooling to separate long genes into smaller pieces for multipool assembly and codon optimization for expression based on the highest organism-specific codon frequency. This software has been successfully used in the design and synthesis of various genes with total length >20 kbp. This program is freely available at http://prime.ibn.a-star.edu.sg. PMID:22328437

  4. Ultrafast intramolecular relaxation dynamics of Mg- and Zn-bacteriochlorophyll a

    Energy Technology Data Exchange (ETDEWEB)

    Kosumi, Daisuke [Osaka City University Advanced Research Institute for Natural Science and Technology, 3-3-138 Sugimoto, Sumiyoshi-ku, Osaka 558-8585 (Japan); CREST/JST, 4-1-8 Honcho, Kawaguchi, Saitama 332-0012 (Japan); Nakagawa, Katsunori; Sakai, Shunsuke [Graduate School of Engineering, Nagoya Institute of Technology, Gokiso-cho, Showa-ku, Nagoya 466-8555 (Japan); Nagaoka, Yuya; Maruta, Satoshi; Sugisaki, Mitsuru [CREST/JST, 4-1-8 Honcho, Kawaguchi, Saitama 332-0012 (Japan); Department of Physics, Graduate School of Science, Osaka City University, 3-3-138 Sugimoto, Sumiyoshi-ku, Osaka 558-8585 (Japan); Dewa, Takehisa [Graduate School of Engineering, Nagoya Institute of Technology, Gokiso-cho, Showa-ku, Nagoya 466-8555 (Japan); PRESTO/JST, 4-1-8 Honcho, Kawaguchi, Saitama 332-0012 (Japan); Nango, Mamoru [The Osaka City University Advanced Research Institute for Natural Science and Technology, 3-3-138 Sugimoto, Sumiyoshi-ku, Osaka 558-8585 (Japan); CREST/JST, 4-1-8 Honcho, Kawaguchi, Saitama 332-0012 (Japan); Graduate School of Engineering, Nagoya Institute of Technology, Gokiso-cho, Showa-ku, Nagoya 466-8555 (Japan); Hashimoto, Hideki [The Osaka City University Advanced Research Institute for Natural Science and Technology, 3-3-138 Sugimoto, Sumiyoshi-ku, Osaka 558-8585 (Japan); CREST/JST, 4-1-8 Honcho, Kawaguchi, Saitama 332-0012 (Japan); Department of Physics, Graduate School of Science, Osaka City University, 3-3-138 Sugimoto, Sumiyoshi-ku, Osaka 558-8585 (Japan)

    2013-07-21

    Ultrafast excited-state dynamics of the photosynthetic pigment (Mg-)bacteriochlorophyll a and its Zn-substituted form were investigated by steady-state absorption/fluorescence and femtosecond pump-probe spectroscopic measurements. The obtained steady-state absorption and fluorescence spectra of bacteriochlorophyll a in solution showed that the central metal compound significantly affects the energy of the Q{sub x} state, but has almost no effect on the Q{sub y} state. Photo-induced absorption spectra were recorded upon excitation of Mg- and Zn-bacteriochlorophyll a into either their Q{sub x} or Q{sub y} state. By comparing the kinetic traces of transient absorption, ground-state beaching, and stimulated emission after excitation to the Q{sub x} or Q{sub y} state, we showed that the Q{sub x} state was substantially incorporated in the ultrafast excited-state dynamics of bacteriochlorophyll a. Based on these observations, the lifetime of the Q{sub x} state was determined to be 50 and 70 fs for Mg- and Zn-bacteriochlorophyll a, respectively, indicating that the lifetime was influenced by the central metal atom due to the change of the energy gap between the Q{sub x} and Q{sub y} states.

  5. Ultrafast intramolecular relaxation dynamics of Mg- and Zn-bacteriochlorophyll a

    International Nuclear Information System (INIS)

    Ultrafast excited-state dynamics of the photosynthetic pigment (Mg-)bacteriochlorophyll a and its Zn-substituted form were investigated by steady-state absorption/fluorescence and femtosecond pump-probe spectroscopic measurements. The obtained steady-state absorption and fluorescence spectra of bacteriochlorophyll a in solution showed that the central metal compound significantly affects the energy of the Qx state, but has almost no effect on the Qy state. Photo-induced absorption spectra were recorded upon excitation of Mg- and Zn-bacteriochlorophyll a into either their Qx or Qy state. By comparing the kinetic traces of transient absorption, ground-state beaching, and stimulated emission after excitation to the Qx or Qy state, we showed that the Qx state was substantially incorporated in the ultrafast excited-state dynamics of bacteriochlorophyll a. Based on these observations, the lifetime of the Qx state was determined to be 50 and 70 fs for Mg- and Zn-bacteriochlorophyll a, respectively, indicating that the lifetime was influenced by the central metal atom due to the change of the energy gap between the Qx and Qy states

  6. Characterization of an FMO Variant of Chlorobaculum tepidum Carrying Bacteriochlorophyll a Esterified by Geranylgeraniol

    Energy Technology Data Exchange (ETDEWEB)

    Wen, Jianzhong [Washington Univ., St. Louis, MO (United States); Harada, Jiro [Ritsumeikan Univ., Shiga (Japan); Buyle, Kenny [Washington Univ., St. Louis, MO (United States); Yuan, Kevin [Washington Univ., St. Louis, MO (United States); Tamiaki, Hitoshi [Ritsumeikan Univ., Shiga (Japan); Oh-oka, Hirozo [Osaka Univ., Suita (Japan); Loomis, Richard A [Washington Univ., St. Louis, MO (United States); Blankenship, R. E. [Washington Univ., St. Louis, MO (United States)

    2010-06-15

    The Fenna-Matthews-Olson light-harvesting antenna (FMO) protein has been a model system for understanding pigment-protein interactions in the energy transfer process in photosynthesis. All previous studies have utilized wild-type FMO proteins from several species. Here we report the purification and characterization of the first FMO protein variant generated via replacement of the esterifying alcohol at the C-17 propionate residue of bacteriochlorophyll (BChl) a, phytol, with geranylgeraniol, which possesses three more double bonds. The FMO protein still assembles with the modified pigment, but both the whole cell absorption and the biochemical purification indicate that the mutant cells contain a much less mature FMO protein. The gene expression was checked using qRT-PCR, and none of the genes encoding BChl a-binding proteins are strongly regulated at the transcriptional level. The smaller amount of the FMO protein in the mutant cell is probably due to the degradation of the apo-FMO protein at different stages after it does not bind the normal pigment. The absorption, fluorescence, and CD spectra of the purified FMO variant protein are similar to those of the wild-type FMO protein except the conformations of most pigments are more heterogeneous, which broadens the spectral bands. Interestingly, the lowest-energy pigment binding site seems to be unchanged and is the only peak that can be well resolved in 77 K absorption spectra. The excited-state lifetime of the variant FMO protein is unchanged from that of the wild type and shows a temperature-dependent modulation similar to that of the wild type. The variant FMO protein is less thermally stable than the wild type. The assembly of the FMO protein and also the implications of the decreased FMO/chlorosome stoichiometry are discussed in terms of the topology of these two antennas on the cytoplasmic membrane.

  7. Energy, genes and evolution: introduction to an evolutionary synthesis.

    Science.gov (United States)

    Lane, Nick; Martin, William F; Raven, John A; Allen, John F

    2013-07-19

    Life is the harnessing of chemical energy in such a way that the energy-harnessing device makes a copy of itself. No energy, no evolution. The 'modern synthesis' of the past century explained evolution in terms of genes, but this is only part of the story. While the mechanisms of natural selection are correct, and increasingly well understood, they do little to explain the actual trajectories taken by life on Earth. From a cosmic perspective-what is the probability of life elsewhere in the Universe, and what are its probable traits?-a gene-based view of evolution says almost nothing. Irresistible geological and environmental changes affected eukaryotes and prokaryotes in very different ways, ones that do not relate to specific genes or niches. Questions such as the early emergence of life, the morphological and genomic constraints on prokaryotes, the singular origin of eukaryotes, and the unique and perplexing traits shared by all eukaryotes but not found in any prokaryote, are instead illuminated by bioenergetics. If nothing in biology makes sense except in the light of evolution, nothing in evolution makes sense except in the light of energetics. This Special Issue of Philosophical Transactions examines the interplay between energy transduction and genome function in the major transitions of evolution, with implications ranging from planetary habitability to human health. We hope that these papers will contribute to a new evolutionary synthesis of energetics and genetics. PMID:23754807

  8. Probing the bacteriochlorophyll binding site by reconstitution of the light-harvesting complex of Rhodospirillum rubrum with bacteriochlorophyll a analogues

    International Nuclear Information System (INIS)

    Structural features of bacteriochlorophyll (BChl) a that are required for binding to the light-harvesting proteins of Rhodospirillum rubrum were determined by testing for reconstitution of the B873 or B820 (structural subunit of B873) light-harvesting complexes with BChl a analogues. The results indicate that the binding site is very specific; of the analogues tested, only derivatives of BChl a with ethyl, phytyl, and geranylgeranyl esterifying alcohols and BChl b (phytyl) successfully reconstituted to form B820-and B873-type complexes. BChl analogues lacking magnesium, the C-3 acetyl group, or the C132 carbomethoxy group did not reconstitute to form B820 or B873. Also unreactive were 132-hydroxy BChl a and 3-acetylchlorophyll a. Competition experiments showed that several of these nonreconstituting analogues significantly slowed BChl a binding to form B820 and blocked BChl a-B873 formation, indicating that the analogues may competitively bind to the protein even though they do not form red-shifted complexes. With the R. rubrum polypeptides, BChl b formed complexes that were further red-shifted than those of BChl a; however, the energies of the red shifts, binding behavior, and circular dichroism (CD) spectra were similar. B873 complexes reconstituted with the geranylgeranyl BChl a derivative, which contains the native esterifying alcohol for R. rubrum, showed in-vivo-like CD features, but the phytyl and ethyl B873 complexes showed inverted CD features in the near infrared. The B820 complex with the ethyl derivative was about 30-fold less stable than the two longer esterifying alcohol derivatives, but all formed stable B873 complexes

  9. Circular dichroism study on the diastereoselective self-assembly of bacteriochlorophyll cs

    Science.gov (United States)

    Balaban, Teodor S.; Holzwarth, Alfred R.; Schaffner, Kurt

    1995-04-01

    Circular dichroism (CD) spectra of self-assembled bacteriochlorophyll cs (BChl cs) aggregates show a pronounced dependency on the solvent, the concentration and on the stereochemistry of the 3 1-hydroxy groups. In n-hexane a psi-type CD is obtained due to the formation of nanostructural aggregates.

  10. Bacteriochlorophyll Aggregates Self-Assembled on Functionalized Gold Nanorod Cores as Mimics of Photosynthetic Chlorosomal Antennae: A Single Molecule Study

    Czech Academy of Sciences Publication Activity Database

    Furumaki, S.; Vácha, František; Hirata, S.; Vácha, M.

    2014-01-01

    Roč. 8, č. 3 (2014), s. 2176-2182. ISSN 1936-0851 Institutional support: RVO:60077344 Keywords : Single molecule spectroscopy * molecular aggregate * bacteriochlorophyll * chromosome Subject RIV: BO - Biophysics Impact factor: 12.881, year: 2014

  11. Transformation of Streptococcus zooepidemicus with Genes Responsible for Polyhydroxybutyrate Synthesis

    Institute of Scientific and Technical Information of China (English)

    吴小明; 高海军; 田格; 陈国强

    2002-01-01

    A procedure for transformation of intact Streptococcus zooepidemicus cells by electroporation was developed through a systematic examination of the effects of various parameters, including growth conditions, electric field strengths used for electroporation, and concentrations of plasmid used for transformation. Efficiencies higher than 104 cfu/μg(cfu, clone forming unit) plasmid DNA were obtained for Streptococcus zooepidemicus H2004 cells. Results demonstrate that the broad-host-range plasmid pDL276 can be replicated in Streptococcus zooepidemicus H2004 and foreign genes responsible for polyhydroxybutyrate (PHB) synthesis inserted into the pDL276 can be successfully expressed in the transformant, in which PHB is detected using the Fourier transform-infrared spectroscopy (FT-IR) method.

  12. PCR-based gene synthesis to produce recombinant proteins for crystallization

    Directory of Open Access Journals (Sweden)

    Byrne-Steele Miranda L

    2008-04-01

    Full Text Available Abstract Background Gene synthesis technologies are an important tool for structural biology projects, allowing increased protein expression through codon optimization and facilitating sequence alterations. Existing methods, however, can be complex and not always reproducible, prompting researchers to use commercial suppliers rather than synthesize genes themselves. Results A PCR-based gene synthesis method, referred to as SeqTBIO, is described to efficiently assemble the coding regions of two novel hyperthermophilic proteins, PAZ (Piwi/Argonaute/Zwille domain, a siRNA-binding domain of an Argonaute protein homologue and a deletion mutant of a family A DNA polymerase (PolA. The gene synthesis procedure is based on sequential assembly such that homogeneous DNA products can be obtained after each synthesis step without extensive manipulation or purification requirements. Coupling the gene synthesis procedure to in vivo homologous recombination techniques allows efficient subcloning and site-directed mutagenesis for error correction. The recombinant proteins of PAZ and PolA were subsequently overexpressed in E. coli and used for protein crystallization. Crystals of both proteins were obtained and they were suitable for X-ray analysis. Conclusion We demonstrate, by using PAZ and PolA as examples, the feasibility of integrating the gene synthesis, error correction and subcloning techniques into a non-automated gene to crystal pipeline such that genes can be designed, synthesized and implemented for recombinant expression and protein crystallization.

  13. Multireference excitation energies for Bacteriochlorophylls A within light harvesting system 2

    DEFF Research Database (Denmark)

    Anda, Andre; Hansen, Thorsten; De Vico, Luca

    2016-01-01

    Light-harvesting system 2 (LH2) of purple bacteria is one of the most popular antenna complexes used to study Nature's way of collecting and channeling solar energy. The dynamics of the absorbed energy is probed by ultrafast spectroscopy. Simulation of these experiments relies on fitting a range of...... parameters to reproduce the spectra. Here, we present a method that can determine key parameters to chemical accuracy. These will eliminate free variables in the modeling, thus reducing the problem. Using MS-RASPT2/RASSCF calculations, we compute excitation energies and transition dipole moments of all...... bacteriochlorophylls in LH2. We find that the excitation energies vary among the bacteriochlorophyll monomers and that they are regulated by the curvature of the macrocycle ring and the dihedral angle of an acetyl moiety. Increasing the curvature lifts the ground state energy, which causes a red shift of the...

  14. Spectral properties of bacteriochlorophyll c in nematic liquid crystal. Part 1. Monomeric forms of dye

    Science.gov (United States)

    Dudkowiak, A.; Francke, C.; Amesz, J.; Planner, A.; Hanyz, I.; Fraçkowiak, D.

    1996-02-01

    The spectroscopic features of bacteriochlorophyll c and bacteriopheophytin c in a nematic liquid crystal matrix have been investigated. Absorption, circular dichroism, fluorescence and time resolved delayed luminescence spectra have been measured. The pigment is introduced to the liquid crystal from a dry and from a hydrated chloroform solution. In both cases the pigment is in the monomeric form. Hydration of the solvent and the presence or absence of the central Mg atom affect the interaction of the pigment molecules with the liquid crystal matrix, changing the fluorescence anisotropy. A model for the bacteriochlorophyll c orientation in the liquid crystal is proposed and the averaged angles between the transition moments and the liquid crystal orientation axis are determined. A slow process (in the microsecond range) of radiative deactivation of energy absorbed by the pigments is observed. This delayed emission could be due to pigment ionization and delayed charge recombination and/or thermal activation from the triplet to the excited singlet state.

  15. Blastomonas aquatica sp. nov., a bacteriochlorophyll-containing bacterium isolated from lake water.

    Science.gov (United States)

    Xiao, Na; Liu, Yongqin; Liu, Xiaobo; Gu, Zhengquan; Jiao, Nianzhi; Liu, Hongcan; Zhou, Yuguang; Shen, Liang

    2015-05-01

    Yellow or orange-to-brown pigmented, ovoid or rod-shaped, Gram-negative staining, aerobic strains PE 4-5(T) and N5-10 m-1 were isolated from brackish water in Lake Peng Co and fresh to brackish water in Lake Namtso on the Tibetan Plateau, China. Bacteriochlorophyll a was produced by the isolates. The predominant cellular fatty acids were C16 : 1, C17 : 1 and C18 : 1 unsaturated fatty acids, C17 : 1ω6c (55.3%), C17 : 1ω8c (13.0%) and C18 : 1ω7c (10.4%) for PE 4-5(T) and C18 : 1ω7c (54.7%) and C16 : 1ω7c (18.0%) for N5-10 m-1. The polar lipid profiles of strains PE 4-5(T) and N5-10 m-1 were composed of diphosphatidylglycerol, phosphatidylcholine (not detected in N5-10 m-1), phosphatidyldimethylethanolamine, phosphatidylethanolamine, phosphatidylmonomethylethanolamine, phosphatidylglycerol, sphingoglycolipid and an unknown phospholipid. The predominant respiratory quinone was ubiquinone Q10 and the DNA G+C content was 66.0 mol% for both strains. The16S rRNA gene sequence of strain PE 4-5(T) shared 99.0% similarity with that of N5-10 m-1, and 97.56% similarity with those of Blastomonas natatoria LMG 17322(T) and Blastomonas ursincola DSM 9006(T), respectively. The DNA-DNA hybridization relatedness between strains PE 4-5(T) and N5-10 m-1 was 79.0 ± 1.0%, but below 70% with the type strains in the genus Blastomonas . Based on the variability of phylogenetic and phenotypic characteristics, the isolates should be classified as representatives of a novel species of the genus Blastomonas; the name Blastomonas aquatica sp. nov. is proposed. The type strain is PE 4-5(T) ( =JCM 30179(T) =CGMCC 1.12851(T)). PMID:25724744

  16. Identification of sugarcane genes involved in the purine synthesis pathway

    Directory of Open Access Journals (Sweden)

    Mario A. Jancso

    2001-12-01

    Full Text Available Nucleotide synthesis is of central importance to all cells. In most organisms, the purine nucleotides are synthesized de novo from non-nucleotide precursors such as amino acids, ammonia and carbon dioxide. An understanding of the enzymes involved in sugarcane purine synthesis opens the possibility of using these enzymes as targets for chemicals which may be effective in combating phytopathogen. Such an approach has already been applied to several parasites and types of cancer. The strategy described in this paper was applied to identify sugarcane clusters for each step of the de novo purine synthesis pathway. Representative sequences of this pathway were chosen from the National Center for Biotechnology Information (NCBI database and used to search the translated sugarcane expressed sequence tag (SUCEST database using the available basic local alignment search tool (BLAST facility. Retrieved clusters were further tested for the statistical significance of the alignment by an implementation (PRSS3 of the Monte Carlo shuffling algorithm calibrated using known protein sequences of divergent taxa along the phylogenetic tree. The sequences were compared to each other and to the sugarcane clusters selected using BLAST analysis, with the resulting table of p-values indicating the degree of divergence of each enzyme within different taxa and in relation to the sugarcane clusters. The results obtained by this strategy allowed us to identify the sugarcane proteins participating in the purine synthesis pathway.A via de síntese de purino nucleotídeos é considerada uma via de central importância para todas as células. Na maioria dos organismos, os purino nucleotídeos são sintetizados ''de novo'' a partir de precursores não-nucleotídicos como amino ácidos, amônia e dióxido de carbono. O conhecimento das enzimas envolvidas na via de síntese de purinas da cana-de-açúcar vai abrir a possibilidade do uso dessas enzimas como alvos no desenho

  17. recA gene product is responsible for inhibition of deoxyribonucleic acid synthesis after ultraviolet irradiation.

    OpenAIRE

    Trgovcević, Z; Petranović, D; Petranović, M; Salaj-Smic, E

    1980-01-01

    Deoxyribonucleic acid synthesis after ultraviolet irradiation was studied in wild-type, uvrA, recB, recA recB, and recA Escherichia coli strains. Inhibition of deoxyribonucleic acid synthesis, which occurs almost immediately after exposing the cells to ultraviolet radiation, depends on the functional gene recA.

  18. recA gene product is responsible for inhibition of deoxyribonucleic acid synthesis after ultraviolet irradiation

    International Nuclear Information System (INIS)

    Deoxyribonucleic acid synthesis after ultraviolet irradiation was studied in wild-type, uvrA, recB, recA, recB, and recA Escherichia coli strains. Inhibition of deoxyribonucleic acid synthesis, which occurs almost immediately after exposing the cells to ultraviolet radiation, depends on the functional gene recA

  19. Functional Analysis of Sinorhizobium meliloti Genes Involved in Biotin Synthesis and Transport

    OpenAIRE

    Entcheva, Plamena; Phillips, Donald A.; Streit, Wolfgang R.

    2002-01-01

    External biotin greatly stimulates bacterial growth and alfalfa root colonization by Sinorhizobium meliloti strain 1021. Several genes involved in responses to plant-derived biotin have been identified in this bacterium, but no genes required for biotin transport are known, and not all loci required for biotin synthesis have been assigned. Searches of the S. meliloti genome database in combination with complementation tests of Escherichia coli biotin auxotrophs indicate that biotin synthesis ...

  20. Combinatorial codon scrambling enables scalable gene synthesis and amplification of repetitive proteins

    Science.gov (United States)

    Tang, Nicholas C.; Chilkoti, Ashutosh

    2016-04-01

    Most genes are synthesized using seamless assembly methods that rely on the polymerase chain reaction (PCR). However, PCR of genes encoding repetitive proteins either fails or generates nonspecific products. Motivated by the need to efficiently generate new protein polymers through high-throughput gene synthesis, here we report a codon-scrambling algorithm that enables the PCR-based gene synthesis of repetitive proteins by exploiting the codon redundancy of amino acids and finding the least-repetitive synonymous gene sequence. We also show that the codon-scrambling problem is analogous to the well-known travelling salesman problem, and obtain an exact solution to it by using De Bruijn graphs and a modern mixed integer linear programme solver. As experimental proof of the utility of this approach, we use it to optimize the synthetic genes for 19 repetitive proteins, and show that the gene fragments are amenable to PCR-based gene assembly and recombinant expression.

  1. Identification of Cell Wall Synthesis Regulatory Genes Controlling Biomass Characteristics and Yield in Rice (Oryza Sativa)

    Energy Technology Data Exchange (ETDEWEB)

    Peng, Zhaohua PEng [Mississippi State University; Ronald, Palmela [UC-Davis; Wang, Guo-Liang [The Ohio State University

    2013-04-26

    This project aims to identify the regulatory genes of rice cell wall synthesis pathways using a cell wall removal and regeneration system. We completed the gene expression profiling studies following the time course from cell wall removal to cell wall regeneration in rice suspension cells. We also completed, total proteome, nuclear subproteome and histone modification studies following the course from cell wall removal and cell wall regeneration process. A large number of differentially expressed regulatory genes and proteins were identified. Meanwhile, we generated RNAi and over-expression transgenic rice for 45 genes with at least 10 independent transgenic lines for each gene. In addition, we ordered T-DNA and transposon insertion mutants for 60 genes from Korea, Japan, and France and characterized the mutants. Overall, we have mutants and transgenic lines for over 90 genes, exceeded our proposed goal of generating mutants for 50 genes. Interesting Discoveries a) Cell wall re-synthesis in protoplasts may involve a novel cell wall synthesis mechanism. The synthesis of the primary cell wall is initiated in late cytokinesis with further modification during cell expansion. Phragmoplast plays an essential role in cell wall synthesis. It services as a scaffold for building the cell plate and formation of a new cell wall. Only one phragmoplast and one new cell wall is produced for each dividing cell. When the cell wall was removed enzymatically, we found that cell wall re-synthesis started from multiple locations simultaneously, suggesting that a novel mechanism is involved in cell wall re-synthesis. This observation raised many interesting questions, such as how the starting sites of cell wall synthesis are determined, whether phragmoplast and cell plate like structures are involved in cell wall re-synthesis, and more importantly whether the same set of enzymes and apparatus are used in cell wall re-synthesis as during cytokinesis. Given that many known cell wall

  2. Recent Progress in Chemical Modifications of Chlorophylls and Bacteriochlorophylls for the Applications in Photodynamic Therapy.

    Science.gov (United States)

    Staron, Jakub; Boron, Bożena; Karcz, Dariusz; Szczygieł, Małgorzata; Fiedor, Leszek

    2015-01-01

    Since photodynamic therapy emerged as a promising cancer treatment, the development of photosensitizers has gained great interest. In this context, the photosynthetic pigments, chlorophylls and bacteriochlorophylls, as excellent natural photosensitizers, attracted much attention. In effect, several (bacterio) chlorophyll-based phototherapeutic agents have been developed and (or are about to) enter the clinics. The aim of this review article is to give a survey of the advances in the synthetic chemistry of these pigments which have been made over the last decade, and which are pertinent to the application of their derivatives as photosensitizers for photodynamic therapy (PDT). The review focuses on the synthetic strategies undertaken to obtain novel derivatives of (bacterio)chlorophylls with both enhanced photosensitizing and tumorlocalizing properties, and also improved photo- and chemical stability. These include modifications of the C- 17-ester moiety, the isocyclic ring, the central binding pocket, and the derivatization of peripheral functionalities at the C-3 and C-7 positions with carbohydrate-, peptide-, and nanoparticle moieties or other residues. The effects of these modifications on essential features of the pigments are discussed, such as the efficiency of reactive oxygen species generation, photostability, phototoxicity and interactions with living organisms. The review is divided into several sections. In the first part, the principles of PDT and photosensitizer action are briefly described. Then the relevant photophysical features of (bacterio)chlorophylls and earlier approaches to their modification are summarized. Next, a more detailed overview of the progress in synthetic methods is given, followed by a discussion of the effects of these modifications on the photophysics of the pigments and on their biological activity. PMID:26282940

  3. Absence of the cbb3 Terminal Oxidase Reveals an Active Oxygen-Dependent Cyclase Involved in Bacteriochlorophyll Biosynthesis in Rhodobacter sphaeroides

    Science.gov (United States)

    Chen, Guangyu E.; Martin, Elizabeth C.; Hunter, C. Neil

    2016-01-01

    ABSTRACT The characteristic green color associated with chlorophyll pigments results from the formation of an isocyclic fifth ring on the tetrapyrrole macrocycle during the biosynthesis of these important molecules. This reaction is catalyzed by two unrelated cyclase enzymes employing different chemistries. Oxygenic phototrophs such as plants and cyanobacteria utilize an oxygen-dependent enzyme, the major component of which is a diiron protein named AcsF, while BchE, an oxygen-sensitive [4Fe-4S] cluster protein, dominates in phototrophs inhabiting anoxic environments, such as the purple phototrophic bacterium Rhodobacter sphaeroides. We identify a potential acsF in this organism and assay for activity of the encoded protein in a strain lacking bchE under various aeration regimes. Initially, cells lacking bchE did not demonstrate AcsF activity under any condition tested. However, on removal of a gene encoding a subunit of the cbb3-type respiratory terminal oxidase, cells cultured under regimes ranging from oxic to micro-oxic exhibited cyclase activity, confirming the activity of the oxygen-dependent enzyme in this model organism. Potential reasons for the utilization of an oxygen-dependent enzyme in anoxygenic phototrophs are discussed. IMPORTANCE The formation of the E ring of bacteriochlorophyll pigments is the least well characterized step in their biosynthesis, remaining enigmatic for over 60 years. Two unrelated enzymes catalyze this cyclization step; O2-dependent and O2-independent forms dominate in oxygenic and anoxygenic phototrophs, respectively. We uncover the activity of an O2-dependent enzyme in the anoxygenic purple phototrophic bacterium Rhodobacter sphaeroides, initially by inactivation of the high-affinity terminal respiratory oxidase, cytochrome cbb3. We propose that the O2-dependent form allows for the biosynthesis of a low level of bacteriochlorophyll under oxic conditions, so that a rapid initiation of photosynthetic processes is possible for

  4. Chemical synthesis and cloning of a gene for human beta-urogastrone.

    OpenAIRE

    J. Smith; Cook, E.; Fotheringham, I; Pheby, S; Derbyshire, R; Eaton, M A; Doel, M; Lilley, D M; Pardon', J.F.; T Patel; Lewis, H.; Bell, L. D.

    1982-01-01

    A DNA duplex coding for the 53 amino acids of human beta-urogastrone has been synthesised. Computer assisted design of the gene included restriction endonuclease sites for plasmid insertion, a termination codon and two triplets coding for lysine at the 5'-end of the structural gene. The synthesis involved preparation of 23 oligodeoxyribonucleotides by phosphotriester procedures coupled to rapid HPLC techniques. The gene was constructed in two halves by enzymatic ligation of the oligonucleotid...

  5. Diverse arrangement of photosynthetic gene clusters in aerobic anoxygenic phototrophic bacteria.

    Directory of Open Access Journals (Sweden)

    Qiang Zheng

    Full Text Available BACKGROUND: Aerobic anoxygenic photototrophic (AAP bacteria represent an important group of marine microorganisms inhabiting the euphotic zone of the ocean. They harvest light using bacteriochlorophyll (BChl a and are thought to be important players in carbon cycling in the ocean. METHODOLOGY/PRINCIPAL FINDINGS: Aerobic anoxygenic phototrophic (AAP bacteria represent an important part of marine microbial communities. Their photosynthetic apparatus is encoded by a number of genes organized in a so-called photosynthetic gene cluster (PGC. In this study, the organization of PGCs was analyzed in ten AAP species belonging to the orders Rhodobacterales, Sphingomonadales and the NOR5/OM60 clade. Sphingomonadales contained comparatively smaller PGCs with an approximately size of 39 kb whereas the average size of PGCs in Rhodobacterales and NOR5/OM60 clade was about 45 kb. The distribution of four arrangements, based on the permutation and combination of the two conserved regions bchFNBHLM-LhaA-puhABC and crtF-bchCXYZ, does not correspond to the phylogenetic affiliation of individual AAP bacterial species. While PGCs of all analyzed species contained the same set of genes for bacteriochlorophyll synthesis and assembly of photosynthetic centers, they differed largely in the carotenoid biosynthetic genes. Spheroidenone, spirilloxanthin, and zeaxanthin biosynthetic pathways were found in each clade respectively. All of the carotenoid biosynthetic genes were found in the PGCs of Rhodobacterales, however Sphingomonadales and NOR5/OM60 strains contained some of the carotenoid biosynthetic pathway genes outside of the PGC. CONCLUSIONS/SIGNIFICANCE: Our investigations shed light on the evolution and functional implications in PGCs of marine aerobic anoxygenic phototrophs, and support the notion that AAP are a heterogenous physiological group phylogenetically scattered among Proteobacteria.

  6. Histamine suppresses gene expression and synthesis of tumor necrosis factor alpha via histamine H2 receptors

    OpenAIRE

    1991-01-01

    Histamine and tumor necrosis factor alpha (TNF-alpha) can each contribute to the pathogenesis of allergic reactions and chronic inflammatory diseases. We now report the effect of histamine on gene expression and total cellular synthesis of TNF-alpha. Lipopolysaccharide (LPS)-induced synthesis of TNF-alpha in peripheral blood mononuclear cells (PBMC) from 18 healthy donors was suppressed by histamine concentrations from 10(-6) to 10(-4) M, levels comparable with those measured in tissues after...

  7. Evolution of pigment synthesis pathways by gene and genome duplication in fish

    Directory of Open Access Journals (Sweden)

    Volff Jean-Nicolas

    2007-05-01

    Full Text Available Abstract Background Coloration and color patterning belong to the most diverse phenotypic traits in animals. Particularly, teleost fishes possess more pigment cell types than any other group of vertebrates. As the result of an ancient fish-specific genome duplication (FSGD, teleost genomes might contain more copies of genes involved in pigment cell development than tetrapods. No systematic genomic inventory allowing to test this hypothesis has been drawn up so far for pigmentation genes in fish, and almost nothing is known about the evolution of these genes in different fish lineages. Results Using a comparative genomic approach including phylogenetic reconstructions and synteny analyses, we have studied two major pigment synthesis pathways in teleost fish, the melanin and the pteridine pathways, with respect to different types of gene duplication. Genes encoding three of the four enzymes involved in the synthesis of melanin from tyrosine have been retained as duplicates after the FSGD. In the pteridine pathway, two cases of duplicated genes originating from the FSGD as well as several lineage-specific gene duplications were observed. In both pathways, genes encoding the rate-limiting enzymes, tyrosinase and GTP-cyclohydrolase I (GchI, have additional paralogs in teleosts compared to tetrapods, which have been generated by different modes of duplication. We have also observed a previously unrecognized diversity of gchI genes in vertebrates. In addition, we have found evidence for divergent resolution of duplicated pigmentation genes, i.e., differential gene loss in divergent teleost lineages, particularly in the tyrosinase gene family. Conclusion Mainly due to the FSGD, teleost fishes apparently have a greater repertoire of pigment synthesis genes than any other vertebrate group. Our results support an important role of the FSGD and other types of duplication in the evolution of pigmentation in fish.

  8. Triplet excited state energies and phosphorescence spectra of (bacterio)chlorophylls.

    Science.gov (United States)

    Hartzler, Daniel A; Niedzwiedzki, Dariusz M; Bryant, Donald A; Blankenship, Robert E; Pushkar, Yulia; Savikhin, Sergei

    2014-07-01

    (Bacterio)Chlorophyll ((B)Chl) molecules play a major role in photosynthetic light-harvesting proteins, and the knowledge of their triplet state energies is essential to understand the mechanisms of photodamage and photoprotection, as the triplet excitation energy of (B)Chl molecules can readily generate highly reactive singlet oxygen. The triplet state energies of 10 natural chlorophyll (Chl a, b, c2, d) and bacteriochlorophyll (BChl a, b, c, d, e, g) molecules and one bacteriopheophytin (BPheo g) have been directly determined via their phosphorescence spectra. Phosphorescence of four molecules (Chl c2, BChl e and g, BPheo g) was characterized for the first time. Additionally, the relative phosphorescence to fluorescence quantum yield for each molecule was determined. The measurements were performed at 77K using solvents providing a six-coordinate environment of the Mg(2+) ion, which allows direct comparison of these (B)Chls. Density functional calculations of the triplet state energies show good correlation with the experimentally determined energies. The correlation determined computationally was used to predict the triplet energies of three additional (B)Chl molecules: Chl c1, Chl f, and BChl f. PMID:24896677

  9. Separation of bacteriochlorophyll homologues from green photosynthetic sulfur bacteria by reversed-phase HPLC.

    Science.gov (United States)

    Borrego, C M; Garcia-Gil, L J

    1994-07-01

    A reversed-phase High Performance Liquid Cromatography (HPLC) method has been developed to accurately separate bacteriochlorophyllsc, d ande homologues in a reasonably short run time of 60 minutes. By using this method, two well-defined groups of bacteriochlorophyll homologue peaks can be discriminated. The first one consists of 4 peaks (min 24 to 30), which corresponds to the four main farnesyl homologues. The second peak subset is formed by a cluster of up to 10 minor peaks (min 33 to 40). These peaks can be related with series of several alcohol esters of the different chlorosome chlorophylls. The number of homologues was, however, quite variable depending on both, the bacteriochlorophyll and the bacterial species. The method hereby described, also provides a good separation of other photosynthetic pigments, either bacterial (Bacteriochlorophylla, chlorobactene, isorenieratene and okenone) or algal ones (Chlorophylla, Pheophytina and β-carotene). A preliminary screening of the homologue composition of several green photosynthetic bacterial species and isolates, has revealed different relative quantitative patterns. These differences seem to be related to physiological aspects rather than to taxonomic ones. The application of the method to the study of natural populations avoids the typical drawbacks on the pigment identification of overlapping eukaryotic and prokaryotic phototrophic microorganisms, giving further information about their physiological status. PMID:24310022

  10. The lamellar spacing in self-assembling bacteriochlorophyll aggregates is proportional to the length of the esterifying alcohol

    Czech Academy of Sciences Publication Activity Database

    Pšenčík, J.; Torkkeli, M.; Župčanová, Anita; Vácha, František; Serimaa, R.E.; Tůma, R.

    2010-01-01

    Roč. 104, 2-3 (2010), s. 211-219. ISSN 0166-8595 R&D Projects: GA ČR GA206/09/0375 Institutional research plan: CEZ:AV0Z50510513 Keywords : Green photosynthetic bacteria * Chlorosome * Bacteriochlorophyll Subject RIV: CE - Biochemistry Impact factor: 2.410, year: 2010

  11. Broadband 2D Electronic Spectroscopy Reveals Coupling Between Dark 1Bu- State of Carotenoid and Qx State of Bacteriochlorophyll

    Directory of Open Access Journals (Sweden)

    Scholes Gregory D.

    2013-03-01

    Full Text Available The study of LH2 protein of purple bacteria by broadband 2D electronic spectroscopy is presented. The dark 1Bu- carotenoid state is directly observed in 2D spectra and its role in carotenoid-bacteriochlorophyll interaction is discussed.

  12. Relationship between the in vivo bacteriochlorophyll fluorescence and the state of the photosynthetic apparatus in purple bacteria

    Czech Academy of Sciences Publication Activity Database

    Bína, David; Litvín, Radek; Vácha, František

    Dordrecht : Springer, 2008 - (Allen, J.; Gantt, E.; Golbeck, J.; Osmond, B.), s. 559-562 ISBN 978-1-4020-6707-5. [International Congress on Photosynthesis/14/. Glasgow (GB), 22.07.2007-27.07.2007] R&D Projects: GA ČR(CZ) GA206/05/2739 Keywords : bacteriochlorophyll * purple bacteria * fluorescence * electric field Subject RIV: BO - Biophysics

  13. Self-assembly and energy transfer in artificial light-harvesting complexes of bacteriochlorophyll c with astaxanthin

    Czech Academy of Sciences Publication Activity Database

    Alster, J.; Polívka, Tomáš; Arellano, J.B.; Hříbek, P.; Vácha, František; Hala, J.; Pšenčík, J.

    2012-01-01

    Roč. 111, 1-2 (2012), s. 193-204. ISSN 0166-8595 R&D Projects: GA ČR GA206/09/0375 Institutional research plan: CEZ:AV0Z50510513 Keywords : light-harvesting * astaxanthin * self-assembly * bacteriochlorophyll aggregates Subject RIV: BO - Biophysics Impact factor: 3.150, year: 2012

  14. [Cloning of structural genes involved in riboflavin synthesis of the yeast Candida famata].

    Science.gov (United States)

    Dmytruk, K V; Abbas, C A; Voronovsky, A Y; Kshanovska, B V; Sybirna, K A; Sybirny, A A

    2004-01-01

    The riboflavin overproducing mutants of the flavinogenic yeast Candida famata isolated by conventional selection methods are used for the industrial production of vitamin B2. Recently, a transformation system was developed for C. famata using the leu2 mutant as a recipient strain and Saccharomyces cerevislae LEU2 gene as a selective marker. In this paper the cloning of C. famata genes for riboflavin synthesis on the basis of developed transformation system for this yeast species is described. Riboflavin autotrophic mutants were isolated from a previously selected C. famata leu2 strain. C. famata genomic DNA library was constructed and used for cloning of the corresponding structural genes for riboflavin synthesis by complementation of the growth defects on a medium without leucine and riboflavin. As a result, the DNA fragments harboring genes RIB1, RIB2, RIB5, RIB6 and RIB7 encoding GTP cyclohydrolase, reductase, dimethylribityllumazine synthase, dihydroxybutanone phosphate synthase and riboflavin synthase, were isolated and subsequently subcloned to the smallest possible fragments. The plasmids with these genes successfully complemented riboflavin auxotrophies of the corresponding mutants of another flavinogenic yeast Pichia guilliermondii. This suggested that C. famata structural genes for riboflavin synthesis and not some of the supressor genes were cloned. PMID:15909421

  15. Ethylene-induced senescence-related gene expression requires protein synthesis

    International Nuclear Information System (INIS)

    We have investigated the effects of inhibiting protein synthesis on the ethylene-induced expression of 3 carnation senescence-related genes, pSR5, pSR8, and pSR12. Treatment of preclimacteric carnation petal discs with 1μg/ml of cycloheximide, a cytoplasmic protein synthesis inhibitor, for 3h inhibited protein synthesis by >80% as quantitated by the incorporation of [35S]methionine into protein. Pre-treatment of petal discs with cycloheximide prevented ethylene-induced SR transcript accumulation. Cycloheximide treatment of petal discs held in air did not result in increased levels of SR mRNA. These results indicate that ethylene does not interact with pre-formed factors but rather that the activation of SR gene expression by ethylene is mediated by labile protein factor(s) synthesized on cytoplasmic ribosomes. Experiments are currently underway to determine if cycloheximide exerts its effect at the transcriptional or post-transcriptional level

  16. Homology of genes for exopolysaccharide synthesis in Rhizobium leguminosarum and effect of cloned exo genes on nodule formation

    International Nuclear Information System (INIS)

    A 5.4 kb BamHI fragment of Rhizobium leguminosarum (R.l.) bv. trifolii TA1 was found to carry genes involved in exopolysaccharide synthesis (exo genes). This fragment was strongly hybridized to the total DNA from DNA from R.l. bv. viciae and bv. phaseoli digested with EcoRI. No homology was found with total DNA of R. melilotic and Rhizobium sp. NGR 234. The exo genes R.l. bv. trifolii TA1 conjugally introduced into R.l. bv. viciae 1302 considerably affected the symbiosis: the nodules induced on vetch were abortive and did not fix nitrogen. On the other hand, Phaseolus beans infected with R.l. bv. trifolii exo genes formed the nitrogen-fixing nodules. It can be conduced that additional copies of exo genes introduced into wild type R.l. strains can disturb the synthesis of acidic exopolysaccharides and affect symbiosis of the plants forming indeterminate nodules, but do not affect symbiosis of the plants forming the determinate nodules. (author). 29 refs, 2 figs, 2 tabs

  17. Expression of fatty acid synthesis genes and fatty acid accumulation in haematococcus pluvialis under different stressors

    Directory of Open Access Journals (Sweden)

    Lei Anping

    2012-03-01

    Full Text Available Abstract Background Biofuel has been the focus of intensive global research over the past few years. The development of 4th generation biofuel production (algae-to-biofuels based on metabolic engineering of algae is still in its infancy, one of the main barriers is our lacking of understanding of microalgal growth, metabolism and biofuel production. Although fatty acid (FA biosynthesis pathway genes have been all cloned and biosynthesis pathway was built up in some higher plants, the molecular mechanism for its regulation in microalgae is far away from elucidation. Results We cloned main key genes for FA biosynthesis in Haematococcus pluvialis, a green microalga as a potential biodiesel feedstock, and investigated the correlations between their expression alternation and FA composition and content detected by GC-MS under different stress treatments, such as nitrogen depletion, salinity, high or low temperature. Our results showed that high temperature, high salinity, and nitrogen depletion treatments played significant roles in promoting microalgal FA synthesis, while FA qualities were not changed much. Correlation analysis showed that acyl carrier protein (ACP, 3-ketoacyl-ACP-synthase (KAS, and acyl-ACP thioesterase (FATA gene expression had significant correlations with monounsaturated FA (MUFA synthesis and polyunsaturated FA (PUFA synthesis. Conclusions We proposed that ACP, KAS, and FATA in H. pluvialis may play an important role in FA synthesis and may be rate limiting genes, which probably could be modified for the further study of metabolic engineering to improve microalgal biofuel quality and production.

  18. Energy transfer in aggregates of bacteriochlorophyll c self-assembled with azulene derivatives.

    Science.gov (United States)

    Matěnová, Martina; Lorelei Horhoiu, Viviana; Dang, Florian-Xuan; Pospíšil, Petr; Alster, Jan; Burda, Jaroslav V; Balaban, Teodor Silviu; Pšenčík, Jakub

    2014-08-21

    Bacteriochlorophyll (BChl) c is the main light-harvesting pigment of certain photosynthetic bacteria. It is found in the form of self-assembled aggregates in the so-called chlorosomes. Here we report the results of co-aggregation experiments of BChl c with azulene and its tailored derivatives. We have performed spectroscopic and quantum chemical characterization of the azulenes, followed by self-assembly experiments. The results show that only azulenes with sufficient hydrophobicity are able to induce aggregation of BChl c. Interestingly, only azulene derivatives possessing a conjugated phenyl ring were capable of efficient (∼50%) excitation energy transfer to BChl molecules. These aggregates represent an artificial light-harvesting complex with enhanced absorption between 220 and 350 nm compared to aggregates of pure BChl c. The results provide insight into the principles of self-assembly of BChl aggregates and suggest an important role of the π-π interactions in efficient energy transfer. PMID:24999619

  19. A method for multiplex gene synthesis employing error correction based on expression.

    Directory of Open Access Journals (Sweden)

    Timothy H-C Hsiau

    Full Text Available Our ability to engineer organisms with new biosynthetic pathways and genetic circuits is limited by the availability of protein characterization data and the cost of synthetic DNA. With new tools for reading and writing DNA, there are opportunities for scalable assays that more efficiently and cost effectively mine for biochemical protein characteristics. To that end, we have developed the Multiplex Library Synthesis and Expression Correction (MuLSEC method for rapid assembly, error correction, and expression characterization of many genes as a pooled library. This methodology enables gene synthesis from microarray-synthesized oligonucleotide pools with a one-pot technique, eliminating the need for robotic liquid handling. Post assembly, the gene library is subjected to an ampicillin based quality control selection, which serves as both an error correction step and a selection for proteins that are properly expressed and folded in E. coli. Next generation sequencing of post selection DNA enables quantitative analysis of gene expression characteristics. We demonstrate the feasibility of this approach by building and testing over 90 genes for empirical evidence of soluble expression. This technique reduces the problem of part characterization to multiplex oligonucleotide synthesis and deep sequencing, two technologies under extensive development with projected cost reduction.

  20. Molecular cloning and expression of Corynebacterium glutamicum genes for amino acid synthesis in Escherichia coli cells

    International Nuclear Information System (INIS)

    Molecular cloning of Corynebacterium glutamicum genes for threonine and lysine synthesis has been done in Escherichia coli cells. The clonal library of EcoRI fragments of chromosomal DNA of C. glutamicum was constructed on the plasmid vector λpSL5. The genes for threonine and lysine synthesis were identified by complementation of E. coli mutations in thrB and lysA genes, respectively. Recombinant plasmids, isolated from independent ThrB+ clone have a common 4.1-kb long EcoRI DNA fragment. Hybrid plasmids isolated from LysA+ transductants of E. coli have common 2.2 and 3.3 kb long EcoRI fragments of C. glutamicum DNA. The hybrid plasmids consistently transduced the markers thrB+ and lysA+. The Southern hybridization analysis showed that the cloned DNA fragments hybridized with the fragments of identical length in C. glutamicum chromosomes

  1. Identification and phylogenetic analysis of heme synthesis genes in trypanosomatids and their bacterial endosymbionts.

    Directory of Open Access Journals (Sweden)

    João M P Alves

    Full Text Available It has been known for decades that some insect-infecting trypanosomatids can survive in culture without heme supplementation while others cannot, and that this capability is associated with the presence of a betaproteobacterial endosymbiont in the flagellate's cytoplasm. However, the specific mechanisms involved in this process remained obscure. In this work, we sequence and phylogenetically analyze the heme pathway genes from the symbionts and from their hosts, as well as from a number of heme synthesis-deficient Kinetoplastida. Our results show that the enzymes responsible for synthesis of heme are encoded on the symbiont genomes and produced in close cooperation with the flagellate host. Our evidence suggests that this synergistic relationship is the end result of a history of extensive gene loss and multiple lateral gene transfer events in different branches of the phylogeny of the Trypanosomatidae.

  2. Molecular cloning and expression of Corynebacterium glutamicum genes for amino acid synthesis in Escherichia coli cells

    Energy Technology Data Exchange (ETDEWEB)

    Beskrovnaya, O.Yu.; Fonshtein, M.Yu.; Kolibaba, L.G.; Yankovskii, N.K.; Debabov, V.G.

    1989-01-01

    Molecular cloning of Corynebacterium glutamicum genes for threonine and lysine synthesis has been done in Escherichia coli cells. The clonal library of EcoRI fragments of chromosomal DNA of C. glutamicum was constructed on the plasmid vector /lambda/pSL5. The genes for threonine and lysine synthesis were identified by complementation of E. coli mutations in thrB and lysA genes, respectively. Recombinant plasmids, isolated from independent ThrB/sup +/ clone have a common 4.1-kb long EcoRI DNA fragment. Hybrid plasmids isolated from LysA/sup +/ transductants of E. coli have common 2.2 and 3.3 kb long EcoRI fragments of C. glutamicum DNA. The hybrid plasmids consistently transduced the markers thrB/sup +/ and lysA/sup +/. The Southern hybridization analysis showed that the cloned DNA fragments hybridized with the fragments of identical length in C. glutamicum chromosomes.

  3. Potential Genes for Regulation of Milk Protein Synthesis in Dairy Goat Mammary Gland

    Institute of Scientific and Technical Information of China (English)

    Chen Dan; Zhang Na; Nan Xue-mei; Li Qing-zhang; Gao Xue-jun

    2016-01-01

    The lactating mammary gland is a prodigious protein-producing factory, but the milk protein synthesis mechanisms are not well understood. The major objective of this paper was to elucidate which genes and pathways were involved in the regulation of milk protein synthesis in the dairy goat mammary gland. Total 36 primiparous Guanzhong dairy goats were allotted in 12 groups according to their mammary development stages: days 90 and 150 of virgin, days 30, 90, and 150 of pregnancy, days 1, 10, 35, and 60 of lactation and days 3, 7, and 21 of involution (three animals per group). Mammary tissue RNA was isolated for quantitative real-time RT-PCR of four casein genes alpha-s1 casein (CSN1S1), alpha-s2 casein (CSN1S2), beta-casein (CSN2) and casein kappa (CSN3), four whey protein genes lactoglobulin (LGB), lactalbumin (LALBA), lactofarrin (LTF), and Whey acidic protein (WAP) and the genes which were potentially to regulate dairy goat milk protein synthesis at the level of transcription or translation [prolactin receptor (PRLR), AKT1, signal transducers and activators of transcription 5 (STAT5), E74-Like Factor 5 (ELF5), eukaryotic translation initiation factor 4E binding protein 1 (EIF4E-BP1), S6kinase (S6K) and caveolin 1]. The results showed that all genes were up-regulated in lactation period. The expressions of PRLR, AKT1, STAT5, ELF5, and S6K were similar to mRNA expressions of milk proteins. Our results indicated that milk protein synthesis in dairy goat mammary gland was possibly regulated by these genes.

  4. Histone gene expression remains coupled to DNA synthesis during in vitro cellular senescence

    International Nuclear Information System (INIS)

    Despite a decrease in the extent to which confluent monolayers of late compared to early passage CF3 human diploid fibroblasts can be stimulated to proliferate, the time course of DNA synthesis onset is similar regardless of the in vitro age of the cells. A parallel and stoichiometric relationship is maintained between the rate of DNA synthesis and the cellular levels of histone mRNA independent of the age of the cell cultures. Furthermore, DNA synthesis and cellular histone mRNA levels decline in a coordinate manner after inhibition of DNA replication by hydroxyurea treatment. These results indicate that while the proliferative activity of human diploid fibroblasts decreases with passage in culture, those cells that retain the ability to proliferate continue to exhibit a tight coupling of DNA replication and histone gene expression

  5. Vibronic coupling explains the ultrafast carotenoid-to-bacteriochlorophyll energy transfer in natural and artificial light harvesters

    OpenAIRE

    Perlík, Václav; Seibt, Joachim; Cranston, Laura J.; Cogdell, Richard J.; Lincoln, Craig N.; Savolainen, Janne; Šanda, František; Mančal, Tomáš; Hauer, Jürgen

    2015-01-01

    The initial energy transfer in photosynthesis occurs between the light-harvesting pigments and on ultrafast timescales. We analyze the carotenoid to bacteriochlorophyll energy transfer in LH2 Marichromatium purpuratum as well as in an artificial light-harvesting dyad system by using transient grating and two-dimensional electronic spectroscopy with 10 fs time resolution. We find that F\\"orster-type models reproduce the experimentally observed 60 fs transfer times, but overestimate coupling co...

  6. Abundance, depth distribution, and composition of aerobic bacteriochlorophyll a-producing bacteria in four basins of the central Baltic Sea

    Czech Academy of Sciences Publication Activity Database

    Salka, I.; Moulisová, Vladimíra; Koblížek, Michal; Jost, G.; Jürgens, K.; Labrenz, M.

    2008-01-01

    Roč. 74, č. 14 (2008), s. 4398-4404. ISSN 0099-2240 R&D Projects: GA ČR GA204/05/0307; GA ČR GA206/07/0241; GA AV ČR 1QS500200570 Institutional research plan: CEZ:AV0Z50200510 Keywords : aerobic bacteriochlorophyll * baltic sea * epifluorescence microscopy Subject RIV: EE - Microbiology, Virology Impact factor: 3.801, year: 2008

  7. Gene Composer: database software for protein construct design, codon engineering, and gene synthesis

    Directory of Open Access Journals (Sweden)

    Mixon Mark

    2009-04-01

    Full Text Available Abstract Background To improve efficiency in high throughput protein structure determination, we have developed a database software package, Gene Composer, which facilitates the information-rich design of protein constructs and their codon engineered synthetic gene sequences. With its modular workflow design and numerous graphical user interfaces, Gene Composer enables researchers to perform all common bio-informatics steps used in modern structure guided protein engineering and synthetic gene engineering. Results An interactive Alignment Viewer allows the researcher to simultaneously visualize sequence conservation in the context of known protein secondary structure, ligand contacts, water contacts, crystal contacts, B-factors, solvent accessible area, residue property type and several other useful property views. The Construct Design Module enables the facile design of novel protein constructs with altered N- and C-termini, internal insertions or deletions, point mutations, and desired affinity tags. The modifications can be combined and permuted into multiple protein constructs, and then virtually cloned in silico into defined expression vectors. The Gene Design Module uses a protein-to-gene algorithm that automates the back-translation of a protein amino acid sequence into a codon engineered nucleic acid gene sequence according to a selected codon usage table with minimal codon usage threshold, defined G:C% content, and desired sequence features achieved through synonymous codon selection that is optimized for the intended expression system. The gene-to-oligo algorithm of the Gene Design Module plans out all of the required overlapping oligonucleotides and mutagenic primers needed to synthesize the desired gene constructs by PCR, and for physically cloning them into selected vectors by the most popular subcloning strategies. Conclusion We present a complete description of Gene Composer functionality, and an efficient PCR-based synthetic gene

  8. Identification of genes for sulfolipid synthesis in primitive red alga Cyanidioschyzon merolae.

    Science.gov (United States)

    Sato, Norihiro; Kobayashi, Satomi; Aoki, Motohide; Umemura, Tomonari; Kobayashi, Isao; Tsuzuki, Mikio

    2016-01-29

    Sulfoquinovosyl diacylglycerol is one of the lipids that construct thylakoid membranes, and is distributed from cyanobacteria to plastids in plants including a red lineage. One of the most primitive red algae, Cyanidioschyzon melorae, similar to cyanobacteria and green plants, possesses homologs of the SQD1 and SQD2 genes that code for UDP-sulfoquinovose and sulfoquinovosyl diacylglycerol synthases, respectively, for the synthesis of sulfoquinovosyl diacylglycerol. We here revealed the structural properties of SQD1 and SQD2 homologs in C. melorae intrinsic to those of the authentic proteins, and verified their enzymatic functions through heterologous expression in cyanobacterial disruptants as to the corresponding genes. The results demonstrated that the system of sulfoquinovosyl diacylglycerol synthesis could have been conserved through evolution of cyanobacteria to plastids in a red lineage, which is compatible with the monophyletic origin of plastids. PMID:26768360

  9. Butyrate induces sLex synthesis by stimulation of selective glycosyltransferase genes

    OpenAIRE

    Radhakrishnan, Prakash; Beum, Paul V.; Tan, Shuhua; Cheng, Pi-Wan

    2007-01-01

    Sialyl Lewis x (sLex) is an important tumor-associated carbohydrate antigen present on the cell surface glycoconjugates involved in leukocyte migration and cancer metastasis. We report the formation of sLex epitope in butyrate-treated human pancreatic adenocarcinoma cells expressing MUC1 and core 2 N-acetylglucosaminyltransferase (C2GnT). Butyrate treatment stimulates not only the transgene but also a group of endogenous glycosyltransferase genes involved in the synthesis of sLex. Current fin...

  10. Oligonucleotide-directed mutagenesis by microscale 'shot-gun' gene synthesis.

    OpenAIRE

    Grundström, T; Zenke, W M; Wintzerith, M; Matthes, H W; Staub, A; Chambon, P

    1985-01-01

    We describe a rapid and efficient microscale method for in vitro site-directed mutagenesis by gene synthesis. Mutants are constructed by "shot-gun ligation" of overlapping synthetic oligonucleotides yielding double stranded synthetic DNA of more than 120 nucleotides in length. The terminal oligonucleotides of the DNA segment to be synthesized are designed to create sticky ends complementary to unique restriction sites of a polylinker present in an M13 vector. The oligonucleotides are hybridiz...

  11. Energy, genes and evolution: introduction to an evolutionary synthesis

    OpenAIRE

    Lane, N.; Martin, W. F.; J. A. Raven; Allen, J. F.

    2013-01-01

    Life is the harnessing of chemical energy in such a way that the energy-harnessing device makes a copy of itself. No energy, no evolution. The ‘modern synthesis’ of the past century explained evolution in terms of genes, but this is only part of the story. While the mechanisms of natural selection are correct, and increasingly well understood, they do little to explain the actual trajectories taken by life on Earth. From a cosmic perspective—what is the probability of life elsewhere in the Un...

  12. Synthesis of bacteriophage-coded gene products during infection of Escherichia coli with amber mutants of T3 and T7 defective in gene 1

    DEFF Research Database (Denmark)

    Issinger, O G; Hausmann, R

    1973-01-01

    During nonpermissive infection by a T7 amber mutant in gene 1 (phage RNA polymerase-deficient), synthesis of the products of the phage genes 3 (endonuclease), 3, 5 (lysozyme), 5 (DNA polymerase), and 17 (serum blocking power) was shown to occur at about half the rate as during wild-type infection...

  13. Attenuation of Mycobacterium tuberculosis by Disruption of a mas-Like Gene or a Chalcone Synthase-Like Gene, Which Causes Deficiency in Dimycocerosyl Phthiocerol Synthesis

    OpenAIRE

    Sirakova, Tatiana D.; Dubey, Vinod S.; Cynamon, Michael H.; Kolattukudy, Pappachan E.

    2003-01-01

    Tuberculosis is one of the leading preventable causes of death. Emergence of drug-resistant tuberculosis makes the discovery of new targets for antimycobacterial drugs critical. The unique mycobacterial cell wall lipids are known to play an important role in pathogenesis, and therefore the genes responsible for their biosynthesis offer potential new targets. To assess the possible role of some of the genes potentially involved in cell wall lipid synthesis, we disrupted a mas-like gene, msl7, ...

  14. Up-Scaled Synthesis and Characterization of Nonviral Gene Delivery Particles for Transient In Vitro and In Vivo Transgene Expression.

    Science.gov (United States)

    Taschauer, Alexander; Geyer, Antonia; Gehrig, Sebastian; Maier, Julia; Sami, Haider; Ogris, Manfred

    2016-06-01

    Polyethylenimine-based polyplexes are promising nonviral gene delivery systems for preclinical and clinical applications. Pipette-based polyplexing is associated with several disadvantages, such as batch-to-batch variability, restriction to smaller volumes, and variable gene delivery results. The present protocol describes syringe-pump-mediated upscaled synthesis of well-defined gene delivery nanoparticles capable of efficient in vitro and in vivo gene delivery. Syringe-pump-based synthesis ensures controlled mixing, upscaling, and reproducible gene delivery. Nanoparticle tracking analysis of the upscaled formulations involved single nanoparticle tracking, thereby generating highly resolved biophysical characterization. Gene delivery performance was investigated by luciferase gene expression in cells and three-dimensional bioluminescence imaging in mice. PMID:27169568

  15. TmPrime: fast, flexible oligonucleotide design software for gene synthesis

    Science.gov (United States)

    Bode, Marcus; Khor, Samuel; Ye, Hongye; Li, Mo-Huang; Ying, Jackie Y.

    2009-01-01

    Herein we present TmPrime, a computer program to design oligonucleotide sets for gene assembly by both ligase chain reaction (LCR) and polymerase chain reaction (PCR). TmPrime offers much flexibility with no constraints on the gene and oligonucleotide lengths. The program divides the long input DNA sequence based on the input desired melting temperature, and dynamically optimizes the length of oligonucleotides to achieve homologous melting temperatures. The output reports the melting temperatures, oligonucleotide sequences and potential formation of secondary structures. Our program also provides functions on sequence pooling to separate long genes into smaller pieces for multi-pool assembly and codon optimization for expression. The software has been successfully used in the design and synthesis of green fluorescent protein fragment (GFPuv) (760 bp), human protein kinase B-2 (PKB2) (1446 bp) and the promoter of human calcium-binding protein A4 (S100A4) (752 bp) using real-time PCR assembly with LCGreen I, which offers a novel approach to compare the efficiency of gene synthesis. The purity of assembled products is successfully estimated with the use of melting curve analysis, which would potentially eliminate the necessity for agarose gel electrophoresis. This program is freely available at http://prime.ibn.a-star.edu.sg. PMID:19515937

  16. Quantification of two forms of green sulfur bacteria in their natural habitat using bacteriochlorophyll fluorescence spectra

    Science.gov (United States)

    Kharcheva, Anastasia V.; Zhiltsova, Anna A.; Lunina, Olga N.; Savvichev, Alexander S.; Patsaeva, Svetlana V.

    2016-04-01

    Detection of phototropic organisms in their natural habitat using optical instruments operating under water is urgently needed for many tasks of ecological monitoring. While fluorescence methods are widely applied nowadays to detect and characterize phytoplankton communities, the techniques for detection and recognition of anoxygenic phototrophs are considered challenging. Differentiation of the forms of anoxygenic green sulfur bacteria in natural water using spectral techniques remains problematic. Green sulfur bacteria could be found in two forms, green-colored (containing BChl d in pigment compound) and brown-colored (containing BChl e), have the special ecological niche in such reservoirs. Separate determination of these microorganisms by spectral methods is complicated because of similarity of spectral characteristics of their pigments. We describe the novel technique of quantification of two forms of green sulfur bacteria directly in water using bacteriochlorophyll fluorescence without pigment extraction. This technique is noninvasive and could be applied in remote mode in the water bodies with restricted water circulation to determine simultaneously concentrations of two forms of green sulfur bacteria in their natural habitat.

  17. The supramolecular organization of self-assembling chlorosomal bacteriochlorophyll c, d, or e mimics.

    Science.gov (United States)

    Jochum, Tobias; Reddy, Chilla Malla; Eichhöfer, Andreas; Buth, Gernot; Szmytkowski, Jedrzej; Kalt, Heinz; Moss, David; Balaban, Teodor Silviu

    2008-09-01

    Bacteriochlorophylls (BChls) c, d, and e are the main light-harvesting pigments of green photosynthetic bacteria that self-assemble into nanostructures within the chlorosomes forming the most efficient antennas of photosynthetic organisms. All previous models of the chlorosomal antennae, which are quite controversially discussed because no single crystals could be grown so far from these organelles, involve a strong hydrogen-bonding interaction between the 3(1) hydroxyl group and the 13(1) carbonyl group. We have synthesized different self-assemblies of BChl c mimics having the same functional groups as the natural counterparts, that is, a hydroxyethyl substituent, a carbonyl group and a divalent metal atom ligated by a tetrapyrrole. These artificial BChl mimics have been shown by single crystal x-ray diffraction to form extended stacks that are packed by hydrophobic interactions and in the absence of hydrogen bonding. Time-resolved photoluminescence proves the ordered nature of the self-assembled stacks. FT-IR spectra show that on self-assembly the carbonyl frequency is shifted by approximately 30 cm(-1) to lower wavenumbers. From the FT-IR data we can infer the proximal interactions between the BChls in the chlorosomes consistent with a single crystal x-ray structure that shows a weak electrostatic interaction between carbonyl groups and the central zinc atom. PMID:18755898

  18. Scrambled self-assembly of bacteriochlorophylls c and e in aqueous Triton X-100 micelles.

    Science.gov (United States)

    Saga, Yoshitaka; Saiki, Tatsuya; Takahashi, Naoya; Shibata, Yutaka; Tamiaki, Hitoshi

    2014-01-01

    Bacteriochlorophyll (BChl) e was coassembled with BChl c in Triton X-100 micelles in aqueous solutions. The Qy absorption bands of the coaggregates were positioned between those of aggregates consisting solely of BChl c or e. The electronic absorption spectra of the coaggregates could not be reproduced by linear combinations of the spectra of the aggregates consisting solely of each pigment, but they were in line with the simulated spectra for the self-aggregates in which both BChls were randomly distributed. These suggest that BChls c and e are not spatially separated; they are homogenously distributed over the self-aggregates to give electronic spectra that are different from those of the aggregate consisting solely of each pigment. Deaggregation of the scrambled self-aggregates by excess Triton X-100 did not produce any spectral components assigned to an aggregate consisting solely of either BChl c or e. Acid-induced decomposition of the scrambled aggregates showed different kinetics from those of the aggregates consisting solely of each pigment. These also support the homogeneous distribution of BChls c and e in the scrambled self-aggregates. These results will be useful to investigate the major light-harvesting antenna systems of green photosynthetic bacteria that contain two kinds of chlorosomal BChls. PMID:24308290

  19. Simulated two-dimensional electronic spectroscopy of the eight-bacteriochlorophyll FMO complex

    Energy Technology Data Exchange (ETDEWEB)

    Yeh, Shu-Hao [Department of Chemistry and Birck Nanotechnology Center, Purdue University, West Lafayette, Indiana 47907 (United States); Kais, Sabre, E-mail: kais@purdue.edu [Department of Chemistry and Birck Nanotechnology Center, Purdue University, West Lafayette, Indiana 47907 (United States); Qatar Environment and Energy Research Institute, Qatar Foundation, Doha (Qatar)

    2014-12-21

    The Fenna-Matthews-Olson (FMO) protein-pigment complex acts as a molecular wire conducting energy between the outer antenna system and the reaction center; it is an important photosynthetic system to study the transfer of excitonic energy. Recent crystallographic studies report the existence of an additional (eighth) bacteriochlorophyll a (BChl a) in some of the FMO monomers. To understand the functionality of this eighth BChl, we simulated the two-dimensional electronic spectra of both the 7-site (apo form) and the 8-site (holo form) variant of the FMO complex from green sulfur bacteria, Prosthecochloris aestuarii. By comparing the spectrum, it was found that the eighth BChl can affect two different excitonic energy transfer pathways: (1) it is directly involved in the first apo form pathway (6 → 3 → 1) by passing the excitonic energy to exciton 6; and (2) it facilitates an increase in the excitonic wave function overlap between excitons 4 and 5 in the second pathway (7 → 4,5 → 2 → 1) and thus increases the possible downward sampling routes across the BChls.

  20. The Complete Genome of Bacillus amyloliquefaciens subsp. plantarum CAU B946 Contains a Gene Cluster for Nonribosomal Synthesis of Iturin A

    OpenAIRE

    Blom, Jochen; Rueckert, Christian; Niu, Ben; Wang, Qi; Borriss, Rainer

    2012-01-01

    The genome of the rhizobacterium Bacillus amyloliquefaciens subsp. plantarum CAU B946 was 4.02 Mb in size and harbored 3,823 genes (coding sequences [CDS]). Nine giant gene clusters were dedicated to nonribosomal synthesis of antimicrobial compounds. Remarkably, strain CAU B946 possessed a gene cluster involved in synthesis of iturin A.

  1. The complete genome of Bacillus amyloliquefaciens subsp. plantarum CAU B946 contains a gene cluster for nonribosomal synthesis of iturin A.

    Science.gov (United States)

    Blom, Jochen; Rueckert, Christian; Niu, Ben; Wang, Qi; Borriss, Rainer

    2012-04-01

    The genome of the rhizobacterium Bacillus amyloliquefaciens subsp. plantarum CAU B946 was 4.02 Mb in size and harbored 3,823 genes (coding sequences [CDS]). Nine giant gene clusters were dedicated to nonribosomal synthesis of antimicrobial compounds. Remarkably, strain CAU B946 possessed a gene cluster involved in synthesis of iturin A. PMID:22408246

  2. MicroRNA-26a/b and their host genes synergistically regulate triacylglycerol synthesis by targeting the INSIG1 gene.

    Science.gov (United States)

    Wang, Hui; Luo, Jun; Zhang, Tianying; Tian, Huibin; Ma, Yue; Xu, Huifen; Yao, Dawei; Loor, Juan J

    2016-05-01

    The microRNA-26 (miR-26) family is known to control adipogenesis in non-ruminants. The genomic loci of miR-26a and miR-26b have been localized in the introns of genes encoding for the proteins of the C-terminal domain RNA polymerase II polypeptide A small phosphatase (CTDSP) family. Insulin-induced gene 1 (INSIG1) encodes a protein with a key role in the regulation of lipogenesis in rodent liver. In the present study, we investigated the synergistic function of the miR-26 family and their host genes in goat mammary epithelial cells (GMEC). Downregulation of miR-26a/b and their host genes in GMEC decreased the expression of genes relate to fatty acid synthesis (PPARG, LXRA, SREBF1, FASN, ACACA, GPAM, LPIN1, DGAT1 and SCD1), triacylglycerol accumulation and unsaturated fatty acid synthesis. Luciferase reporter assays confirmed INSIG1 as a direct target of miR-26a/b. Furthermore, inhibition of the CTDSP family also downregulated the expression of INSIG1. Taken together, our findings highlight a functional association of miR-26a/b, their host genes and INSIG1, and provide new insights into the regulatory network controlling milk fat synthesis in GMEC. The data indicate that targeting this network via nutrition might be important for regulating milk fat synthesis in ruminants. PMID:27002347

  3. A simple and accurate two-step long DNA sequences synthesis strategy to improve heterologous gene expression in pichia.

    Directory of Open Access Journals (Sweden)

    Jiang-Ke Yang

    Full Text Available In vitro gene chemical synthesis is a powerful tool to improve the expression of gene in heterologous system. In this study, a two-step gene synthesis strategy that combines an assembly PCR and an overlap extension PCR (AOE was developed. In this strategy, the chemically synthesized oligonucleotides were assembled into several 200-500 bp fragments with 20-25 bp overlap at each end by assembly PCR, and then an overlap extension PCR was conducted to assemble all these fragments into a full length DNA sequence. Using this method, we de novo designed and optimized the codon of Rhizopus oryzae lipase gene ROL (810 bp and Aspergillus niger phytase gene phyA (1404 bp. Compared with the original ROL gene and phyA gene, the codon-optimized genes expressed at a significantly higher level in yeasts after methanol induction. We believe this AOE method to be of special interest as it is simple, accurate and has no limitation with respect to the size of the gene to be synthesized. Combined with de novo design, this method allows the rapid synthesis of a gene optimized for expression in the system of choice and production of sufficient biological material for molecular characterization and biotechnological application.

  4. Candida famata (Debaryomyces hansenii) DNA sequences containing genes involved in riboflavin synthesis.

    Science.gov (United States)

    Voronovsky, Andriy Y; Abbas, Charles A; Dmytruk, Kostyantyn V; Ishchuk, Olena P; Kshanovska, Barbara V; Sybirna, Kateryna A; Gaillardin, Claude; Sibirny, Andriy A

    2004-11-01

    Previously cloned Candida famata (Debaryomyces hansenii) strain VKM Y-9 genomic DNA fragments containing genes RIB1 (codes for GTP cyclohydrolase II), RIB2 (encodes specific reductase), RIB5 (codes for dimethylribityllumazine synthase), RIB6 (encodes dihydroxybutanone phosphate synthase) and RIB7 (codes for riboflavin synthase) were sequenced. The derived amino acid sequences of C. famata RIB genes showed extensive homology to the corresponding sequences of riboflavin synthesis enzymes of other yeast species. The highest identity was observed to homologues of D. hansenii CBS767, as C. famata is the anamorph of this hemiascomycetous yeast. The D. hansenii CBS767 RIB3 gene encoding specific deaminase was cloned. This gene successfully complemented riboflavin auxotrophy of the rib3 mutant of flavinogenic yeast, Pichia guilliermondii. Putative iron-responsive elements (potential sites for binding of the transcription factors Fep1p or Aft1p and Aft2p) were found in the upstream regions of some C. famata and D. hansenii RIB genes. The sequences of C. famata RIB genes have been submitted to the EMBL data library under Accession Nos AJ810169-AJ810173. PMID:15543522

  5. Genetic mapping of the regulator gene determining enterotoxin synthesis in Vibrio cholerae

    International Nuclear Information System (INIS)

    Data on the genetic mapping of mutation tox-7 (the mutation affecting the synthesis of the cholera toxin) were obtained by conjugation crosses between the atoxigenic donor strain Vibrio cholerae Eltor and the toxigenic recipient strain V. cholera classica. The molecular and genetic analysis of the Tox- recombinants indicated that, when the synthesis of the cholera toxin is disrupted in these strains, the tox-7 mutation (which impairs the regulator gene tox) is gained. Close linkage between the tox-7 and pur-63 mutations was established (during the selection procedure there was 81.1% combined transfer with respect to marker pur-63 situated in the donor strain chromosome more proximal than mutation tox-7). The markers were localized in the following order in the region under investigation: asp-cys-nal-pur-61-trp-his-pur-63-tox-7-ile

  6. Genomic characterization of ribitol teichoic acid synthesis in Staphylococcus aureus: genes, genomic organization and gene duplication

    Directory of Open Access Journals (Sweden)

    Lu Lingyi

    2006-04-01

    Full Text Available Abstract Background Staphylococcus aureus or MRSA (Methicillin Resistant S. aureus, is an acquired pathogen and the primary cause of nosocomial infections worldwide. In S. aureus, teichoic acid is an essential component of the cell wall, and its biosynthesis is not yet well characterized. Studies in Bacillus subtilis have discovered two different pathways of teichoic acid biosynthesis, in two strains W23 and 168 respectively, namely teichoic acid ribitol (tar and teichoic acid glycerol (tag. The genes involved in these two pathways are also characterized, tarA, tarB, tarD, tarI, tarJ, tarK, tarL for the tar pathway, and tagA, tagB, tagD, tagE, tagF for the tag pathway. With the genome sequences of several MRSA strains: Mu50, MW2, N315, MRSA252, COL as well as methicillin susceptible strain MSSA476 available, a comparative genomic analysis was performed to characterize teichoic acid biosynthesis in these S. aureus strains. Results We identified all S. aureus tar and tag gene orthologs in the selected S. aureus strains which would contribute to teichoic acids sythesis.Based on our identification of genes orthologous to tarI, tarJ, tarL, which are specific to tar pathway in B. subtilis W23, we also concluded that tar is the major teichoic acid biogenesis pathway in S. aureus. Further analyses indicated that the S. aureus tar genes, different from the divergon organization in B. subtilis, are organized into several clusters in cis. Most interesting, compared with genes in B. subtilis tar pathway, the S. aureus tar specific genes (tarI,J,L are duplicated in all six S. aureus genomes. Conclusion In the S. aureus strains we analyzed, tar (teichoic acid ribitol is the main teichoic acid biogenesis pathway. The tar genes are organized into several genomic groups in cis and the genes specific to tar (relative to tag: tarI, tarJ, tarL are duplicated. The genomic organization of the S. aureus tar pathway suggests their regulations are different when

  7. Integrating gene synthesis and microfluidic protein analysis for rapid protein engineering.

    Science.gov (United States)

    Blackburn, Matthew C; Petrova, Ekaterina; Correia, Bruno E; Maerkl, Sebastian J

    2016-04-20

    The capability to rapidly design proteins with novel functions will have a significant impact on medicine, biotechnology and synthetic biology. Synthetic genes are becoming a commodity, but integrated approaches have yet to be developed that take full advantage of gene synthesis. We developed a solid-phase gene synthesis method based on asymmetric primer extension (APE) and coupled this process directly to high-throughput, on-chip protein expression, purification and characterization (via mechanically induced trapping of molecular interactions, MITOMI). By completely circumventing molecular cloning and cell-based steps, APE-MITOMI reduces the time between protein design and quantitative characterization to 3-4 days. With APE-MITOMI we synthesized and characterized over 400 zinc-finger (ZF) transcription factors (TF), showing that although ZF TFs can be readily engineered to recognize a particular DNA sequence, engineering the precise binding energy landscape remains challenging. We also found that it is possible to engineer ZF-DNA affinity precisely and independently of sequence specificity and thatin silicomodeling can explain some of the observed affinity differences. APE-MITOMI is a generic approach that should facilitate fundamental studies in protein biophysics, and protein design/engineering. PMID:26704969

  8. Constitutive synthesis of a transport function encoded by the Thiobacillus ferrooxidans merC gene cloned in Escherichia coli.

    OpenAIRE

    Kusano, T; Ji, G Y; Inoue, C; Silver, S

    1990-01-01

    Mercuric reductase activity determined by the Thiobacillus ferrooxidans merA gene (cloned and expressed constitutively in Escherichia coli) was measured by volatilization of 203Hg2+. (The absence of a merR regulatory gene in the cloned Thiobacillus mer determinant provides a basis for the constitutive synthesis of this system.) In the absence of the Thiobacillus merC transport gene, the mercury volatilization activity was cryptic and was not seen with whole cells but only with sonication-disr...

  9. Functional conservation analysis and expression modes of grape anthocyanin synthesis genes responsive to low temperature stress.

    Science.gov (United States)

    Zhang, Cheng; Jia, Haifeng; Wu, Weimin; Wang, Xicheng; Fang, Jinggui; Wang, Chen

    2015-12-10

    In grape cultivation, low temperature generally increases the expression of genes involved in synthesis of anthocyanin. In this study, multi-type structural analysis of the proteins encoded by five anthocyanin biosynthesis genes VvF3H, VvPAL, VvCHS3, VvCHS2 and VvLDOX, in addition to nine of their homologous genes revealed that proteins in grapevine shared a high similarity with that in kiwi, red orange and some other species in which the biosynthesis of anthocyanin significantly influenced by low temperature as proved by previous studies. Low temperature regulatory elements were also found in the promoter region of the grapevine genes VvCHS2, VvPAL and VvF3H. These findings indicate that the functions of anthocyanin biosynthesis genes in grapevine are conservative and might be sensitive to low temperature. In order to identify the specific expression patterns of the five anthocyanin biosynthesis genes and the changes of polyphenols, anthocyanins and flavonoids under low temperature stress. The transcription analysis of the five genes and the content of polyphenols, anthocyanins and flavonoids in grape skins were examined, by using Vitis vinifera L. cv. 'Yongyou 1' and 'Juxing' berries as experimental material and treated at 4°C and 25°C for 24h, 48 h, 72 h and 96 h. The results showed that low temperature greatly enhanced the expression of the five anthocyanin biosynthesis genes. Low temperature greatly slowed down the decomposition of polyphenol, anthocyanin, and flavonoid in grape skins. Our study also found that cv. 'Juxing' responded more sensitively to low temperature than cv. 'Yongyou 1'. All the findings would provide a basis for further study on the mechanism of anthocyanin biosynthesis under environmental stress. PMID:26253159

  10. Synthesis and In Vitro Testing of New Potent Polyacridine-Melittin Gene Delivery Peptides

    OpenAIRE

    Baumhover, Nicholas J.; Anderson, Kevin; Fernandez, Christian A.; Rice, Kevin G.

    2010-01-01

    The combination of a polyacridine peptide modified with a melittin fusogenic peptide results in a potent gene transfer agent. Polyacridine peptides of the general formula (Acr-X)n-Cys were prepared by solid phase peptide synthesis, where Acr is Lys modified on its ε-amine with acridine, X is Arg, Leu or Lys and n is 2, 3 or 4 repeats. The Cys residue was modified by either a maleimide-melittin or a thiolpyridine-Cys-melittin fusogenic peptide resulting in reducible or non-reducible polyacridi...

  11. Concurrent synthesis and release of nod-gene-inducing flavonoids from alfalfa roots

    International Nuclear Information System (INIS)

    Flavonoid signals from alfalfa (Medicago sativa L.) induce transcription of nodulation (nod) genes in Rhizobium meliloti. Alfalfa roots release three major nod-gene inducers: 4',7-dihydroxyflavanone, 4',7-dihydroxyflavone, and 4,4'-dihydroxy-2'-methoxychalcone. The objective of the present study was to define temporal relationships between synthesis and exudation for those flavonoids. Requirements for concurrent flavonoid biosynthesis were assessed by treating roots of intact alfalfa seedlings with [U-14C]-L-phenylalanine in the presence or absence of the phenylalanine ammonia-lyase inhibitor L-2-aminoxy-3-phenylpropionic acid (AOPP). In the absence of AOPP, each of the three flavonoids in exudates contained 14C. In the presence of AOPP, 14C labeling and release of all the exuded nod-gene inducers were reduced significantly. AOPP inhibited labeling and release of the strongest nod-gene inducer, methoxychalcone, by more than 90%. The release process responsible for exudation of nod-gene inducers appears to be specific rather than a general phenomenon such as a sloughing off of cells during root growth

  12. Identification of the bchP gene, encoding geranylgeranyl reductase in Chlorobaculum tepidum

    DEFF Research Database (Denmark)

    Gomez Maqueo Chew, Aline; Frigaard, Niels-Ulrik; Bryant, Donald A

    2008-01-01

    The Chlorobaculum tepidum genome contains two paralogous genes, CT2256 and CT1232, whose products are members of the FixC dehydrogenase superfamily and have sequence similarity to geranylgeranyl reductases. Each gene was insertionally inactivated, and the resulting mutants were characterized. CT2......2256 encodes geranylgeranyl reductase (BchP); CT1232 is not involved in bacteriochlorophyll or chlorophyll biosynthesis....

  13. Genes involved in carnitine synthesis and carnitine uptake are up-regulated in the liver of sows during lactation

    OpenAIRE

    Rosenbaum, Susann; Ringseis, Robert; Most, Erika; Hillen, Sonja; Becker, Sabrina; Erhardt, Georg; Reiner, Gerald; Eder, Klaus

    2013-01-01

    BACKGROUND:Convincing evidence exist that carnitine synthesis and uptake of carnitine into cells is regulated by peroxisome proliferator-activated receptor alpha (PPARA), a transcription factor which is physiologically activated during fasting or energy deprivation. Sows are typically in a negative energy balance during peak lactation. We investigated the hypothesis that genes involved in carnitine synthesis and uptake in the liver of sows are up-regulated during peak lactation. FINDINGS:Tra...

  14. Noisy translation of a single gene: fluctuations in protein synthesis from a single template

    CERN Document Server

    Garai, Ashok; Ramakrishnan, T V

    2008-01-01

    We introduce some new quantitative measures of fluctuations in the process of synthesis of proteins from a single messenger RNA (mRNA) template. We calculate the statistical distributions of these fluctuating quantities and extract the strength of the corresponding translational noise. For these calculations we use a model that captures both the mechano-chemistry of each individual ribosome as well as their steric interactions in ribosome traffic on the same mRNA track. By comparing our results for a specific gene of the {\\it Escherichia coli} bacteria with those for the corresponding homogeneous mRNA template, we demonstrate the effects of the sequence inhomogeneities of real genes on the fluctuations and noise. We also suggest {\\it in-vitro} laboratory experiments for testing our theoretical predictions.

  15. [Synthesis of GnRH analogs and their application in targeted gene delivery systems].

    Science.gov (United States)

    Iablokova, T V; Chelushkin, P S; Dorosh, M Iu; Efremov, A M; Orlov, S V; Burov, S V

    2012-01-01

    A set of GnRH analogues containing nuclear localization signal (NLS) of SV-40 virus large T-antigen have been synthesized using solid phase peptide synthesis and chemical ligation technique. Selective chemical ligation was achieved as a result of hydrazone formation in the course of interaction between NLS hydrazide and GnRH analog modified by pyruvic acid. The efficiency of synthesized peptide carriers was demonstrated in experiments with human cancer cells transfected by reporter luciferase and beta-galactosidase genes or suicide HSV-1 thymidine kinase gene. It was shown that selectivity of action on cancer cells can be achieved as a result of peptide/DNA complex penetration through the cell membrane by GnRH receptor-mediated endocytosis pathway. PMID:22792703

  16. Gene duplication and an accelerated evolutionary rate in 11S globulin genes are associated with higher protein synthesis in dicots as compared to monocots

    OpenAIRE

    Li Chun; Li Meng; Dunwell Jim M; Zhang Yuan-Ming

    2012-01-01

    Abstract Background Seed storage proteins are a major source of dietary protein, and the content of such proteins determines both the quantity and quality of crop yield. Significantly, examination of the protein content in the seeds of crop plants shows a distinct difference between monocots and dicots. Thus, it is expected that there are different evolutionary patterns in the genes underlying protein synthesis in the seeds of these two groups of plants. Results Gene duplication, evolutionary...

  17. Graphene oxide-cationic polymer conjugates: Synthesis and application as gene delivery vectors.

    Science.gov (United States)

    Teimouri, Mohsen; Nia, Azadeh Hashem; Abnous, Khalil; Eshghi, Hossein; Ramezani, Mohammad

    2016-01-01

    Nanomedicine as the interface between nanotechnology and medical sciences is a new area that has attracted the attention of vast groups of researchers. Carbon nanomaterials are common platform for synthesis of nanoparticles for biomedical applications due to their low cytotoxicity and feasible internalization into mammalian cell lines (Yang et al., 2007; Arora et al., 2014; Oh and Park, 2014). Synthesis of vectors based on various cationic polymers polyethylenimine (PEI), polypropylenimine (PPI) and polyamidoamine (PAMAM) and their derivatives were considered as a strategy for transferring plasmid DNA and treatment of genetic diseases. Considering the low cytotoxicity of graphene, chemical modification of its surface has led to fabrication of novel gene delivery systems based on graphene and graphene oxide. Herein we report the synthesis of three groups of vectors based on conjugation of graphene oxide (GO) with alkylated derivatives of three different cationic polymers (polyethylenimine (PEI), polypropylenimine (PPI) and polyamidoamine (PAMAM)) through different linkers including surface carboxyl group, glycine and spermidine. Two main challenges in design of gene delivery vectors is decreasing cytotoxicity while improving the transfection efficiency. All synthesized vectors showed significantly lower cellular toxicity compared to bare polymer. A plasmid encoding green fluorescent protein (GFP) was used to evaluate the transfection efficiency of nanoparticles both qualitatively using live cell fluorescent imaging and quantitatively using flow cytometry and each vector was compared to its polymer base. Most successful conjugation strategy was observed in the case of PEI conjugates among which most efficient vector was PEI-GO conjugate bearing glycine linker. This vector was 9 fold more effective in terms of the percent of EGFP transfected cells. PMID:27072918

  18. Effects of Long Chain Fatty Acid Synthesis and Associated Gene Expression in Microalga Tetraselmis sp.

    Directory of Open Access Journals (Sweden)

    T. Catalina Adarme-Vega

    2014-06-01

    Full Text Available With the depletion of global fish stocks, caused by high demand and effective fishing techniques, alternative sources for long chain omega-3 fatty acids are required for human nutrition and aquaculture feeds. Recent research has focused on land-based cultivation of microalgae, the primary producers of omega-3 fatty acids in the marine food web. The effect of salinity on fatty acids and related gene expression was studied in the model marine microalga, Tetraselmis sp. M8. Correlations were found for specific fatty acid biosynthesis and gene expression according to salinity and the growth phase. Low salinity was found to increase the conversion of C18:4 stearidonic acid (SDA to C20:4 eicosatetraenoic acid (ETA, correlating with increased transcript abundance of the Δ-6-elongase-encoding gene in salinities of 5 and 10 ppt compared to higher salinity levels. The expression of the gene encoding β-ketoacyl-coenzyme was also found to increase at lower salinities during the nutrient deprivation phase (Day 4, but decreased with further nutrient stress. Nutrient deprivation also triggered fatty acids synthesis at all salinities, and C20:5 eicosapentaenoic acid (EPA increased relative to total fatty acids, with nutrient starvation achieving a maximum of 7% EPA at Day 6 at a salinity of 40 ppt.

  19. Vibronic coupling explains the ultrafast carotenoid-to-bacteriochlorophyll energy transfer in natural and artificial light harvesters.

    Science.gov (United States)

    Perlík, Václav; Seibt, Joachim; Cranston, Laura J; Cogdell, Richard J; Lincoln, Craig N; Savolainen, Janne; Šanda, František; Mančal, Tomáš; Hauer, Jürgen

    2015-06-01

    The initial energy transfer steps in photosynthesis occur on ultrafast timescales. We analyze the carotenoid to bacteriochlorophyll energy transfer in LH2 Marichromatium purpuratum as well as in an artificial light-harvesting dyad system by using transient grating and two-dimensional electronic spectroscopy with 10 fs time resolution. We find that Förster-type models reproduce the experimentally observed 60 fs transfer times, but overestimate coupling constants, which lead to a disagreement with both linear absorption and electronic 2D-spectra. We show that a vibronic model, which treats carotenoid vibrations on both electronic ground and excited states as part of the system's Hamiltonian, reproduces all measured quantities. Importantly, the vibronic model presented here can explain the fast energy transfer rates with only moderate coupling constants, which are in agreement with structure based calculations. Counterintuitively, the vibrational levels on the carotenoid electronic ground state play the central role in the excited state population transfer to bacteriochlorophyll; resonance between the donor-acceptor energy gap and the vibrational ground state energies is the physical basis of the ultrafast energy transfer rates in these systems. PMID:26049454

  20. Vibronic coupling explains the ultrafast carotenoid-to-bacteriochlorophyll energy transfer in natural and artificial light harvesters

    Science.gov (United States)

    Perlík, Václav; Seibt, Joachim; Cranston, Laura J.; Cogdell, Richard J.; Lincoln, Craig N.; Savolainen, Janne; Šanda, František; Mančal, Tomáš; Hauer, Jürgen

    2015-06-01

    The initial energy transfer steps in photosynthesis occur on ultrafast timescales. We analyze the carotenoid to bacteriochlorophyll energy transfer in LH2 Marichromatium purpuratum as well as in an artificial light-harvesting dyad system by using transient grating and two-dimensional electronic spectroscopy with 10 fs time resolution. We find that Förster-type models reproduce the experimentally observed 60 fs transfer times, but overestimate coupling constants, which lead to a disagreement with both linear absorption and electronic 2D-spectra. We show that a vibronic model, which treats carotenoid vibrations on both electronic ground and excited states as part of the system's Hamiltonian, reproduces all measured quantities. Importantly, the vibronic model presented here can explain the fast energy transfer rates with only moderate coupling constants, which are in agreement with structure based calculations. Counterintuitively, the vibrational levels on the carotenoid electronic ground state play the central role in the excited state population transfer to bacteriochlorophyll; resonance between the donor-acceptor energy gap and the vibrational ground state energies is the physical basis of the ultrafast energy transfer rates in these systems.

  1. Spectral heterogeneity and carotenoid-to-bacteriochlorophyll energy transfer in LH2 light-harvesting complexes from Allochromatium vinosum.

    Science.gov (United States)

    Magdaong, Nikki M; LaFountain, Amy M; Hacking, Kirsty; Niedzwiedzki, Dariusz M; Gibson, George N; Cogdell, Richard J; Frank, Harry A

    2016-02-01

    Photosynthetic organisms produce a vast array of spectral forms of antenna pigment-protein complexes to harvest solar energy and also to adapt to growth under the variable environmental conditions of light intensity, temperature, and nutrient availability. This behavior is exemplified by Allochromatium (Alc.) vinosum, a photosynthetic purple sulfur bacterium that produces different types of LH2 light-harvesting complexes in response to variations in growth conditions. In the present work, three different spectral forms of LH2 from Alc. vinosum, B800-820, B800-840, and B800-850, were isolated, purified, and examined using steady-state absorption and fluorescence spectroscopy, and ultrafast time-resolved absorption spectroscopy. The pigment composition of the LH2 complexes was analyzed by high-performance liquid chromatography, and all were found to contain five carotenoids: lycopene, anhydrorhodovibrin, spirilloxanthin, rhodopin, and rhodovibrin. Spectral reconstructions of the absorption and fluorescence excitation spectra based on the pigment composition revealed significantly more spectral heterogeneity in these systems compared to LH2 complexes isolated from other species of purple bacteria. The data also revealed the individual carotenoid-to-bacteriochlorophyll energy transfer efficiencies which were correlated with the kinetic data from the ultrafast transient absorption spectroscopic experiments. This series of LH2 complexes allows a systematic exploration of the factors that determine the spectral properties of the bound pigments and control the rate and efficiency of carotenoid-to-bacteriochlorophyll energy transfer. PMID:26048106

  2. Vibronic coupling explains the ultrafast carotenoid-to-bacteriochlorophyll energy transfer in natural and artificial light harvesters

    Energy Technology Data Exchange (ETDEWEB)

    Perlík, Václav; Seibt, Joachim; Šanda, František; Mančal, Tomáš [Institute of Physics, Faculty of Mathematics and Physics, Charles University in Prague, Ke Karlovu 5, Prague 121 16 (Czech Republic); Cranston, Laura J.; Cogdell, Richard J. [Institute of Molecular Cell and System Biology, College of Medical, Veterinary and Life Sciences, University of Glasgow, Glasgow Biomedical Research Centre, 120 University Place, Glasgow G12 8TA, Scotland (United Kingdom); Lincoln, Craig N.; Hauer, Jürgen, E-mail: juergen.hauer@tuwien.ac.at [Photonics Institute, Vienna University of Technology, Gusshausstrasse 27, 1040 Vienna (Austria); Savolainen, Janne [Department of Physical Chemistry II, Ruhr-University Bochum, 44780 Bochum (Germany)

    2015-06-07

    The initial energy transfer steps in photosynthesis occur on ultrafast timescales. We analyze the carotenoid to bacteriochlorophyll energy transfer in LH2 Marichromatium purpuratum as well as in an artificial light-harvesting dyad system by using transient grating and two-dimensional electronic spectroscopy with 10 fs time resolution. We find that Förster-type models reproduce the experimentally observed 60 fs transfer times, but overestimate coupling constants, which lead to a disagreement with both linear absorption and electronic 2D-spectra. We show that a vibronic model, which treats carotenoid vibrations on both electronic ground and excited states as part of the system’s Hamiltonian, reproduces all measured quantities. Importantly, the vibronic model presented here can explain the fast energy transfer rates with only moderate coupling constants, which are in agreement with structure based calculations. Counterintuitively, the vibrational levels on the carotenoid electronic ground state play the central role in the excited state population transfer to bacteriochlorophyll; resonance between the donor-acceptor energy gap and the vibrational ground state energies is the physical basis of the ultrafast energy transfer rates in these systems.

  3. Vibronic coupling explains the ultrafast carotenoid-to-bacteriochlorophyll energy transfer in natural and artificial light harvesters

    International Nuclear Information System (INIS)

    The initial energy transfer steps in photosynthesis occur on ultrafast timescales. We analyze the carotenoid to bacteriochlorophyll energy transfer in LH2 Marichromatium purpuratum as well as in an artificial light-harvesting dyad system by using transient grating and two-dimensional electronic spectroscopy with 10 fs time resolution. We find that Förster-type models reproduce the experimentally observed 60 fs transfer times, but overestimate coupling constants, which lead to a disagreement with both linear absorption and electronic 2D-spectra. We show that a vibronic model, which treats carotenoid vibrations on both electronic ground and excited states as part of the system’s Hamiltonian, reproduces all measured quantities. Importantly, the vibronic model presented here can explain the fast energy transfer rates with only moderate coupling constants, which are in agreement with structure based calculations. Counterintuitively, the vibrational levels on the carotenoid electronic ground state play the central role in the excited state population transfer to bacteriochlorophyll; resonance between the donor-acceptor energy gap and the vibrational ground state energies is the physical basis of the ultrafast energy transfer rates in these systems

  4. Selective Protein Extraction from Chlorobium tepidum Chlorosomes Using Detergents. Evidence That CsmA Forms Multimers and Binds Bacteriochlorophyll a

    DEFF Research Database (Denmark)

    Bryant, Donald A; Vassilieva, Elena V; Frigaard, Niels-Ulrik; Li, Hui

    2002-01-01

    Chlorosomes of the photosynthetic green sulfur bacterium Chlorobium tepidum consist of bacteriochlorophyll (BChl) c aggregates that are surrounded by a lipid-protein monolayer envelope that contains ten different proteins. Chlorosomes also contain a small amount of BChl a, but the organization an...

  5. Homeobox Genes and Melatonin Synthesis: Regulatory Roles of the Cone-Rod Homeobox Transcription Factor in the Rodent Pineal Gland

    OpenAIRE

    Kristian Rohde; Morten Møller; Martin Fredensborg Rath

    2014-01-01

    Nocturnal synthesis of melatonin in the pineal gland is controlled by a circadian rhythm in arylalkylamine N-acetyltransferase (AANAT) enzyme activity. In the rodent, Aanat gene expression displays a marked circadian rhythm; release of norepinephrine in the gland at night causes a cAMP-based induction of Aanat transcription. However, additional transcriptional control mechanisms exist. Homeobox genes, which are generally known to encode transcription factors controlling developmental processe...

  6. Prevalence of gca, a gene involved in synthesis of A-band common antigen polysaccharide in Pseudomonas aeruginosa.

    OpenAIRE

    Currie, H L; Lightfoot, J; Lam, J S

    1995-01-01

    Two distinct forms of lipopolysaccharide are expressed by Pseudomonas aeruginosa. These forms are known as the A band and the B band. In an attempt to obtain a better understanding of A-band lipopolysaccharide synthesis, a previously isolated A-band gene known as the gca gene (GDP-D-mannose conversion protein for A-band common antigen polysaccharide) was sequenced and analyzed. Previous protein expression data from our laboratory, along with nucleotide sequence analysis from the present study...

  7. Aquatic contaminants alter genes involved in neurotransmitter synthesis and gonadotropin release in largemouth bass

    International Nuclear Information System (INIS)

    Many aquatic contaminants potentially affect the central nervous system, however the underlying mechanisms of how toxicants alter normal brain function are not well understood. The objectives of this study were to compare the effects of emerging and prevalent environmental contaminants on the expression of brain transcripts with a role in neurotransmitter synthesis and reproduction. Adult male largemouth bass (Micropterus salmoides) were injected once for a 96 h duration with control (water or oil) or with one of two doses of a single chemical to achieve the following body burdens (μg/g): atrazine (0.3 and 3.0), toxaphene (10 and 100), cadmium (CdCl2) (0.000067 and 0.00067), polychlorinated biphenyl (PCB) 126 (0.25 and 2.5), and phenanthrene (5 and 50). Partial largemouth bass gene segments were cloned for enzymes involved in neurotransmitter (glutamic acid decarboxylase 65, GAD65; tyrosine hydroxylase) and estrogen (brain aromatase; CYP19b) synthesis for real-time PCR assays. In addition, neuropeptides regulating feeding (neuropeptide Y) and reproduction (chicken GnRH-II, cGnRH-II; salmon GnRH, sGnRH) were also investigated. Of the chemicals tested, only cadmium, PCB 126, and phenanthrene showed any significant effects on the genes tested, while atrazine and toxaphene did not. Cadmium (0.000067 μg/g) significantly increased cGnRH-II mRNA while PCB 126 (0.25 μg/g) decreased GAD65 mRNA. Phenanthrene decreased GAD65 and tyrosine hydroxylase mRNA levels at the highest dose (50 μg/g) but increased cGnRH-II mRNA at the lowest dose (5 μg/g). CYP19b, NPY, and sGnRH mRNA levels were unaffected by any of the treatments. A hierarchical clustering dendrogram grouped PCB 126 and phenanthrene more closely than other chemicals with respect to the genes tested. This study demonstrates that brain transcripts important for neurotransmitter synthesis neuroendocrine function are potential targets for emerging and prevalent aquatic contaminants.

  8. Aquatic contaminants alter genes involved in neurotransmitter synthesis and gonadotropin release in largemouth bass

    Energy Technology Data Exchange (ETDEWEB)

    Martyniuk, Christopher J. [Department of Physiological Sciences and Center for Environmental and Human Toxicology, University of Florida, Gainesville, FL 32611 (United States); Sanchez, Brian C. [Department of Forestry and Natural Resources and School of Civil Engineering, 195 Marsteller St., Purdue University, West Lafayette, IN 47907 (United States); Szabo, Nancy J.; Denslow, Nancy D. [Department of Physiological Sciences and Center for Environmental and Human Toxicology, University of Florida, Gainesville, FL 32611 (United States); Sepulveda, Maria S., E-mail: mssepulv@purdue.edu [Department of Forestry and Natural Resources and School of Civil Engineering, 195 Marsteller St., Purdue University, West Lafayette, IN 47907 (United States)

    2009-10-19

    Many aquatic contaminants potentially affect the central nervous system, however the underlying mechanisms of how toxicants alter normal brain function are not well understood. The objectives of this study were to compare the effects of emerging and prevalent environmental contaminants on the expression of brain transcripts with a role in neurotransmitter synthesis and reproduction. Adult male largemouth bass (Micropterus salmoides) were injected once for a 96 h duration with control (water or oil) or with one of two doses of a single chemical to achieve the following body burdens ({mu}g/g): atrazine (0.3 and 3.0), toxaphene (10 and 100), cadmium (CdCl{sub 2}) (0.000067 and 0.00067), polychlorinated biphenyl (PCB) 126 (0.25 and 2.5), and phenanthrene (5 and 50). Partial largemouth bass gene segments were cloned for enzymes involved in neurotransmitter (glutamic acid decarboxylase 65, GAD65; tyrosine hydroxylase) and estrogen (brain aromatase; CYP19b) synthesis for real-time PCR assays. In addition, neuropeptides regulating feeding (neuropeptide Y) and reproduction (chicken GnRH-II, cGnRH-II; salmon GnRH, sGnRH) were also investigated. Of the chemicals tested, only cadmium, PCB 126, and phenanthrene showed any significant effects on the genes tested, while atrazine and toxaphene did not. Cadmium (0.000067 {mu}g/g) significantly increased cGnRH-II mRNA while PCB 126 (0.25 {mu}g/g) decreased GAD65 mRNA. Phenanthrene decreased GAD65 and tyrosine hydroxylase mRNA levels at the highest dose (50 {mu}g/g) but increased cGnRH-II mRNA at the lowest dose (5 {mu}g/g). CYP19b, NPY, and sGnRH mRNA levels were unaffected by any of the treatments. A hierarchical clustering dendrogram grouped PCB 126 and phenanthrene more closely than other chemicals with respect to the genes tested. This study demonstrates that brain transcripts important for neurotransmitter synthesis neuroendocrine function are potential targets for emerging and prevalent aquatic contaminants.

  9. Aquatic contaminants alter genes involved in neurotransmitter synthesis and gonadotropin release in largemouth bass.

    Science.gov (United States)

    Martyniuk, Christopher J; Sanchez, Brian C; Szabo, Nancy J; Denslow, Nancy D; Sepúlveda, Maria S

    2009-10-19

    Many aquatic contaminants potentially affect the central nervous system, however the underlying mechanisms of how toxicants alter normal brain function are not well understood. The objectives of this study were to compare the effects of emerging and prevalent environmental contaminants on the expression of brain transcripts with a role in neurotransmitter synthesis and reproduction. Adult male largemouth bass (Micropterus salmoides) were injected once for a 96 h duration with control (water or oil) or with one of two doses of a single chemical to achieve the following body burdens (microg/g): atrazine (0.3 and 3.0), toxaphene (10 and 100), cadmium (CdCl(2)) (0.000067 and 0.00067), polychlorinated biphenyl (PCB) 126 (0.25 and 2.5), and phenanthrene (5 and 50). Partial largemouth bass gene segments were cloned for enzymes involved in neurotransmitter (glutamic acid decarboxylase 65, GAD65; tyrosine hydroxylase) and estrogen (brain aromatase; CYP19b) synthesis for real-time PCR assays. In addition, neuropeptides regulating feeding (neuropeptide Y) and reproduction (chicken GnRH-II, cGnRH-II; salmon GnRH, sGnRH) were also investigated. Of the chemicals tested, only cadmium, PCB 126, and phenanthrene showed any significant effects on the genes tested, while atrazine and toxaphene did not. Cadmium (0.000067 microg/g) significantly increased cGnRH-II mRNA while PCB 126 (0.25 microg/g) decreased GAD65 mRNA. Phenanthrene decreased GAD65 and tyrosine hydroxylase mRNA levels at the highest dose (50 microg/g) but increased cGnRH-II mRNA at the lowest dose (5 microg/g). CYP19b, NPY, and sGnRH mRNA levels were unaffected by any of the treatments. A hierarchical clustering dendrogram grouped PCB 126 and phenanthrene more closely than other chemicals with respect to the genes tested. This study demonstrates that brain transcripts important for neurotransmitter synthesis neuroendocrine function are potential targets for emerging and prevalent aquatic contaminants. PMID:19781795

  10. Synthesis and properties of new chlorin and bacteriochlorin photosensitizers

    Science.gov (United States)

    Mironov, Andrei F.

    1996-01-01

    A series of novel sensitizers, which absorb in the range of 660 - 820 nm, derived from natural occurring chlorophyll and bacteriochlorophyll was synthesized. Biomass of blue-green algae Spirulina platensis was used to prepare chlorophyll a derivatives, and biomass of purple bacteria Rhodobacter capsulatus was applied for preparation of bacteriochlorophyll a. The influence of different substituents on spectral characteristics and the amphipility of the sensitizer was investigated. The route for the synthesis of porphyrin macrocycle with the spacer that bears the isothiocyanate group capable for binding with proteins was proposed. Photophysical properties of chlorin p6, purpurin 18 and their esters in different solvents are investigated. Accumulation of two chlorins in the model Erlich tumor was studied.

  11. Novel Method of Cell-Free In Vitro Synthesis of the Human Fibroblast Growth Factor 1 Gene

    Directory of Open Access Journals (Sweden)

    Peijun Zuo

    2010-01-01

    Full Text Available Recombinant DNA projects generally involve cell-based gene cloning. However, because template DNA is not always readily available, in vitro chemical synthesis of complete genes from DNA oligonucleotides is becoming the preferred method for cloning. This article describes a new, rapid procedure based on Taq polymerase for the precise assembly of DNA oligonucleotides to yield the complete human fibroblast growth factor 1 (FGF1 gene, which is 468 bp long and has a G+C content of 51.5%. The new method involved two steps: (1 the design of the DNA oligonucleotides to be assembled and (2 the assembly of multiple oligonucleotides by PCR to generate the whole FGF1 gene. The procedure lasted a total of only 2 days, compared with 2 weeks for the conventional procedure. This method of gene synthesis is expected to facilitate various kinds of complex genetic engineering projects that require rapid gene amplification, such as cell-free whole-DNA library construction, as well as the construction of new genes or genes that contain any mutation, restriction site, or DNA tag.

  12. Cloning and mapping of infA, the gene for protein synthesis initiation factor IF1.

    OpenAIRE

    Sands, J F; Cummings, H S; Sacerdot, C; Dondon, L; Grunberg-Manago, M; Hershey, J W

    1987-01-01

    The gene for translation initiation factor IF1, infA, has been identified by using two synthetic oligonucleotides to screen a Charon 30 library of Escherichia coli DNA. A recombinant lambda phage, C1921, was purified from a plaque positive for both probes. A 2.8 kb BglII fragment and a 2.0 kb HindIII fragment isolated from C1921 were subcloned into the BamHI and HindIII sites of pBR322 to yield pTB7 and pTH2 respectively. Synthesis of IF1 in maxicells transformed with pTB7 or pTH2 indicates t...

  13. Antiaging Gene Klotho Regulates Adrenal CYP11B2 Expression and Aldosterone Synthesis.

    Science.gov (United States)

    Zhou, Xiaoli; Chen, Kai; Wang, Yongjun; Schuman, Mariano; Lei, Han; Sun, Zhongjie

    2016-06-01

    Deficiency of the antiaging gene Klotho (KL) induces renal damage and hypertension through unknown mechanisms. In this study, we assessed whether KL regulates expression of CYP11B2, a key rate-limiting enzyme in aldosterone synthesis, in adrenal glands. We found that haplodeficiency of KL(+/-) in mice increased the plasma level of aldosterone by 16 weeks of age, which coincided with spontaneous and persistent elevation of BP. Blockade of aldosterone actions by eplerenone reversed KL deficiency-induced hypertension and attenuated the kidney damage. Protein expression of CYP11B2 was upregulated in adrenal cortex of KL(+/-) mice. KL and CYP11B2 proteins colocalized in adrenal zona glomerulosa cells. Silencing of KL upregulated and overexpression of KL downregulated CYP11B2 expression in human adrenocortical cells. Notably, silencing of KL decreased expression of SF-1, a negative transcription factor of CYP11B2, but increased phosphorylation of ATF2, a positive transcription factor of CYP11B2, which may contribute to upregulation of CYP11B2 expression. Therefore, these results show that KL regulates adrenal CYP11B2 expression. KL deficiency-induced spontaneous hypertension and kidney damage may be partially attributed to the upregulation of CYP11B2 expression and aldosterone synthesis. PMID:26471128

  14. Sugar-mediated semidian oscillation of gene expression in the cassava storage root regulates starch synthesis

    Energy Technology Data Exchange (ETDEWEB)

    Jansson, Christer; Baguma, Yona; Sun, Chuanxin; Boren, Mats; Olsson, Helena; Rosenqvist, Sara; Mutisya, Joel; Rubaihayo, Patrick R.; Jansson, Christer

    2008-01-15

    Starch branching enzyme (SBE) activity in the cassava storage root exhibited a diurnal fluctuation, dictated by a transcriptional oscillation of the corresponding SBE genes. The peak of SBE activity coincided with the onset of sucrose accumulation in the storage, and we conclude that the oscillatory mechanism keeps the starch synthetic apparatus in the storage root sink in tune with the flux of sucrose from the photosynthetic source. When storage roots were uncoupled from the source, SBE expression could be effectively induced by exogenous sucrose. Turanose, a sucrose isomer that cannot be metabolized by plants, mimicked the effect of sucrose, demonstrating that downstream metabolism of sucrose was not necessary for signal transmission. Also glucose and glucose-1-P induced SBE expression. Interestingly, induction by sucrose, turanose and glucose but not glucose-1-P sustained an overt semidian (12-h) oscillation in SBE expression and was sensitive to the hexokinase (HXK) inhibitor glucosamine. These results suggest a pivotal regulatory role for HXK during starch synthesis. Abscisic acid (ABA) was another potent inducer of SBE expression. Induction by ABA was similar to that of glucose-1-P in that it bypassed the semidian oscillator. Both the sugar and ABA signaling cascades were disrupted by okadaic acid, a protein phosphatase inhibitor. Based on these findings, we propose a model for sugar signaling in regulation of starch synthesis in the cassava storage root.

  15. A single gene directs synthesis of a precursor protein with beta- and alpha-amylase activities in Bacillus polymyxa.

    OpenAIRE

    Uozumi, N; Sakurai, K.(Theoretical Particle Physics and Cosmology Group, Department of Physics, King’s College London, WC2R 2LS, London, UK); Sasaki, T.; Takekawa, S.; Yamagata, H; Tsukagoshi, N; Udaka, S

    1989-01-01

    The Bacillus polymyxa amylase gene comprises 3,588 nucleotides. The mature amylase comprises 1,161 amino acids with a molecular weight of 127,314. The gene appeared to be divided into two portions by the direct-repeat sequence located at almost the middle of the gene. The 5' region upstream of the direct-repeat sequence was shown to be responsible for the synthesis of beta-amylase. The 3' region downstream of the direct-repeat sequence contained four sequences homologous with those in other a...

  16. EFFECT OF HYPOXIA ON DNA SYNTHESIS AND C-MYC GENE EXPRESSION OF PULMONARY ARTERY SMOOTH MUSCLE CELLS

    Institute of Scientific and Technical Information of China (English)

    罗兰; 李世强; 蔡英年

    1996-01-01

    The neonate is particularly susceptible to the development of hypoxie pulmonary hypertension. The present study was undertaken to observe the effect of hypoxia on DNA synthesis and c-mye gene expressionbetween newborn calf and adult bovine PASMC in vitro DNA synthesis measured by 3H-TdR incorporation was increased after hypoxie challenge for 24h. Hypoxia enhanced the increment in 3H-TdR incorporationinduced by EGF. Northern blot analysis revealed that PASMC cultured in both normoxia and hypoxia expressed c-mye gene transcript of 2.2Kb ,but there is a higher 2.2Kb mRNA expression in hypoxie PASMC than that in normoxia. We speculate that newborn calf PASMC exhibited potential response to hypoxia than adult,which was augmented by EGF. Enhanced c-myc gene expression may lead to a great understanding of the mechanism of PASMC growth in the development of pulmonary hypertension.

  17. Polyploid genome of Camelina sativa revealed by isolation of fatty acid synthesis genes

    Directory of Open Access Journals (Sweden)

    Shewmaker Christine K

    2010-10-01

    Full Text Available Abstract Background Camelina sativa, an oilseed crop in the Brassicaceae family, has inspired renewed interest due to its potential for biofuels applications. Little is understood of the nature of the C. sativa genome, however. A study was undertaken to characterize two genes in the fatty acid biosynthesis pathway, fatty acid desaturase (FAD 2 and fatty acid elongase (FAE 1, which revealed unexpected complexity in the C. sativa genome. Results In C. sativa, Southern analysis indicates the presence of three copies of both FAD2 and FAE1 as well as LFY, a known single copy gene in other species. All three copies of both CsFAD2 and CsFAE1 are expressed in developing seeds, and sequence alignments show that previously described conserved sites are present, suggesting that all three copies of both genes could be functional. The regions downstream of CsFAD2 and upstream of CsFAE1 demonstrate co-linearity with the Arabidopsis genome. In addition, three expressed haplotypes were observed for six predicted single-copy genes in 454 sequencing analysis and results from flow cytometry indicate that the DNA content of C. sativa is approximately three-fold that of diploid Camelina relatives. Phylogenetic analyses further support a history of duplication and indicate that C. sativa and C. microcarpa might share a parental genome. Conclusions There is compelling evidence for triplication of the C. sativa genome, including a larger chromosome number and three-fold larger measured genome size than other Camelina relatives, three isolated copies of FAD2, FAE1, and the KCS17-FAE1 intergenic region, and three expressed haplotypes observed for six predicted single-copy genes. Based on these results, we propose that C. sativa be considered an allohexaploid. The characterization of fatty acid synthesis pathway genes will allow for the future manipulation of oil composition of this emerging biofuel crop; however, targeted manipulations of oil composition and general

  18. Organization and expression of the Co1D-CA23 plasmid genes associated with colicin synthesis

    International Nuclear Information System (INIS)

    The authors have investigated the organization and regulation of the functioning of colicin genes, the immunity protein, and lysis protein of the colicinogenic plasmid Co1D-CA23. In addition they have analyzed the polypeptides synthesized in minicells carrying plasmid Co1D, its Th5 mutants, and the recombinant plasmids obtained on cloning of the EcoRV fragments of Co1D on vector BR325. They have determined the position of the promoter of the colicin gene and the direction of its transcription. Furthermore they were able to show that the gene determining cell immunity to colicin D is transcribed independently of the colicin gene from its own SOS-independent promoter. Treatment of the cells carrying plasmid Co1D with mitomycin C leads to the induction of synthesis of not only colicin but also of a protein with a molecular weight of 10 kdalton, causing under these conditions the death and lysis of the cells. Together with colicin, this protein is detected in the culture liquid on lysis of the cells. Plasmid mutations impairing the synthesis of the lysis protein inhibit the release of colicin into the medium. They have shown that the genes of colicin and the lysis protein are arranged into one operon, the lysis gene being transcribed after the colicin gene. They have proposed a genetic map for plasmid Co1D-CA23

  19. Terminator Operon Reporter: combining a transcription termination switch with reporter technology for improved gene synthesis and synthetic biology applications.

    Science.gov (United States)

    Zampini, Massimiliano; Mur, Luis A J; Rees Stevens, Pauline; Pachebat, Justin A; Newbold, C James; Hayes, Finbarr; Kingston-Smith, Alison

    2016-01-01

    Synthetic biology is characterized by the development of novel and powerful DNA fabrication methods and by the application of engineering principles to biology. The current study describes Terminator Operon Reporter (TOR), a new gene assembly technology based on the conditional activation of a reporter gene in response to sequence errors occurring at the assembly stage of the synthetic element. These errors are monitored by a transcription terminator that is placed between the synthetic gene and reporter gene. Switching of this terminator between active and inactive states dictates the transcription status of the downstream reporter gene to provide a rapid and facile readout of the accuracy of synthetic assembly. Designed specifically and uniquely for the synthesis of protein coding genes in bacteria, TOR allows the rapid and cost-effective fabrication of synthetic constructs by employing oligonucleotides at the most basic purification level (desalted) and without the need for costly and time-consuming post-synthesis correction methods. Thus, TOR streamlines gene assembly approaches, which are central to the future development of synthetic biology. PMID:27220405

  20. Vibronic coupling explains the ultrafast carotenoid-to-bacteriochlorophyll energy transfer in natural and artificial light harvesters

    CERN Document Server

    Perlík, Václav; Cranston, Laura J; Cogdell, Richard J; Lincoln, Craig N; Savolainen, Janne; Šanda, František; Mančal, Tomáš; Hauer, Jürgen

    2015-01-01

    The initial energy transfer in photosynthesis occurs between the light-harvesting pigments and on ultrafast timescales. We analyze the carotenoid to bacteriochlorophyll energy transfer in LH2 Marichromatium purpuratum as well as in an artificial light-harvesting dyad system by using transient grating and two-dimensional electronic spectroscopy with 10 fs time resolution. We find that F\\"orster-type models reproduce the experimentally observed 60 fs transfer times, but overestimate coupling constants, which leads to a disagreement with both linear absorption and electronic 2D-spectra. We show that a vibronic model, which treats carotenoid vibrations on both electronic ground and excited state as part of the system's Hamiltonian, reproduces all measured quantities. Importantly, the vibronic model presented here can explain the fast energy transfer rates with only moderate coupling constants, which are in agreement with structure based calculations. Counterintuitively, the vibrational levels on the carotenoid el...

  1. Synthesis of multi-functional large pore mesoporous silica nanoparticles as gene carriers

    International Nuclear Information System (INIS)

    The development of functional nanocarriers that can enhance the cellular delivery of a variety of nucleic acid agents is important in many biomedical applications such as siRNA therapy. We report the synthesis of large pore mesoporous silica nanoparticles (LPMSN) loaded with iron oxide and covalently modified by polyethyleneimine (denoted PEI-Fe-LPMSN) as carriers for gene delivery. The LPMSN have a particle size of ∼200 nm and a large pore size of 11 nm. The large pore size is essential for the formation of large iron oxide nanoparticles to increase the magnetic properties and the adsorption capacity of siRNA molecules. The magnetic property facilitates the cellular uptake of nanocarriers under an external magnetic field. PEI is covalently grafted on the silica surface to enhance the nanocarriers’ affinity against siRNA molecules and to improve gene silencing performance. The PEI-Fe-LPMSN delivered siRNA–PLK1 effectively into osteosarcoma cancer cells, leading to cell viability inhibition of 80%, higher compared to the 50% reduction when the same dose of siRNA was delivered by a commercial product, oligofectamine. (paper)

  2. Association of triacylglyceride content and transcript abundance of genes involving in lipid synthesis of nitrogen deficient Phaeodactylum tricornutum

    Science.gov (United States)

    Zhang, Lin; Han, Jichang; Yang, Guanpin; Zhu, Baohua; Pan, Kehou

    2014-03-01

    Phaeodactylum tricornutum is a diatom that is rich in lipids. Recently, it has received much attention as a feedstock for biodiesel production. Nitrogen deficiency is widely known to increase the content of neutral lipids (mainly triacylglycerides, or TAGs) of microalgae, including P. tricornutum, but the mechanism is unclear. In this study, we deciphered the correlations between TAG content and nine key enzymatic genes involved in lipid synthesis in P. tricornutum. After being cultured under nitrogen-free conditions for 0, 4, 24, 48, 72, 120, and 168 h, the TAG contents of P. tricornutum cells were assayed and the transcript abundances of the target genes were monitored by quantitative real-time PCR. The results show that the abundances of four target gene transcripts ( LACS3, G3PDH2, G3PDH3, and G3PDH5) were positively correlated with TAG content, indicating that these genes may be involved in TAG synthesis in P. tricornutum. The findings improve our understanding of the metabolic network and regulation of lipid synthesis and will guide the future genetic improvement of the TAG content of P. tricornutum.

  3. Association Mapping of Cell Wall Synthesis Regulatory Genes and Cell Wall Quality in Switchgrass

    Energy Technology Data Exchange (ETDEWEB)

    Bartley, Laura [Univ. of Oklahoma, Norman, OK (United States). Dept. of Microbiology and Plant Biology; Wu, Y. [Oklahoma State Univ., Stillwater, OK (United States); Zhu, L. [Oklahoma State Univ., Stillwater, OK (United States); Brummer, E. C. [Noble Foundation, Ardmore, OK (United States); Saha, M. [Noble Foundation, Ardmore, OK (United States)

    2016-05-31

    markers might be used to select switchgrass genotypes with improved composition in breeding programs for biofuel and forage production. Because the SSAC continues to be characterized by collaborators in the bioenergy community, the data generated will be used to identify additional markers in higher resolution genotyping data to approach identifying the genes and alleles that cause natural variation in switchgrass cell wall quality. For example, these markers can be surveyed in the 2100-member Oklahoma Southern and Northern Lowland switchgrass collections that this project also characterized. An orthogonal approach to biodiversity studies, using comparative functional genomics permits systematic querying of how much regulatory information is likely to be transferable from dicots to grasses and use of accumulated functional genomics resources for better-characterized grass species, such as rice, itself a biomass source in global agriculture and in certain regions. The project generated and tested a number of specific hypotheses regarding cell wall transcription factors and enzymes of grasses. To aid identification of cell wall regulators, the project assembled a novel, highdepth and -quality gene association network using a general linearized model scoring system to combine rice gene network data. Using known or putative orthologs of Arabidopsis cell wall biosynthesis genes and regulators, the project pulled from this network a cell wall sub-network that includes 96 transcription factors. Reverse genetics of a co-ortholog of the Arabidopsis MYB61 transcription factor in rice revealed that this regulatory node has evolved the ability to regulate grass-specific cell wall synthesis enzymes. A transcription factor with such activity has not been previously characterized to our knowledge, representing a major conclusion of this work. Changes in gene expression in a protoplast-based assay demonstrated positive or negative roles in cell wall regulation for eleven other

  4. Synthesis, cloning and expression of a novel pre-miniproinsulin analogue gene in Escherichia coli

    Directory of Open Access Journals (Sweden)

    Ahmed A. Abolliel

    2015-09-01

    Full Text Available In the present study, a novel pre-miniproinsulin analogue was designed to have a short 9 residue sequence replacing the 35 residue C-chain, one lysine and one arginine added to the C-terminus of the B-chain in combination with glycine and arginine substitution at A21 and B29, respectively, and a 16-residue fusion partner comprising the pentapeptide sequence (PSDKP of the N-terminus of human tumor necrosis factor-α (TNF-α, 6 histidine residues for Ni2+ chelated affinity purification and a pentapeptide ending with methionine for ease of chemical cleavage fused at the N-terminus. Homology modeling of the designed protein against miniproinsulin (protein databank file 1 efeA as a template showed that the distance between the α-carbons of the C-terminus of the B-chain and the N-terminus of the A-chain did not change; the root-mean-square deviation of the backbone atoms between the structures of modeled miniproinsulin and miniproinsulin template was 0.000 Å. DNA sequencing of the synthesized gene showed 100% identity with theoretical sequence. The gene was constructed taking into account the codon preference of Escherichia coli (CAI value 0.99 in order to increase the expression rate of the DNA in the host strain. The designed gene was synthesized using DNA synthesis technology and then cloned into the expression plasmid pET-24a(+ and propagated in E. coli strain JM109. Gene expression was successful in two E. coli strains: namely JM109(DE3 and BL21(DE3pLysS. SDS–PAGE analysis was carried out to check protein size and to check and optimize expression. Rapid screening and purification of the resulting protein was carried out by Ni–NTA technology. The identity of the expressed protein was verified by immunological detection method of western blot using polyclonal rabbit antibody against insulin.

  5. Synthesis of a gene for the HIV transactivator protein TAT by a novel single stranded approach involving in vivo gap repair.

    OpenAIRE

    Adams, S E; Johnson, I D; Braddock, M; Kingsman, A J; Kingsman, S M; Edwards, R M

    1988-01-01

    The synthesis of a gene for the HIV TAT protein is described using a novel approach that capitalises on the ability to synthesise oligonucleotides of greater than 100 bp in length. It involves the synthesis of large oligomers covering one strand of the desired gene in its entirety and the use of small complementary bridging and adapter oligonucleotides to direct the assembly and cloning of the large oligomers. After ligation to the cloning vector the partially single stranded intermediate is ...

  6. Potency of Individual Bile Acids to Regulate Bile Acid Synthesis and Transport Genes in Primary Human Hepatocyte Cultures

    OpenAIRE

    Liu, Jie; LU, Hong; Lu, Yuan-Fu; Lei, Xiaohong; Cui, Julia Yue; Ellis, Ewa; Strom, Stephen C.; Klaassen, Curtis D.

    2014-01-01

    Bile acids (BAs) are known to regulate their own homeostasis, but the potency of individual bile acids is not known. This study examined the effects of cholic acid (CA), chenodeoxycholic acid (CDCA), deoxycholic acid (DCA), lithocholic acid (LCA) and ursodeoxycholic acid (UDCA) on expression of BA synthesis and transport genes in human primary hepatocyte cultures. Hepatocytes were treated with the individual BAs at 10, 30, and 100μM for 48 h, and RNA was extracted for real-time PCR analysis. ...

  7. Plant Anthocyanin Synthesis and Gene Regulation%植物花青素合成与基因调控

    Institute of Scientific and Technical Information of China (English)

    马延蕊; 张金文; 梁慧光; 柳永强

    2012-01-01

    文章在阐述植物花青素生物化学合成的基础上,综述了植物花色素苷合成的基因调控及环境、激素化学物质等外在因子对花青素基因调控的影响。结果表明:在植物花青素代谢中,温度、光照、紫外线、施肥状况、激素水平等因素能诱发花青素合成的调节基因或反义基因的表达,从而诱导或抑制了花青素的合成。在调控基因中,一些对花青素合成的结构蛋白表达产生促进作用,一些则具有抑制效应。不同外在因子激活或抑制调控基因的种类与数量不同,因此,产生了不同的花青素组型或相同组型的不同配比,使植物器官表现不同的颜色。%This paper aims to explain the biochemistry of anthocyanin synthesis based on an overview of plant anthocyanin synthesis genes and environmental factors in the regulation of anthocyanin metabolism. The results show that: ① The metabolism of anthocyanins in plants is affected by the temperature, light, ultraviolet, fertilization status, hormone levels and other factors, which affect the military anthocyanin biosynthetic genes, and then induce or inhibit the synthesis of anthocyanins. ② In the regulation of genes, some of the structural genes of anthocyanin synthesis showed promoting effect, while others showed inhibitory effect. At different environ- mental conditions, the regulation of gene activation and inhibition of the amount of different regulatory genes that anthocyanin accumulation is different, and cause different colors of plant-organs production. ③ In different environmental factors or hor-mones induced to produce the same or different regulation of gene expression changes in regulatory genes, resulting in several different anthocyanins or anthocyanin ratio changes, so that the color of plant organs in different colors.

  8. Effect of Carotenoids and Monogalactosyl Diglyceride on Bacteriochlorophyll c Aggregates in Aqueous Buffer: Implications for the Self-assembly of Chlorosomes

    OpenAIRE

    Klinger, Pavel; Arellano, Juan B.; Vácha, Frantisek; Hála, Jan; Psencik, Jakub

    2004-01-01

    Aggregation of bacteriochlorophyll (BChl) c from chlorosomes, the main light-harvesting complex of green bacteria, has been studied in aqueous buffer. Unlike other chlorophyll-like molecules, BChl c is rather soluble in aqueous buffer, forming dimers. When BChl c is mixed with carotenoids (Car), the BChl c Qy transition is further redshifted, in respect to that of monomers and dimers. The results suggest that Car are incorporated in the aggregates and induce further aggregation of BChl c. The...

  9. High throughput gene complementation screening permits identification of a mammalian mitochondrial protein synthesis (ρ(-)) mutant.

    Science.gov (United States)

    Potluri, Prasanth; Procaccio, Vincent; Scheffler, Immo E; Wallace, Douglas C

    2016-08-01

    To identify nuclear DNA (nDNA) oxidative phosphorylation (OXPHOS) gene mutations using cultured cells, we have developed a complementation system based on retroviral transduction with a full length cDNA expression library and selection for OXHOS function by growth in galactose. We have used this system to transduce the Chinese hamster V79-G7 OXPHOS mutant cell line with a defect in mitochondrial protein synthesis. The complemented cells were found to have acquired the cDNA for the bS6m polypeptide of the small subunit of the mitochondrial ribosome. bS6m is a 14 kDa polypeptide located on the outside of the mitochondrial 28S ribosomal subunit and interacts with the rRNA. The V79-G7 mutant protein was found to harbor a methionine to threonine missense mutation at codon 13. The hamster bS6m null mutant could also be complemented by its orthologs from either mouse or human. bS6m protein tagged at its C-terminus by HA, His or GFP localized to the mitochondrion and was fully functional. Through site-directed mutagenesis we identified the probable RNA interacting residues of the bS6m peptide and tested the functional significance of mammalian specific C-terminal region. The N-terminus of the bS6m polypeptide functionally corresponds to that of the prokaryotic small ribosomal subunit, but deletion of C-terminal residues along with the zinc ion coordinating cysteine had no functional effect. Since mitochondrial diseases can result from hundreds to thousands of different nDNA gene mutations, this one step viral complementation cloning may facilitate the molecular diagnosis of a range of nDNA mitochondrial disease mutations. This article is part of a Special Issue entitled 'EBEC 2016: 19th European Bioenergetics Conference, Riva del Garda, Italy, July 2-6, 2016', edited by Prof. Paolo Bernardi. PMID:26946086

  10. Synthesis and Gene Silencing Properties of siRNAs Containing Terminal Amide Linkages

    Directory of Open Access Journals (Sweden)

    Maria Gaglione

    2014-01-01

    Full Text Available The active components of the RNAi are 21 nucleotides long dsRNAs containing a 2 nucleotide overhang at the 3′ end, carrying 5′-phosphate and 3′-hydroxyl groups (siRNAs. Structural analysis revealed that the siRNA is functionally bound at both ends to RISC. Terminal modifications are considered with interest as the introduction of chemical moieties interferes with the 3′ overhang recognition by the PAZ domain and the 5′-phosphate recognition by the MID and PIWI domains of RISC. Herein, we report the synthesis of modified siRNAs containing terminal amide linkages by introducing hydroxyethylglycine PNA (hegPNA moieties at 5′, and at 3′ positions and on both terminals. Results of gene silencing studies highlight that some of these modifications are compatible with the RNAi machinery and markedly increase the resistance to serum-derived nucleases even after 24 h of incubation. Molecular docking simulations were attained to give at atomistic level a clearer picture of the effect of the most performing modifications on the interactions with the human Argonaute 2 PAZ, MID, and PIWI domains. This study adds another piece to the puzzle of the heterogeneous chemical modifications that can be attained to enhance the silencing efficiency of siRNAs.

  11. Effect of Diet Supplementation on the Expression of Bovine Genes Associated with Fatty Acid Synthesis and Metabolism

    Directory of Open Access Journals (Sweden)

    Sandeep J. Joseph

    2010-03-01

    Full Text Available Conjugated linoleic acids (CLA are of important nutritional and health benefit to human. Food products of animal origin are their major dietary source and their concentration increases with high concentrate diets fed to animals. To examine the effects of diet supplementation on the expression of genes related to lipid metabolism, 28 Angus steers were fed either pasture only, pasture with soybean hulls and corn oil, pasture with corn grain, or high concentrate diet. At slaughter, samples of subcutaneous adipose tissue were collected, from which RNA was extracted. Relative abundance of gene expression was measured using Affymetrix GeneChip Bovine Genome array. An ANOVA model nested within gene was used to analyze the background adjusted, normalized average difference of probe-level intensities. To control experiment wise error, a false discovery rate of 0.01 was imposed on all contrasts. Expression of several genes involved in the synthesis of enzymes related to fatty acid metabolism and lipogenesis such as stearoyl-CoA desaturase (SCD, fatty acid synthetase (FASN, lipoprotein lipase (LPL, fatty-acyl elongase (LCE along with several trancription factors and co-activators involved in lipogenesis were found to be differentially expressed. Confirmatory RT-qPCR was done to validate the microarray results, which showed satisfactory correspondence between the two platforms. Results show that changes in diet by increasing dietary energy intake by supplementing high concentrate diet have effects on the transcription of genes encoding enzymes involved in fat metabolism which in turn has effects on fatty acid content in the carcass tissue as well as carcass quality. Corn supplementation either as oil or grain appeared to significantly alter the expression of genes directly associated with fatty acid synthesis.

  12. Potency of Individual Bile Acids to Regulate Bile Acid Synthesis and Transport Genes in Primary Human Hepatocyte Cultures

    Science.gov (United States)

    Liu, Jie; Lu, Hong; Lu, Yuan-Fu; Lei, Xiaohong; Cui, Julia Yue; Ellis, Ewa; Strom, Stephen C.; Klaassen, Curtis D.

    2014-01-01

    Bile acids (BAs) are known to regulate their own homeostasis, but the potency of individual bile acids is not known. This study examined the effects of cholic acid (CA), chenodeoxycholic acid (CDCA), deoxycholic acid (DCA), lithocholic acid (LCA) and ursodeoxycholic acid (UDCA) on expression of BA synthesis and transport genes in human primary hepatocyte cultures. Hepatocytes were treated with the individual BAs at 10, 30, and 100μM for 48 h, and RNA was extracted for real-time PCR analysis. For the classic pathway of BA synthesis, BAs except for UDCA markedly suppressed CYP7A1 (70–95%), the rate-limiting enzyme of bile acid synthesis, but only moderately (35%) down-regulated CYP8B1 at a high concentration of 100μM. BAs had minimal effects on mRNA of two enzymes of the alternative pathway of BA synthesis, namely CYP27A1 and CYP7B1. BAs increased the two major target genes of the farnesoid X receptor (FXR), namely the small heterodimer partner (SHP) by fourfold, and markedly induced fibroblast growth factor 19 (FGF19) over 100-fold. The BA uptake transporter Na+-taurocholate co-transporting polypeptide was unaffected, whereas the efflux transporter bile salt export pump was increased 15-fold and OSTα/β were increased 10–100-fold by BAs. The expression of the organic anion transporting polypeptide 1B3 (OATP1B3; sixfold), ATP-binding cassette (ABC) transporter G5 (ABCG5; sixfold), multidrug associated protein-2 (MRP2; twofold), and MRP3 (threefold) were also increased, albeit to lesser degrees. In general, CDCA was the most potent and effective BA in regulating these genes important for BA homeostasis, whereas DCA and CA were intermediate, LCA the least, and UDCA ineffective. PMID:25055961

  13. Identification of reference genes suitable for qRT-PCR in grapevine and application for the study of the expression of genes involved in pterostilbene synthesis.

    Science.gov (United States)

    Gamm, Magdalena; Héloir, Marie-Claire; Kelloniemi, Jani; Poinssot, Benoît; Wendehenne, David; Adrian, Marielle

    2011-04-01

    The recent publication of the grapevine genome sequence facilitates the use of qRT-PCR to study gene expression changes. For this approach, reference genes are commonly used to normalize data and their stability of expression should be systematically validated. Among grapevine defenses is the production of the antimicrobial stilbenic phytoalexins, notably the highly fungitoxic pterostilbene, which plays a crucial role in grapevine interaction with Plasmopara viticola and Botrytis cinerea. As a resveratrol O-methyltransferase (ROMT) gene involved in pterostilbene synthesis was recently identified, we investigated the accumulation of the corresponding transcripts to those of two other stilbene biosynthesis related genes phenylalanine ammonia lyase (PAL) and stilbene synthase (STS) in response to pathogen infection. Using three computer-based statistical methods and C(t) values or LRE method generated values as input data, we have first identified two reference genes (VATP16 and 60SRP) suitable for normalization of qPCR expression data obtained in grapevine leaves and berries infected by P. viticola and B. cinerea, respectively. Next, we have highlighted that the expression of ROMT is induced in P. viticola-infected leaves and also in B. cinerea-infected berries, confirming the involvement of pterostilbene in grapevine defenses. PMID:21340517

  14. Homeobox Genes and Melatonin Synthesis: Regulatory Roles of the Cone-Rod Homeobox Transcription Factor in the Rodent Pineal Gland

    Directory of Open Access Journals (Sweden)

    Kristian Rohde

    2014-01-01

    Full Text Available Nocturnal synthesis of melatonin in the pineal gland is controlled by a circadian rhythm in arylalkylamine N-acetyltransferase (AANAT enzyme activity. In the rodent, Aanat gene expression displays a marked circadian rhythm; release of norepinephrine in the gland at night causes a cAMP-based induction of Aanat transcription. However, additional transcriptional control mechanisms exist. Homeobox genes, which are generally known to encode transcription factors controlling developmental processes, are also expressed in the mature rodent pineal gland. Among these, the cone-rod homeobox (CRX transcription factor is believed to control pineal-specific Aanat expression. Based on recent advances in our understanding of Crx in the rodent pineal gland, we here suggest that homeobox genes play a role in adult pineal physiology both by ensuring pineal-specific Aanat expression and by facilitating cAMP response element-based circadian melatonin production.

  15. Temporal aspects of DNA and RNA synthesis during human immunodeficiency virus infection: Evidence for differential gene expression

    International Nuclear Information System (INIS)

    The kinetics of retroviral DNA and RNA synthesis are parameters vital to understanding viral growth, especially for human immunodeficiency virus (HIV), which encodes several of its own regulatory genes. The authors have established a single-cycle growth condition for HIV in H9 cells, a human CD4+ lymphocyte line. The full-length viral linear DNA is first detectable by 4 h postinfection. During a one-step growth of HIV, amounts of viral DNA gradually increase until 8 to 12 h postinfection and then decrease. The copy number of unintegrated viral DNA is not extraordinarily high even at its peak. Most strikingly, there is a temporal program of RNA accumulation: the earliest RNA is greatly enriched in the 2-kilobase subgenomic mRNA species, while the level of 9.2-kilobase RNA which is both genomic RNA and mRNA remains low until after 24 h of infection. Virus production begins at about 24 h postinfection. Thus, viral DNA synthesis is as rapid as for other retroviruses, but viral RNA synthesis involves temporal alteration in the species that accumulate, presumably as a consequence of viral regulatory genes

  16. Probing the Role of the Eighth Bacteriochlorophyll in holo-FMO Complex by Simulated Two-Dimensional Electronic Spectroscopy

    CERN Document Server

    Yeh, Shu-Hao

    2014-01-01

    The Fenna-Matthews-Olson (FMO) protein-pigment complex acts as a molecular wire between the outer antenna system and the reaction center (RC); it is an important model system to study the excitonic energy transfer. Recent crystallographic studies report the existence of an additional (eighth) bacteriochlorophyll a (BChl a). To understand the functionality of this eighth BChl, we simulated the two-dimensional electronic spectra of both the 7-site (apo form) and the 8-site (holo form) variant of the FMO complex from green sulfur bacteria, Prosthecochloris aestuarii. By comparing the difference between the spectrum, it was found that the eighth BChl can affect two different excitonic energy transfer pathways, these being: (1) directly involve in the first pathway 6 $\\rightarrow$ 3 $\\rightarrow$ 1 of the apo form model by passing the excitonic energy to exciton 6; and (2) increase the excitonic wave function overlap between excitons 4 and 5 in the second pathway (7 $\\rightarrow$ 4,5 $\\rightarrow$ 2 $\\rightarrow$ ...

  17. A Robust and Versatile Method of Combinatorial Chemical Synthesis of Gene Libraries via Hierarchical Assembly of Partially Randomized Modules

    Science.gov (United States)

    Popova, Blagovesta; Schubert, Steffen; Bulla, Ingo; Buchwald, Daniela; Kramer, Wilfried

    2015-01-01

    A major challenge in gene library generation is to guarantee a large functional size and diversity that significantly increases the chances of selecting different functional protein variants. The use of trinucleotides mixtures for controlled randomization results in superior library diversity and offers the ability to specify the type and distribution of the amino acids at each position. Here we describe the generation of a high diversity gene library using tHisF of the hyperthermophile Thermotoga maritima as a scaffold. Combining various rational criteria with contingency, we targeted 26 selected codons of the thisF gene sequence for randomization at a controlled level. We have developed a novel method of creating full-length gene libraries by combinatorial assembly of smaller sub-libraries. Full-length libraries of high diversity can easily be assembled on demand from smaller and much less diverse sub-libraries, which circumvent the notoriously troublesome long-term archivation and repeated proliferation of high diversity ensembles of phages or plasmids. We developed a generally applicable software tool for sequence analysis of mutated gene sequences that provides efficient assistance for analysis of library diversity. Finally, practical utility of the library was demonstrated in principle by assessment of the conformational stability of library members and isolating protein variants with HisF activity from it. Our approach integrates a number of features of nucleic acids synthetic chemistry, biochemistry and molecular genetics to a coherent, flexible and robust method of combinatorial gene synthesis. PMID:26355961

  18. Chalcone Synthase (CHS) Gene Suppression in Flax Leads to Changes in Wall Synthesis and Sensing Genes, Cell Wall Chemistry and Stem Morphology Parameters.

    Science.gov (United States)

    Zuk, Magdalena; Działo, Magdalena; Richter, Dorota; Dymińska, Lucyna; Matuła, Jan; Kotecki, Andrzej; Hanuza, Jerzy; Szopa, Jan

    2016-01-01

    The chalcone synthase (CHS) gene controls the first step in the flavonoid biosynthesis. In flax, CHS down-regulation resulted in tannin accumulation and reduction in lignin synthesis, but plant growth was not affected. This suggests that lignin content and thus cell wall characteristics might be modulated through CHS activity. This study investigated the possibility that CHS affects cell wall sensing as well as polymer content and arrangement. CHS-suppressed and thus lignin-reduced plants showed significant changes in expression of genes involved in both synthesis of components and cell wall sensing. This was accompanied by increased levels of cellulose and hemicellulose. CHS-reduced flax also showed significant changes in morphology and arrangement of the cell wall. The stem tissue layers were enlarged averagely twofold compared to the control, and the number of fiber cells more than doubled. The stem morphology changes were accompanied by reduction of the crystallinity index of the cell wall. CHS silencing induces a signal transduction cascade that leads to modification of plant metabolism in a wide range and thus cell wall structure. PMID:27446124

  19. Chalcone Synthase (CHS) Gene Suppression in Flax Leads to Changes in Wall Synthesis and Sensing Genes, Cell Wall Chemistry and Stem Morphology Parameters

    Science.gov (United States)

    Zuk, Magdalena; Działo, Magdalena; Richter, Dorota; Dymińska, Lucyna; Matuła, Jan; Kotecki, Andrzej; Hanuza, Jerzy; Szopa, Jan

    2016-01-01

    The chalcone synthase (CHS) gene controls the first step in the flavonoid biosynthesis. In flax, CHS down-regulation resulted in tannin accumulation and reduction in lignin synthesis, but plant growth was not affected. This suggests that lignin content and thus cell wall characteristics might be modulated through CHS activity. This study investigated the possibility that CHS affects cell wall sensing as well as polymer content and arrangement. CHS-suppressed and thus lignin-reduced plants showed significant changes in expression of genes involved in both synthesis of components and cell wall sensing. This was accompanied by increased levels of cellulose and hemicellulose. CHS-reduced flax also showed significant changes in morphology and arrangement of the cell wall. The stem tissue layers were enlarged averagely twofold compared to the control, and the number of fiber cells more than doubled. The stem morphology changes were accompanied by reduction of the crystallinity index of the cell wall. CHS silencing induces a signal transduction cascade that leads to modification of plant metabolism in a wide range and thus cell wall structure. PMID:27446124

  20. Short- and long-term influences of calcitonin gene-related peptide on the synthesis of acetylcholinesterase in mammalian myotubes

    OpenAIRE

    da Costa, Valter Luiz; Lapa, Antonio José; Godinho, Rosely O.

    2001-01-01

    The present study analyses the short- (15 min – 2 h) and long-term (24 – 48 h) influences of calcitonin gene-related peptide (CGRP) on acetylcholinesterase (AChE) expression in the rat cultured skeletal muscle and the signal transduction events underlying CGRP actions.To assess the effect of CGRP on AChE synthesis, myotubes were pre-exposed to the irreversible AChE inhibitor diisopropyl fluorophosphate (DFP) and treated with CGRP or forskolin, an adenylyl cyclase (AC) activator. Treatment of ...

  1. Increased Production of Fatty Acids and Triglycerides in Aspergillus oryzae by Enhancing Expressions of Fatty Acid Synthesis-Related Genes

    Energy Technology Data Exchange (ETDEWEB)

    Tamano, Koichi; Bruno, Kenneth S.; Karagiosis, Sue A.; Culley, David E.; Deng, Shuang; Collett, James R.; Umemura, Myco; Koike, Hideaki; Baker, Scott E.; Machida, Masa

    2013-01-01

    Microbial production of fats and oils is being developedas a means of converting biomass to biofuels. Here we investigate enhancing expression of enzymes involved in the production of fatty acids and triglycerides as a means to increase production of these compounds in Aspergillusoryzae. Examination of the A.oryzaegenome demonstrates that it contains twofatty acid synthases and several other genes that are predicted to be part of this biosynthetic pathway. We enhancedthe expressionof fatty acid synthesis-related genes by replacing their promoters with thepromoter fromthe constitutively highly expressedgene tef1. We demonstrate that by simply increasing the expression of the fatty acid synthasegenes we successfullyincreasedtheproduction of fatty acids and triglyceridesby more than two fold. Enhancement of expression of the fatty acid pathway genes ATP-citrate lyase and palmitoyl-ACP thioesteraseincreasedproductivity to a lesser extent.Increasing expression ofacetyl-CoA carboxylase caused no detectable change in fatty acid levels. Increases in message level for each gene were monitored usingquantitative real-time RT-PCR. Our data demonstrates that a simple increase in the abundance of fatty acid synthase genes can increase the detectable amount of fatty acids.

  2. The expression of Arabidopsis glutamate dehydrogenase gene gdh2 is induced under the influence of tetrapyrrole synthesis inhibitor norflurazon

    Directory of Open Access Journals (Sweden)

    E.Yu. Garnik

    2013-11-01

    Full Text Available The gdh2 gene encoding beta-subunit of glutamate dehydrogenase in Arabidopsis belongs to diurnal-regulated genes. Its expression is highly increased in the dark and reduced to minimal rates at the day light. Some sugar-responsive regulatory pathways are known to be involved in the gdh2 light repression, but the specific mechanisms of this regulation are unknown. In our experiments expression of gdh2 gene increased 6-11 fold in Arabidopsis seedlings grown in presence of the tetrapyrrole synthesis inhibitor norflurazon. The increasing rate depended on the light intensity and did not correlate with the induction of ROS marker genes. This observation can be explained by both a low glucose level in the cells treated with norflurazon and absence of repression by the chloroplast-to-nucleus retrograde pathways because of chloroplast dysfunction. We assume that the diurnal regulation of gdh2 gene expression involves not only sugar-dependent, but also chloroplast-to-nucleus regulatory signals.

  3. On the biphoton excitation of the fluorescence of the bacteriochlorophyll molecules of purple photosynthetic bacteria by powerful near IR femto-picosecond pulses

    International Nuclear Information System (INIS)

    The authors of a number of experimental works detected nonresonance biphoton excitation of bacteriochlorophyll molecules, which represent the main pigment in the light-absorbing natural “antenna” complexes of photosynthesizing purple bacteria, by femtosecond IR pulses (1250–1500 nm). They believe that IR quanta excite hypothetic forbidden levels of the pigments of these bacteria in the double frequency range 625–750 nm. We propose and ground an alternative triplet mechanism to describe this phenomenon. According to our hypothesis, the mechanism of biphoton excitation of molecules by IR quanta can manifest itself specifically, through high triplet levels of molecules in the high fields induced by femtosecond-picosecond laser pulses.

  4. The nature of coherences in the B820 bacteriochlorophyll dimer revealed by two-dimensional electronic spectroscopy.

    Science.gov (United States)

    Ferretti, Marco; Novoderezhkin, Vladimir I; Romero, Elisabet; Augulis, Ramunas; Pandit, Anjali; Zigmantas, Donatas; van Grondelle, Rienk

    2014-06-01

    Light-harvesting in photosynthesis is determined by the excitonic interactions in disordered antennae and the coupling of collective electronic excitations to fast nuclear motions, producing efficient energy transfer with a complicated interplay between exciton and vibrational coherences. Two-dimensional electronic spectroscopy (2DES) is a powerful tool to study the presence of these coherences in photosynthetic complexes. However, the unambiguous assignment of the nature of the observed coherences is still under debate. In this paper we apply 2DES to an excitonically coupled bacteriochlorophyll dimer, the B820 subunit of the light harvesting complex 1 (LH1-RC) of R. rubrum G9. Fourier analysis of the measured kinetics and modeling of the spectral responses in a complete basis of electronic and vibrational states allow us to distinguish between pure vibrational, mixed exciton-vibrational (vibronic), and predominantly exciton coherences. The mixed coherences have been found in a wide range of oscillation frequencies, whereas exciton coherences give the biggest contributions for the frequencies in the 400-550 cm(-1) range, corresponding to the exciton splitting energy of the B820 dimer. Significant exciton coherences are also present at higher frequencies, i.e., up to 800 cm(-1), which are determined by realizations of the disorder with a large energy gap between the two pigments (which increases the apparent value of the exciton splitting). Although the B820 dimer is a model system, the approach presented here represents a basis for further analyses of more complicated systems, providing a tool for studying the interplay between electronic and vibrational coherences in disordered photosynthetic antennae and reaction centres. PMID:24430275

  5. Hexanol-induced order-disorder transitions in lamellar self-assembling aggregates of bacteriochlorophyll c in Chlorobium tepidum chlorosomes.

    Science.gov (United States)

    Arellano, Juan B; Torkkeli, Mika; Tuma, Roman; Laurinmäki, Pasi; Melø, Thor B; Ikonen, Teemu P; Butcher, Sarah J; Serimaa, Ritva E; Psencík, Jakub

    2008-03-01

    Chlorosomes are light-harvesting complexes of green photosynthetic bacteria. Chlorosomes contain bacteriochlorophyll (BChl) c, d, or e aggregates that exhibit strong excitonic coupling. The short-range order, which is responsible for the coupling, has been proposed to be augmented by pigment arrangement into undulated lamellar structures with spacing between 2 and 3 nm. Treatment of chlorosomes with hexanol reversibly converts the aggregated chlorosome chlorophylls into a form with spectral properties very similar to that of the monomer. Although this transition has been extensively studied, the structural basis remains unclear due to variability in the obtained morphologies. Here we investigated hexanol-induced structural changes in the lamellar organization of BChl c in chlorosomes from Chlorobium tepidum by a combination of X-ray scattering, electron cryomicroscopy, and optical spectroscopy. At a low hexanol/pigment ratio, the lamellae persisted in the presence of hexanol while the short-range order and exciton interactions between chlorin rings were effectively eliminated, producing a monomer-like absorption. The result suggested that hexanol hydroxyls solvated the chlorin rings while the aliphatic tail partitioned into the hydrophobic part of the lamellar structure. This partitioning extended the chlorosome along its long axis. Further increase of the hexanol/pigment ratio produced round pigment-hexanol droplets, which lost all lamellar order. After hexanol removal the spectral properties were restored. In the samples treated under the high hexanol/pigment ratio, lamellae reassembled in small domains after hexanol removal while the shape and long-range order were irreversibly lost. Thus, all the interactions required for establishing the short-range order by self-assembly are provided by BChl c molecules alone. However, the long-range order and overall shape are imposed by an external structure, e.g., the proteinaceous chlorosome baseplate. PMID:18197717

  6. Different effects of identical symmetry-related mutations near the bacteriochlorophyll dimer in the photosynthetic reaction center of Rhodobacter sphaeroides.

    Science.gov (United States)

    Vasilieva, L G; Fufina, T Y; Gabdulkhakov, A G; Shuvalov, V A

    2015-06-01

    In the bacterial photosynthetic reaction center (RC), asymmetric protein environment of the bacteriochlorophyll (BChl) dimer largely determines the photophysical and photochemical properties of the primary electron donor. Previously, we noticed significant differences in properties of Rhodobacter sphaeroides RCs with identical mutations in symmetry-related positions - I(M206)H and I(L177)H. The substitution I(L177)H resulted in covalent binding of BChl PA with the L-subunit, as well as in 6-coordination of BChl BB, whereas in RC I(M206)H no such changes of pigment-protein interactions were found. In addition, the yield of RC I(M206)H after its isolation from membranes was significantly lower than the yield of RC I(L177)H. This study shows that replacement of amino acid residues in the M203-M206 positions near BChls PB and BA by symmetry-related residues from the L-subunit near BChls PA and BB leads to further decrease in RC amount in the membranes associated obviously with poor assembly of the complex. Introduction of a new hydrogen bond between BChl PB and its protein environment by means of the F(M197)H mutation stabilized the mutant RC but did not affect its low yield. We suggest that the mutation I(M206)H and substitution of amino acid residues in M203-M205 positions could disturb glycolipid binding on the RC surface near BChl BA that is important for stable assembly of the complex in the membrane. PMID:26531011

  7. Genes involved in the synthesis of the exopolysaccharide methanolan by the obligate methylotroph Methylobacillus sp strain 12S.

    Science.gov (United States)

    Yoshida, Takako; Ayabe, Yuko; Yasunaga, Masaaki; Usami, Yusuke; Habe, Hiroshi; Nojiri, Hideaki; Omori, Toshio

    2003-02-01

    Methylobacillus sp. strain 12S produces an exopolysaccharide (EPS), methanolan, composed of glucose, mannose and galactose. Twenty-four ORFs flanking a Tn5 insertion site in an EPS-deficient mutant were identified, and 21 genes (epsCBAKLDEFGHIJMNOPQRSTU) were predicted to participate in methanolan synthesis on the basis of the features of the primary sequence. Gene disruption analyses revealed that epsABCEFGIJNOP and epsR are required for methanolan synthesis, whereas epsKD and epsH are not essential. EpsFG and EpsE showed homology with Wzc (chain length regulator) and Wza (export protein) of group 1 capsule-producing Escherichia coli, suggesting that methanolan was synthesized via a Wzy-like biosynthesis system. This possibility was supported by the fact that the putative hydropathy profiles of EpsH and EpsM were similar to those of Wzx and Wzy, which are also involved in the flipping of the repeating unit in the cytoplasmic membrane and the polymerization of the capsule in the Wzy-dependent system. EpsBJNOP and EpsR are probably glycosyltransferases involved in the synthesis of the repeating unit onto the lipid carrier. In particular, EpsB appeared to catalyse the initial transfer of the glucose moiety. On the basis of their predicted location in the cells, it is proposed that EpsI and EpsL are involved in methanolan export to the cell surface. E. coli strains expressing EpsQ, EpsS and EpsT showed enhanced activities of GDP-mannose pyrophosphorylase, UDP-galactose 4-epimerase and UDP-glucose pyrophosphorylase, respectively, revealing that they were responsible for the production of the activated compositional sugars of methanolan. EpsU contains a conserved a lytic transglycosylase motif, indicating that it could participate in the degradation of polysaccharides. EpsA and EpsK, which have conserved DNA-binding and cAMP-binding motifs, respectively, were deduced to be transcriptional regulators. In particular, EpsA seems to positively regulate the transcription of

  8. The rnc Gene Promotes Exopolysaccharide Synthesis and Represses the vicRKX Gene Expressions via MicroRNA-Size Small RNAs in Streptococcus mutans

    Science.gov (United States)

    Mao, Meng-Ying; Yang, Ying-Ming; Li, Ke-Zeng; Lei, Lei; Li, Meng; Yang, Yan; Tao, Xiang; Yin, Jia-Xin; Zhang, Ru; Ma, Xin-Rong; Hu, Tao

    2016-01-01

    Dental caries is a biofilm-dependent disease that largely relies on the ability of Streptococcus mutans to synthesize exopolysaccharides. Although the rnc gene is suggested to be involved in virulence mechanisms in many other bacteria, the information regarding it in S. mutans is very limited. Here, using deletion or overexpression mutant assay, we demonstrated that rnc in S. mutans significantly positively regulated exopolysaccharide synthesis and further altered biofilm formation. Meanwhile, the cariogenecity of S. mutans was decreased by deletion of rnc in a specific pathogen-free (SPF) rat model. Interestingly, analyzing the expression at mRNA level, we found the downstream vic locus was repressed by rnc in S. mutans. Using deep sequencing and bioinformatics analysis, for the first time, three putative microRNA-size small RNAs (msRNAs) targeting vicRKX were predicted in S. mutans. The expression levels of these msRNAs were negatively correlated with vicRKX but positively correlated with rnc, indicating rnc probably repressed vicRKX expression through msRNAs at the post-transcriptional level. In all, the results present that rnc has a potential role in the regulation of exopolysaccharide synthesis and can affect vicRKX expressions via post-transcriptional repression in S. mutans. This study provides an alternative avenue for further research aimed at preventing caries. PMID:27242713

  9. Specific genes involved in synthesis and editing of heparan sulfate proteoglycans show altered expression patterns in breast cancer

    International Nuclear Information System (INIS)

    The expression of a specific set of genes controls the different structures of heparan sulfate proteoglycans (HSPGs), which are involved in the growth, invasion and metastatic properties of cancerous cells. The purpose of this study is to increase knowledge of HSPG alterations in breast cancer. Twenty-three infiltrating ductal adenocarcinomas (IDCs), both metastatic and non-metastatic were studied. A transcriptomic approach to the structure of heparan sulfate (HS) chains was used, employing qPCR to analyze both the expression of the enzymes involved in their biosynthesis and editing, as well as the proteoglycan core proteins. Since some of these proteoglycans can also carry chondroitin sulfate chains, we extended the study to include the genes involved in the biosynthesis of these glycosaminoglycans. Histochemical techniques were also used to analyze tissular expression of particular genes showing significant expression differences, of potential interest. No significant change in transcription was detected in approximately 70% of analyzed genes. However, 13 demonstrated changes in both tumor types (40% showing more intense deregulation in the metastatic), while 5 genes showed changes only in non-metastatic tumors. Changes were related to 3 core proteins: overexpression of syndecan-1 and underexpression of glypican-3 and perlecan. HS synthesis was affected by lower levels of some 3-O-sulfotransferase transcripts, the expression of NDST4 and, only in non metastatic tumors, higher levels of extracellular sulfatases. Furthermore, the expression of chondroitin sulfate also was considerably affected, involving both the synthesis of the saccharidic chains and sulfations at all locations. However, the pro-metastatic enzyme heparanase did not exhibit significant changes in mRNA expression, although in metastatic tumors it appeared related to increased levels of the most stable form of mRNA. Finally, the expression of heparanase 2, which displays anti-metastatic features

  10. Response of Fatty Acid Synthesis Genes to the Binding of Human Salivary Amylase by Streptococcus gordonii

    OpenAIRE

    Nikitkova, Anna E.; Haase, Elaine M.; Vickerman, M Margaret; Gill, Steven R.; Scannapieco, Frank A.

    2012-01-01

    Streptococcus gordonii, an important primary colonizer of dental plaque biofilm, specifically binds to salivary amylase via the surface-associated amylase-binding protein A (AbpA). We hypothesized that a function of amylase binding to S. gordonii may be to modulate the expression of chromosomal genes, which could influence bacterial survival and persistence in the oral cavity. Gene expression profiling by microarray analysis was performed to detect genes in S. gordonii strain CH1 that were di...

  11. The Microphthalmia Transcription Factor (Mitf) Controls Expression of the Ocular Albinism Type 1 Gene: Link between Melanin Synthesis and Melanosome Biogenesis

    OpenAIRE

    Vetrini, Francesco; Auricchio, Alberto; Du, Jinyan; Angeletti, Barbara; Fisher, David E.; Ballabio, Andrea; Marigo, Valeria

    2004-01-01

    Melanogenesis is the process that regulates skin and eye pigmentation. Albinism, a genetic disease causing pigmentation defects and visual disorders, is caused by mutations in genes controlling either melanin synthesis or melanosome biogenesis. Here we show that a common transcriptional control regulates both of these processes. We performed an analysis of the regulatory region of Oa1, the murine homolog of the gene that is mutated in the X-linked form of ocular albinism, as Oa1's function af...

  12. Down-regulation of SMT3A gene expression in association with DNA synthesis induction after X-ray irradiation in nevoid basal cell carcinoma syndrome (NBCCS) cells

    International Nuclear Information System (INIS)

    Fibroblast cells derived from nevoid basal carcinoma syndrome (NBCCS) patients show increased levels of DNA synthesis after X-ray irradiation. Genes, whose expression is modulated in association with the DNA synthesis induction, were searched by using PCR-based mRNA differential display analysis in one of the NBCCS cell lines, NBCCS1 cells. Decreased levels of SMT3A gene expression were found in X-ray-irradiated NBCCS1 cells. This decrease was also shown by RT-PCR analysis in another cell line, NBCCS3 cells. In addition to NBCCS cells, normal fibroblast cells showed the DNA synthesis induction after X-ray irradiation when they were treated with antisense oligonucleotides (AO) for SMT3A. However, treatment of normal fibroblasts with the random oligonucleotides (RO) resulted in decreased levels of DNA synthesis after X-ray irradiation. Thus, down-regulation of SMT3A gene expression may be involved in the DNA synthesis induction after X-ray irradiation in the NBCCS cells at least tested

  13. Down-regulation of SMT3A gene expression in association with DNA synthesis induction after X-ray irradiation in nevoid basal cell carcinoma syndrome (NBCCS) cells

    Energy Technology Data Exchange (ETDEWEB)

    Sugaya, Shigeru [Department of Environmental Biochemistry, Graduate School of Medicine, Chiba University, 1-8-1 Inohana, Chuo-ku, Chiba 260-8670 (Japan); Nakanishi, Hiroshi [Department of Clinical Molecular Biology, Graduate School of Medicine, Chiba University, 1-8-1 Inohana, Chuo-ku, Chiba 260-8670 (Japan); Tanzawa, Hideki [Department of Clinical Molecular Biology, Graduate School of Medicine, Chiba University, 1-8-1 Inohana, Chuo-ku, Chiba 260-8670 (Japan); Sugita, Katsuo [Department of Clinical Medicine, Faculty of Education, Chiba University, 1-33 Yayoi, Inage-ku, Chiba 263-8522 (Japan); Kita, Kazuko [Department of Environmental Biochemistry, Graduate School of Medicine, Chiba University, 1-8-1 Inohana, Chuo-ku, Chiba 260-8670 (Japan); Suzuki, Nobuo [Department of Environmental Biochemistry, Graduate School of Medicine, Chiba University, 1-8-1 Inohana, Chuo-ku, Chiba 260-8670 (Japan)]. E-mail: nobuo@faculty.chiba-u.jp

    2005-10-15

    Fibroblast cells derived from nevoid basal carcinoma syndrome (NBCCS) patients show increased levels of DNA synthesis after X-ray irradiation. Genes, whose expression is modulated in association with the DNA synthesis induction, were searched by using PCR-based mRNA differential display analysis in one of the NBCCS cell lines, NBCCS1 cells. Decreased levels of SMT3A gene expression were found in X-ray-irradiated NBCCS1 cells. This decrease was also shown by RT-PCR analysis in another cell line, NBCCS3 cells. In addition to NBCCS cells, normal fibroblast cells showed the DNA synthesis induction after X-ray irradiation when they were treated with antisense oligonucleotides (AO) for SMT3A. However, treatment of normal fibroblasts with the random oligonucleotides (RO) resulted in decreased levels of DNA synthesis after X-ray irradiation. Thus, down-regulation of SMT3A gene expression may be involved in the DNA synthesis induction after X-ray irradiation in the NBCCS cells at least tested.

  14. Early gene expression in bacteriophage T7. I. In vivo synthesis, inactivation, and translational utilization of early mRNA's.

    Science.gov (United States)

    Hercules, K; Jovanovich, S; Sauerbrier, W

    1976-02-01

    In vivo decay rates for the individual T7 early mRNA species were determined. The physical half-lives, measured at 37 C, range from 1.1 min for gene 0.7 RNA to 4.5 min for gene 0.3 RNA. Physical half-lives, as observed after rifampin inhibition of RNA synthesis and polyacylamide electrophoresis of RNAs, are approximately 30% longer than functional half-lives, as observed by 14C-labeled amino acid uptake into individual T7 early proteins. The different RNA species are synthesized at grossly different rates, 0.3 RNA at four times the rate of 1.0 RNA, 0.7 RNA at twice the rate, and 1.1 and 1.3 RNAs at about the same or a slightly lower rate than 1.0 RNA. Rho-factor-mediated termination of transcription behind genes 0.3, 0.7, and perhaps behind 1.0 is inferred from these data. The in vivo translational utilization of the individual T7 early-message species was found to vary by not more than a factor of 2. PMID:1255850

  15. Effects of partial deletion of the wzm and wzt genes on lipopolysaccharide synthesis and virulence of Brucella abortus S19.

    Science.gov (United States)

    Wang, Xiuran; Wang, Lin; Lu, Tiancheng; Yang, Yanling; Chen, Si; Zhang, Rui; Lang, Xulong; Yan, Guangmou; Qian, Jing; Wang, Xiaoxu; Meng, Lingyi; Wang, Xinglong

    2014-06-01

    Brucellosis is a worldwide human and animal infectious disease, and the effective methods of its control are immunisation of animals by vaccination and elimination. Brucella abortus S19 is one of the popular vaccines with virulence in the control of cattle Brucellosis. In the present study, allelic exchange plasmids of wzm and wzt genes and partial knockout mutants of wzm and wzt were constructed to evaluate the resulting difference in virulence of B. abortus S19. PCR analysis revealed that the target genes were knocked out. The mutants were rough mutants and they could be differentiated from natural infection by the Rose Bengal plate and standard agglutination tests. The molecular weights of lipopolysaccharides of the Δwzm and Δwzt mutants were clustered between 25 and 40 kDa, and 30 and 35 kDa separately, and were markedly different from those in B. abortus S19. The virulence of B. abortus Δwzm and Δwzt was decreased compared with that of B. abortus S19 in mice. All these results identified that there were several differences between the wzm and wzt genes on lipopolysaccharide synthesis and on the virulence of B. abortus. PMID:24718931

  16. Expression of auxin synthesis gene tms1 under control of tuber-specific promoter enhances potato tuberization in vitro

    Institute of Scientific and Technical Information of China (English)

    Oksana O Kolachevskaya; Valeriya V Alekseeva; Lidiya I Sergeeva; Elena B Rukavtsova; Irina A Getman; Dick Vreugdenhil; Yaroslav I Buryanov; Georgy A Romanov

    2015-01-01

    Phytohormones, auxins in particular, play an important role in plant development and productivity. Earlier data showed positive impact of exogenous auxin on potato (Solanum tuberosum L.) tuberization. The aim of this study was to generate potato plants with increased auxin level predominantly in tubers. To this end, a pBinB33-tms1 vector was constructed harboring the Agrobacterium auxin biosynthesis gene tms1 fused to tuber-specific promoter of the class I patatin gene (B33-promoter) of potato. Among numerous independently generated B33:tms1 lines, those without visible differences from control were selected for detailed studies. In the majority of transgenic lines, tms1 gene transcription was detected, mostly in tubers rather than in shoots. Indoleacetic acid (IAA) content in tubers and the auxin tuber-to-shoot ratio were increased in tms1-expressing transformants. The organ-specific increase in auxin synthesis in B33:tms1-transformants accelerated and intensified the process of tuber formation, reduced the dose of carbohydrate supply required for in vitro tuber-ization, and decreased the photoperiodic dependence of tuber initiation. Overall, a positive correlation was observed between tms1 expression, IAA content in tubers, and stimulation of tuber formation. The revealed proper-ties of B33:tms1 transformants imply an important role for auxin in potato tuberization and offer prospects to magnify potato productivity by a moderate organ-specific enhance-ment of auxin content.

  17. Synthesis and skin gene analysis of 4'-acetoxy-resveratrol (4AR), therapeutic potential for dermal applications.

    Science.gov (United States)

    Lephart, Edwin D; Acerson, Mark J; Andrus, Merritt B

    2016-07-15

    Resveratrol (RV) 1, a plant polyphenol, has proven effective in commercial products yet drawbacks include low bioavailability due to rapid metabolism. Structural modifications have led to a 4'-acetoxy analog 2 (4AR) now produced using a selective one-step esterification reaction. The one-step synthesis is shown together with expression of skin genes using human dermal models to establish 4AR 2 benefits to skin health. 4AR 2 at 1% in qPCR experiments using a human skin model significantly increased gene expression of the anti-aging factor, SIRT 1 by over 3.3-fold, extracellular matrix proteins collagen III, IV, elastin and tissue inhibitors of metalloproteinases (TIMP 1, 2), anti-oxidants CAT, LOX, superoxide dismutase (SOD 1, 2), metallothioneins (MT1H, MT1H), skin aging biomarkers fibrillin (FBN1), laminin (LAMB1), proliferating cell nuclear antigen (PCNA), skin growth factors (HBEGF, IGF1, NGF and TGF). 4AR 2 also decreased gene expression of inflammatory and skin-aging molecules (IL-1, IL-6, IL-8, COX-2, TNGRSF) and S100 calcium binding proteins A8, A9. These findings suggest that 4AR 2 has potential for topically treatment and prevention of skin aging. PMID:27265258

  18. Cloning and Characterization of the Phytoene Desaturase(pds) Gene-a Key Enzyme for Carotenoids Synthesis in Dunaliella (Chlorophyta)

    Institute of Scientific and Technical Information of China (English)

    SUN Guohua; SUI Zhenghong; ZHANG Xuecheng

    2008-01-01

    The unicellular green alga Dunaliella is outstanding for its ability of massive accumulation of carotenoids. To elucidate the carotenoids synthesis pathway in this alga, phytoene desaturase (pals) gene cDNA together with its DNA sequences were isolated and their structures and functions analyzed. The full-length pds cDNA of 2290 bp (GenBank Accession No. DQ243892) was de- duced from RACE results, including untranslated 21 bp 5'- and 520 bp 3'- flanking regions and an open reading frame of 582 amino acids, coding a protein of 64.196 kDa. The DNA sequence of 2908 bp (GenBank Accession No. DQ845248) including five introns was obtained. The fifth intron was uncompleted and complex, including two bases' perfect repeats (GT)10 and large different-sized repeats within the last 400 bp. The Southern blot hybridization result demonstrated that this gene occurred as a single copy in this species, and the quantitative RT-PCR result showed that the transcription of this gene was constitutive. The evolutional significance ofpds was discussed.

  19. Biosynthesis of unnatural bacteriochlorophyll c derivatives esterified with α,ω-diols in the green sulfur photosynthetic bacterium Chlorobaculum tepidum.

    Science.gov (United States)

    Nishimori, Risato; Mizoguchi, Tadashi; Tamiaki, Hitoshi; Kashimura, Shigenori; Saga, Yoshitaka

    2011-09-13

    Unnatural bacteriochlorophyll (BChl) c derivatives possessing a hydroxy group at the terminus of a hydrocarbon chain at the 17-propionate were biosynthesized in the green sulfur photosynthetic bacterium Chlorobaculum tepidum. Addition of exogenous 1,8-octanediol, 1,12-dodecanediol, and 1,16-hexadecanediol in acetone to liquid cultures resulted in accumulation of BChl c monoesterified with the corresponding diols. The relative ratios of the novel BChl c derivatives esterified with 1,8-, 1,12-, and 1,16-diols to totally producing BChl c were 8.2, 50.2, and 57.6% in the cells grown with additive α,ω-diols at concentrations of 1.5, 0.06, and 0.06 mM, respectively, at the final concentration. The homologue composition of BChl c derivatives esterified with these α,ω-diols was similar to that of original, coexisting BChl c esterified with farnesol (BChl c(F)), suggesting that esterification of α,ω-diols occurred at the last step of the BChl c biosynthetic pathway by BChl c synthase, BchK, in the same manner as in BChl c(F). Chlorosomes, which were isolated from cells grown in the presence of exogenous α,ω-diols, contained a ratio and a composition of BChl c derivatives esterified with the diols similar to those in the whole cells, indicating that these BChl c derivatives were actually present in chlorosomes. Q(y) absorption bands of C. tepidum cells containing the novel BChl c derivatives were shifted to a shorter wavelength, although their bandwidths were analogous to those of cells obtained by normal cultivation. Circular dichroism spectra of cells that had BChl c derivatives esterified with α,ω-diols exhibited S-shaped signals in the Q(y) region, whose polarities were the reverse of those of cells grown in the normal medium and by supplementation with neat acetone as a control experiment. These spectral features of C. tepidum possessing BChl c derivatives esterified with α,ω-diols imply that the novel BChl c derivatives possessing a hydroxy group at the

  20. Cinnamic acid 4-hydroxylase of sorghum [Sorghum biocolor (L.) Moench] gene SbC4H1 restricts lignin synthesis in Arabidopsis

    Science.gov (United States)

    Cinnamic acid 4-hydroxylase (C4H) is the first hydroxylase enzyme of the phenylpropanoid pathway, and its content and activity affects the lignin synthesis. In this study, we isolated a C4H gene SbC4H1 from the suppression subtractive hybridization library of brown midrib (bmr) mutants of Sorghum b...

  1. Emotionally controlled decision-making and a gene variant related to serotonin synthesis in women with borderline personality disorder.

    Science.gov (United States)

    Maurex, Liselotte; Zaboli, Ghazal; Wiens, Stefan; Asberg, Marie; Leopardi, Rosario; Ohman, Arne

    2009-02-01

    The Iowa Gambling Task (IGT) was used to examine (i) social decision-making in women with borderline personality disorder (BPD), and (ii) the relationship between impaired decision-making and the tryptophan hydroxylase-1 (TPH-1) gene, involved in serotonin synthesis. Forty-two women with BPD and a history of suicide attempts were genotyped, and the frequency of a TPH-1 haplotype previously uniquely associated with BPD was calculated. The BPD group scored significantly lower than a control group in the IGT. Furthermore, the TPH-1 haplotype displayed a significantly higher frequency in BPD participants with impaired decision making, compared to BPD participants with normal scores. These findings suggest that impaired decision-making as determined by the IGT is a feature of BPD and may be (i) associated with serotonin dysfunction, and (ii) possibly relevant for suicidal behavior. PMID:18826425

  2. Collagen synthesis promoting pullulan-PEI-ascorbic acid conjugate as an efficient anti-cancer gene delivery vector.

    Science.gov (United States)

    Ambattu, Lizebona August; Rekha, M R

    2015-08-01

    Cationized pullulan (pullulan-PEI; PP) was synthesized and further modified with an anti-oxidant molecule, ascorbic acid (PPAA) at various ratios. The nanoplexes formed at an optimum ratio of 4:1 was within a size of 150nm and had a zeta potential of 9-14mV. The nanoplexes at this ratio was used for further investigations. The cell internalization and transfection efficiency of these nanoplexes were determined in presence of serum. The internalization and transfection efficiency were found to be unaffected by the presence of fetal bovine serum. Another interesting observation was that this polymer was found to have collagen synthesis promoting property. The collagen synthesis effect of these polymers was quantified and observed that PPAA3 promoted the highest. Transfection efficiency was evaluated by assessing the p53 gene expression in C6 rat glioma cells and cell death was quantified to be 96% by flow cytometry, thus establishing the high efficacy of this polymer. PMID:25933522

  3. A Method for Multiplex Gene Synthesis Employing Error Correction Based on Expression

    OpenAIRE

    Timothy H-C Hsiau; David Sukovich; Phillip Elms; Prince, Robin N.; Tobias Strittmatter; Paul Ruan; Bo Curry; Paige Anderson; Jeff Sampson; J Christopher Anderson

    2015-01-01

    Our ability to engineer organisms with new biosynthetic pathways and genetic circuits is limited by the availability of protein characterization data and the cost of synthetic DNA. With new tools for reading and writing DNA, there are opportunities for scalable assays that more efficiently and cost effectively mine for biochemical protein characteristics. To that end, we have developed the Multiplex Library Synthesis and Expression Correction (MuLSEC) method for rapid assembly, error correcti...

  4. Automating gene library synthesis by structure-based combinatorial protein engineering: examples from plant sesquiterpene synthases.

    Science.gov (United States)

    Dokarry, Melissa; Laurendon, Caroline; O'Maille, Paul E

    2012-01-01

    Structure-based combinatorial protein engineering (SCOPE) is a homology-independent recombination method to create multiple crossover gene libraries by assembling defined combinations of structural elements ranging from single mutations to domains of protein structure. SCOPE was originally inspired by DNA shuffling, which mimics recombination during meiosis, where mutations from parental genes are "shuffled" to create novel combinations in the resulting progeny. DNA shuffling utilizes sequence identity between parental genes to mediate template-switching events (the annealing and extension of one parental gene fragment on another) in PCR reassembly reactions to generate crossovers and hence recombination between parental genes. In light of the conservation of protein structure and degeneracy of sequence, SCOPE was developed to enable the "shuffling" of distantly related genes with no requirement for sequence identity. The central principle involves the use of oligonucleotides to encode for crossover regions to choreograph template-switching events during PCR assembly of gene fragments to create chimeric genes. This approach was initially developed to create libraries of hybrid DNA polymerases from distantly related parents, and later developed to create a combinatorial mutant library of sesquiterpene synthases to explore the catalytic landscapes underlying the functional divergence of related enzymes. This chapter presents a simplified protocol of SCOPE that can be integrated with different mutagenesis techniques and is suitable for automation by liquid-handling robots. Two examples are presented to illustrate the application of SCOPE to create gene libraries using plant sesquiterpene synthases as the model system. In the first example, we outline how to create an active-site library as a series of complex mixtures of diverse mutants. In the second example, we outline how to create a focused library as an array of individual clones to distil minimal combinations of

  5. Cloning, sequencing, and expression of Bacillus subtilis genes involved in ATP-dependent nuclease synthesis.

    OpenAIRE

    Kooistra, J; Venema, G

    1991-01-01

    The genes encoding the subunits of the Bacillus subtilis ATP-dependent nuclease (add genes) have been cloned. The genes were located on an 8.8-kb SalI-SmaI chromosomal DNA fragment. Transformants of a recBCD deletion mutant of Escherichia coli with plasmid pGV1 carrying this DNA fragment showed ATP-dependent nuclease activity. Three open reading frames were identified on the 8.8-kb SalI-SmaI fragment, which could encode three proteins with molecular masses of 135 (AddB protein), 141 (AddA pro...

  6. Lipoprotein Lipase, Tissue Expression and Effects on Genes Related to Fatty Acid Synthesis in Goat Mammary Epithelial Cells

    Directory of Open Access Journals (Sweden)

    Wang-Sheng Zhao

    2014-12-01

    Full Text Available Lipoprotein lipase (LPL serves as a central factor in hydrolysis of triacylglycerol and uptake of free fatty acids from the plasma. However, there are limited data concerning the action of LPL on the regulation of milk fat synthesis in goat mammary gland. In this investigation, we describe the cloning and sequencing of the LPL gene from Xinong Saanen dairy goat mammary gland, along with a study of its phylogenetic relationships. Sequence analysis showed that goat LPL shares similarities with other species including sheep, bovine, human and mouse. LPL mRNA expression in various tissues determined by RT-qPCR revealed the highest expression in white adipose tissue, with lower expression in heart, lung, spleen, rumen, small intestine, mammary gland, and kidney. Expression was almost undetectable in liver and muscle. The expression profiles of LPL gene in mammary gland at early, peak, mid, late lactation, and the dry period were also measured. Compared with the dry period, LPL mRNA expression was markedly greater at early lactation. However, compared with early lactation, the expression was lower at peak lactation and mid lactation. Despite those differences, LPL mRNA expression was still greater at peak, mid, and late lactation compared with the dry period. Using goat mammary epithelial cells (GMEC, the in vitro knockdown of LPL via shRNA or with Orlistat resulted in a similar degree of down-regulation of LPL (respectively. Furthermore, knockdown of LPL was associated with reduced mRNA expression of SREBF1, FASN, LIPE and PPARG but greater expression of FFAR3. There was no effect on ACACA expression. Orlistat decreased expression of LIPE, FASN, ACACA, and PPARG, and increased FFAR3 and SREBF1 expression. The pattern of LPL expression was similar to the changes in milk fat percentage in lactating goats. Taken together, results suggest that LPL may play a crucial role in fatty acid synthesis.

  7. The role of the erythroid-specific delta-aminolevulinate synthase gene expression in erythroid heme synthesis.

    Science.gov (United States)

    Meguro, K; Igarashi, K; Yamamoto, M; Fujita, H; Sassa, S

    1995-08-01

    Using antisense technology, the effects of suppressed gene expression of the erythroid-specific delta-aminolevulinate (ALA) synthase (ALAS-E) on heme synthesis, expression of mRNAs encoding an erythroid-specific transcription factor NF-E2, other heme pathway enzymes, and beta-globin were examined in murine erythroleukemia (MEL) cells. In MEL cells in which an antisense ALAS-E RNA was expressed (AS clone), sense ALAS-E mRNA levels in both untreated and dimethylsulfoxide (DMSO)-treated cells were decreased compared with their respective controls. Heme synthesis in AS clones was decreased in proportion to the suppressed levels of ALAS-E mRNA. In addition, mRNAs for ALA dehydratase, porphobilinogen deaminase, ferrochelatase (FeC), and beta-globin were also decreased in AS clones. There was a strong correlation between the level of ALAS-E mRNA and most of the mRNAs of the heme pathway enzymes and beta-globin. There was a decrease in the mRNA level of p45, but not of mafK, which are the large and the small subunits of NF-E2, respectively, in AS clones. Treatment of AS cells with hemin and ALA in the presence of DMSO partially restored the suppressed mRNA levels for beta-globin and FeC and heme content, respectively. These findings thus indicate that heme formation, which is determined by the level of ALAS-E, plays an essential role on gene expression of many proteins necessary for erythroid development. PMID:7620186

  8. Microarray-based analysis of the differential expression of melanin synthesis genes in dark and light-muzzle Korean cattle.

    Directory of Open Access Journals (Sweden)

    Sang Hwan Kim

    Full Text Available The coat color of mammals is determined by the melanogenesis pathway, which is responsible for maintaining the balance between black-brown eumelanin and yellow-reddish pheomelanin. It is also believed that the color of the bovine muzzle is regulated in a similar manner; however, the molecular mechanism underlying pigment deposition in the dark-muzzle has yet to be elucidated. The aim of the present study was to identify melanogenesis-associated genes that are differentially expressed in the dark vs. light muzzle of native Korean cows. Using microarray clustering and real-time polymerase chain reaction techniques, we observed that the expression of genes involved in the mitogen-activated protein kinase (MAPK and Wnt signaling pathways is distinctively regulated in the dark and light muzzle tissues. Differential expression of tyrosinase was also noticed, although the difference was not as distinct as those of MAPK and Wnt. We hypothesize that emphasis on the MAPK pathway in the dark-muzzle induces eumelanin synthesis through the activation of cAMP response element-binding protein and tyrosinase, while activation of Wnt signaling counteracts this process and raises the amount of pheomelanin in the light-muzzle. We also found 2 novel genes (GenBank No. NM-001076026 and XM-588439 with increase expression in the black nose, which may provide additional information about the mechanism of nose pigmentation. Regarding the increasing interest in the genetic diversity of cattle stocks, genes we identified for differential expression in the dark vs. light muzzle may serve as novel markers for genetic diversity among cows based on the muzzle color phenotype.

  9. Streptococcus pneumoniae arginine synthesis genes promote growth and virulence in pneumococcal meningitis

    NARCIS (Netherlands)

    J.R. Piet; M. Geldhoff; B.D.C. van Schaik; M.C. Brouwer; M. Valls Seron; M.E. Jakobs; K. Schipper; Y. Pannekoek; A.H. Zwinderman; T. van der Poll; A.H.C. van Kampen; F. Baas; A van der Ende; D. van de Beek

    2014-01-01

    Streptococcus pneumoniae (pneumococcus) is a major human pathogen causing pneumonia, sepsis and bacterial meningitis. Using a clinical phenotype based approach with bacterial whole-genome sequencing we identified pneumococcal arginine biosynthesis genes to be associated with outcome in patients with

  10. Genetic variation in genes of the fatty acid synthesis pathway and breast cancer risk

    DEFF Research Database (Denmark)

    Campa, Daniele; McKay, James; Sinilnikova, Olga;

    2009-01-01

    FASN) is related to breast cancer risk and body-mass index (BMI) by studying 1,294 breast cancer cases and 2,452 controls from the European Prospective Investigation on Cancer (EPIC). We resequenced the FAS gene and combined information of SNPs found by resequencing and SNPs from public databases....... Using a tagging approach and selecting 20 SNPs, we covered all the common genetic variation of these genes. In this study we were not able to find any statistically significant association between the SNPs in the FAS, ChREBP and SREPB-1 genes and an increased risk of breast cancer overall and by......Fatty acid synthase (FAS) is the major enzyme of lipogenesis. It catalyzes the NADPH-dependent condensation of acetyl-CoA and malonyl-CoA to produce palmitic acid. Transcription of the FAS gene is controlled synergistically by the transcription factors ChREBP (carbohydrate response element...

  11. TmPrime: fast, flexible oligonucleotide design software for gene synthesis

    OpenAIRE

    Bode, Marcus; Khor, Samuel; Ye, Hongye; Li, Mo-Huang; Ying, Jackie Y.

    2009-01-01

    Herein we present TmPrime, a computer program to design oligonucleotide sets for gene assembly by both ligase chain reaction (LCR) and polymerase chain reaction (PCR). TmPrime offers much flexibility with no constraints on the gene and oligonucleotide lengths. The program divides the long input DNA sequence based on the input desired melting temperature, and dynamically optimizes the length of oligonucleotides to achieve homologous melting temperatures. The output reports the melting temperat...

  12. Lipopolysaccharide derived from the digestive tract activates inflammatory gene expression and inhibits casein synthesis in the mammary glands of lactating dairy cows.

    Science.gov (United States)

    Zhang, Kai; Chang, Guangjun; Xu, Tianle; Xu, Lei; Guo, Junfei; Jin, Di; Shen, Xiangzhen

    2016-03-01

    To meet the nutrition requirements of lactation, dairy cows are usually fed a high concentrate diet (HC). However, high-grain feeding causes subacute ruminal acidosis (SARA), a metabolic disorder that causes milk protein depression. This study aimed to investigate the effect of lipopolysaccharide (LPS) released in the rumen on inflammatory gene expression and casein synthesis in mammary glands of lactating dairy cows fed a HC diet. We found that milk protein was significantly decreased in the HC group after 15 weeks of feeding. Overall, LPS concentrations in the rumen fluid, lacteal artery and vein were increased in the HC group. Transcriptome microarray was used to evaluate alterations in the signaling pathway in mammary glands. Signaling pathways involved in inflammatory responses were activated, whereas those involved in protein synthesis were inhibited in the HC group. mRNA expression involved in inflammatory responses, including that of TLR4, NF-кB and pro-inflammatory genes, was increased in the HC group, while αs1-casein (CSN1S1), β-casein (CSN2), mTOR and S6K gene expression were decreased. Moreover, protein expression was consistent with the corresponding gene expression. After feeding with an HC diet, LPS derived from the rumen increased inflammatory gene expression and inhibited casein synthesis in the mammary glands of lactating dairy cows fed a HC diet. PMID:26893357

  13. Gene regulatory and metabolic adaptation processes of Dinoroseobacter shibae DFL12T during oxygen depletion.

    Science.gov (United States)

    Laass, Sebastian; Kleist, Sarah; Bill, Nelli; Drüppel, Katharina; Kossmehl, Sebastian; Wöhlbrand, Lars; Rabus, Ralf; Klein, Johannes; Rohde, Manfred; Bartsch, Annekathrin; Wittmann, Christoph; Schmidt-Hohagen, Kerstin; Tielen, Petra; Jahn, Dieter; Schomburg, Dietmar

    2014-05-01

    Metabolic flexibility is the key to the ecological success of the marine Roseobacter clade bacteria. We investigated the metabolic adaptation and the underlying changes in gene expression of Dinoroseobacter shibae DFL12(T) to anoxic life by a combination of metabolome, proteome, and transcriptome analyses. Time-resolved studies during continuous oxygen depletion were performed in a chemostat using nitrate as the terminal electron acceptor. Formation of the denitrification machinery was found enhanced on the transcriptional and proteome level, indicating that D. shibae DFL12(T) established nitrate respiration to compensate for the depletion of the electron acceptor oxygen. In parallel, arginine fermentation was induced. During the transition state, growth and ATP concentration were found to be reduced, as reflected by a decrease of A578 values and viable cell counts. In parallel, the central metabolism, including gluconeogenesis, protein biosynthesis, and purine/pyrimidine synthesis was found transiently reduced in agreement with the decreased demand for cellular building blocks. Surprisingly, an accumulation of poly-3-hydroxybutanoate was observed during prolonged incubation under anoxic conditions. One possible explanation is the storage of accumulated metabolites and the regeneration of NADP(+) from NADPH during poly-3-hydroxybutanoate synthesis (NADPH sink). Although D. shibae DFL12(T) was cultivated in the dark, biosynthesis of bacteriochlorophyll was increased, possibly to prepare for additional energy generation via aerobic anoxygenic photophosphorylation. Overall, oxygen depletion led to a metabolic crisis with partly blocked pathways and the accumulation of metabolites. In response, major energy-consuming processes were reduced until the alternative respiratory denitrification machinery was operative. PMID:24648520

  14. Activation of anthocyanin synthesis genes by white light in eggplant hypocotyl tissues, and identification of an inducible P-450 cDNA

    International Nuclear Information System (INIS)

    Eggplant seedlings (Solanum melongena) grown under red light irradiation showed a normal morphology with green, fully expanded cotyledons. When the seedlings grown under red light were irradiated with ultraviolet-containing white light, anthocyanin synthesis was induced in the hypocotyl tissues, especially when a UV light supplement was added. The accumulation of pigments was closely associated with the expression of genes involved in flavonoid synthesis. These genes include chalcone synthase (CHS) and dihydroflavonol 4-reductase (DFR). Using subtracted probes, which had been enriched for the accumulated mRNA, one white light-responsive cDNA was identified as being a P450 gene by comparison with database sequences. The maximal amino acid homology this cDNA had with other P450s was 36%. This was with CYP71 from avocado (Persea americana). Thus it represents a new P-450 family, which has been named CYP75. The mRNA of this gene was localized in the hypocotyl tissues of eggplant seedlings, which had been white light-irradiated. The transcript was accumulated by changing the light source, as in the case of other flavonoid biosynthesis genes. In delphinidin producing petunia plants, the mRNAs corresponding to the eggplant P-450 and flavonoid biosynthesis genes such as CHS and DFR were most abundant during the mid stage of flower bud development, but could not be detected in leaf tissues. These results suggest that this P-450 gene encodes a hydroxylating enzyme involved in flavonoid biosynthesis. (author)

  15. Synthesis of Electroneutralized Amphiphilic Copolymers with Peptide Dendrons for Intramuscular Gene Delivery.

    Science.gov (United States)

    Pu, Linyu; Wang, Jiali; Li, Na; Chai, Qiuxia; Irache, Juan M; Wang, Gang; Tang, James Zhenggui; Gu, Zhongwei

    2016-06-01

    Intramuscular gene delivery materials are of great importance in plasmid-based gene therapy system, but there is limited information so far on how to design and synthesize them. A previous study showed that the peptide dendron-based triblock copolymer with its components arranged in a reversed biomembrane architecture could significantly increase intramuscular gene delivery and expression. Herein, we wonder whether copolymers with biomembrane-mimicking arrangement may have similar function on intramuscular gene delivery. Meanwhile, it is of great significance to uncover the influence of electric charge and molecular structure on the function of the copolymers. To address the issues, amphiphilic triblock copolymers arranged in hydrophilic-hydrophobic-hydrophilic structure were constructed despite the paradoxical characteristics and difficulties in synthesizing such hydrophilic but electroneutral molecules. The as-prepared two copolymers, dendronG2(l-lysine-OH)-poly propylene glycol2k(PPG2k)-dendronG2(l-lysine-OH) (rL2PL2) and dendronG3(l-lysine-OH)-PPG2k-dendronG3(l-lysine-OH) (rL3PL3), were in similar structure but had different hydrophilic components and surface charges, thus leading to different capabilities in gene delivery and expression in skeletal muscle. rL2PL2 was more efficient than Pluronic L64 and rL3PL3 when mediating luciferase, β-galactosidase, and fluorescent protein expressions. Furthermore, rL2PL2-mediated growth-hormone-releasing hormone expression could significantly induce mouse body weight increase in the first 21 days after injection. In addition, both rL2PL2 and rL3PL3 showed good in vivo biosafety in local and systemic administration. Altogether, rL2PL2-mediated gene expression in skeletal muscle exhibited applicable potential for gene therapy. The study revealed that the molecular structure and electric charge were critical factors governing the function of the copolymers for intramuscular gene delivery. It can be concluded that, combined

  16. Expression of cytoskeletal and matrix genes following exposure to ionizing radiation: Dose-rate effects and protein synthesis requirements

    International Nuclear Information System (INIS)

    Experiments were designed to examine the effects of radiation dose-rate and of the protein synthesis inhibitor cycloheximide on expression of cytoskeletal elements (γ- and β-actin and α-tubulin) and matrix elements (fibronectin) in Syrian hamster embryo cells. Past work from our laboratory had already demonstrated optimum time points and doses for examination of radiation effects on accumulation of specific transcripts. Our results here demonstrated little effect of dose-rate for JANUS fission spectrum neutrons when comparing expression of either α-tubulin or fibronectin genes. Past work had already documented similar results for expression of actin transcripts. Effects of cycloheximide, however, revealed several interesting and novel findings: (1) Cycloheximide repressed accumulation of α-tubulin following exposure to high dose-rate neutrons or γ rays; this did not occur following similar low dose-rate exposure (2) Cycloheximide did not affect accumulation of mRNA for actin genes. Cycloheximide abrogated the moderate induction of fibronectin-mRNA which occurred following exposure to γ rays and high dose-rate neutrons. These results suggest a role for labile proteins in the maintenance of α-tubulin and fibronectin mRNA accumulation following exposure to ionizing radiation. In addition, they suggest that the cellular/molecular response to low dose-rate neutrons may be different from the response to high dose-rate neutrons

  17. Expression of cytoskeletal and matrix genes following exposure to ionizing radiation: Dose-rate effects and protein synthesis requirements

    International Nuclear Information System (INIS)

    Experiments were designed to examine the effects Of radiation dose-rate and of the protein synthesis inhibitor cycloheximide on expression of cytoskeletal elements (γ- and β-actin and α-tubulin) and matrix elements (fibronectin) in Syrian hamster embryo cells. Past work from our laboratory had already demonstrated optimum time points and doses for examination of radiation effects on accumulation of specific transcripts. Our results here demonstrated little effect of dose-rate for JANUS fission spectrum neutrons when comparing expression of either α-tubulin or fibronectin genes. Past work had already documented similar results for expression of actin transcripts. Effects of cycloheximide revealed that cycloheximide repressed accumulation of α-tubulin following exposure to high dose-rate neutrons or γ rays; this did not occur following similar low dose-rate exposure. (2) Cycloheximide did not affect accumulation of MRNA for actin genes; and that cycloheximide abrogated the moderate induction of fibronectin-mRNA which occurred following exposure to γ rays and high dose-rate neutrons. These results suggest a role for labile proteins in the maintenance of α-tubulin and fibronectin MRNA accumulation following exposure to ionizing radiation. in addition, they suggest that the cellular/molecular response to low dose-rate neutrons may be different from the response to high dose-rate neutrons

  18. Development of Novel Polymeric Materials for Gene Therapy and pH-Sensitive Drug Delivery: Modeling, Synthesis, Characterization, and Analysis

    Energy Technology Data Exchange (ETDEWEB)

    Brian Curtis Anderson

    2002-08-27

    The underlying theme of this thesis is the use of polymeric materials in bioapplications. Chapters 2-5 either develop a fundamental understanding of current materials used for bioapplications or establish protocols and procedures used in characterizing and synthesizing novel materials. In chapters 6 and 7 these principles and procedures are applied to the development of materials to be used for gene therapy and drug delivery. Chapter one is an introduction to the ideas that will be necessary to understand the subsequent chapters, as well as a literature review of these topics. Chapter two is a paper that has been published in the ''Journal of Controlled Release'' that examines the mechanism of drug release from a polymer gel, as well as experimental design suggestions for the evaluation of water soluble drug delivery systems. Chapter three is a paper that has been published in the ''Journal of Pharmaceutical Sciences'' that discusses the effect ionic salts have on properties of the polymer systems examined in chapter two. Chapter four is a paper published in the Materials Research Society Fall 2000 Symposium Series dealing with the design and synthesis of a pH-sensitive polymeric drug delivery device. Chapter five is a paper that has been published in the journal ''Biomaterials'' proposing a novel polymer/metal composite for use as a biomaterial in hip arthroplasty surgery. Chapter six is a paper that will appear in an upcoming volume of the Journal ''Biomaterials'' dealing with the synthesis of a novel water soluble cationic polymer with possible applications in non-viral gene therapy. Chapter seven is a paper that has been submitted to ''Macromolecules'' discussing several novel block copolymers based on poly(ethylene glycol) and poly(diethylamino ethyl methacrylate) that possess both pH-sensitive and temperature sensitive properties. Chapter eight contains a

  19. H2O2-Activated Up-Regulation of Glutathione in Arabidopsis Involves Induction of Genes Encoding Enzymes Involved in Cysteine Synthesis in the Chloroplast

    Institute of Scientific and Technical Information of China (English)

    Guillaume Queval; Dorothée Thominet; Hélène Vanacker; Myroslawa Miginiac-Maslow; Bertrand Gakière; Graham Noctor

    2009-01-01

    Glutathione is a key player in cellular redox homeostasis and, therefore, in the response to H2O2, but the factors regulating oxidation-activated glutathione synthesis are still unclear. We investigated H2O2-induced glutathione synthesis in a conditional Arabidopsis catalase-deficient mutant (cat2). Plants were grown from seed at elevated CO2 for 5 weeks, then transferred to air in either short-day or long-day conditions. Compared to cat2 at elevated CO2 or wild-type plants in any condition, transfer of cat2 to air in both photoperiods caused measurable oxidation of the leaf glutathione pool within hours. Oxidation continued on subsequent days and was accompanied by accumulation of glutathione. This effect was stronger in cat2 transferred to air in short days, and was not linked to appreciable increases in the extractable activities of or transcripts encoding enzymes involved in the committed pathway of glutathione synthesis. In contrast, it was accompanied by increases in serine, O-acetylserine, and cysteine. These changes in metabolites were accompanied by induction of genes encoding adenosine phosphosulfate reductase (APR), particularly APR3, as well as a specific serine acetyltransferase gene (SAT2.1) encoding a chloroplastic SAT. Marked induction of these genes was only observed in cat2 transferred to air in short-day conditions, where cysteine and glutathione accumulation was most dramatic. Unlike other SAT genes, which showed negligible induction in cat2, the relative abundance of APR and SAT2.1 transcripts was closely correlated with marker transcripts for H2O2 signaling. Together, the data underline the importance of cysteine synthesis in oxidant-induced up-regulation of glutathione synthesis and suggest that the chloroplast makes an important contribution to cysteine production under these circumstances.

  20. The Halloween genes code for cytochrome P450 enzymes mediating synthesis of the insect molting hormone

    DEFF Research Database (Denmark)

    Rewitz, Kim; Rybczynski, Robert; Warren, James T.; Gilbert, Lawrence I.

    2006-01-01

    The developmental events occurring during molting and metamorphosis of insects are controlled by precisely timed changes in levels of ecdysteroids, the molting hormones. The final four sequential hydroxylations of steroid precursors into the active ecdysteroid of insects, 20-hydroxyecdysone (20E......), are mediated by four cytochrome P450 enzymes, encoded by genes in the Halloween family. Orthologs of the Drosophila Halloween genes phantom (phm: CYP306A1), disembodied (dib: CYP302A1), shadow (sad: CYP315A1) and shade (shd: CYP314A1) were obtained from the endocrinological model insect, the tobacco...... hornworm Manduca sexta. Expression of these genes was studied and compared with changes in ecdysteroid titer that controls transition from the larval to pupal stage. Phm, dib and sad (mediating the final hydroxylations in the biosynthesis of ecdysone; E) were selectively expressed in the prothoracic gland...

  1. Synthesis of a new conjugated polymer for DNA alkylation and gene regulation.

    Science.gov (United States)

    Nie, Chenyao; Zhu, Chunlei; Feng, Liheng; Lv, Fengting; Liu, Libing; Wang, Shu

    2013-06-12

    A new polyfluorene derivative containing pendent alkylating chlorambucil (PFP-Cbl) was synthesized and characterized. Under direct incubation with DNA in vitro, PFP-Cbl could undergo an efficient DNA alkylating reaction and induce DNA cross-linking. In vitro transcription and translation experiment exhibited that the PFP-Cbl significantly down-regulated the gene expression of luciferase reporter plasmid. The down-regulation of gene expression was also verified through the transfection experiment of p-EGFP plasmid, which showed decreased green fluorescent protein (GFP) in cells. Meanwhile, the self-luminous property of PFP-Cbl could make it able to trace the internalized PFP-Cbl and plasmid complexes resulted from cross-linking in cells by fluorescent microscopy. Combining the features of alkylating function, multivalent binding sites, and fluorescent characteristics, PFP-Cbl provides a new insight in the area of gene regulation and extends the new applications of conjugated polymers (CPs). PMID:23548104

  2. Self-assembly of natural light-harvesting bacteriochlorophylls of green sulfur photosynthetic bacteria in silicate capsules as stable models of chlorosomes.

    Science.gov (United States)

    Saga, Yoshitaka; Akai, Sho; Miyatake, Tomohiro; Tamiaki, Hitoshi

    2006-01-01

    Naturally occurring bacteriochlorophyll(BChl)s-c, -d, and -e from green sulfur photosynthetic bacteria were self-assembled in an aqueous solution in the presence of octadecyltriethoxysilane and tetraethoxysilane, followed by polycondensation of the alkoxysilanes by incubation for 50 h at 25 degrees C. The resulting BChl self-assemblies in silicate capsules exhibited visible absorption and circular dichroism spectra similar to the corresponding natural light-harvesting systems (chlorosomes) of green sulfur bacteria. Dynamic light scattering measurements indicated that the silicate capsules had an average hydrodynamic diameter of several hundred nanometers. BChl self-aggregates in silicate capsules were significantly stable to a nonionic surfactant Triton X-100, which was apt to decompose the BChl aggregates to their monomeric form, compared with conventional micelle systems. BChls in silicate capsules were more tolerant to demetalation of the central magnesium under acidic conditions than the natural systems. PMID:16848406

  3. Association of bacteriochlorophyll a with the CsmA protein in chlorosomes of the photosynthetic green filamentous bacterium Chloroflexus aurantiacus

    DEFF Research Database (Denmark)

    Sakuragi, Y; Frigaard, N-U; Shimada, K;

    1999-01-01

    . Subsequent proteinase K treatment of these HD-treated chlorosomes caused digestion of CsmA and a simultaneous decrease of the BChl a absorption band. Based on these results, we suggest that CsmA is associated with BChl a in the chlorosomes. This suggestion was supported by the measured stoichiometric ratio......The protein assumed to be associated with bacteriochlorophyll (BChl) a in chlorosomes from the photosynthetic green filamentous bacterium Chloroflexus aurantiacus was investigated by alkaline treatment, proteolytic digestion and a new treatment using 1-hexanol, sodium cholate and Triton X-100. Upon...... alkaline treatment, only the 5.7 kDa CsmA protein was removed from the chlorosomes among six proteins detected by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) analysis, concomitantly with the disappearance of BChl a absorption at 795 nm. Trypsin treatment removed two proteins with...

  4. On the biphoton excitation of the fluorescence of the bacteriochlorophyll molecules of purple photosynthetic bacteria by powerful near IR femto-picosecond pulses

    Energy Technology Data Exchange (ETDEWEB)

    Borisov, A. Yu., E-mail: borissov@belozersky.msu.ru [Moscow State University, Belozersky Institute of Physicochemical Biology (Russian Federation)

    2011-11-15

    The authors of a number of experimental works detected nonresonance biphoton excitation of bacteriochlorophyll molecules, which represent the main pigment in the light-absorbing natural 'antenna' complexes of photosynthesizing purple bacteria, by femtosecond IR pulses (1250-1500 nm). They believe that IR quanta excite hypothetic forbidden levels of the pigments of these bacteria in the double frequency range 625-750 nm. We propose and ground an alternative triplet mechanism to describe this phenomenon. According to our hypothesis, the mechanism of biphoton excitation of molecules by IR quanta can manifest itself specifically, through high triplet levels of molecules in the high fields induced by femtosecond-picosecond laser pulses.

  5. Heme Synthesis by Plastid Ferrochelatase I Regulates Nuclear Gene Expression in Plants

    Science.gov (United States)

    Woodson, Jesse D.; Perez-Ruiz, Juan M.; Chory, Joanne

    2016-01-01

    Summary Chloroplast signals regulate hundreds of nuclear genes during development and in response to stress, but little is known of the signals or signal transduction mechanisms of plastid-to-nucleus (retrograde) signaling [1, 2]. In Arabidopsis thaliana, genetic studies using norflurazon (NF), an inhibitor of carotenoid biosynthesis, have identified five GUN (genomes uncoupled) genes, implicating the tetrapyrrole pathway as a source of a retrograde signal. Loss of function of any of these GUN genes leads to increased expression of photosynthesis-associated nuclear genes (PhANGs) when chloroplast development has been blocked by NF [3, 4]. Here we present a new Arabidopsis gain-of-function mutant, gun6-1D, with a similar phenotype. The gun6-1Dmutant overexpresses the conserved plastid ferrochelatase 1 (FC1, heme synthase). Genetic and biochemical experiments demonstrate that increased flux through the heme branch of the plastid tetrapyrrole biosynthetic pathway increases PhANG expression. The second conserved plant ferrochelatase, FC2, colocalizes with FC1, but FC2 activity is unable to increase PhANG expression in undeveloped plastids. These data suggest a model in which heme, specifically produced by FC1, may be used as a retrograde signal to coordinate PhANG expression with chloroplast development. PMID:21565502

  6. Excitonic polarons in quasi-one-dimensional LH1 and LH2 bacteriochlorophyll a antenna aggregates from photosynthetic bacteria: A wavelength-dependent selective spectroscopy study

    International Nuclear Information System (INIS)

    Spectral characteristics of the optically excited states in the ring-shaped quasi-one-dimensional aggregates comprising 18 and 32 tightly coupled bacteriochlorophyll a molecules have been investigated using selective spectroscopy methods and theoretical modelling of the data. Distinguished by the lowest electronic transition energies in the LH2 and LH1 antenna complexes these aggregates govern the functionally important ultrafast funneling of solar excitation energy in the photosynthetic membranes of purple bacteria. It was found by using a sophisticated differential fluorescence line narrowing method that exciton-phonon coupling in terms of the dimensionless Huang-Rhys factor is strong in these systems, justifying an excitonic polaron theoretical approach for the data analysis. Although we reached this qualitative conclusion already previously, in this work essential dependence of the exciton-phonon coupling strength and reorganization energy on excitation wavelength as well as on excitation light fluence has been established. We then show that these results corroborate with the properties of excitonic polarons in diagonally disordered ensembles of the aggregates. Furthermore, the weighted density of states of the phonon modes, which is an important characteristic of dynamical systems interacting with their surroundings, was derived. Its shape, being similar for all studied circular aggregates, deviates significantly from a reference profile describing local response of a protein to the Qy electronic transition in a single bacteriochlorophyll a molecule. Similarities of the data for regular and B800 deficient mutant LH2 complexes indicate that the B800 pigments have no direct influence on the electronic states of the B850 aggregate system. Consistent set of model parameters was determined, unambiguously implying that excitonic polarons, rather than bare excitons are proper lowest-energy optical excitations in the LH1 and LH2 antenna complexes

  7. The synthesis and expression of a gene for the complement factor C5a

    International Nuclear Information System (INIS)

    A gene coding for the 74 amino acid human complement factor C5a has been chemically synthesized, cloned and expressed in E. coli. The 253 base-pair fragment was constructed from 16 oligodeoxyribonucleotides using the phosphoramidite chemistry. It was assembled in two steps: the assembly of three sub-fragments from the oligonucleotides and the final assembly from the three subfragments. The gene was cloned into a pBR322 derived plasmid containing the lac UV5-D up-promoter mutation. Expression was quanitified by immunoassay and by measuring competition for binding of human 125labelled C5a to the receptors on polymorphonuclear leukocytes. The C5a now being obtained possesses a 40-fold higher specific activity (C5atotal protein) than C5a from complement-activated human serum. This should avoid the problem of contamination of intact C5a with the des-Arg form of the molecule found in serum

  8. Polyketide synthesis genes associated with toxin production in two species of Gambierdiscus (Dinophyceae)

    OpenAIRE

    Kohli, Gurjeet S.; John, Uwe; Figueroa, Rosa I.; Rhodes, Lesley L.; Harwood, D. Tim; Groth, Marco; Bolch, Christopher J. S.; Murray, Shauna A.

    2015-01-01

    Background Marine microbial protists, in particular, dinoflagellates, produce polyketide toxins with ecosystem-wide and human health impacts. Species of Gambierdiscus produce the polyether ladder compounds ciguatoxins and maitotoxins, which can lead to ciguatera fish poisoning, a serious human illness associated with reef fish consumption. Genes associated with the biosynthesis of polyether ladder compounds are yet to be elucidated, however, stable isotope feeding studies of such compounds co...

  9. Polyploid genome of Camelina sativa revealed by isolation of fatty acid synthesis genes

    OpenAIRE

    Shewmaker Christine K; Goldstein Elianna; Schroeder Jesara; Comai Luca; Beilstein Mark; Ditt Renata F; Hutcheon Carolyn; Nguyen Van Thu; De Rocher Jay; Kiser Jack

    2010-01-01

    Abstract Background Camelina sativa, an oilseed crop in the Brassicaceae family, has inspired renewed interest due to its potential for biofuels applications. Little is understood of the nature of the C. sativa genome, however. A study was undertaken to characterize two genes in the fatty acid biosynthesis pathway, fatty acid desaturase (FAD) 2 and fatty acid elongase (FAE) 1, which revealed unexpected complexity in the C. sativa genome. Results In C. sativa, Southern analysis indicates the p...

  10. Sugar-mediated semidian oscillation of gene expression in the cassava storage root regulates starch synthesis

    OpenAIRE

    Baguma, Yona; Sun, Chuanxin; Borén, Mats; Olsson, Helena; Rosenqvist, Sara; Mutisya, Joel; Patrick R. Rubaihayo; Jansson, Christer

    2008-01-01

    Starch branching enzyme (SBE) activity in the cassava storage root exhibited a diurnal fluctuation, dictated by a transcriptional oscillation of the corresponding SBE genes. The peak of SBE activity coincided with the onset of sucrose accumulation in the storage, and we conclude that the oscillatory mechanism keeps the starch synthetic apparatus in the storage root sink in tune with the flux of sucrose from the photosynthetic source. When storage roots were uncoupled from the source, SBE expr...

  11. Aliphatic glucosinolate synthesis and gene expression changes in gamma-irradiated cabbage.

    Science.gov (United States)

    Banerjee, Aparajita; Rai, Archana N; Penna, Suprasanna; Variyar, Prasad S

    2016-10-15

    Glucosinolates, found principally in the plant order Brassicales, are modulated by different post-harvest processing operations. Among these, ionizing radiation, a non-thermal process, has gained considerable interest for ensuring food security and safety. In gamma-irradiated cabbage, enhanced sinigrin, a major glucosinolate, has been reported. However, the molecular basis of such a radiation induced effect is not known. Herein, the effect of radiation processing on the expression of glucosinolate biosynthetic genes was investigated. RT-PCR based expression analysis of seven glucosinolate biosynthetic pathway genes (MYB28, CYP79F1, CYP83A1, SUR1, UGT74B1, SOT18 and TGG1) showed that CYP83A1, MYB28, UGT74B1, CYP79F1 and SUR1 were up-regulated in irradiated cabbage. The content of jasmonates, signalling molecules involved in glucosinolate induction was, however, unaffected in irradiated cabbage suggesting their non-involvement in glucosinolate induction during radiation processing. This is the first report on the effect of gamma irradiation on the expression of glucosinolate biosynthetic genes in vegetables. PMID:27173540

  12. A Potential Benefit of Albinism in Astyanax Cavefish: Downregulation of the oca2 Gene Increases Tyrosine and Catecholamine Levels as an Alternative to Melanin Synthesis

    OpenAIRE

    Helena Bilandžija; Li Ma; Amy Parkhurst; Jeffery, William R

    2013-01-01

    Albinism, the loss of melanin pigmentation, has evolved in a diverse variety of cave animals but the responsible evolutionary mechanisms are unknown. In Astyanax mexicanus, which has a pigmented surface dwelling form (surface fish) and several albino cave-dwelling forms (cavefish), albinism is caused by loss of function mutations in the oca2 gene, which operates during the first step of the melanin synthesis pathway. In addition to albinism, cavefish have evolved differences in behavior, incl...

  13. Synthesis and characterization of a pH-sensitive shielding system for polycation gene carriers

    Institute of Scientific and Technical Information of China (English)

    2010-01-01

    To increase the in vivo stability of polycation gene carriers,a pH-sensitive shielding system,γ-benzyl L-glutamate-co-glutamate acid polymer(PGA(60)(60 refers to the molar ratio of glutamate acid in the polymer)),was synthesized and characterized.PGA(60) showed pH sensitivity at about pH 6.0.PGA(60) shielded the positive charge of DNA/PEI(1:1) complexes.Gel retardation assay showed that no DNA-strand exchange with PGA(60) occurred after PGA(60) was added to DNA/PEI complexes at different proportions.MTT cytotoxicity tests demonstrated that neither PGA(60) nor DNA/PEI/PGA(60) ternary complexes had cytotoxicity at the test concentration.The transfection efficiency was improved when the positive charge was partly shielded by PGA(60).Because of the charge repulsion between the surface of cells and ternary complex particles,there was almost no transfection efficiency when the zeta potential of ternary complexes turned to negative.Because of the suitable pH sensitive range,PGA(60) may be a potential shielding system for polycation gene carriers to be used in vivo.

  14. Synthesis of the human insulin gene: protein expression, scaling up and bioactivity.

    Science.gov (United States)

    Redwan, El-Rashdy M; Matar, Saleh M; El-Aziz, Gamal Abd; Serour, Ehab A

    2008-01-01

    Optimized Synthetic human insulin gene was preferred to easy of cloning, plasmid stability, and protein expression away from the native sequence and its rare codons. Two steps to obtain the insulin, so we assembled the gene of 293 bp using a battery of overlapped synthetic oligos, then cloned into pET101directional TOPO expression vector downstream to the T7 promoter. The proinsulin products were produced as inclusion bodies in E. coli at a level of 10%. The batch cultivation of the strain yielded 6 g/L, while the high cell density of fed-batch cultivation yielded 46 g/L. The proinsulin purification yielded 110 mg/gram cell weight, and 1.3 mg/gram of a bioactive insulin. The native insulin was generated by enzymatic conversion of chemically processed proinsulin. The produced insulin was matched with that of a commercial aqueous version at a level of enzyme immunoassys, SDS-PAGE, RP-HPLC, and bioactivity. The present results showed that the produced insulin has a comparable biochemical and potency similar to that of commercial one. PMID:18080908

  15. Synthesis and Evaluation of Tetramethylguanidinium-Polyethylenimine Polymers as Efficient Gene Delivery Vectors

    Directory of Open Access Journals (Sweden)

    Manohar Mahato

    2014-01-01

    Full Text Available Previously, we demonstrated that 6-(N,N,N′,N′-tetramethylguanidinium chloride-hexanoyl-polyethylenimine (THP polymers exhibited significantly enhanced transfection efficiency and cell viability. Here, in the present study, we have synthesized a series of N,N,N′,N′-tetramethylguanidinium-polyethylenimine (TP1-TP5 polymers via a single-step reaction involving peripheral primary amines of bPEI and varying amounts of 2-(1H-benzotriazol-1-yl-1,1,3,3-tetramethyluronium hexafluorophosphate (HBTU. These polymers were found to interact efficiently with negatively charged pDNA and formed stable complexes in the size range of ~240–450 nm. Acid-base titration profiles revealed improved buffering capacity of TP polymers as compared to bPEI. Transfection and cytotoxicity assays performed with TP/pDNA complexes on HEK293, CHO, and HeLa cells showed significantly higher transfection efficiency and cell viability with one of the complexes, TP2/pDNA complex, exhibited the highest transfection efficiency (~1.4–2.3-fold outcompeting native bPEI and the commercially available transfection reagent, Lipofectamine 2000. Compared to previously reported THP polymers, the transfection efficiency of TP/pDNA complexes was found to be lower, as examined by flow cytometry. These results highlight the importance of the hydrophobic C-6 linker in THP polymers in forming compact nanostructures with pDNA, which might lead to efficient uptake and internalization of the complexes; however, the projected TP polymers offer an advantage of their rapid and economical one-step synthesis.

  16. Synthesis and evaluation of cationic nanomicelles for in vitro and in vivo gene delivery

    Science.gov (United States)

    Mandke, Rhishikesh Subhash

    The goal of proposed study was to contribute towards the development of a nano size, high efficiency and low toxicity non-viral polymeric vector for gene delivery in vitro and in vivo. A series of fatty acid grafted low-molecular-weight chitosan (N-acyl LMWCs) were synthesized, purified and characterized for their physicochemical properties using various analytical techniques such as infrared spectroscopy, elemental analysis and dynamic light scattering. The formulation parameters including pH, sonication duration, and filtration altered the physicochemical characteristics of N-acyl LMWC nanomicelles. The acyl chain length and degree of unsaturation in fatty acids also had an impact on the physicochemical properties and the transfection efficiency of nanomicelles. N-acyl LMWC nanomicelles showed efficient in vitro transfection as visualized and quantified using a reporter plasmid (encoding green fluorescent protein), and therapeutic plasmids (encoding for interleukin-4 and interleukin-10), respectively. The in vitro transfection efficiencies of N-acyl LMWCs with 18:1 and 18:2 grafts (oleic and linoleic acids) were comparable with FuGENERTM HD (marketed non-viral vector) but were ˜8-fold and 35-fold higher as compared to LMWC and naked DNA, respectively. The in vivo transfection efficiency of N-acyl LMWC to deliver plasmids individually encoding IL-4 and IL-10 as well as a bicistronic plasmid encoding both IL-4 and IL-10 was studied in a multiple, low-dose streptozotocin induced diabetic mouse model. The transfection efficiency of pDNA/N-acyl LMWC polyplexes injected via intramuscular route showed significant improvement (p<0.05) over passive (naked DNA) or positive (FuGENE HD) controls. Additionally, a sustained and efficient expression of IL-4 and IL-10 was observed, accompanied by a reduction in interferon-gamma (INF-gamma), and tumor necrosis factor-alpha (TNF-alpha) levels. The pancreas of pDNA/N-acyl LMWC polyplex treated animals exhibited protection from

  17. Sequences more than 500 base pairs upstream of the human U3 small nuclear RNA gene stimulate the synthesis of U3 RNA in frog oocytes

    International Nuclear Information System (INIS)

    Small nuclear RNA (snRNA) genes contain strong promoters capable of initiating transcription once every 4 s. Studies on the human U1 snRNA gene, carried out in other laboratories, showed that sequences within 400 bp of the 5' flanking region are sufficient for maximal levels of transcription both in vivo and in frog oocytes [reviewed in Dahlberg and Lund (1988)]. The authors studied the expression of a human U3 snRNA gene by injecting 5' deletion mutants into frog oocytes. The results show that sequences more than 500 bp upstream of the U3 snRNA gene have a 2-3-fold stimulatory effect on the U3 snRNA synthesis. These results indicate that the human U3 snRNA gene is different from human U1 snRNA gene in containing regulatory elements more than 500 bp upstream. The U3 snRNA gene upstream sequences contain an AluI homologous sequence in the -1,200 region; these AluI sequences were transcribed in vitro and in frog oocytes but were not detectable in Hela cells

  18. Fe2+ chelator proferrorosamine A: a gene cluster of Erwinia rhapontici P45 involved in its synthesis and its impact on growth of Erwinia amylovora CFBP1430.

    Science.gov (United States)

    Born, Yannick; Remus-Emsermann, Mitja N P; Bieri, Marco; Kamber, Tim; Piel, Jörn; Pelludat, Cosima

    2016-02-01

    Proferrorosamine A (proFRA) is an iron (Fe2+) chelator produced by the opportunistic plant pathogen Erwinia rhapontici P45. To identify genes involved in proFRA synthesis, transposon mutagenesis was performed. The identified 9.3 kb gene cluster, comprising seven genes, designated rosA-rosG, encodes proteins that are involved in proFRA synthesis. Based on gene homologies, a biosynthetic pathway model for proFRA is proposed. To obtain a better understanding of the effect of proFRA on non-proFRA producing bacteria, E. rhapontici P45 was co-cultured with Erwinia amylovora CFBP1430, a fire-blight-causing plant pathogen. E. rhapontici P45, but not corresponding proFRA-negative mutants, led to a pink coloration of E. amylovora CFBP1430 colonies on King's B agar, indicating accumulation of the proFRA-iron complex ferrorosamine, and growth inhibition in vitro. By saturating proFRA-containing extracts with Fe2+, the inhibitory effect was neutralized, suggesting that the iron-chelating capability of proFRA is responsible for the growth inhibition of E. amylovora CFBP1430. PMID:26732708

  19. Synthesis of water-based cationic polyurethane for antibacterial and gene delivery applications.

    Science.gov (United States)

    Wu, Geng-Hsi; Hsu, Shan-Hui

    2016-10-01

    Cationic polymers are often used as antimicrobial materials and transfection reagents. Water-based process could reduce environmental pollution and prevent the risk of solvent residue in the final product. In this study, waterborne biodegradable cationic polyurethane (WCPU) was synthesized by reacting polycaprolactone (PCL diol), isophorone diisocyanate (IPDI), and N-methyldiethanolamine (N-MDEA) under 75°C. An aqueous dispersion of WCPU nanoparticles (NPs) could be acquired by vigorous stirring under acidic condition. The particles in the dispersion had an average size of ∼80nm and a zeta potential of ∼60mV. When cast into films, the contact angle of the film was ∼67° and the zeta potential was ∼16mV. WCPU NPs demonstrated excellent antibacterial activity against Escherichia coli (E. coli) and Staphylococcus aureus (S. aureus) (100% inhibition with a contact time of 3h). Meanwhile, the antibacterial ratio of WCPU films to E. coli and S. aureus reached 100% after 24h of contact. Moreover, WCPU NPs could be used as a transfection reagent without significant toxicity for concentrations less than 1000μg/mL and showed the ability to condensate plasmid DNA. The transfection efficiency for HEK293T cells and hBMSCs was ∼60% and ∼30% at 48h, respectively, after the transfection. Therefore, the WCPU synthesized in this study has potential antibacterial and gene delivery applications. PMID:27451371

  20. Synthesis and Characterization of Layered Double Hydroxides and Their Potential as Nonviral Gene Delivery Vehicles

    Science.gov (United States)

    Balcomb, Blake; Singh, Moganavelli; Singh, Sooboo

    2015-01-01

    Layered double hydroxides (LDHs) exhibit characteristic anion-exchange chemistry making them ideal carriers of negatively charged molecules like deoxyribonucleic acid (DNA). In this study, hydrotalcite (Mg−Al) and hydrotalcite-like compounds (Mg−Fe, Zn−Al, and Zn−Fe), also known as LDHs, were evaluated for their potential application as a carrier of DNA. LDHs were prepared by coprecipitation at low supersaturation and characterized by Powder X-ray diffraction (XRD), infrared (IR), Raman, and inductively coupled plasma—optical emission spectroscopy, scanning electron microscopy (SEM), and transmission electron microscopy (TEM). XRD patterns showed strong and sharp diffraction peaks for the (003) and (006) planes indicating well-ordered crystalline materials. TEM images yielded irregular circular to hexagonal-shaped particles of 50–250 nm in size. Varying degrees of DNA binding was observed for all the compounds, and nuclease digestion studies revealed that the LDHs afford some degree of protection to the bound DNA. Minimal toxicity was observed in human embryonic kidney (HEK293), cervical cancer (HeLa) and hepatocellular carcinoma (HepG2) cell lines with most showing a cell viability in excess of 80 %. All LDH complexes promoted significant levels of luciferase gene expression, with the DNA:Mg−Al LDHs proving to be the most efficient in all cell lines. PMID:25969811

  1. Multiple Types of 8-Vinyl Reductases for (Bacterio)Chlorophyll Biosynthesis Occur in Many Green Sulfur Bacteria ▿

    OpenAIRE

    Liu, Zhenfeng; Bryant, Donald A.

    2011-01-01

    Two 8-vinyl reductases, BciA and BciB, have been identified in chlorophototrophs. The bciA gene of Chlorobaculum tepidum was replaced with genes similar to bciB from other green sulfur bacteria. Pigment analyses of the complemented strains showed that the bciB homologs encode 8-vinyl reductases similar to those of cyanobacteria.

  2. Recent advances in genes involved in secondary metabolite synthesis, hyphal development, energy metabolism and pathogenicity in Fusarium graminearum (teleomorph Gibberella zeae).

    Science.gov (United States)

    Geng, Zongyi; Zhu, Wei; Su, Hao; Zhao, Yong; Zhang, Ke-Qin; Yang, Jinkui

    2014-01-01

    The ascomycete fungus, Fusarium graminearum (teleomorph Gibberella zeae), is the most common causal agent of Fusarium head blight (FHB), a devastating disease for cereal crops worldwide. F. graminearum produces ascospores (sexual spores) and conidia (asexual spores), which can serve as disease inocula of FHB. Meanwhile, Fusarium-infected grains are often contaminated with mycotoxins such as trichothecenes (TRIs), fumonisins, and zearalenones, among which TRIs are related to the pathogenicity of F. graminearum, and these toxins are hazardous to humans and livestock. In recent years, with the complete genome sequencing of F. graminearum, an increasing number of functional genes involved in the production of secondary metabolites, hyphal differentiation, sexual and asexual reproduction, virulence and pathogenicity have been identified from F. graminearum. In this review, the secondary metabolite synthesis, hyphal development and pathogenicity related genes in F. graminearum were thoroughly summarized, and the genes associated with secondary metabolites, sexual reproduction, energy metabolism, and pathogenicity were highlighted. PMID:24389085

  3. Gene

    Data.gov (United States)

    U.S. Department of Health & Human Services — Gene integrates information from a wide range of species. A record may include nomenclature, Reference Sequences (RefSeqs), maps, pathways, variations, phenotypes,...

  4. Control of placental alkaline phosphatase gene expression in HeLa cells: induction of synthesis by prednisolone and sodium butyrate

    International Nuclear Information System (INIS)

    HeLa S3 cells produce an alkaline phosphatase indistinguishable from the enzyme from human term placenta. The phosphatase activity in these cells was induced by both prednisolone and sodium butyrate. Both agents stimulated de novo synthesis of the enzyme. The increase in phosphatase activity paralleled the increase in immunoactivity and biosynthesis of placental alkaline phosphatase. The fully processed phosphatase monomer in control, prednisolone-treated or butyrate-treated cells was a 64.5 K polypeptide, measured by both incorporation of L-[35S]methionine into enzyme protein and active-site labeling. The 64.5K polypeptide was formed by the incorporation of additional N-acetylneuraminic acid moieties to a precursor polypeptide of 61.5K. However, this biosynthetic pathway was identified only in butyrate-treated cells. In prednisolone-treated cells, the processing of 61.5K to 64.5K monomer was accelerated, and the presence of the 61.5 precursor could only be detected by either neuraminidase or monensin treatment. Phosphatase mRNA which comigrated with the term placental alkaline phosphatase mRNA of 2.7 kilobases was induced in the presence of either prednisolone or butyrate. Alkaline phosphatase mRNA is untreated HeLa S3 cells migrated slightly faster than the term placental alkaline phosphatase mRNA. Butyrate also induced a second still faster migrating alkaline phosphatase mRNA. Both prednisolone and butyrate increased the steady-state levels of placental alkaline phosphatase mRNA. The data indicate that the increase in phosphatase mRNA by prednisolone and butyrate resulted in the induction of alkaline phosphatase activity and biosynthesis in HeLa S3 cells. Furthermore, both agents induced the expression of different alkaline phosphatase gene transcripts without altering its protein product

  5. Development of novel polymeric materials for gene therapy and pH-sensitive drug delivery: Modeling, synthesis, characterization, and analysis

    Science.gov (United States)

    Anderson, Brian Curtis

    The aim of this work was to obtain a fundamental understanding of drug release mechanisms from polymers that undergo thermoreversible gelation and to synthesize new polymers based on these that exhibit both pH and temperature sensitivity. Novel block and random copolymers with cationic character have been developed for drug delivery and gene therapy applications. The development of these materials began with a study of the mechanism of drug release from poly(ethylene oxide) (PEO) and poly(propylene oxide) (PPO) block copolymers. This study revealed the release rates of drugs from water-soluble hydrogels composed of the PEO-PPO-PEO block copolymer PluronicRTM F127 was dictated almost solely by the rate of interfacial dissolution at the water/gel interface. A setup was designed to measure drug release from such soluble systems in order to avoid confounding hydrodynamic effects as a result of shear on the delicate polymer/gel interface. This study was followed by a complementary analysis of the effect ionic salts play in the phase transitions and drug release profiles in aqueous F127 solutions. In an attempt to incorporate pH sensitivity into such drug release systems, several block copolymers of poly(N,N-diethylaminoethyl methacrylate) (PDEAEM), PEO and PPO were synthesized via anionic polymerization. Diblock materials (PEO-b-PDEAEM), either with or without a carboxylic acid endcap, were synthesized and characterized. Tablet dissolution experiments demonstrated pH-sensitivity in their drug release profiles relative to PEO tablets. Pentablock materials (PDEAEM-b-PEO-b-PPO- b-PEO-b-PDEAEM) were synthesized that maintain the thermoreversible gelation and micellization properties of F127 while introducing pH-dependent release from aqueous gels of the copolymer. This is the first example of non-crosslinked materials that exhibit both pH- and temperature-sensitive behavior. Using a similar synthesis route, random copolymers of PDEAEM and poly(poly(ethylene glycol) methyl

  6. A potential benefit of albinism in Astyanax cavefish: downregulation of the oca2 gene increases tyrosine and catecholamine levels as an alternative to melanin synthesis.

    Directory of Open Access Journals (Sweden)

    Helena Bilandžija

    Full Text Available Albinism, the loss of melanin pigmentation, has evolved in a diverse variety of cave animals but the responsible evolutionary mechanisms are unknown. In Astyanax mexicanus, which has a pigmented surface dwelling form (surface fish and several albino cave-dwelling forms (cavefish, albinism is caused by loss of function mutations in the oca2 gene, which operates during the first step of the melanin synthesis pathway. In addition to albinism, cavefish have evolved differences in behavior, including feeding and sleep, which are under the control of the catecholamine system. The catecholamine and melanin synthesis pathways diverge after beginning with the same substrate, L-tyrosine. Here we describe a novel relationship between the catecholamine and melanin synthesis pathways in Astyanax. Our results show significant increases in L-tyrosine, dopamine, and norepinephrine in pre-feeding larvae and adult brains of Pachón cavefish relative to surface fish. In addition, norepinephrine is elevated in cavefish adult kidneys, which contain the teleost homologs of catecholamine synthesizing adrenal cells. We further show that the oca2 gene is expressed during surface fish development but is downregulated in cavefish embryos. A key finding is that knockdown of oca2 expression in surface fish embryos delays the development of pigmented melanophores and simultaneously increases L-tyrosine and dopamine. We conclude that a potential evolutionary benefit of albinism in Astyanax cavefish may be to provide surplus L-tyrosine as a precursor for the elevated catecholamine synthesis pathway, which could be important for adaptation to the challenging cave environment.

  7. NATURAL MUTATION IN THE GENE OF RESPONSE REGULATOR BgrR RESULTING IN REPRESSION OF Bac PROTEIN SYNTHESIS, A PATHOGENICITY FACTOR OF STREPTOCOCCUS AGALACTIAE

    Directory of Open Access Journals (Sweden)

    A. S. Rozhdestvenskaya

    2013-01-01

    Full Text Available Abstract. Streptococcus agalactiae can cause variety of diseases of newborns and adults. For successful colonization of different human tissues and organs as well as for suppression of the host immune system S. agalactiae expresses numerous virulence factors. For coordinated expression of the virulence genes S. agalactiae employs regulatory molecules including regulatory proteins of two-component systems. Results of the present study demonstrated that in S. agalactiae strain A49V the natural mutation in the brgR gene encoding for BgrR regulatory protein, which is component of regulatory system BgrRS, resulted in the repression of Bac protein synthesis, a virulence factor of S. agalactiae. A single nucleotide deletion in the bgrR gene has caused a shift of the reading frame and the changes in the primary, secondary and tertiary structures of the BgrR protein. The loss of functional activity of BgrR protein in A49V strain and repression of Bac protein synthesis have increased virulence of the strain in experimental animal streptococcal infection.

  8. Effect of carotenoids and monogalactosyl diglyceride on bacteriochlorophyll c aggregates in aqueous buffer: implications for the self-assembly of chlorosomes.

    Science.gov (United States)

    Klinger, Pavel; Arellano, Juan B; Vácha, Frantisek; Hála, Jan; Psencík, Jakub

    2004-01-01

    Aggregation of bacteriochlorophyll (BChl) c from chlorosomes, the main light-harvesting complex of green bacteria, has been studied in aqueous buffer. Unlike other chlorophyll-like molecules, BChl c is rather soluble in aqueous buffer, forming dimers. When BChl c is mixed with carotenoids (Car), the BChl c Qy transition is further redshifted, in respect to that of monomers and dimers. The results suggest that Car are incorporated in the aggregates and induce further aggregation of BChl c. The redshift of the BChl c Qy band is proportional to the Car concentration. In contrast, the mixture of bacteriochlorophyllide (BChlide) c, which lacks the nonpolar esterifying alcohol, does not form aggregates with Car in aqueous buffer or nonpolar solvents. Instead, the position of the BChlide c Qy transition remains unshifted in respect to that of the monomeric molecule, and Car precipitates with the course of time in aqueous buffer. Similar effects on both BChl c and BChlide c are also observed when monogalactosyl diglyceride (MGDG), which forms the monolayer envelope of chlorosomes, is used instead of (or together with) Car. The results show that the hydrophobic interactions of the BChl c esterifying alcohols with themselves and the nonpolar carbon skeleton of Car, or the fatty acid tails of MGDG, are essential driving forces for BChl aggregation in chlorosomes. PMID:15623345

  9. Self-assembly of [Et,Et]-bacteriochlorophyll cF on highly oriented pyrolytic graphite revealed by scanning tunneling microscopy.

    Science.gov (United States)

    Möltgen, H; Kleinermanns, K; Jesorka, A; Schaffner, K; Holzwarth, A R

    2002-06-01

    The chlorosomal light-harvesting antennae of green phototrophic bacteria consist of large supramolecular aggregates of bacteriochlorophyll c (BChl c). The supramolecular structure of (3(1)-R/S)-BChl c on highly oriented pyrolytic graphite (HOPG) and molybdenum disulfide (MoS2) has been investigated by scanning tunneling microscopy (STM). On MoS2, we observed single BChl c molecules, dimers or tetramers, depending on the polarity of the solvent. On HOPG, we observed extensive self-assembly of the dimers and tetramers. We propose C=O...H-O...Mg bonding networks for the observed dimer chains, in agreement with former ultraviolet-visible and infrared spectroscopic work. The BChl c moieties in the tetramers are probably linked by four C=O...H-O hydrogen bonds to form a circle and further stabilized by Mg...O-H bondings to underlying BChl c layers. The tetramers form highly ordered, distinct chains and extended two-dimensional networks. We investigated semisynthetic chlorins for comparison by STM but observed that only BChl c self-assembles to well-structured large aggregates on HOPG. The results on the synthetic chlorins support our structure proposition. PMID:12081324

  10. Ultra-broadband 2D electronic spectroscopy of carotenoid-bacteriochlorophyll interactions in the LH1 complex of a purple bacterium

    International Nuclear Information System (INIS)

    We investigate the excitation energy transfer (EET) pathways in the photosynthetic light harvesting 1 (LH1) complex of purple bacterium Rhodospirillum rubrum with ultra-broadband two-dimensional electronic spectroscopy (2DES). We employ a 2DES apparatus in the partially collinear geometry, using a passive birefringent interferometer to generate the phase-locked pump pulse pair. This scheme easily lends itself to two-color operation, by coupling a sub-10 fs visible pulse with a sub-15-fs near-infrared pulse. This unique pulse combination allows us to simultaneously track with extremely high temporal resolution both the dynamics of the photoexcited carotenoid spirilloxanthin (Spx) in the visible range and the EET between the Spx and the B890 bacterio-chlorophyll (BChl), whose Qx and Qy transitions peak at 585 and 881 nm, respectively, in the near-infrared. Global analysis of the one-color and two-color 2DES maps unravels different relaxation mechanisms in the LH1 complex: (i) the initial events of the internal conversion process within the Spx, (ii) the parallel EET from the first bright state S2 of the Spx towards the Qx state of the B890, and (iii) the internal conversion from Qx to Qy within the B890

  11. Ultra-broadband 2D electronic spectroscopy of carotenoid-bacteriochlorophyll interactions in the LH1 complex of a purple bacterium.

    Science.gov (United States)

    Maiuri, Margherita; Réhault, Julien; Carey, Anne-Marie; Hacking, Kirsty; Garavelli, Marco; Lüer, Larry; Polli, Dario; Cogdell, Richard J; Cerullo, Giulio

    2015-06-01

    We investigate the excitation energy transfer (EET) pathways in the photosynthetic light harvesting 1 (LH1) complex of purple bacterium Rhodospirillum rubrum with ultra-broadband two-dimensional electronic spectroscopy (2DES). We employ a 2DES apparatus in the partially collinear geometry, using a passive birefringent interferometer to generate the phase-locked pump pulse pair. This scheme easily lends itself to two-color operation, by coupling a sub-10 fs visible pulse with a sub-15-fs near-infrared pulse. This unique pulse combination allows us to simultaneously track with extremely high temporal resolution both the dynamics of the photoexcited carotenoid spirilloxanthin (Spx) in the visible range and the EET between the Spx and the B890 bacterio-chlorophyll (BChl), whose Qx and Qy transitions peak at 585 and 881 nm, respectively, in the near-infrared. Global analysis of the one-color and two-color 2DES maps unravels different relaxation mechanisms in the LH1 complex: (i) the initial events of the internal conversion process within the Spx, (ii) the parallel EET from the first bright state S2 of the Spx towards the Qx state of the B890, and (iii) the internal conversion from Qx to Qy within the B890. PMID:26049453

  12. WST11, a novel water-soluble bacteriochlorophyll derivative; cellular uptake, pharmacokinetics, biodistribution and vascular-targeted photodynamic activity using melanoma tumors as a model.

    Science.gov (United States)

    Mazor, Ohad; Brandis, Alexander; Plaks, Vicki; Neumark, Eran; Rosenbach-Belkin, Varda; Salomon, Yoram; Scherz, Avigdor

    2005-01-01

    WST11 is a novel negatively charged water-soluble palladium-bacteriochlorophyll derivative that was developed for vascular-targeted photodynamic therapy (VTP) in our laboratory. The in vitro results suggest that WST11 cellular uptake, clearance and phototoxicity are mediated by serum albumin trafficking. In vivo, WST11 was found to clear rapidly from the circulation (t1/2=1.65 min) after intravenous bolus injection in the mouse, whereas a longer clearance time (t1/2=7.5 min) was noted in rats after 20 min of infusion. The biodistribution of WST11 in mouse tissues indicates hepatic clearance (t1/2=20 min), with minor (kidney, lung and spleen) or no intermediary accumulation in other tissues. As soon as 1 h after injection, WST11 had nearly cleared from the body of the mouse, except for a temporal accumulation in the lungs from which it cleared within 40 min. On the basis of these results, we set the VTP protocol for a short illumination period (5 min), delivered immediately after WST11 injection. On subjecting M2R melanoma xenografts to WST11-VTP, we achieved 100% tumor flattening at all doses and a 70% cure with 9 mg/kg and a light exposure dose of 100 mW/cm2. These results provide direct evidence that WST11 is an effective agent for VTP and provide guidelines for further development of new candidates. PMID:15623318

  13. Ultra-broadband 2D electronic spectroscopy of carotenoid-bacteriochlorophyll interactions in the LH1 complex of a purple bacterium

    Energy Technology Data Exchange (ETDEWEB)

    Maiuri, Margherita [CNR-IFN, Dipartimento di Fisica, Politecnico di Milano, P.zza L. da Vinci 32, Milano 20133 (Italy); Department of Chemistry, Princeton University, Washington Road, Princeton, New Jersey 08544 (United States); Réhault, Julien; Polli, Dario; Cerullo, Giulio, E-mail: giulio.cerullo@polimi.it [CNR-IFN, Dipartimento di Fisica, Politecnico di Milano, P.zza L. da Vinci 32, Milano 20133 (Italy); Carey, Anne-Marie; Hacking, Kirsty; Cogdell, Richard J. [Glasgow Biomedical Research Centre, IBLS, University of Glasgow, 126 Place, Glasgow G12 8TA, Scotland (United Kingdom); Garavelli, Marco [Dipartimento di Chimica “G. Ciamician,” Università di Bologna, Via Selmi 2, IT-40126 Bologna (Italy); CNRS, Institut de Chimie de Lyon, École Normale Supérieure de Lyon, Université de Lyon, 46 Allée d’Italie, F-69364 Lyon Cedex 07 (France); Lüer, Larry [Madrid Institute for Advanced Studies, IMDEA Nanociencia, Madrid (Spain)

    2015-06-07

    We investigate the excitation energy transfer (EET) pathways in the photosynthetic light harvesting 1 (LH1) complex of purple bacterium Rhodospirillum rubrum with ultra-broadband two-dimensional electronic spectroscopy (2DES). We employ a 2DES apparatus in the partially collinear geometry, using a passive birefringent interferometer to generate the phase-locked pump pulse pair. This scheme easily lends itself to two-color operation, by coupling a sub-10 fs visible pulse with a sub-15-fs near-infrared pulse. This unique pulse combination allows us to simultaneously track with extremely high temporal resolution both the dynamics of the photoexcited carotenoid spirilloxanthin (Spx) in the visible range and the EET between the Spx and the B890 bacterio-chlorophyll (BChl), whose Q{sub x} and Q{sub y} transitions peak at 585 and 881 nm, respectively, in the near-infrared. Global analysis of the one-color and two-color 2DES maps unravels different relaxation mechanisms in the LH1 complex: (i) the initial events of the internal conversion process within the Spx, (ii) the parallel EET from the first bright state S{sub 2} of the Spx towards the Q{sub x} state of the B890, and (iii) the internal conversion from Q{sub x} to Q{sub y} within the B890.

  14. [Gene expression of the key enzymes controlling starch synthesis and metabolism in rice grain endosperm under effects of high temperature after anthesis].

    Science.gov (United States)

    Zhong, Lian-Jin; Dong, Hu; Cai, Xiao-Bo; Feng, Yan-Ning; Ren, Ping; Cheng, Fang-Min

    2012-03-01

    Taking an early-season indica cultivar 'Jiazao 935' whose grain quality was sensitive to temperature as test material, and by using artificial climatic chamber and real-time fluorescence quantitative PCR (FQ-PCR), this paper studied the relative expression amount and its dynamic changes of ten isoform genes of the key enzymes controlling starch synthesis and metabolism in rice grain endosperm, including sbe1, sbe3, and sbe4 of starch branching enzyme (SBE), isal, isa2, isa3, and pul of starch debranching enzyme (DBE), and Wx, sss1, and sss2a of starch synthase (SS), at the mean daily temperature 22 and 32 degrees C after anthesis. There existed obvious differences in the expression patterns of these genes under the high temperature stress, and the expression patterns were isoform-dependent. The relative expression amount of sbe1 and sbe3 under high temperature decreased significantly, and both of the genes were the sensitive isoform genes of SBE to high temperature stress. Among the DBE genes, pul was the isoform gene with high expression level, being more sensitive to high temperature stress than isa1, isa2, and isa3. Among the SS genes, sss2a had a significantly lower relative expression amount than sss1 and Wx, but sss2a and sss1 were more sensitive to high temperature than Wx, suggesting that sss2a and sss1 could be the important genes that adjusted the starch structure in rice endosperm under high temperature stress, especially at the middle and late grain filling stages. PMID:22720620

  15. Vitamin B12 regulates photosystem gene expression via the CrtJ antirepressor AerR in Rhodobacter capsulatus

    OpenAIRE

    Cheng, Zhuo; Li, Keran; Hammad, Loubna A.; Karty, Jonathan A.; Bauer, Carl E.

    2014-01-01

    The tetrapyrroles heme, bacteriochlorophyll and cobalamin (B12) exhibit a complex interrelationship regarding their synthesis. In this study, we demonstrate that AerR functions as an antirepressor of the tetrapyrrole regulator CrtJ. We show that purified AerR contains B12 that is bound to a conserved histidine (His145) in AerR. The interaction of AerR to CrtJ was further demonstrated in vitro by pull down experiments using AerR as bait and quantified using microscale thermophoresis. DNase I D...

  16. Discovery of single-nucleotide mutations in genes related to rice starch synthesis and herbicide resistance by using self-made CEL I extracts

    International Nuclear Information System (INIS)

    The foundation of CEL I, a specific nuclease isolated from celery, makes the detection of point mutations to be easy and robust and it is essential nowadays in TILLING. However, large amounts of CEL I are consumed in TILLING and its extraction process is time-consuming. Furthermore, the high cost both in isolation and application of commercial CEL I Kit is an albatross for scientists in developing countries. Herein is presented a rapid method for detection of single-nucleotide mutations in rice genes by using self-made CEL I extracts. After tests on mismatch cleavage activity of CEL I extracts at different extraction steps, it was proved that CEL I extracts after clarification and dialysis are sufficiently enriched in mismatch cleavage activity. By optimization of factors related to mismatch cleavage activity, we found that CEL I extract made by ourself showed same function for mismatch cleavage as the commercial CEL I and established a feasible and effective method for detecting point mutation. Understanding and manipulating genetic variation is paramount to elucidating gene function, identifying genes, breeding, and conserving natural diversity. The general applicability of CEL I makes it great potential for detecting and understanding genetic variation in rice. By the method of mutation detection we set up using self-made CEL I, we found single-nucleotide mutations of some rice genes, such as waxy, SSIIa (starch synthase IIa) and als (acetolactate synthase), related to rice starch synthesis or herbicide resistance. The single-based variation (T/G or A/G) were detected both in first intron of waxy gene and 8th extron of SSIIa gene. For als gene, we found the single-nucleotide mutation at the position about 700bp and 400bp in the 1.5kb fragment amplified from different varieties and M2 plants respectively. (author)

  17. p38 MAPKs regulate the expression of genes in the dopamine synthesis pathway through phosphorylation of NR4A nuclear receptors.

    Science.gov (United States)

    Sekine, Yusuke; Takagahara, Shuichi; Hatanaka, Ryo; Watanabe, Takeshi; Oguchi, Haruka; Noguchi, Takuya; Naguro, Isao; Kobayashi, Kazuto; Tsunoda, Makoto; Funatsu, Takashi; Nomura, Hiroshi; Toyoda, Takeshi; Matsuki, Norio; Kuranaga, Erina; Miura, Masayuki; Takeda, Kohsuke; Ichijo, Hidenori

    2011-09-01

    In Drosophila, the melanization reaction is an important defense mechanism against injury and invasion of microorganisms. Drosophila tyrosine hydroxylase (TH, also known as Pale) and dopa decarboxylase (Ddc), key enzymes in the dopamine synthesis pathway, underlie the melanin synthesis by providing the melanin precursors dopa and dopamine, respectively. It has been shown that expression of Drosophila TH and Ddc is induced in various physiological and pathological conditions, including bacterial challenge; however, the mechanism involved has not been fully elucidated. Here, we show that ectopic activation of p38 MAPK induces TH and Ddc expression, leading to upregulation of melanization in the Drosophila cuticle. This p38-dependent melanization was attenuated by knockdown of TH and Ddc, as well as by that of Drosophila HR38, a member of the NR4A family of nuclear receptors. In mammalian cells, p38 phosphorylated mammalian NR4As and Drosophila HR38 and potentiated these NR4As to transactivate a promoter containing NR4A-binding elements, with this transactivation being, at least in part, dependent on the phosphorylation. This suggests an evolutionarily conserved role for p38 MAPKs in the regulation of NR4As. Thus, p38-regulated gene induction through NR4As appears to function in the dopamine synthesis pathway and may be involved in immune and stress responses. PMID:21878507

  18. Sequence of the bchG gene from Chloroflexus aurantiacus: relationship between chlorophyll synthase and other polyprenyltransferases

    Science.gov (United States)

    Lopez, J. C.; Ryan, S.; Blankenship, R. E.

    1996-01-01

    The sequence of the Chloroflexus aurantiacus open reading frame thought to be the C. aurantiacus homolog of the Rhodobacter capsulatus bchG gene is reported. The BchG gene product catalyzes esterification of bacteriochlorophyllide a by geranylgeraniol-PPi during bacteriochlorophyll a biosynthesis. Homologs from Arabidopsis thaliana, Synechocystis sp. strain PCC6803, and C. aurantiacus were identified in database searches. Profile analysis identified three related polyprenyltransferase enzymes which attach an aliphatic alcohol PPi to an aromatic substrate. This suggests a broader relationship between chlorophyll synthases and other polyprenyltransferases.

  19. The mouse pink-eyed dilution allele of the P-gene greatly inhibits eumelanin but not pheomelanin synthesis.

    Science.gov (United States)

    Hirobe, Tomohisa; Ito, Shosuke; Wakamatsu, Kazumasa

    2011-02-01

    The mouse pink-eyed dilution (p) locus is known to control eumelanin synthesis, melanosome morphology, and tyrosinase activity in melanocytes. However, it has not been fully determined whether the mutant allele, p affects pheomelanin synthesis. Effects of the p allele on eumelanin and phemelanin synthesis were investigated by chemical analysis of dorsal hairs of 5-week-old mice obtained from the F(2) generations (black, pink-eyed black, recessive yellow, pink-eyed recessive yellow, agouti, and pink-eyed agouti) between C57BL/10JHir (B10)-congenic pink-eyed black mice (B10-p/p) and recessive yellow (B10-Mc1r(e)/Mc1r(e)) or agouti (B10-A/A) mice. The eumelanin content was dramatically (>20-fold) decreased in pink-eyed black and pink-eyed agouti mice, whereas the pheomelanin content did not decrease in pink-eyed black, pink-eyed recessive yellow, or pink-eyed agouti mice compared to the corresponding P/- mice. These results suggest that the pink-eyed dilution allele greatly inhibits eumelanin synthesis, but not pheomelanin synthesis. PMID:21232027

  20. The absence of the luxS gene increases swimming motility and flagella synthesis in Escherichia coli K12

    International Nuclear Information System (INIS)

    Research highlights: → This paper provides the first evidence that luxS deletion enhances swimming motility and flagella synthesis in Escherichia coli K12 based on motility, transcriptome, and scanning electron microscopy analyses. → A conceptual genetic regulatory network underlying the increased flagella synthesis was constructed based on the transcriptome and network component analyses, and previously known regulatory relations. → The genetic regulatory network suggests that the increased flagella synthesis and motility might be contributed to by increased flhDC transcription level and/or decreased c-di-GMP concentration in luxS-deficient E. coli. -- Abstract: Despite the significant role of S-ribosylhomocysteinase (LuxS) in the activated methyl cycle pathway and quorum sensing, the connectivity between luxS and other cellular functions remains incomplete. Herein, we show that luxS deletion significantly increases swimming motility and flagella synthesis in Escherichia coli K12 using motility, transcriptome, and scanning electron microscopy assays. Further, based on the transcriptome and network component analyses, and known regulatory relations, we propose a conceptual genetic regulatory network underlying the increased flagella synthesis in response to luxS deletion.

  1. Visual gene developer: a fully programmable bioinformatics software for synthetic gene optimization

    OpenAIRE

    McDonald Karen; Jung Sang-Kyu

    2011-01-01

    Abstract Background Direct gene synthesis is becoming more popular owing to decreases in gene synthesis pricing. Compared with using natural genes, gene synthesis provides a good opportunity to optimize gene sequence for specific applications. In order to facilitate gene optimization, we have developed a stand-alone software called Visual Gene Developer. Results The software not only provides general functions for gene analysis and optimization along with an interactive user-friendly interfac...

  2. The site-directed mutation I(L177)H in Rhodobacter sphaeroides reaction center affects coordination of P(A) and B(B) bacteriochlorophylls.

    Science.gov (United States)

    Vasilieva, L G; Fufina, T Y; Gabdulkhakov, A G; Leonova, M M; Khatypov, R A; Shuvalov, V A

    2012-08-01

    To explore the influence of the I(L177)H single mutation on the properties of the nearest bacteriochlorophylls (BChls), three reaction centers (RCs) bearing double mutations were constructed in the photosynthetic purple bacterium Rhodobacter sphaeroides, and their properties and pigment content were compared with those of the correspondent single mutant RCs. Each pair of the mutations comprised the amino acid substitution I(L177)H and another mutation altering histidine ligand of BChl P(A) or BChl B(B). Contrary to expectations, the double mutation I(L177)H+H(L173)L does not bring about a heterodimer RC but causes a 46nm blue shift of the long-wavelength P absorbance band. The histidine L177 or a water molecule were suggested as putative ligands for P(A) in the RC I(L177)H+H(L173)L although this would imply a reorientation of the His backbone and additional rearrangements in the primary donor environment or even a repositioning of the BChl dimer. The crystal structure of the mutant I(L177)H reaction center determined to a resolution of 2.9Å shows changes at the interface region between the BChl P(A) and the monomeric BChl B(B). Spectral and pigment analysis provided evidence for β-coordination of the BChl B(B) in the double mutant RC I(L177)H+H(M182)L and for its hexacoordination in the mutant reaction center I(L177)H. Computer modeling suggests involvement of two water molecules in the β-coordination of the BChl B(B). Possible structural consequences of the L177 mutation affecting the coordination of the two BChls P(A) and B(B) are discussed. This article is part of a Special Issue entitled: Photosynthesis Research for Sustainability: from Natural to Artificial. PMID:22365928

  3. Transgenesis of humanized fat1 promotes n-3 polyunsaturated fatty acid synthesis and expression of genes involved in lipid metabolism in goat cells.

    Science.gov (United States)

    Fan, Yixuan; Ren, Caifang; Wang, Zhibo; Jia, Ruoxin; Wang, Dan; Zhang, Yanli; Zhang, Guomin; Wan, Yongjie; Huang, Mingrui; Wang, Feng

    2016-01-15

    The n-3 fatty acid desaturase gene fat1 codes for the n-3 desaturase enzyme, which can convert n-6 polyunsaturated fatty acids (PUFAs) to n-3 PUFAs. The n-3 PUFAs are essential components required for normal cellular function and have preventive and therapeutic effects on many diseases. Goat is an important domestic animal for human consumption of meat and milk. To elevate the concentrations of n-3 PUFAs and examine the regulatory mechanism of fat1 in PUFA metabolism in goat cells, we successfully constructed a humanized fat1 expression vector and confirmed the efficient expression of fat1 in goat ear skin-derived fibroblast cells (GEFCs) by qRT-PCR and Western blot analysis. Fatty acid analysis showed that fat1 overexpression significantly increased the levels of total n-3 PUFAs and decreased the levels of total n-6 PUFAs in GEFCs. In addition, qRT-PCR results indicate that the FADS1 and FADS2 desaturase genes, ELOV2 and ELOV5 elongase genes, ACO and CPT1 oxidation genes, and PPARa and PPARγ transcription factors are up-regulated, and transcription factors of SREBP-1c gene are down-regulated in the fat1 transgenic goat cells. Overall, fat1-overexpression resulted in an increase in the n-3 fatty acids and altered expression of PUFA synthesis related genes in GEFCs. This work lays a foundation for both the production of fat1 transgenic goats and further study of the mechanism of fat1 function in the PUFAs metabolism. PMID:26474750

  4. PEI-PEG-Chitosan Copolymer Coated Iron Oxide Nanoparticles for Safe Gene Delivery: synthesis, complexation, and transfection**

    OpenAIRE

    Kievit, Forrest M.; Veiseh, Omid; Bhattarai, Narayan; Fang, Chen; Gunn, Jonathan W.; Lee, Donghoon; Ellenbogen, Richard G.; Olson, James M.; Zhang, Miqin

    2009-01-01

    Gene therapy offers the potential of mediating disease through modification of specific cellular functions of target cells. However, effective transport of nucleic acids to target cells with minimal side effects remains a challenge despite the use of unique viral and non-viral delivery approaches. Here we present a non-viral nanoparticle gene carrier that demonstrates effective gene delivery and transfection both in vitro and in vivo. The nanoparticle system (NP-CP-PEI) is made of a superpara...

  5. Translational enhancement of recombinant protein synthesis in transgenic silkworms by a 5'-untranslated region of polyhedrin gene of Bombyx mori Nucleopolyhedrovirus.

    Science.gov (United States)

    Iizuka, Masashi; Tomita, Masahiro; Shimizu, Katsuhiko; Kikuchi, Yutaka; Yoshizato, Katsutoshi

    2008-06-01

    Previously, we established a method to produce recombinant proteins (r-proteins) in cocoons of germline transgenic silkworms, and showed that a step(s) in post-transcription processes was rate-limiting in obtaining a high yield of r-proteins. In this study, we examined whether the 5'-untranslated region (5'-UTR) of the polyhedrin gene (pol) of nucleopolyhedrovirus (NPV) has a translational enhancer activity in the r-protein expression by middle silk gland (MSG) cells of silkworm Bombyx mori (Bm). Sericin 1 gene (ser1) promoter-driven transformation vectors were constructed in which pol5'-UTRs of NPVs isolated from four different species, Bm, Spodoptera frugiperda, Ectropis oblique, and Malacosoma neustria, were each placed upstream of a reporter gene. Transient expression assays in MSGs showed that these pol5'-UTRs all enhanced the protein expression of reporter genes, and the pol5'-UTR of Bm NPV (pol5'-UTR/Bm) was the most effective among them. Thus, transgenic silkworms were generated, which bore the ser1 promoter-driven His-tagged secretory EGFP (sEGFP-His) gene under the control of pol5'-UTR/Bm. The synthesis of sEGFP-His proteins in MSGs of the transgenic worms was approximately 1.5-fold higher than that in those bearing null vectors. However, its mRNA expression levels were 67% of the control worms, indicating that the pol5'-UTR/Bm specifically enhanced the translational level. In conclusion, pol5'-UTR/Bm increased the yield of r-protein production in transgenic silkworms by enhancing the translational activity and this 5'-UTR could be useful for the mass production of r-proteins in germline transgenic silkworms. PMID:18640598

  6. Map-Based Cloning of Seed Dormancy1-2 Identified a Gibberellin Synthesis Gene Regulating the Development of Endosperm-Imposed Dormancy in Rice.

    Science.gov (United States)

    Ye, Heng; Feng, Jiuhuan; Zhang, Lihua; Zhang, Jinfeng; Mispan, Muhamad S; Cao, Zhuanqin; Beighley, Donn H; Yang, Jianchang; Gu, Xing-You

    2015-11-01

    Natural variation in seed dormancy is controlled by multiple genes mapped as quantitative trait loci in major crop or model plants. This research aimed to clone and characterize the Seed Dormancy1-2 (qSD1-2) locus associated with endosperm-imposed dormancy and plant height in rice (Oryza sativa). qSD1-2 was delimited to a 20-kb region, which contains OsGA20ox2 and had an additive effect on germination. Naturally occurring or induced loss-of-function mutations of the gibberellin (GA) synthesis gene enhanced seed dormancy and also reduced plant height. Expression of this gene in seeds (including endospermic cells) during early development increased GA accumulation to promote tissue morphogenesis and maturation programs. The mutant allele prevalent in semidwarf cultivars reduced the seed GA content by up to 2-fold at the early stage, which decelerated tissue morphogenesis including endosperm cell differentiation, delayed abscisic acid accumulation by a shift in the temporal distribution pattern, and postponed dehydration, physiological maturity, and germinability development. As the endosperm of developing seeds dominates the moisture equilibrium and desiccation status of the embryo in cereal crops, qSD1-2 is proposed to control primary dormancy by a GA-regulated dehydration mechanism. Allelic distribution of OsGA20ox2, the rice Green Revolution gene, was associated with the indica and japonica subspeciation. However, this research provided no evidence that the primitive indica- and common japonica-specific alleles at the presumably domestication-related locus functionally differentiate in plant height and seed dormancy. Thus, the evolutionary mechanism of this agriculturally important gene remains open for discussion. PMID:26373662

  7. Mutant huntingtin activates Nrf2-responsive genes and impairs dopamine synthesis in a PC12 model of Huntington's disease

    Directory of Open Access Journals (Sweden)

    den Dunnen Johan T

    2008-10-01

    Full Text Available Abstract Background Huntington's disease is a progressive autosomal dominant neurodegenerative disorder that is caused by a CAG repeat expansion in the HD or Huntington's disease gene. Although micro array studies on patient and animal tissue provide valuable information, the primary effect of mutant huntingtin will inevitably be masked by secondary processes in advanced stages of the disease. Thus, cell models are instrumental to study early, direct effects of mutant huntingtin. mRNA changes were studied in an inducible PC12 model of Huntington's disease, before and after aggregates became visible, to identify groups of genes that could play a role in the early pathology of Huntington's disease. Results Before aggregation, up-regulation of gene expression predominated, while after aggregates became visible, down-regulation and up-regulation occurred to the same extent. After aggregates became visible there was a down-regulation of dopamine biosynthesis genes accompanied by down-regulation of dopamine levels in culture, indicating the utility of this model to identify functionally relevant pathways. Furthermore, genes of the anti-oxidant Nrf2-ARE pathway were up-regulated, possibly as a protective mechanism. In parallel, we discovered alterations in genes which may result in increased oxidative stress and damage. Conclusion Up-regulation of gene expression may be more important in HD pathology than previously appreciated. In addition, given the pathogenic impact of oxidative stress and neuroinflammation, the Nrf2-ARE signaling pathway constitutes a new attractive therapeutic target for HD.

  8. Metabolic consequences of knocking out UGT85B1, the gene encoding the glucosyltransferase required for synthesis of dhurrin in Sorghum bicolor (L. Moench).

    Science.gov (United States)

    Blomstedt, Cecilia K; O'Donnell, Natalie H; Bjarnholt, Nanna; Neale, Alan D; Hamill, John D; Møller, Birger Lindberg; Gleadow, Roslyn M

    2016-02-01

    Many important food crops produce cyanogenic glucosides as natural defense compounds to protect against herbivory or pathogen attack. It has also been suggested that these nitrogen-based secondary metabolites act as storage reserves of nitrogen. In sorghum, three key genes, CYP79A1, CYP71E1 and UGT85B1, encode two Cytochrome P450s and a glycosyltransferase, respectively, the enzymes essential for synthesis of the cyanogenic glucoside dhurrin. Here, we report the use of targeted induced local lesions in genomes (TILLING) to identify a line with a mutation resulting in a premature stop codon in the N-terminal region of UGT85B1. Plants homozygous for this mutation do not produce dhurrin and are designated tcd2 (totally cyanide deficient 2) mutants. They have reduced vigor, being dwarfed, with poor root development and low fertility. Analysis using liquid chromatography-mass spectrometry (LC-MS) shows that tcd2 mutants accumulate numerous dhurrin pathway-derived metabolites, some of which are similar to those observed in transgenic Arabidopsis expressing the CYP79A1 and CYP71E1 genes. Our results demonstrate that UGT85B1 is essential for formation of dhurrin in sorghum with no co-expressed endogenous UDP-glucosyltransferases able to replace it. The tcd2 mutant suffers from self-intoxication because sorghum does not have a feedback mechanism to inhibit the initial steps of dhurrin biosynthesis when the glucosyltransferase activity required to complete the synthesis of dhurrin is lacking. The LC-MS analyses also revealed the presence of metabolites in the tcd2 mutant which have been suggested to be derived from dhurrin via endogenous pathways for nitrogen recovery, thus indicating which enzymes may be involved in such pathways. PMID:26493517

  9. Design of a minimal polypeptide unit for bacteriochlorophyll binding and self-assembly based on photosynthetic bacterial light-harvesting proteins.

    Science.gov (United States)

    Noy, Dror; Dutton, P Leslie

    2006-02-21

    We introduce LH1beta24, a minimal 24 amino acid polypeptide that binds and assembles bacteriochlorophylls (BChls) in micelles of octyl beta-glucoside (OG) into complexes with spectral properties that resemble those of B820, a universal intermediate in the assembly of native purple bacterial light-harvesting complexes (LHs). LH1beta24 was designed by a survey of sequences and crystal structures of bacterial LH proteins from different organisms combined with currently available information from in vitro reconstitution studies and genetically modified LHs in vivo. We took as a template for the design sphbeta31, a truncated 31 amino acid analogue of the native beta-apoprotein from the core LH complex of Rhodobacter sphaeroides. This peptide self-assembles with BChls to form B820 and, upon cooling and lowering OG concentration, forms red-shifted B850 spectral species that are considered analogous to native LH complexes. We find that LH1beta24 self-assembles with BChl in OG to form homodimeric B820-type subunits comprising two LH1beta24 and two BChl molecules per subunit. We demonstrate, by modeling the structure using the highly homologous structure of LH2 from Rhodospirillum molischianum, that it has the minimal size for BChl binding. Additionally, we have compared the self-assembly of sphbeta31 and LH1beta24 with BChls and discovered that the association enthalpies and entropies of both species are similar to those measured for native LH1 from Rhodospirillum rubrum. However, sphbeta31 readily aggregates into intermediate higher oligomeric species and further to form B850 species; moreover, the assembly process of these oligomers is not reversible, and they are apparently large nonspecific BChl-peptide coaggregates rather than well-defined nativelike LH complexes. Similar aggregates were observed during LH1beta24 assembly, but these were formed less readily and required lower temperatures than sphbeta31. In view of these results, we reevaluate previous in vitro

  10. Identification of the genes affecting the regulation of riboflavin synthesis in the flavinogenic yeast Pichia guilliermondii using insertion mutagenesis

    OpenAIRE

    Boretsky, Yuriy R.; Pynyaha, Yuriy V.; Boretsky, Volodymyr Y.; Fedorovych, Dariya V.; Fayura, Lyubov R.; Protchenko, Olha; Philpott, Caroline C.; Andriy A Sibirny

    2011-01-01

    Pichia guilliermondii is a representative of a group of so-called flavinogenic yeast species that overproduce riboflavin (vitamin B2) in response to iron limitation. Using insertion mutagenesis, we isolated P. guilliermondii mutants overproducing riboflavin. Analysis of nucleotide sequence of recombination sites revealed that insertion cassettes integrated into the genome disrupting P. guilliermondii genes similar to the VMA1 gene of Ashbya gossypii and Saccharomyces cerevisiae and FES1 and F...

  11. Odoriferous Defensive stink gland transcriptome to identify novel genes necessary for quinone synthesis in the red flour beetle, Tribolium castaneum.

    Directory of Open Access Journals (Sweden)

    Jianwei Li

    Full Text Available Chemical defense is one of the most important traits, which endow insects the ability to conquer a most diverse set of ecological environments. Chemical secretions are used for defense against anything from vertebrate or invertebrate predators to prokaryotic or eukaryotic parasites or food competitors. Tenebrionid beetles are especially prolific in this category, producing several varieties of substituted benzoquinone compounds. In order to get a better understanding of the genetic and molecular basis of defensive secretions, we performed RNA sequencing in a newly emerging insect model, the red flour beetle Tribolium castaneum (Coleoptera: Tenebrionidae. To detect genes that are highly and specifically expressed in the odoriferous gland tissues that secret defensive chemical compounds, we compared them to a control tissue, the anterior abdomen. 511 genes were identified in different subtraction groups. Of these, 77 genes were functionally analyzed by RNA interference (RNAi to recognize induced gland alterations morphologically or changes in gland volatiles by gas chromatography-mass spectrometry. 29 genes (38% presented strong visible phenotypes, while 67 genes (87% showed alterations of at least one gland content. Three of these genes showing quinone-less (ql phenotypes - Tcas-ql VTGl; Tcas-ql ARSB; Tcas-ql MRP - were isolated, molecularly characterized, their expression identified in both types of the secretory glandular cells, and their function determined by quantification of all main components after RNAi. In addition, microbe inhibition assays revealed that a quinone-free status is unable to impede bacterial or fungal growth. Phylogenetic analyses of these three genes indicate that they have evolved independently and specifically for chemical defense in beetles.

  12. Genome Sequencing of Xanthomonas vasicola Pathovar vasculorum Reveals Variation in Plasmids and Genes Encoding Lipopolysaccharide Synthesis, Type-IV Pilus and Type-III Secretion Effectors

    Directory of Open Access Journals (Sweden)

    Arthur Wasukira

    2014-03-01

    Full Text Available Xanthomonas vasicola pathovar vasculorum (Xvv is the bacterial agent causing gumming disease in sugarcane. Here, we compare complete genome sequences for five isolates of Xvv originating from sugarcane and one from maize. This identified two distinct types of lipopolysaccharide synthesis gene clusters among Xvv isolates: one is similar to that of Xanthomonas axonopodis pathovar citri (Xac and is probably the ancestral type, while the other is similar to those of the sugarcane-inhabiting species, Xanthomonas sacchari. Four of six Xvv isolates harboured sequences similar to the Xac plasmid, pXAC47, and showed a distinct Type-IV pilus (T4P sequence type, whereas the T4P locus of the other two isolates resembled that of the closely related banana pathogen, Xanthomonas campestris pathovar musacearum (Xcm. The Xvv isolate from maize has lost a gene encoding a homologue of the virulence effector, xopAF, which was present in all five of the sugarcane isolates, while xopL contained a premature stop codon in four out of six isolates. These findings shed new light on evolutionary events since the divergence of Xvv and Xcm, as well as further elucidating the relationships between the two closely related pathogens.

  13. A var gene upstream element controls protein synthesis at the level of translation initiation in Plasmodium falciparum.

    Science.gov (United States)

    Brancucci, Nicolas M B; Witmer, Kathrin; Schmid, Christoph; Voss, Till S

    2014-01-01

    Clonally variant protein expression in the malaria parasite Plasmodium falciparum generates phenotypic variability and allows isogenic populations to adapt to environmental changes encountered during blood stage infection. The underlying regulatory mechanisms are best studied for the major virulence factor P. falciparum erythrocyte membrane protein 1 (PfEMP1). PfEMP1 is encoded by the multicopy var gene family and only a single variant is expressed in individual parasites, a concept known as mutual exclusion or singular gene choice. var gene activation occurs in situ and is achieved through the escape of one locus from epigenetic silencing. Singular gene choice is controlled at the level of transcription initiation and var 5' upstream (ups) sequences harbour regulatory information essential for mutually exclusive transcription as well as for the trans-generational inheritance of the var activity profile. An additional level of control has recently been identified for the var2csa gene, where an mRNA element in the 5' untranslated region (5' UTR) is involved in the reversible inhibition of translation of var2csa transcripts. Here, we extend the knowledge on post-transcriptional var gene regulation to the common upsC type. We identified a 5' UTR sequence that inhibits translation of upsC-derived mRNAs. Importantly, this 5' UTR element efficiently inhibits translation even in the context of a heterologous upstream region. Further, we found var 5' UTRs to be significantly enriched in uAUGs which are known to impair the efficiency of protein translation in other eukaryotes. Our findings suggest that regulation at the post-transcriptional level is a common feature in the control of PfEMP1 expression in P. falciparum. PMID:24937593

  14. A var gene upstream element controls protein synthesis at the level of translation initiation in Plasmodium falciparum.

    Directory of Open Access Journals (Sweden)

    Nicolas M B Brancucci

    Full Text Available Clonally variant protein expression in the malaria parasite Plasmodium falciparum generates phenotypic variability and allows isogenic populations to adapt to environmental changes encountered during blood stage infection. The underlying regulatory mechanisms are best studied for the major virulence factor P. falciparum erythrocyte membrane protein 1 (PfEMP1. PfEMP1 is encoded by the multicopy var gene family and only a single variant is expressed in individual parasites, a concept known as mutual exclusion or singular gene choice. var gene activation occurs in situ and is achieved through the escape of one locus from epigenetic silencing. Singular gene choice is controlled at the level of transcription initiation and var 5' upstream (ups sequences harbour regulatory information essential for mutually exclusive transcription as well as for the trans-generational inheritance of the var activity profile. An additional level of control has recently been identified for the var2csa gene, where an mRNA element in the 5' untranslated region (5' UTR is involved in the reversible inhibition of translation of var2csa transcripts. Here, we extend the knowledge on post-transcriptional var gene regulation to the common upsC type. We identified a 5' UTR sequence that inhibits translation of upsC-derived mRNAs. Importantly, this 5' UTR element efficiently inhibits translation even in the context of a heterologous upstream region. Further, we found var 5' UTRs to be significantly enriched in uAUGs which are known to impair the efficiency of protein translation in other eukaryotes. Our findings suggest that regulation at the post-transcriptional level is a common feature in the control of PfEMP1 expression in P. falciparum.

  15. Gene expression

    International Nuclear Information System (INIS)

    We prepared probes for isolating functional pieces of the metallothionein locus. The probes enabled a variety of experiments, eventually revealing two mechanisms for metallothionein gene expression, the order of the DNA coding units at the locus, and the location of the gene site in its chromosome. Once the switch regulating metallothionein synthesis was located, it could be joined by recombinant DNA methods to other, unrelated genes, then reintroduced into cells by gene-transfer techniques. The expression of these recombinant genes could then be induced by exposing the cells to Zn2+ or Cd2+. We would thus take advantage of the clearly defined switching properties of the metallothionein gene to manipulate the expression of other, perhaps normally constitutive, genes. Already, despite an incomplete understanding of how the regulatory switch of the metallothionein locus operates, such experiments have been performed successfully

  16. Synthesis of 2'-deoxy-2'-[18F]-fluoro-5-iodo-1-β-D-arabinofuranosyluracil ([18F]-FIAU) and micro-PET imaging of suicide gene expression in tumor-bearing nude mice

    International Nuclear Information System (INIS)

    Herpes simplex virus type-1 thymidine kinase (HSV1-tk) is being used as a suicide gene for gene therapy of cancer. An in vivo method to assess the HSV1-tk enzyme activity after gene transfer is desirable to monitor gene expression as an indicator of gene delivery. Imaging of the HSV1-tk reporter gene along with various reporter probes is of current interest. We originally developed [18F]-FHPG and [18F]-FHBG for PET imaging of HSV1-tk gene expression and demonstrated that [18F]-FHBG is more useful than [18F]-FHPG for this purpose. [124I]-FIAU has been shown to be a potential PET imaging agent for HSV1-tk gene expression, and is superior to [18F]-FHPG and [18F]-FHBG. We also demonstrated that radiolabeled FMAU can be used as a marker for HSV-tk gene expression, and is superior to [18F]-FHPG and [18F]-FHBG. Earlier we reported a synthesis for 2'-deoxy-2'-[18F]fluoro-5-methyl-1-β-D-arabinofuranosyluracil ([18F]-FMAU) and some other 5-substituted nucleosides. We have synthesized now [18F]-FIAU, used the tracer for micro-PET imaging of suicide gene expression in tumor-bearing nude mice, and compared the results with earlier studies using [14C]-FMAU. (orig.)

  17. The Phytohormone Ethylene Enhances Cellulose Production, Regulates CRP/FNRKx Transcription and Causes Differential Gene Expression within the Bacterial Cellulose Synthesis Operon of Komagataeibacter (Gluconacetobacter) xylinus ATCC 53582

    Science.gov (United States)

    Augimeri, Richard V.; Strap, Janice L.

    2015-01-01

    Komagataeibacter (formerly Gluconacetobacter) xylinus ATCC 53582 is a plant-associated model organism for bacterial cellulose (BC) biosynthesis. This bacterium inhabits the carposphere where it interacts with fruit through the bi-directional transfer of phytohormones. The majority of research regarding K. xylinus has been focused on identifying and characterizing structural and regulatory factors that control BC biosynthesis, but its ecophysiology has been generally overlooked. Ethylene is a phytohormone that regulates plant development in a variety of ways, but is most commonly known for its positive role on fruit ripening. In this study, we utilized ethephon (2-chloroethylphosphonic acid) to produce in situ ethylene to investigate the effects of this phytohormone on BC production and the expression of genes known to be involved in K. xylinus BC biosynthesis (bcsA, bcsB, bcsC, bcsD, cmcAx, ccpAx and bglAx). Using pellicle assays and reverse transcription quantitative polymerase chain reaction (RT-qPCR), we demonstrate that ethephon-derived ethylene enhances BC directly in K. xylinus by up-regulating the expression of bcsA and bcsB, and indirectly though the up-regulation of cmcAx, ccpAx, and bglAx. We confirm that IAA directly decreases BC biosynthesis by showing that IAA down-regulates bcsA expression. Similarly, we confirm that ABA indirectly influences BC biosynthesis by showing it does not affect the expression of bcs operon genes. In addition, we are the first to report the ethylene and indole-3-acetic acid (IAA) induced differential expression of genes within the bacterial cellulose synthesis (bcs) operon. Using bioinformatics we have identified a novel phytohormone-regulated CRP/FNRKx transcription factor and provide evidence that it influences BC biosynthesis in K. xylinus. Lastly, utilizing current and previous data, we propose a model for the phytohormone-mediated fruit-bacteria interactions that K. xylinus experiences in nature. PMID:26733991

  18. The phytohormone ethylene enhances bacterial cellulose production, regulates CRP/FNRKx transcription and causes differential gene expression within the cellulose synthesis operon of Komagataeibacter (Gluconacetobacter xylinus ATCC 53582

    Directory of Open Access Journals (Sweden)

    Richard Vincent Augimeri

    2015-12-01

    Full Text Available Komagataeibacter (formerly Gluconacetobacter xylinus ATCC 53582 is a plant-associated model organism for bacterial cellulose (BC biosynthesis. This bacterium inhabits the carposphere where it interacts with fruit through the bi-directional transfer of phytohormones. The majority of research regarding K. xylinus has been focused on identifying and characterizing structural and regulatory factors that control BC biosynthesis, but its ecophysiology has been generally overlooked. Ethylene is a phytohormone that regulates plant development in a variety of ways, but is most commonly known for its positive role on fruit ripening. In this study, we utilized ethephon (2-chloroethylphosphonic acid to produce in situ ethylene to investigate the effects of this phytohormone on BC production and the expression of genes known to be involved in K. xylinus BC biosynthesis (bcsA, bcsB, bcsC, bcsD, cmcAx, ccpAx and bglAx. Using pellicle assays and reverse transcription quantitative polymerase chain reaction (RT-qPCR, we demonstrate that ethephon-derived ethylene enhances BC directly in K. xylinus by up-regulating the expression of bcsA and bcsB, and indirectly though the up-regulation of cmcAx, ccpAx and bglAx. We confirm that IAA directly decreases BC biosynthesis by showing that IAA down-regulates bcsA expression. Similarly, we confirm that ABA indirectly influences BC biosynthesis by showing it does not affect the expression of bcs operon genes. In addition, we are the first to report the ethylene and indole-3-acetic acid (IAA induced differential expression of genes within the bacterial cellulose synthesis (bcs operon. Using bioinformatics we have identified a novel phytohormone-regulated CRP/FNRKx transcription factor and provide evidence that it influences BC biosynthesis in K. xylinus. Lastly, utilizing current and previous data, we propose a model for the phytohormone-mediated fruit-bacteria interactions that K. xylinus experiences in nature.

  19. The Phytohormone Ethylene Enhances Cellulose Production, Regulates CRP/FNRKx Transcription and Causes Differential Gene Expression within the Bacterial Cellulose Synthesis Operon of Komagataeibacter (Gluconacetobacter) xylinus ATCC 53582.

    Science.gov (United States)

    Augimeri, Richard V; Strap, Janice L

    2015-01-01

    Komagataeibacter (formerly Gluconacetobacter) xylinus ATCC 53582 is a plant-associated model organism for bacterial cellulose (BC) biosynthesis. This bacterium inhabits the carposphere where it interacts with fruit through the bi-directional transfer of phytohormones. The majority of research regarding K. xylinus has been focused on identifying and characterizing structural and regulatory factors that control BC biosynthesis, but its ecophysiology has been generally overlooked. Ethylene is a phytohormone that regulates plant development in a variety of ways, but is most commonly known for its positive role on fruit ripening. In this study, we utilized ethephon (2-chloroethylphosphonic acid) to produce in situ ethylene to investigate the effects of this phytohormone on BC production and the expression of genes known to be involved in K. xylinus BC biosynthesis (bcsA, bcsB, bcsC, bcsD, cmcAx, ccpAx and bglAx). Using pellicle assays and reverse transcription quantitative polymerase chain reaction (RT-qPCR), we demonstrate that ethephon-derived ethylene enhances BC directly in K. xylinus by up-regulating the expression of bcsA and bcsB, and indirectly though the up-regulation of cmcAx, ccpAx, and bglAx. We confirm that IAA directly decreases BC biosynthesis by showing that IAA down-regulates bcsA expression. Similarly, we confirm that ABA indirectly influences BC biosynthesis by showing it does not affect the expression of bcs operon genes. In addition, we are the first to report the ethylene and indole-3-acetic acid (IAA) induced differential expression of genes within the bacterial cellulose synthesis (bcs) operon. Using bioinformatics we have identified a novel phytohormone-regulated CRP/FNRKx transcription factor and provide evidence that it influences BC biosynthesis in K. xylinus. Lastly, utilizing current and previous data, we propose a model for the phytohormone-mediated fruit-bacteria interactions that K. xylinus experiences in nature. PMID:26733991

  20. Heterologous Gene Expression in Lactococcus lactis subsp. lactis : Synthesis, Secretion, and Processing of the Bacillus subtilis Neutral Protease

    NARCIS (Netherlands)

    Guchte, Maarten van de; Kodde, Jan; Vossen, Jos M.B.M. van der; Kok, Jan; Venema, Gerard

    1990-01-01

    The Bacillus subtilis nprE gene lacking its own promoter sequence was inserted in the lactococcal expression vector pMG36e. Upon introduction of the recombinant plasmid into Lactococcus lactis subsp. lactis strain MG1363, neutral protease activity could be visualized by the appearance of large clear

  1. The cobY gene of the archaeon Halobacterium sp. strain NRC-1 is required for de novo cobamide synthesis.

    Science.gov (United States)

    Woodson, J D; Peck, R F; Krebs, M P; Escalante-Semerena, J C

    2003-01-01

    Genetic and nutritional analyses of mutants of the extremely halophilic archaeon Halobacterium sp. strain NRC-1 showed that open reading frame (ORF) Vng1581C encodes a protein with nucleoside triphosphate:adenosylcobinamide-phosphate nucleotidyltransferase enzyme activity. This activity was previously associated with the cobY gene of the methanogenic archaeon Methanobacterium thermoautotrophicum strain DeltaH, but no evidence was obtained to demonstrate the direct involvement of this protein in cobamide biosynthesis in archaea. Computer analysis of the Halobacterium sp. strain NRC-1 ORF Vng1581C gene and the cobY gene of M. thermoautotrophicum strain DeltaH showed the primary amino acid sequence of the proteins encoded by these two genes to be 35% identical and 48% similar. A strain of Halobacterium sp. strain NRC-1 carrying a null allele of the cobY gene was auxotrophic for cobinamide-GDP, a known intermediate of the late steps of cobamide biosynthesis. The auxotrophic requirement for cobinamide-GDP was corrected when a wild-type allele of cobY was introduced into the mutant strain, demonstrating that the lack of cobY function was solely responsible for the observed block in cobamide biosynthesis in this archaeon. The data also show that Halobacterium sp. strain NRC-1 possesses a high-affinity transport system for corrinoids and that this archaeon can synthesize cobamides de novo under aerobic growth conditions. To the best of our knowledge this is the first genetic and nutritional analysis of cobalamin biosynthetic mutants in archaea. PMID:12486068

  2. Identification of genes and pathways involved in the synthesis of Mead acid (20:3n-9), an indicator of essential fatty acid deficiency.

    Science.gov (United States)

    Ichi, Ikuyo; Kono, Nozomu; Arita, Yuka; Haga, Shizuka; Arisawa, Kotoko; Yamano, Misato; Nagase, Mana; Fujiwara, Yoko; Arai, Hiroyuki

    2014-01-01

    In mammals, 5,8,11-eicosatrienoic acid (Mead acid, 20:3n-9) is synthesized from oleic acid during a state of essential fatty acid deficiency (EFAD). Mead acid is thought to be produced by the same enzymes that synthesize arachidonic acid and eicosapentaenoic acid, but the genes and the pathways involved in the conversion of oleic acid to Mead acid have not been fully elucidated. The levels of polyunsaturated fatty acids in cultured cells are generally very low compared to those in mammalian tissues. In this study, we found that cultured cells, such as NIH3T3 and Hepa1-6 cells, have significant levels of Mead acid, indicating that cells in culture are in an EFAD state under normal culture conditions. We then examined the effect of siRNA-mediated knockdown of fatty acid desaturases and elongases on the level of Mead acid, and found that knockdown of Elovl5, Fads1, or Fads2 decreased the level of Mead acid. This and the measured levels of possible intermediate products for the synthesis of Mead acid such as 18:2n-9, 20:1n-9 and 20:2n-9 in the knocked down cells indicate two pathways for the synthesis of Mead acid: pathway 1) 18:1n-9→(Fads2)→18:2n-9→(Elovl5)→20:2n-9→(Fads1)→20:3n-9 and pathway 2) 18:1n-9→(Elovl5)→20:1n-9→(Fads2)→20:2n-9→(Fads1)→20:3n-9. PMID:24184513

  3. A var Gene Upstream Element Controls Protein Synthesis at the Level of Translation Initiation in Plasmodium falciparum

    OpenAIRE

    Brancucci, Nicolas M. B.; Kathrin Witmer; Christoph Schmid; Voss, Till S.

    2014-01-01

    Clonally variant protein expression in the malaria parasite Plasmodium falciparum generates phenotypic variability and allows isogenic populations to adapt to environmental changes encountered during blood stage infection. The underlying regulatory mechanisms are best studied for the major virulence factor P. falciparum erythrocyte membrane protein 1 (PfEMP1). PfEMP1 is encoded by the multicopy var gene family and only a single variant is expressed in individual parasites, a concept known as ...

  4. Retinoic acid response element in the human alcohol dehydrogenase gene ADH3: implications for regulation of retinoic acid synthesis.

    OpenAIRE

    Duester, G; Shean, M L; McBride, M S; Stewart, M J

    1991-01-01

    Retinoic acid regulation of one member of the human class I alcohol dehydrogenase (ADH) gene family was demonstrated, suggesting that the retinol dehydrogenase function of ADH may play a regulatory role in the biosynthetic pathway for retinoic acid. Promoter activity of human ADH3, but not ADH1 or ADH2, was shown to be activated by retinoic acid in transient transfection assays of Hep3B human hepatoma cells. Deletion mapping experiments identified a region in the ADH3 promoter located between...

  5. The cobY Gene of the Archaeon Halobacterium sp. Strain NRC-1 Is Required for De Novo Cobamide Synthesis

    OpenAIRE

    Woodson, J. D.; Peck, R. F.; Krebs, M P; Escalante-Semerena, J C

    2003-01-01

    Genetic and nutritional analyses of mutants of the extremely halophilic archaeon Halobacterium sp. strain NRC-1 showed that open reading frame (ORF) Vng1581C encodes a protein with nucleoside triphosphate:adenosylcobinamide-phosphate nucleotidyltransferase enzyme activity. This activity was previously associated with the cobY gene of the methanogenic archaeon Methanobacterium thermoautotrophicum strain ΔH, but no evidence was obtained to demonstrate the direct involvement of this protein in c...

  6. Amended Description of the Genes for Synthesis of Actinomyces naeslundii T14V Type 1 Fimbriae and Associated Adhesin▿ †

    OpenAIRE

    CHEN, PING; Cisar, John O.; Hess, Sonja; Ho, Jenny T. C.; Leung, Kai P.

    2007-01-01

    The type 1 fimbriae of Actinomyces naeslundii T14V mediate adhesion of this gram-positive species to the tooth surface. The present findings show that the locus for type 1 fimbria production in this strain includes three genes, fimQ for a minor fimbrial subunit that appears to be an adhesin, fimP for the major structural subunit, and srtC1 for a type 1 fimbria-specific sortase involved in the assembly of these structures.

  7. Glucan synthesis in the genus Lactobacillus: isolation and characterization of glucansucrase genes, enzymes and glucan products from six different strains

    OpenAIRE

    Kralj, S.; van Geel-Schutten, G.H.; Dondorff, M.M.G.; Kirsanovs, S.; van der Maarel, M.J.E.C.; Dijkhuizen, L.

    2004-01-01

    Members of the genera Streptococcus and Leuconostoc synthesize various α-glucans (dextran, alternan and mutan). In Lactobacillus, until now, the only glucosyltransferase (GTF) enzyme that has been characterized is gtfA of Lactobacillus reuteri 121, the first GTF enzyme synthesizing a glucan (reuteran) that contains mainly α-(1→4) linkages together with α-(1→6) and α-(1→4,6) linkages. Recently, partial sequences of glucansucrase genes were detected in other members of the genus Lactobacillus. ...

  8. Synthesis and evaluation of diethylethylamine–chitosan for gene delivery: composition effects on the in vitro transfection efficiency

    International Nuclear Information System (INIS)

    Chitosan has been indicated as a safe and promising polycation vector for gene delivery. However its low transfection efficiency has been a challenging obstacle for its application. To address this limitation, we synthesized chitosan derivatives which had increasing amounts of diethylethylamine groups (DEAE) attached to the chitosan main chain. The plasmid DNA VR1412 (pDNA), encoding the ß-galactosidase (ß-gal) reporter gene was used to prepare nanoparticles with the chitosan derivatives, and the transfection studies were performed with HeLa cells. By means of dynamic light scattering and zeta potential measurements, it was shown that diethylethylamine–chitosan derivatives (DEAEx–CH) were able to condense DNA into small particles having a surface charge depending on the polymer/DNA ratio (N/P ratio). Nanoparticles prepared with derivatives containing 15 and 25% of DEAE groups (DEAE15–CH and DEAE25–CH) exhibited transfection efficiencies ten times higher than that observed with deacetylated chitosan (CH). For derivatives with higher degrees of substitution (DS), transfection efficiency decreased. The most effective carriers showed low cytotoxicity and good transfection activities at low charge ratios (N/P). Vectors with low DS were easily degraded in the presence of lysozyme at physiological conditions in vitro and the nontoxicity displayed by these vectors opens up new opportunities in the design of DEAE–chitosan-based nanoparticles for gene delivery. (paper)

  9. 酵母菌合成酯类化合物关键酶基因的研究进展%Study on key enzyme genes of yeast for esters synthesis

    Institute of Scientific and Technical Information of China (English)

    庄世文; 付俊淑; 黄金海

    2012-01-01

    酯类化合物是发酵食品的重要香气成分之一,酵母发酵是酯类化合物产生的主要来源。在分析酵母菌酯类生物合成途径的基础上,对合成途径中的关键酶基因,包括脂肪酶/酯酶基因、醇酰基转移酶基因和醇脱氢酶基因及其酶作用进展作了综述。%Esters are one of the most important flavor compounds in fermented food and mainly synthesized by yeasts when fermenting.The key enzyme genes of yeast for esters synthesis, including lipase/esterase genes, alcohol acetyltransferase genes and alcohol dehydrogenase genes, based on the biosynthetic pathways of esters production were introduced.

  10. Identification of the genes affecting the regulation of riboflavin synthesis in the flavinogenic yeast Pichia guilliermondii using insertion mutagenesis.

    Science.gov (United States)

    Boretsky, Yuriy R; Pynyaha, Yuriy V; Boretsky, Volodymyr Y; Fedorovych, Dariya V; Fayura, Lyubov R; Protchenko, Olha; Philpott, Caroline C; Sibirny, Andriy A

    2011-05-01

    Pichia guilliermondii is a representative of a group of so-called flavinogenic yeast species that overproduce riboflavin (vitamin B(2)) in response to iron limitation. Using insertion mutagenesis, we isolated P. guilliermondii mutants overproducing riboflavin. Analysis of nucleotide sequence of recombination sites revealed that insertion cassettes integrated into the genome disrupting P. guilliermondii genes similar to the VMA1 gene of Ashbya gossypii and Saccharomyces cerevisiae and FES1 and FRA1 genes of S. cerevisiae. The constructed P. guilliermondiiΔvma1-17 mutant possessed five- to sevenfold elevated riboflavin production and twofold decreased iron cell content as compared with the parental strain. Pichia guilliermondiiΔfra1-45 mutant accumulated 1.8-2.2-fold more iron in the cells and produced five- to sevenfold more riboflavin as compared with the parental strain. Both Δvma1-17 and Δfes1-77 knockout strains could not grow at 37 °C in contrast to the wild-type strain and the Δfra1-45 mutant. Increased riboflavin production by the wild-type strain was observed at 37 °C. Although the Δfes1-77 mutant did not overproduce riboflavin, it showed partial complementation when crossed with previously isolated P. guilliermondii riboflavin-overproducing mutant rib80-22. Complementation analysis revealed that Δvma1-17 and Δfra1-45 mutants are distinct from previously reported riboflavin-producing mutants hit1-1, rib80-22 and rib81-31 of this yeast. PMID:21261808

  11. Synthesis of mesoporous SiO2–ZnO nanocapsules: encapsulation of small biomolecules for drugs and “SiOZO-plex” for gene delivery

    International Nuclear Information System (INIS)

    This work presents a new synthesis of mesoporous SiO2–ZnO composite nanocapsules with sizes of 90–150 nm and represents their applications in encapsulation of small biomolecules (fluorescent molecules, drugs, and DNA) for uses in medical biotechnology (e.g., drug and gene delivery) for the first time. The nanocapsule size and morphology have been confirmed through the HRSEM and HRTEM. The mesoporous structure of the novel materials has been confirmed through both BET and HRTEM, and the pore diameter observed to be ca. 2–8 nm with an average diameter of 5.1 nm. The BET surface area of mesoporous SiO2–ZnO was found to be ∼230 m2 g−1. Three different types of pores were detected through HRTEM: type-I, normal pores in silica matrix, pore with ZnO nanoparticles at the boundary (type-II) and type-III, the pores with tiny ZnO nanoparticles (∼5–7 nm) inside them. To demonstrate the biocompatibility and cell viability of the nanocapsules, normal and cancerous lymphocyte cells have been chosen and investigated in a systematic way. Fluorescent dye (Rhodamine 6G), anticancer drug e.g., Doxorubicin (DOX) were loaded in all types of pores, and EtBr-labeled DNA molecules were loaded efficiently into the mesopores of second and third types of the composite nanocapsules to manifest the characteristic of mesoporous, and to find out its loading efficacy. The release kinetics of Rhodamine 6G and DOX were studied. The results highlight the potential of novel functional mesoporous SiO2–ZnO nanoparticles for using as the carrier of drugs and formation of “SiOZO-plex”, a complex of mesoporous SiO2–ZnO with DNA for gene delivery applications.Graphical Abstract

  12. Synthesis of mesoporous SiO{sub 2}-ZnO nanocapsules: encapsulation of small biomolecules for drugs and 'SiOZO-plex' for gene delivery

    Energy Technology Data Exchange (ETDEWEB)

    Kumar, Vijay Bhooshan [School of Engineering Sciences and Technology, University of Hyderabad (India); Annamanedi, Madhavi [School of Life Sciences, University of Hyderabad, Department of Animal Sciences (India); Prashad, Muvva Durga [University of Hyderabad, Centre for Nanoscience and Nanotechnology (India); Arunasree, Kalle M. [School of Life Sciences, University of Hyderabad, Department of Animal Sciences (India); Mastai, Yitzhak; Gedanken, Aharon, E-mail: gedanken@mail.biu.ac.il [Bar-Ilan University, Department of Chemistry, Institute for Nanotechnology and Advanced Materials (Israel); Paik, Pradip, E-mail: ppse@uohyd.ernet.in [School of Engineering Sciences and Technology, University of Hyderabad (India)

    2013-09-15

    This work presents a new synthesis of mesoporous SiO{sub 2}-ZnO composite nanocapsules with sizes of 90-150 nm and represents their applications in encapsulation of small biomolecules (fluorescent molecules, drugs, and DNA) for uses in medical biotechnology (e.g., drug and gene delivery) for the first time. The nanocapsule size and morphology have been confirmed through the HRSEM and HRTEM. The mesoporous structure of the novel materials has been confirmed through both BET and HRTEM, and the pore diameter observed to be ca. 2-8 nm with an average diameter of 5.1 nm. The BET surface area of mesoporous SiO{sub 2}-ZnO was found to be {approx}230 m{sup 2} g{sup -1}. Three different types of pores were detected through HRTEM: type-I, normal pores in silica matrix, pore with ZnO nanoparticles at the boundary (type-II) and type-III, the pores with tiny ZnO nanoparticles ({approx}5-7 nm) inside them. To demonstrate the biocompatibility and cell viability of the nanocapsules, normal and cancerous lymphocyte cells have been chosen and investigated in a systematic way. Fluorescent dye (Rhodamine 6G), anticancer drug e.g., Doxorubicin (DOX) were loaded in all types of pores, and EtBr-labeled DNA molecules were loaded efficiently into the mesopores of second and third types of the composite nanocapsules to manifest the characteristic of mesoporous, and to find out its loading efficacy. The release kinetics of Rhodamine 6G and DOX were studied. The results highlight the potential of novel functional mesoporous SiO{sub 2}-ZnO nanoparticles for using as the carrier of drugs and formation of 'SiOZO-plex', a complex of mesoporous SiO{sub 2}-ZnO with DNA for gene delivery applications.Graphical Abstract.

  13. Knockdown of the coenzyme Q synthesis gene Smed-dlp1 affects planarian regeneration and tissue homeostasis.

    Science.gov (United States)

    Shiobara, Yumiko; Harada, Chiaki; Shiota, Takeshi; Sakamoto, Kimitoshi; Kita, Kiyoshi; Tanaka, Saeko; Tabata, Kenta; Sekie, Kiyoteru; Yamamoto, Yorihiro; Sugiyama, Tomoyasu

    2015-12-01

    The freshwater planarian is a model organism used to study tissue regeneration that occupies an important position among multicellular organisms. Planarian genomic databases have led to the identification of genes that are required for regeneration, with implications for their roles in its underlying mechanism. Coenzyme Q (CoQ) is a fundamental lipophilic molecule that is synthesized and expressed in every cell of every organism. Furthermore, CoQ levels affect development, life span, disease and aging in nematodes and mice. Because CoQ can be ingested in food, it has been used in preventive nutrition. In this study, we investigated the role of CoQ in planarian regeneration. Planarians synthesize both CoQ9 and rhodoquinone 9 (RQ9). Knockdown of Smed-dlp1, a trans-prenyltransferase gene that encodes an enzyme that synthesizes the CoQ side chain, led to a decrease in CoQ9 and RQ9 levels. However, ATP levels did not consistently decrease in these animals. Knockdown animals exhibited tissue regression and curling. The number of mitotic cells decreased in Smed-dlp1 (RNAi) animals. These results suggested a failure in physiological cell turnover and stem cell function. Accordingly, regenerating planarians died from lysis or exhibited delayed regeneration. Interestingly, the observed phenotypes were partially rescued by ingesting food supplemented with α-tocopherol. Taken together, our results suggest that oxidative stress induced by reduced CoQ9 levels affects planarian regeneration and tissue homeostasis. PMID:26516985

  14. A novel amidase from Brevibacterium epidermidis ZJB-07021: gene cloning, refolding and application in butyrylhydroxamic acid synthesis.

    Science.gov (United States)

    Ruan, Li-Tao; Zheng, Ren-Chao; Zheng, Yu-Guo

    2016-08-01

    A novel amidase gene (bami) was cloned from Brevibacterium epidermidis ZJB-07021 by combination of degenerate PCR and high-efficiency thermal asymmetric interlaced PCR (hiTAIL-PCR). The deduced amino acid sequence showed low identity (≤55 %) with other reported amidases. The bami gene was overexpressed in Escherichia coli, and the resultant inclusion bodies were refolded and purified to homogeneity with a recovery of 22.6 %. Bami exhibited a broad substrate spectrum towards aliphatic, aromatic and heterocyclic amides, and showed the highest acyl transfer activity towards butyramide with specific activity of 1331.0 ± 24.0 U mg(-1). Kinetic analysis demonstrated that purified Bami exhibited high catalytic efficiency (414.9 mM(-1) s(-1)) for acyl transfer of butyramide, with turnover number (K cat) of 3569.0 s(-1). Key parameters including pH, substrate/co-substrate concentration, reaction temperature and catalyst loading were investigated and the Bami showed maximum acyl transfer activity at 50 °C, pH 7.5. Enzymatic catalysis of 200 mM butyramide with 15 μg mL(-1) purified Bami was completed in 15 min with a BHA yield of 88.1 % under optimized conditions. The results demonstrated the great potential of Bami for the production of a variety of hydroxamic acids. PMID:27276936

  15. Glucan synthesis in the genus Lactobacillus: isolation and characterization of glucansucrase genes, enzymes and glucan products from six different strains.

    Science.gov (United States)

    Kralj, S; van Geel-Schutten, G H; Dondorff, M M G; Kirsanovs, S; van der Maarel, M J E C; Dijkhuizen, L

    2004-11-01

    Members of the genera Streptococcus and Leuconostoc synthesize various alpha-glucans (dextran, alternan and mutan). In Lactobacillus, until now, the only glucosyltransferase (GTF) enzyme that has been characterized is gtfA of Lactobacillus reuteri 121, the first GTF enzyme synthesizing a glucan (reuteran) that contains mainly alpha-(1-->4) linkages together with alpha-(1-->6) and alpha-(1-->4,6) linkages. Recently, partial sequences of glucansucrase genes were detected in other members of the genus Lactobacillus. This paper reports, for the first time, isolation and characterization of dextransucrase and mutansucrase genes and enzymes from various Lactobacillus species and the characterization of the glucan products synthesized, which mainly have alpha-(1-->6)- and alpha-(1-->3)-glucosidic linkages. The four GTF enzymes characterized from three different Lb. reuteri strains are highly similar at the amino acid level, and consequently their protein structures are very alike. Interestingly, these four Lb. reuteri GTFs have relatively large N-terminal variable regions, containing RDV repeats, and relatively short putative glucan-binding domains with conserved and less-conserved YG-repeating units. The three other GTF enzymes, isolated from Lactobacillus sakei, Lactobacillus fermentum and Lactobacillus parabuchneri, contain smaller variable regions and larger putative glucan-binding domains compared to the Lb. reuteri GTF enzymes. PMID:15528655

  16. Herbal compound 861 regulates mRNA expression of collagen synthesis- and degradation-related genes in human hepatic stellate cells

    Institute of Scientific and Technical Information of China (English)

    Lin Wang; Bao-En Wang; Jian Wang; Pei-Gen Xiao; Xue-Hai Tan

    2008-01-01

    AIM: To identify the role of herbal compound 861 (Cpd 861) in the regulation of mRNA expression of collagen synthesis- and degradation-related genes in human hepatic stellate cells (HSCs).METHODS: mRNA levels of collagen types I and III, matrix metalloproteinase 1 (MMP-1), matrix metalloproteinase 2 (MMP-2), membrane type-1 matrix metalloproteinase (MT1-MMP), tissue inhibitor of metalloproteinase 1 (TIMP-1), and transforming growth factor β1 (TGF-βi) in cultured-activated HSCs treated with Cpd 861 or interferon-γ (IFN-γ) were determined by real-time PCR.RESULTS: Both Cpd 861 and IFN-γ reduced the mRNA levels of collagen type Ⅲ, MMP-2 and TGF-βl. Moreover, Cpd 861 significantly enhanced the MMP-1 mRNA levels while down-regulated the TIMP-1 mRNA expression, increasing the ratio of MMP-1 to TIMP-1 to (6.3 + 0.3)-fold compared to the control group.CONCLUSION: The anti-fibrosis function of Cpd 861 may be mediated by both decreased interstitial collagen sythesis by inhibiting the transcription of collagen type in and TGF-pi and increased degradation of these collagens by up-regulating MMP-1 and down-regulating TIMP-1 mRNA levels.

  17. Homogeneous synthesis of quaternized chitin in NaOH/urea aqueous solution as a potential gene vector.

    Science.gov (United States)

    Peng, Na; Ai, Ziye; Fang, Zehong; Wang, Yanfeng; Xia, Zhiping; Zhong, Zibiao; Fan, Xiaoli; Ye, Qifa

    2016-10-01

    Water-soluble quaternized chitins (QCs) were homogeneously synthesized by reacting chitin with (3-chloro-2-hydroxypropyl) trimethylammonium chloride (CHPTAC) in 8wt% NaOH/4wt% urea aqueous solutions. The chemical structure and solution properties of the quaternized chitins were characterized by (1)H NMR, FT-IR, elemental analysis, dynamic light scattering (DLS) and zeta potential measurements. The results demonstrated that the water-soluble QCs, with a degree of substitution (DS) values of 0.27-0.54, could be obtained by varying the concentration of chitin, the molar ratio of CHPTAC to chitin unit, and the reaction time at room temperature (25°C). Two QCs (DS=0.36 and 0.54) were selected and studied as gene carriers. Agarose gel retardation assay revealed that both QCs could condense DNA efficiently when N/P ratio>3. The results of particle size and zeta potential indicated that both QCs had a good ability of condensing plasmid DNA into compact nanoparticles with the size of 100-200nm and zeta potential of +18 to +36mV. Compared to polyethylenimine (PEI, 25kDa), the QCs exhibited outstanding low cytotoxicity. Transfection efficiencies of the QCs/DNA complexes were measured using pGL-3 encoding luciferase as the foreign DNA, and the QCs/DNA complexes showed effective transfection efficiencies in 293T cells. These results revealed that the QCs prepared in NaOH/urea aqueous solutions could be used as promising non-viral gene carriers owing to their excellent characteristics. PMID:27312628

  18. Synthesis of Mannosylated Polyethylenimine and Its Potential Application as Cell-Targeting Non-Viral Vector for Gene Therapy

    Directory of Open Access Journals (Sweden)

    Ying Hu

    2014-10-01

    Full Text Available Mannose polyethylenimine with a molecular weight of 25 k (Man-PEI25k was synthesized via a phenylisothiocyanate bridge using mannopyranosylphenyl isothiocyanate as a coupling reagent, and characterized by 1H NMR (nuclear magnetic resonance and FT-IR (Fourier transform infrared spectroscopy analysis. Spherical nanoparticles were formed with diameters of 80–250 nm when the copolymer was mixed with DNA at various charge ratios of copolymer/DNA (N/P. Gel electrophoresis demonstrated that the DNA had been condensed and retained by the PEI derivates at low N/P ratios. The Man-PEI25k/DNA complexes were less cytotoxic than the PEI complexes with a molecular weight of 25 k (PEI25k at the same N/P ratio. Laser scan confocal microscopy and flow cytometry confirmed that the Man-PEI25k/DNA complexes gave higher cell uptake efficiency in (Dendritic cells DC2.4 cells than HeLa cells. The transfection efficiency of Man-PEI25k was higher than that of PEI25k towards DC2.4 cells. These results indicated that Man-PEI25k could be used as a potential DC-targeting non-viral vector for gene therapy.

  19. Improved ethyl caproate production of Chinese liquor yeast by overexpressing fatty acid synthesis genes with OPI1 deletion.

    Science.gov (United States)

    Chen, Yefu; Luo, Weiwei; Gong, Rui; Xue, Xingxiang; Guan, Xiangyu; Song, Lulu; Guo, Xuewu; Xiao, Dongguang

    2016-09-01

    During yeast fermentation, ethyl esters play a key role in the development of the flavor profiles of Chinese liquor. Ethyl caproate, an ethyl ester eliciting apple-like flavor, is the characteristic flavor of strong aromatic liquor, which is the best selling liquor in China. In the traditional fermentation process, ethyl caproate is mainly produced at the later fermentation stage by aroma-producing yeast, bacteria, and mold in a mud pit instead of Saccharomyces cerevisiae at the expense of grains and fermentation time. To improve the production of ethyl caproate by Chinese liquor yeast (S. cerevisiae) with less food consumption and shorter fermentation time, we constructed three recombinant strains, namely, α5-ACC1ΔOPI1, α5-FAS1ΔOPI1, and α5-FAS2ΔOPI1 by overexpressing acetyl-CoA carboxylase (ACC1), fatty acid synthase 1 (FAS1), and fatty acid synthase 2 (FAS2) with OPI1 (an inositol/choline-mediated negative regulatory gene) deletion, respectively. In the liquid fermentation of corn hydrolysate, the contents of ethyl caproate produced by α5-ACC1ΔOPI1, α5-FAS1ΔOPI1, and α5-FAS2ΔOPI1 increased by 0.40-, 1.75-, and 0.31-fold, correspondingly, compared with the initial strain α5. The contents of other fatty acid ethyl esters (FAEEs) (C8:0, C10:0, C12:0) also increased. In comparison, the content of FAEEs produced by α5-FAS1ΔOPI1 significantly improved. Meanwhile, the contents of acetyl-CoA and ethyl acetate were enhanced by α5-FAS1ΔOPI1. Overall, this study offers a promising platform for the development of pure yeast culture fermentation of Chinese strong aromatic liquor without the use of a mud pit. PMID:27344573

  20. Gene synthesis, bacterial expression, and 1H NMR spectroscopic studies of the rat outer mitochondrial membrane cytochrome b5.

    Science.gov (United States)

    Rivera, M; Barillas-Mury, C; Christensen, K A; Little, J W; Wells, M A; Walker, F A

    1992-12-01

    The gene coding for the water-soluble domain of the outer mitochondrial membrane cytochrome b5 (OM cytochrome b5) from rat liver has been synthetized and expressed in Escherichia coli. The DNA sequence was obtained by back-translating the known amino acid sequence [Lederer, F., Ghrir, R., Guiard, B., Cortial, S., & Ito, A. (1983) Eur. J. Biochem. 132, 95-102]. The recombinant OM cytochrome b5 was characterized by UV-visible, EPR, and 1H NMR spectroscopy. The UV-visible and EPR spectra of the OM cytochrome b5 are almost identical to the ones obtained from the overexpressed rat microsomal cytochrome b5 [Bodman, S. B. V., Schyler, M. A., Jollie, D. R., & Sligar, S. G. (1986) Proc. Natl. Acad. Sci. U.S.A. 83, 9443-9447]. The one-dimensional 1H NMR spectrum of the OM cytochrome b5 indicates that the rhombic perturbation of the ferric center is essentially identical to that in the microsomal beef, rabbit, chicken, and rat cytochromes b5. Two-dimensional 1H NMR spectroscopy (NOESY) and one-dimensional NOE difference spectroscopy were used to assign the contact-shifted resonances that correspond to each of the two isomers that result from the rotation of the heme around its alpha-gamma-meso axis. The assignment of the resonances allowed the determination of the heme orientation ratio in the OM cytochrome b5, which was found to be 1.0 +/- 0.1. It is noteworthy that the two cytochromes b5 that have similar populations of the two heme isomers (large heme disorder) originate from the rat liver. PMID:1333795

  1. Synthesis and Characterization of Cationic Glycidyl-Based Poly(aminoester-Folic Acid Targeting Conjugates and Study on Gene Delivery

    Directory of Open Access Journals (Sweden)

    Yu Che Hsiao

    2012-07-01

    Full Text Available A new poly(aminoester (EPAE-FA containing folic acid and amino groups in the backbone and side chain was synthesized. EPAE-FA self-assembled readily with the plasmid DNA (pCMV-βgal in HEPES buffer and was characterized by dynamic light scattering, zeta potential, fluorescence images, and XTT cell viability assays. To evaluate the transfection effect of graft ratio of FA on the EPAE system, EPAE-FA polymers with two different graft ratios (EPAE-FA12k and EPAE-FA14k were also prepared. This study found that all EPAE-FA polymers were able to bind plasmid DNA and yielded positively charged complexes with nano-sized particles ( < 200 nm. To assess the transfection efficiency mediated by EPAE and EPAE-FA polymers, we performed in vitro transfection activity assays using FR-negative (COS-7 and FR-positive (HeLa cells. The EPAE-FA12k/DNA and EPAE-FA14k/DNA complexes were able to transfect HeLa cell in vitro with higher transfection efficiency than PEI25k/DNA at the similar weight ratio. These results demonstrated that the introduction of FA into EPAE system had a significant effect on transferring ability for FR-positive cells (HeLa. Examination of the cytotoxicity of PEI25k and EPAE-FA system revealed that EPAE-FA system had lower cytotoxicity. In this paper, EPAE-FA seemed to be a novel cationic poly(aminoester for gene delivery and an interesting candidate for further study.

  2. The Reaumuria trigyna leucoanthocyanidin dioxygenase (RtLDOX) gene complements anthocyanidin synthesis and increases the salt tolerance potential of a transgenic Arabidopsis LDOX mutant.

    Science.gov (United States)

    Zhang, Huirong; Du, Chao; Wang, Yan; Wang, Jia; Zheng, Linlin; Wang, Yingchun

    2016-09-01

    Reaumuria trigyna is a typical, native desert halophyte that grows under extreme conditions in Inner Mongolia. In a previous transcriptomic profiling analysis, flavonoid pathway-related genes in R. trigyna showed significant differences in transcript abundance under salt stress. Leucoanthocyanidin dioxygenase (LDOX, EC 1.14.11.19) is one of three dioxygenases in the flavonoid pathway that catalyzes the formation of anthocyanidins from leucoanthocyanidins. In this study, we cloned the full-length cDNA of R. trigyna LDOX (RtLDOX), and found RtLDOX recombinant protein was able to replace flavanone-3-hydroxylase (F3H, EC 1.14.11.9), another dioxygenase in the flavonoid pathway, to convert naringenin to dihydrokaempferol in vitro. R. trigyna LDOX can complement the Arabidopsis LDOX mutant transparent testa11 (tt11-11), which has reduced proanthocyanin (PA) and anthocyanin levels in seeds, to accumulate these two compounds. Thus, RtLDOX acts as a multifunctional dioxygenase to effect the synthesis of PA and anthocyanins and can perform F3H dioxygenase activities in the flavonoid biosynthesis pathway. The RtLDOX promoter harbored many cis-acting elements that might be recognized and bound by transcription factors related to stress response. RtLDOX expression was strongly increased under salt stress, and RtLDOX transgenic Arabidopsis mutant under NaCl stress accumulated the content of flavonoids leading to an increased antioxidant activities and plant biomass. These results suggest that RtLDOX as a multifunctional dioxygenase in flavonoid biosynthesis involves in enhancing plant response to NaCl stress. PMID:27219053

  3. Translocator protein (Tspo) gene promoter-driven green fluorescent protein synthesis in transgenic mice: an in vivo model to study Tspo transcription.

    Science.gov (United States)

    Wang, Hui-Jie; Fan, Jinjiang; Papadopoulos, Vassilios

    2012-11-01

    Translocator protein (TSPO), previously known as the peripheral-type benzodiazepine receptor, is a ubiquitous drug- and cholesterol-binding protein primarily found in the outer mitochondrial membrane as part of a mitochondrial cholesterol transport complex. TSPO is present at higher levels in steroid-synthesizing and rapidly proliferating tissues and its biological role has been mainly linked to mitochondrial function, steroidogenesis and cell proliferation/apoptosis. Aberrant TSPO levels have been linked to multiple diseases, including cancer, endocrine disorders, brain injury, neurodegeneration, ischemia-reperfusion injury and inflammatory diseases. Investigation of the functions of this protein in vitro and in vivo have been mainly carried out using high-affinity drug ligands, such as isoquinoline carboxamides and benzodiazepines and more recently, gene silencing methods. To establish a model to study the regulation of Tspo transcription in vivo, we generated a transgenic mouse model expressing green fluorescent protein (GFP) from Aequorea coerulescens under control of the Tspo promoter region (Tspo-AcGFP). The expression profiles of Tspo-AcGFP, endogenous TSPO and Tspo mRNA were found to be well-correlated. Tspo-AcGFP synthesis in the transgenic mice was seen in almost every tissue examined and as with TSPO in wild-type mice, Tspo-AcGFP was highly expressed in steroidogenic cells of the endocrine and reproductive systems, epithelial cells of the digestive system, skeletal muscle and other organs. In summary, this transgenic Tspo-AcGFP mouse model recapitulates endogenous Tspo expression patterns and could be a useful, tractable tool for monitoring the transcriptional regulation and function of Tspo in live animal experiments. PMID:22868914

  4. Nitrophenols isolated from diesel exhaust particles regulate steroidogenic gene expression and steroid synthesis in the human H295R adrenocortical cell line

    International Nuclear Information System (INIS)

    Studies of nitrophenols isolated from diesel exhaust particles (DEPs), 3-methyl-4-nitrophenol (PNMC) and 4-nitro-3-phenylphenol (PNMPP) have revealed that these chemicals possess estrogenic and anti-androgenic activity in vitro and in vivo and that PNMC accumulate in adrenal glands in vivo. However, the impacts of exposure to these compounds on adrenal endocrine disruption and steroidogenesis have not been investigated. To elucidate the non-receptor mediated effects of PNMC and PNMPP, we investigated the production of the steroid hormones progesterone, cortisol, testosterone, and estradiol-17β and modulation of nine major enzyme genes involved in the synthesis of steroid hormones (CYP11A, CYP11B1, CYP17, CYP19, 17βHSD1, 17βHSD4, CYP21, 3βHSD2, StAR) in human adrenal H295R cells supplied with cAMP. Exposure to 10-7 to 10-5 M PNMC and 1 mM 8-Br-cAMP for 48 h decreased testosterone, cortisol, and estradiol-17β levels and increased progesterone secretion. At 10-5 M, PNMC with 1 mM 8-Br-cAMP significantly stimulated expression of the 17βHSD4 and significantly suppressed expression of 3βHSD2. In comparison, 10-7 to 2 x 10-5 M PNMPP with 1 mM 8-Br-cAMP for 48 h decreased concentrations of estradiol-17β, increased progesterone levels, but did not affect testosterone and cortisol secretion due to the significant suppression of CYP17 and the non-significant but obvious suppression of CYP19. Our results clarified steroidogenic enzymes as candidates responsible for the inhibition or stimulation for the production of steroid hormones in the steroidogenic pathway, thus providing the first experimental evidence for multiple mechanisms of disruption of endocrine pathways by these nitrophenols

  5. Synthesis of stocks and phenotypic effects of dwarf and bantam sex-linked major genes in egg-type chickens

    Directory of Open Access Journals (Sweden)

    Randolfo William Silvestre Custódio

    1997-09-01

    Full Text Available The present study describes the production of stocks segregating dwarf (dw, bantam (dwB and normal (dw+ alleles, as well as the characters, shank length, adult body weight, age at sexual maturity and egg production. Heterozygous K dw+/k dwB sires were mated to normal (dw+ dams to produce stock D6.a, and mated to dwB females to produce stock D6.b. Stock D4.a came from mating F1 heterozygous dwB dw sires to dwarf Leghorns. In a third series of matings, 7/8 Sebright and 1/8 dw-Leghorn dwB dw sires were crossed to three groups of dams of different genotypes. The progeny of the normal (dw+, dwarf (dw, and bantam (dwB dams were designated as stocks D4.b, D4.c and D4.d, respectively. The dw+ dams were White Leghorn strain cross females. The difference between the rate of laying of normal (69.7% and their bantam sisters (68.6% was not statistically significant when the average 32-week body weight of the dw+ sisters was 1,897 g. However, when the 32-week body weight of the normal daughters from the same sires and smaller dams was around 1,646 g, the difference between the rate of laying of the normal (78.1% and their bantam sisters (75.9% was significant (P Neste trabalho descreve-se a obtenção de seis plantéis experimentais, segregantes para os alelos ligados ao sexo, dwarf (dw, bantam (dwB e normal (dw+, em galinhas para ovos. Comparações entre desempenhos entre plantéis hemizigóticos para genes maiores no locus dwarf também foram efetuadas com relação a comprimento de canela medida na 16ª semana de idade, peso do corpo adulto, produção de ovos e maturidade sexual. Machos heterozigotos (K dw+/ k dwB foram acasalados com fêmeas normais (dw+ e com fêmeas bantam (dwB. As progênies fêmeas, nos referidos acasalamentos, foram denominadas de D6.a e D6.b, respectivamente. Galos 7/8 Sebright e 1/8 dw-Leghorn heterozigotos (dwB dw foram acasalados com três grupos de galinhas de diferentes genótipos (dw+, dw e dwB. As progênies das m

  6. Effect of triclosan, triclocarban, 2,2',4,4'-tetrabromodiphenyl ether, and bisphenol A on the iodide uptake, thyroid peroxidase activity, and expression of genes involved in thyroid hormone synthesis.

    Science.gov (United States)

    Wu, Yuanfeng; Beland, Frederick A; Fang, Jia-Long

    2016-04-01

    Triclosan, triclocarban, 2,2',4,4'-tetrabromodiphenyl ether (BDE-47), and bisphenol A (BPA) have been reported to disturb thyroid hormone (TH) homeostasis. We have examined the effects of these chemicals on sodium/iodide symporter (NIS)-mediated iodide uptake and the expression of genes involved in TH synthesis in rat thyroid follicular FRTL-5 cells, and on the activity of thyroid peroxidase (TPO) using rat thyroid microsomes. All four chemicals inhibited NIS-mediated iodide uptake in a concentration-dependent manner. A decrease in the iodide uptake was also observed in the absence of sodium iodide. Kinetic studies showed that all four chemicals were non-competitive inhibitors of NIS, with the order of Ki values being triclosantriclocarbantriclocarban. BDE-47 decreased the level of Tpo, while BPA altered the expression of all six genes. Triclosan and triclocarban inhibited the activity of TPO at 166 and >300 μM, respectively. Neither BDE-47 nor BPA affected TPO activity. In conclusion, triclosan, triclocarban, BDE-47, and BPA inhibited iodide uptake, but had differential effects on the expression of TH synthesis-related genes and the activity of TPO. PMID:26827900

  7. Histamine enhances interleukin (IL)-1-induced IL-1 gene expression and protein synthesis via H2 receptors in peripheral blood mononuclear cells. Comparison with IL-1 receptor antagonist.

    OpenAIRE

    Vannier, E; Dinarello, C A

    1993-01-01

    Histamine and IL-1 have been implicated in the pathogenesis of chronic inflammatory diseases, such as pulmonary allergic reactions and rheumatoid arthritis. We therefore investigated whether histamine modulated the synthesis of IL-1 beta. Human PBMC were stimulated with IL-1 alpha (10 ng/ml) in the absence or presence of histamine (10(-9)-10(-4) M). Histamine alone did not induce protein synthesis or mRNA accumulation for IL-1 beta. IL-1 alpha-induced IL-1 beta synthesis was enhanced two to t...

  8. Roles of the relA+ gene and of 4-thiouridine in near-ultraviolet (334nm) radiation inhibition of induced synthesis of tryptophanase in Escherichia coli B/r

    International Nuclear Information System (INIS)

    Near-ultraviolet radiation (near UV; 300-380 nm) is known to inhibit the induced synthesis of tryptophanase by tryptophan in Escherichia coli, showing an action spectrum similar to that for near-UV-induced growth delay. It has now been shown that a relA mutant of E. coli B/r exhibits 50% as much monochromatic near-UV (334 nm) inhibition of tryptophanase induction as the wild type, and that a mutant lacking 4-thiouridine, an unusual nucleoside in tRNA, exhibits + gene product; growth delay appears not to be primarily involved. (author)

  9. Two-step stimulation of B lymphocytes to enter DNA synthesis: synergy between anti-immunoglobulin antibody and cytochalasin on expression of c-myc and a G1-specific gene.

    OpenAIRE

    Buckler, A J; Rothstein, T. L.; Sonenshein, G E

    1988-01-01

    Previously we demonstrated that stimulation of resting murine splenic B lymphocytes with goat anti-mouse immunoglobulin antibody (GaMIg) plus cytochalasin D (CD) led to DNA synthesis; GaMIg and CD added simultaneously, or GaMIg added before CD, induced this response (T. L. Rothstein, J. Immunol. 136:813-816, 1986). Cells similarly treated with GaMIg or CD alone did not enter S phase. Here we have measured the effects of this two-signal stimulation on the c-myc, 2F1, and gamma-actin genes. The...

  10. Total synthesis and expression of a gene for the alpha-subunit of bovine rod outer segment guanine nucleotide-binding protein (transducin).

    OpenAIRE

    Sakmar, T P; Khorana, H G

    1988-01-01

    To facilitate structure-function studies by site-specific mutagenesis, we have synthesized a gene for the alpha-subunit of the bovine rod outer segment (ROS) guanine nucleotide-binding protein (transducin). The gene codes for the native amino acid sequence and contains, by design, 38 unique restriction sites which are uniformly spaced. This enables mutagenesis in any part of the gene by restriction fragment replacement. The gene is 1076 base pairs in length. It was constructed from 44 synthet...

  11. Single base mutation in the proα2(I) collagen gene that causes efficient splicing of RNA from exon 27 to exon 29 and synthesis of a shortened but in-frame proα2(I) chain

    International Nuclear Information System (INIS)

    Previous observations demonstrated that a lethal variant of osteogenesis imperfecta had two altered alleles for proα2(I) chains of type I procollagen. One mutation produced a nonfunctioning allele in that there was synthesis of mRNA but no detectable synthesis of proα2(I) chains from the allele. The mutation in the other allele caused synthesis of shortened proα2(I) chains that lacked most or all of the 18 amino acids encoded by exon 28. Subclones of the proα2(I) gene were prepared from the proband's DNA and the DNA sequence was determined for a 582-base-pair (bp) region that extended from the last 30 bp of intervening sequence 26 to the first 26 bp of intervening sequence 29. Data from six independent subclones demonstrated that all had the same sequence as a previously isolated normal clone for the proα2(I) gene except that four subclones had a single base mutation at the 3' end of intervening sequence 27. The mutation was a substitution of guanine for adenine that changed the universal consensus sequence for the 3' splicing site of RNA from -AG- to -GG-. S1 nuclease experiments demonstrated that about half the proα2(I) mRNA in the proband's fibroblasts was abnormally spliced and that the major species of abnormal proα2(I) mRNA was completely spliced from the last codon of exon 27 to the first codon of exon 29. The mutation is apparently unique among RNA splicing mutations of mammalian systems in producing a shortened polypeptide chain that is in-frame in terms of coding sequences, that is used in the subunit assembly of a protein, and that contributes to a lethal phenotype

  12. [Insertional mutation in the AZOBR_p60120 gene is accompanied by defects in the synthesis of lipopolysaccharide and calcofluor-binding polysaccharides in the bacterium Azospirillum brasilense Sp245].

    Science.gov (United States)

    Katsy, E I; Prilipov, A G

    2015-03-01

    In the bacterium Azospirillum brasilense Sp245, extracellular calcofluor-binding polysaccharides (Cal+ phenotype) and two types of lipopolysaccharides, LPSI and LPSII, were previously identified. These lipopolysaccharides share the same repeating O-polysaccharide unit but have different antigenic structures and different charges of their O-polysaccharides and/or core oligosaccharides. Several dozens of predicted genes involved in the biosynthesis of polysaccharides have been localized in the AZOBR_p6 plasmid of strain Sp245 (GenBank accession no. HE577333). In the present work, it was demonstrated that an artificial transposon Omegon-Km had inserted into the central region of the AZOBR_p60120 gene in the A. brasilense Sp245 LPSI- Cal- KM252 mutant. In A. brasilense strain Sp245, this plasmid gene encodes a putative glycosyltransferase containing conserved domains characteristic of the enzymes participating in the synthesis of O-polysaccharides and capsular polysaccharides (accession no. YP004987664). In mutant KM252, a respective predicted protein is expected to be completely inactivated. As a result of the analysis of the EcoRI fragment of the AZOBR_p6 plasmid, encompassing the AZOBR_p60120 gene and a number of other loci, novel data on the structure of AZOBR_p6 were obtained: an approximately 5-kb gap (GenBank accession no. KM189439) was closed in the nucleotide sequence of this plasmid. PMID:26027369

  13. Colistin-resistant, lipopolysaccharide-deficient Acinetobacter baumannii responds to lipopolysaccharide loss through increased expression of genes involved in the synthesis and transport of lipoproteins, phospholipids, and poly-β-1,6-N-acetylglucosamine.

    Science.gov (United States)

    Henry, Rebekah; Vithanage, Nuwan; Harrison, Paul; Seemann, Torsten; Coutts, Scott; Moffatt, Jennifer H; Nation, Roger L; Li, Jian; Harper, Marina; Adler, Ben; Boyce, John D

    2012-01-01

    We recently demonstrated that colistin resistance in Acinetobacter baumannii can result from mutational inactivation of genes essential for lipid A biosynthesis (Moffatt JH, et al., Antimicrob. Agents Chemother. 54:4971-4977). Consequently, strains harboring these mutations are unable to produce the major Gram-negative bacterial surface component, lipopolysaccharide (LPS). To understand how A. baumannii compensates for the lack of LPS, we compared the transcriptional profile of the A. baumannii type strain ATCC 19606 to that of an isogenic, LPS-deficient, lpxA mutant strain. The analysis of the expression profiles indicated that the LPS-deficient strain showed increased expression of many genes involved in cell envelope and membrane biogenesis. In particular, upregulated genes included those involved in the Lol lipoprotein transport system and the Mla-retrograde phospholipid transport system. In addition, genes involved in the synthesis and transport of poly-β-1,6-N-acetylglucosamine (PNAG) also were upregulated, and a corresponding increase in PNAG production was observed. The LPS-deficient strain also exhibited the reduced expression of genes predicted to encode the fimbrial subunit FimA and a type VI secretion system (T6SS). The reduced expression of genes involved in T6SS correlated with the detection of the T6SS-effector protein AssC in culture supernatants of the A. baumannii wild-type strain but not in the LPS-deficient strain. Taken together, these data show that, in response to total LPS loss, A. baumannii alters the expression of critical transport and biosynthesis systems associated with modulating the composition and structure of the bacterial surface. PMID:22024825

  14. Fundamental studies on the synthesis, characterization, stabilization, 3-D scaffolds, and trafficking mechanisms of nano-structured calcium phosphates (NanoCaPs) for non-viral gene delivery

    Science.gov (United States)

    Olton, Dana

    Non-viral transfer of therapeutic genes into mammalian cells represents a potentially viable approach to (1) treat and cure acute and chronic genetically transferred congenital disorders and to (2) aid in tissue regeneration. Non-viral vectors have been praised for their potential to circumvent some of the limitations associated with viral vectors including immunogenicity, cytotoxicity and insertional mutagenesis. Among the various types of non-viral gene delivery vectors, nano-structured ceramic particles, particularly, particles of calcium phosphate (CaP) remain an attractive option because of their safety, biocompatibility, biodegradability, ease of handling as well as their adsorptive capacity for DNA. CaP-DNA complexes have been used in vitro since the 1970s and have recently been tested in vivo. However, despite CaPs' extensive use, concerns still remain regarding the synthesis and colloidal instability of this vector. Also, towards the development of a more efficient gene delivery agent, there is a need to understand the mechanisms involved in both the cellular uptake as well as in the subsequent intracellular processing of CaP-DNA complexes. Moreover, although significant advances have been made in the synthesis and design of tissue engineered constructs, the development of a safe, effective scaffold has yet to be realized. As such, the focus of this thesis has been to address these four concerns. In this work, we begin by presenting a novel aqueous-based approach to synthesize nano-particles of CaP (NanoCaPs). Our results show that this approach generates nano-crystalline hydroxyapatite particles. When tested in vitro, transfection of these complexes resulted in higher, more consistent levels of gene expression when compared to particles synthesized via manual mixing. The optimized forms of these particles both effectively bound (90% efficient) and condensed (70% efficient) plasmid DNA (pDNA) and possessed negative zeta potentials of approximately -20m

  15. Essential Bacillus subtilis genes

    DEFF Research Database (Denmark)

    Kobayashi, K.; Ehrlich, S.D.; Albertini, A.;

    2003-01-01

    To estimate the minimal gene set required to sustain bacterial life in nutritious conditions, we carried out a systematic inactivation of Bacillus subtilis genes. Among approximate to4,100 genes of the organism, only 192 were shown to be indispensable by this or previous work. Another 79 genes were...... predicted to be essential. The vast majority of essential genes were categorized in relatively few domains of cell metabolism, with about half involved in information processing, one-fifth involved in the synthesis of cell envelope and the determination of cell shape and division, and one-tenth related to...... cell energetics. Only 4% of essential genes encode unknown functions. Most essential genes are present throughout a wide range of Bacteria, and almost 70% can also be found in Archaea and Eucarya. However, essential genes related to cell envelope, shape, division, and respiration tend to be lost from...

  16. The Reacquisition of Biotin Prototrophy in Saccharomyces cerevisiae Involved Horizontal Gene Transfer, Gene Duplication and Gene Clustering

    OpenAIRE

    Hall, Charles; Dietrich, Fred S

    2007-01-01

    The synthesis of biotin, a vitamin required for many carboxylation reactions, is a variable trait in Saccharomyces cerevisiae. Many S. cerevisiae strains, including common laboratory strains, contain only a partial biotin synthesis pathway. We here report the identification of the first step necessary for the biotin synthesis pathway in S. cerevisiae. The biotin auxotroph strain S288c was able to grow on media lacking biotin when BIO1 and the known biotin synthesis gene BIO6 were introduced t...

  17. Suppression of a deletion mutation in the gene encoding essential PBP2b reveals a new lytic transglycosylase involved in peripheral peptidoglycan synthesis in Streptococcus pneumoniae D39.

    Science.gov (United States)

    Tsui, Ho-Ching Tiffany; Zheng, Jiaqi J; Magallon, Ariel N; Ryan, John D; Yunck, Rachel; Rued, Britta E; Bernhardt, Thomas G; Winkler, Malcolm E

    2016-06-01

    In ellipsoid-shaped ovococcus bacteria, such as the pathogen Streptococcus pneumoniae (pneumococcus), side-wall (peripheral) peptidoglycan (PG) synthesis emanates from midcells and is catalyzed by the essential class B penicillin-binding protein PBP2b transpeptidase (TP). We report that mutations that inactivate the pneumococcal YceG-domain protein, Spd_1346 (renamed MltG), remove the requirement for PBP2b. ΔmltG mutants in unencapsulated strains accumulate inactivation mutations of class A PBP1a, which possesses TP and transglycosylase (TG) activities. The 'synthetic viable' genetic relationship between Δpbp1a and ΔmltG mutations extends to essential ΔmreCD and ΔrodZ mutations that misregulate peripheral PG synthesis. Remarkably, the single MltG(Y488D) change suppresses the requirement for PBP2b, MreCD, RodZ and RodA. Structural modeling and comparisons, catalytic-site changes and an interspecies chimera indicate that pneumococcal MltG is the functional homologue of the recently reported MltG endo-lytic transglycosylase of Escherichia coli. Depletion of pneumococcal MltG or mltG(Y488D) increases sphericity of cells, and MltG localizes with peripheral PG synthesis proteins during division. Finally, growth of Δpbp1a ΔmltG or mltG(Y488D) mutants depends on induction of expression of the WalRK TCS regulon of PG hydrolases. These results fit a model in which MltG releases anchored PG glycan strands synthesized by PBP1a for crosslinking by a PBP2b:RodA complex in peripheral PG synthesis. PMID:26933838

  18. Gene activity during germination of spores of the fern, Onoclea sensibilis: RNA and protein synthesis and the role of stored mRNA

    Science.gov (United States)

    Raghavan, V.

    1991-01-01

    Pattern of 3H-uridine incorporation into RNA of spores of Onoclea sensibilis imbibed in complete darkness (non-germinating conditions) and induced to germinate in red light was followed by oligo-dT cellulose chromatography, gel electrophoresis coupled with fluorography and autoradiography. In dark-imbibed spores, RNA synthesis was initiated about 24 h after sowing, with most of the label accumulating in the high mol. wt. poly(A) -RNA fraction. There was no incorporation of the label into poly(A) +RNA until 48 h after sowing. In contrast, photo-induced spores began to synthesize all fractions of RNA within 12 h after sowing and by 24 h, incorporation of 3H-uridine into RNA of irradiated spores was nearly 70-fold higher than that into dark-imbibed spores. Protein synthesis, as monitored by 3H-arginine incorporation into the acid-insoluble fraction and by autoradiography, was initiated in spores within 1-2 h after sowing under both conditions. Autoradiographic experiments also showed that onset of protein synthesis in the cytoplasm of the germinating spore is independent of the transport of newly synthesized nuclear RNA. One-dimensional sodium dodecyl sulphate-polyacrylamide gel electrophoresis of 35S-methionine-labelled proteins revealed a good correspondence between proteins synthesized in a cell-free translation system directed by poly(A) +RNA of dormant spores and those synthesized in vivo by dark-imbibed and photo-induced spores. These results indicate that stored mRNAs of O. sensibilis spores are functionally competent and provide templates for the synthesis of proteins during dark-imbibition and germination.

  19. Expression of ABA synthesis and metabolism genes under different irrigation strategies and atmospheric VPDs is associated with stomatal conductance in grapevine (Vitis vinifera L. cv Cabernet Sauvignon)

    OpenAIRE

    Speirs, Jim; Binney, Allan; Collins, Marisa; Edwards, Everard; Loveys, Brian

    2013-01-01

    The influence of different levels of irrigation and of variation in atmospheric vapour pressure deficit (VPD) on the synthesis, metabolism, and transport of abscisic acid (ABA) and the effects on stomatal conductance were examined in field-grown Cabernet Sauvignon grapevines. Xylem sap, leaf tissue, and root tissue were collected at regular intervals during two seasons in conjunction with measurements of leaf water potential (Ψleaf) and stomatal conductance (gs). The different irrigation leve...

  20. The glutathione synthesis gene Gclm modulates amphiphilic polymer-coated CdSe/ZnS quantum dot-induced lung inflammation in mice.

    Directory of Open Access Journals (Sweden)

    Lisa A McConnachie

    Full Text Available Quantum dots (QDs are unique semi-conductor fluorescent nanoparticles with potential uses in a variety of biomedical applications. However, concerns exist regarding their potential toxicity, specifically their capacity to induce oxidative stress and inflammation. In this study we synthesized CdSe/ZnS core/shell QDs with a tri-n-octylphosphine oxide, poly(maleic anhydride-alt-1-tetradecene (TOPO-PMAT coating and assessed their effects on lung inflammation in mice. Previously published in vitro data demonstrated these TOPO-PMAT QDs cause oxidative stress resulting in increased expression of antioxidant proteins, including heme oxygenase, and the glutathione (GSH synthesis enzyme glutamate cysteine ligase (GCL. We therefore investigated the effects of these QDs in vivo in mice deficient in GSH synthesis (Gclm +/- and Gclm -/- mice. When mice were exposed via nasal instillation to a TOPO-PMAT QD dose of 6 µg cadmium (Cd equivalents/kg body weight, neutrophil counts in bronchoalveolar lavage fluid (BALF increased in both Gclm wild-type (+/+ and Gclm heterozygous (+/- mice, whereas Gclm null (-/- mice exhibited no such increase. Levels of the pro-inflammatory cytokines KC and TNFα increased in BALF from Gclm +/+ and +/- mice, but not from Gclm -/- mice. Analysis of lung Cd levels suggested that QDs were cleared more readily from the lungs of Gclm -/- mice. There was no change in matrix metalloproteinase (MMP activity in any of the mice. However, there was a decrease in whole lung myeloperoxidase (MPO content in Gclm -/- mice, regardless of treatment, relative to untreated Gclm +/+ mice. We conclude that in mice TOPO-PMAT QDs have in vivo pro-inflammatory properties, and the inflammatory response is dependent on GSH synthesis status. Because there is a common polymorphism in humans that influences GCLM expression, these findings imply that humans with reduced GSH synthesis capabilities may be more susceptible to the pro-inflammatory effects of QDs.

  1. Inhibitors of Fatty Acid Synthesis Induce PPAR α -Regulated Fatty Acid β -Oxidative Genes: Synergistic Roles of L-FABP and Glucose

    OpenAIRE

    Huan Huang; McIntosh, Avery L.; Martin, Gregory G.; Petrescu, Anca D.; Landrock, Kerstin K.; Danilo Landrock; Kier, Ann B.; Friedhelm Schroeder

    2013-01-01

    While TOFA (acetyl CoA carboxylase inhibitor) and C75 (fatty acid synthase inhibitor) prevent lipid accumulation by inhibiting fatty acid synthesis, the mechanism of action is not simply accounted for by inhibition of the enzymes alone. Liver fatty acid binding protein (L-FABP), a mediator of long chain fatty acid signaling to peroxisome proliferator-activated receptor-α (PPARα) in the nucleus, was found to bind TOFA and its activated CoA th...

  2. Influence of environmental factors on growth and pigment synthesis by purple thiobacteria

    Directory of Open Access Journals (Sweden)

    Y. О. Pavlova

    2007-12-01

    Full Text Available The influence of different environmental factors on growth and pigment biosynthesis by particular strains of purple thiobacteria was investigated. These strains belong to the genus Chromatium, Thiocystis, Thiocapsa and Lamprocystis and were isolated from Yavoriv sulphur mine. Calcium, magnesium, manganese, iron and sodium chloride should be included in the medium for optimal growth of these bacteria. Addition of these elements entails increasing the biomass production and synthesis of carotenoids and bacteriochlorophyll a. Initial concentration of inoculum and electron donor has essential influence on growth of purple thiobacteria. Early in the development of culture, sulphide was oxidized, and then the growth impairment and destruction of cells under exposure of light were observed. For the optimization of bacteria growth the electron donor (sulphide must be added many times during the cultivation process in the concentration, which is not exceed an inhibition dose. The additional bringing of the electron donor in the medium promotes the raise of cells’ biomass. The acetate introduction in the medium has positive influence on the pigments’ biosynthesis. The essential factor of growth and pigments’ biosynthesis is the light intensity. Peak gain of the culture growth was observed under 400 lx. The amplification of light exposure is accompanied by the decrease of growth and content of pigments in cells. Oxygen inhibits the synthesis of pigments in all strains

  3. Genes and Gene Therapy

    Science.gov (United States)

    ... correctly, a child can have a genetic disorder. Gene therapy is an experimental technique that uses genes to ... or prevent disease. The most common form of gene therapy involves inserting a normal gene to replace an ...

  4. In vitro and in vivo application of RNA interference for targeting genes involved in peritrophic matrix synthesis in a lepidopteran system

    Institute of Scientific and Technical Information of China (English)

    Umut Toprak; Doug Baldwin; Martin Erlandson; Cedric Gillott; Stephanie Harris; Dwayne D.Hegedus

    2013-01-01

    The midgut of most insects is lined with a semipermeable acellular tube,the peritrophic matrix(PM),composed of chitin and proteins.Although various genes encoding PM proteins have been characterized,our understanding of their roles in PM structure and function is very limited.One promising approach for obtaining functional information is RNA interference,which has been used to reduce the levels of specific mRNAs using double-stranded RNAs administered to larvae by either injection or feeding.Although this method is well documented in dipterans and coleopterans,reports of its success in lepidopterans are varied.In the current study,the silencing midgut genes encoding PM proteins(insect intestinal mucin 1,insect intestinal mucin 4,PM protein l)and the chitin biosynthetic or modifying enzymes(chitin synthase-B and chitin deacetylase 1)in a noctuid lepidopteran,Mamestra configurata,was examined in vitro and in vivo.In vitro studies in primary midgut epithelial cell preparations revealed an acute and rapid silencing(by 24 h)for the gene encoding chitin deacetylase 1 and a slower rate of silencing(by 72 h)for the gene encoding PM protein 1.Genes encoding insect intestinal mucins were slightly silenced by 72 h,whereas no silencing was detected for the gene encoding chitin synthase-B.In vivo experiments focused on chitin deacetylase 1,as the gene was silenced to the greatest extent in vitro.Continuous feeding of neonates and fourth instar larvae with double-stranded RNA resulted in silencing of chitin deacetylase 1 by 24 and 36 h,respectively.Feeding a single dose to neonates also resulted in silencing by 24 h.The current study demonstrates that genes encoding PM proteins can be silenced and outlines conditions for RNA interference by per os feeding in lepidopterans.

  5. The genes encoding the biotin carboxyl carrier protein and biotin carboxylase subunits of Bacillus subtilis acetyl coenzyme A carboxylase, the first enzyme of fatty acid synthesis.

    OpenAIRE

    P. Marini(GANIL); Li, S J; Gardiol, D; Cronan, J E; De Mendoza, D

    1995-01-01

    The genes encoding two subunits of acetyl coenzyme A carboxylase, biotin carboxyl carrier protein, and biotin carboxylase have been cloned from Bacillus subtilis. DNA sequencing and RNA blot hybridization studies indicated that the B. subtilis accB homolog which encodes biotin carboxyl carrier protein, is part of an operon that includes accC, the gene encoding the biotin carboxylase subunit of acetyl coenzyme A carboxylase.

  6. Expanding the modern synthesis.

    Science.gov (United States)

    Wallace, Rodrick

    2010-10-01

    The Modern Evolutionary Synthesis formalizes the role of variation, heredity, differential reproduction and mutation in population genetics. Here we explore a mathematical structure, based on the asymptotic limit theorems of communication theory, that instantiates the punctuated dynamic relations of organisms with their embedding environments, including the possibility of the transfer of heritage information between different classes of organism. The approach applies a standard coevolutionary argument to genes, environment, and gene expression reconfigured as interacting information sources. In essence, we provide something of a formal roadmap for the modernization of the Modern Synthesis, making applications to both relatively rapid evolutionary punctuated equilibrium and to the conservation of ecological interactions across deep evolutionary time. PMID:20965439

  7. Cloning and sequencing of hfq (host factor required for synthesis of bacteriophage Q beta RNA gene of Salmonella Typhimurium isolated from poultry

    Directory of Open Access Journals (Sweden)

    Parthasarathi Behera

    2015-05-01

    Full Text Available Aim: The aim was to clone and sequence hfq gene of Salmonella Typhimurium strain PM-45 and compare its sequence with hfq gene of other serovar of Salmonella. Materials and Methods: Salmonella Typhimurium strain PM-45 was procured from the G. B. Pant University of Agriculture and Technology, Pantnagar, India. The genomic DNA was isolated from Salmonella Typhimurium. Hfq gene was polymerase chain reaction (PCR amplified from the DNA using specific primers, which was subsequently cloned into pET32a vector and transformed into Escherichia coli BL21 pLys cells. The recombinant plasmid was isolated and subjected to restriction enzyme digestion as well as PCR. The clone was then sequenced. The sequence was analyzed and submitted in GenBank. Results: PCR produced an amplicon of 309 bp. Restriction digestion of the recombinant plasmid released the desired insert. The hfq sequence shows 100% homology with similar sequences from other Salmonella Typhimurium isolates. Both nucleotide and amino acid sequences are highly conserved. The submitted sequence is having Genbank accession no KM998764. Conclusion: Hfq, the hexameric RNA binding protein is one of the most important post-transcriptional regulator of bacteria. The sequence of hfq gene of Salmonella Typhimurium is highly conserved within and between Salmonella enterica serovars. This gene sequence is probably under heavy selection pressure to maintain the conformational integrity of its product in spite of its being not a survival gene.

  8. Comparative transcriptional profiling of melatonin synthesis and catabolic genes indicates the possible role of melatonin in developmental and stress responses in rice

    Directory of Open Access Journals (Sweden)

    Yunxie eWei

    2016-05-01

    Full Text Available As a well-known animal hormone, melatonin (N-acetyl-5-methoxytryptamine is also involved in multiple plant biological processes, especially in various stress responses. Rice is one of the most important crops, and melatonin is taken in by many people everyday from rice. However, the transcriptional profiling of melatonin-related genes in rice is largely unknown. In this study, the expression patterns of 11 melatonin related genes in rice in different periods, tissues, in response to different treatments were synthetically analyzed using published microarray data. These results suggest that the melatonin-related genes may play important and dual roles in rice developmental stages. We highlight the commonly regulation of rice melatonin-related genes by abscisic acid (ABA, jasmonic acid (JA, various abiotic stresses and pathogen infection, indicating the possible role of these genes in multiple stress responses and underlying crosstalks of plant hormones, especially ABA and JA. Taken together, this study may provide insight into the association among melatonin biosynthesis and catabolic pathway, plant development and stress responses in rice. The profile analysis identified candidate genes for further functional characterization in circadian rhythm and specific stress responses.

  9. THE EXPERIENCE OF THE TRANSFORMATION OF SOME CULTIVATED PLANTS WITH THE GENE UGT ENCODING THE SYNTHESIS OF UDPG-TRANSFERASE IN ORDER TO CHANGE THE HORMONAL STATUS

    Directory of Open Access Journals (Sweden)

    Rekoslavskaya N.I.

    2012-08-01

    Full Text Available The gene ugt/iaglu was isolated from cDNA library obtained from seedlings of Zea mays L. Positive clones prepared by Lambda ZAPII (Stratagene, USA procedure were screened via western blot with antibodies to UDPG-transferase from corn endosperm raised in rabbit serum. The plasmid pBluescript harboring the gene ugt/iaglu was placed into Escherichia coli (E.coli DH5a under T7/T3 promoter. The gene ugt/iaglu was sequenced and the size was determined as much as 1740 bp. The UDPG-transferase or by trivial name Indoleacetic acid (IAA - glucose synthase (IAGlu-synthase binds IAA with glucose from UDPG thereby making the temporary inactivation and storing of this phytohormone which is capable to be released after the demand from cells. Several cultivated plants were used for transfromation with the gene ugt/iaglu from corn: tomato, potato, lettuce, egg-plant, pepper, strawberry, cucumber, squash, aspen, poplar, pine and others. All plants transformed with the gene ugt/iaglu showed fast growth, better flowering and harvest. The insertion and expression of the gene ugt/iaglu was confirmed in transformed tomato, potato and aspen with PCR, RT-PCR, southern and northern blottings. The contents of free IAA and its bound form IAGlu were higher as much as twice in tomato, potato and aspen transformed with the gene ugt/iaglu. The harvest of tomato was 3-4 times higher in transgenic tomato. The amount of potato tubers and their whole masses were 1.5 - 2 times higher in transgenic potato of several varieties in comparison to control.

  10. Post-irradiation replication and repair in UV-irradiated cells of Proteus mirabilis depends on protein synthesis and a functioning rec+ gene

    International Nuclear Information System (INIS)

    The amount of and the molecular weight of newly synthesized DNA (piDNA) as well as its repair after UV irradiation in excision-proficient strains of P.mirabilis and E.coli K12 have been compared. A fraction of post-replication repair (PRR) in P.mirabilis is found to be dependent on de novo protein synthesis after UV irradiation. Pre-irradiation by UV and pre-treatment with nalidixic acid increase the efficiency of post-irradiation replication and PRR even in the presence of chloramphenicol. An inducible repair function in P.mirabilis is supposed to stimulate post-irradiation replication and repair. (author)

  11. Synthesis and properties of a novel biodegradable poly(ester amine copolymer based on poly(L-lactide and low molecular weight polyethylenimine for gene delivery

    Directory of Open Access Journals (Sweden)

    Guo QF

    2011-08-01

    Full Text Available Qing Fa Guo, Ting Ting Liu, Xi Yan, Xiu Hong Wang, Shuai Shi, Feng Luo, Zhi Yong QianState Key Laboratory of Biotherapy and Cancer Center, West China Hospital, West China Medical School, Sichuan University, Chengdu, People's Republic of ChinaBackground: Gene therapy is a promising approach to the treatment of a wide range of diseases. The development of efficient and adequate gene delivery systems could be one of the most important factors. Polyethyleneimine, a cationic polymer, is one of the most successful and widely used vectors for nonviral transfection in vitro and in vivo.Methods: A novel biodegradable poly(ester amine copolymer (PEA was successfully prepared from low molecular weight polyethylenimine (PEI, 2000 Da and poly(L-lactide copolymers.Results: According to the results of agarose gel electrophoresis, particle size and zeta potential measurement, and transfection efficiency, the PEA copolymers showed a good ability to condense plasmid DNA effectively into nanocomplexes with a small particle size (≤150 nm and moderate zeta potential (≥10 mV at an appropriate polymeric carrier/DNA weight ratio. Compared with high molecular weight PEI (25kDa, the PEA obtained showed relatively high gene transfection efficiency as well as low cytotoxicity in vitro.Conclusion: These results indicate that such PEA might have potential application as a gene delivery system.Keywords: polyethylenimine, poly(L-lactide, gene delivery, cytotoxicity, transfection efficiency

  12. CSD2, CSD3, and CSD4, genes required for chitin synthesis in Saccharomyces cerevisiae: the CSD2 gene product is related to chitin synthases and to developmentally regulated proteins in Rhizobium species and Xenopus laevis.

    OpenAIRE

    Bulawa, C E

    1992-01-01

    In Saccharomyces cerevisiae, chitin forms the primary division septum and the bud scar in the walls of vegetative cells. Three chitin synthetic activities have been detected. Two of them, chitin synthase I and chitin synthase II, are not required for synthesis of most of the chitin present in vivo. Using a novel screen, I have identified three mutations, designated csd2, csd3, and csd4, that reduce levels of chitin in vivo by as much as 10-fold without causing any obvious perturbation of cell...

  13. Exopolysaccharide Biosynthesis in Lactococcus lactis NIZO B40: Functional Analysis of the Glycosyltransferase Genes Involved in Synthesis of the Polysaccharide Backbone

    OpenAIRE

    Kranenburg, Richard van; van Swam, Iris I.; Marugg, Joey D.; Kleerebezem, Michiel; de Vos, Willem M.

    1999-01-01

    We used homologous and heterologous expression of the glycosyltransferase genes of the Lactococcus lactis NIZO B40 eps gene cluster to determine the activity and substrate specificities of the encoded enzymes and established the order of assembly of the trisaccharide backbone of the exopolysaccharide repeating unit. EpsD links glucose-1-phosphate from UDP-glucose to a lipid carrier, EpsE and EpsF link glucose from UDP-glucose to lipid-linked glucose, and EpsG links galactose from UDP-galactos...

  14. Synthesis and maturation of lambda receptor in Escherichia coli K-12: in vivo and in vitro expression of gene lamB under lac promoter control.

    OpenAIRE

    Marchal, C.; Perrin, D; Hedgpeth, J; Hofnung, M

    1980-01-01

    The lambda receptor is an outer membrane protein from Escherichia coli K-12 lamB, its structural gene, is part of the maltose regulon. We have cloned this gene in a phage so that it is under the control of the lac promoter. The phage was devised in such a way that it can infect lamB mutants and that chromosomal lamB mutations can be transferred to it. In vivo, the lambda receptor is expressed under lac promoter control and is exported normally to the outer membrane, independently of the expre...

  15. Synthesis of 2'-deoxy-2'-[{sup 18}F]-fluoro-5-ethyl-1-{beta}-D-arabinofuranosyluracil ([{sup 18}F]-FEAU) and micro-PET imaging of HSV-tk gene expression in tumor-bearing nude mice

    Energy Technology Data Exchange (ETDEWEB)

    Alauddin, M.M.; Shahinian, A.; Park, R.; Tohme, M.; Fissekis, J.D.; Conti, P.S. [Univ. of Southern California, Los Angeles, CA (United States). PET Imaging Science Center

    2004-07-01

    Herpes simplex virus type-1 thymidine kinase (HSV1-tk) is being used as a suicide gene for gene therapy of cancer. An in vivo method to assess the HSV1-tk enzyme activity after gene transfer is desirable to monitor gene expression as an indicator of gene delivery. Imaging of the HSV1-tk reporter gene along with various reporter probes is of current interest. We originally developed [{sup 18}F]-FHPG and [{sup 18}F]-FHBG for PET imaging of HSV1-tk gene expression and demonstrated that [{sup 18}F]-FHBG is more useful than [{sup 18}F]-FHPG for this purpose. [{sup 124}I]-FIAU has been shown to be a potential PET imaging agent for HSV1-tk gene expression, and is superior to [{sup 18}F]-FHPG and [{sup 18}F]-FHBG. We also demonstrated that radiolabeled FMAU can be used as a marker for HSV-tk gene expression, and is superior to [{sup 18}F]-FHPG and [{sup 18}F]-FHBG. Earlier we reported a synthesis for 2'-deoxy-2'-[{sup 18}F]fluoro-5-methyl-1-{beta}-D-arabinofuranosyluracil ([{sup 18}F]-FMAU) and some other 5-substituted nucleosides. We have synthesized now [{sup 18}F]-FEAU, used the tracer for micro-PET imaging of suicide gene expression in tumor-bearing nude mice, and compared the results with earlier studies using [{sup 14}C]-FMAU. (orig.)

  16. Synthesis of 2'-deoxy-2'-[{sup 18}F]-fluoro-5-iodo-1-{beta}-D-arabinofuranosyluracil ([{sup 18}F]-FIAU) and micro-PET imaging of suicide gene expression in tumor-bearing nude mice

    Energy Technology Data Exchange (ETDEWEB)

    Alauddin, M.M.; Shahinian, A.; Park, R.; Tohme, M.; Fissekis, J.D.; Conti, P.S. [Univ. of Southern California, Los Angeles, CA (United States). PET Imaging Science Center

    2004-07-01

    Herpes simplex virus type-1 thymidine kinase (HSV1-tk) is being used as a suicide gene for gene therapy of cancer. An in vivo method to assess the HSV1-tk enzyme activity after gene transfer is desirable to monitor gene expression as an indicator of gene delivery. Imaging of the HSV1-tk reporter gene along with various reporter probes is of current interest. We originally developed [{sup 18}F]-FHPG and [{sup 18}F]-FHBG for PET imaging of HSV1-tk gene expression and demonstrated that [{sup 18}F]-FHBG is more useful than [{sup 18}F]-FHPG for this purpose. [{sup 124}I]-FIAU has been shown to be a potential PET imaging agent for HSV1-tk gene expression, and is superior to [{sup 18}F]-FHPG and [{sup 18}F]-FHBG. We also demonstrated that radiolabeled FMAU can be used as a marker for HSV-tk gene expression, and is superior to [{sup 18}F]-FHPG and [{sup 18}F]-FHBG. Earlier we reported a synthesis for 2'-deoxy-2'-[{sup 18}F]fluoro-5-methyl-1-{beta}-D-arabinofuranosyluracil ([{sup 18}F]-FMAU) and some other 5-substituted nucleosides. We have synthesized now [{sup 18}F]-FIAU, used the tracer for micro-PET imaging of suicide gene expression in tumor-bearing nude mice, and compared the results with earlier studies using [{sup 14}C]-FMAU. (orig.)

  17. DNA Synthesis, Assembly and Applications in Synthetic Biology

    OpenAIRE

    Ma, Siying; Tang, Nicholas; Tian, Jingdong

    2012-01-01

    The past couple of years saw exciting new developments in microchip-based gene synthesis technologies. Such technologies hold the potential for significantly increasing the throughput and decreasing the cost of gene synthesis. Together with more efficient enzymatic error correction and genome assembly methods, these new technologies are pushing the field of synthetic biology to a higher level.

  18. 8-Methoxypsoralen DNA interstrand cross-linking of the ribosomal RNA genes in Tetrahymena thermophila. Distribution, repair and effect on rRNA synthesis

    DEFF Research Database (Denmark)

    Fengquin, X; Nielsen, Henrik; Zhen, W;

    1993-01-01

    The distribution and repair of 8-methoxypsoralen-DNA interstrand cross-links in the ribosomal RNA genes (rDNA) in Tetrahymena thermophila have been studied in vivo by Southern blot analysis. It is found that the cross-links at a density of < or = 1/2 x 10(4) base pairs (bp) are distributed equall...

  19. A REVIEW ON THE CAPSULAR POLYSACCHARIDE SYNTHESIS GENES OF STREPTOCOCCUS AGALACTIAE%无乳链球菌(Streptococcus agalactiae)荚膜多糖合成基因研究进展

    Institute of Scientific and Technical Information of China (English)

    汪开毓; 黄锦炉; 肖丹; 王均; 黄凌远

    2013-01-01

    Streptococcus agalactiae is an important zoonotic pathogen in human, animal and fish. The capsular poly-saccharides on the cell surface have been recognized as virulence factors, of which the forming processes are regulated by the capsular polysaccharide biosynthesis (cps) genes. The capsular polysaccharide biosynthesis genes exist in capsular polysaccharide operon of S. agalactiae genome, which were predicted to participate in the origination of capsular polysaccharide synthesis, polymerization of oligosaccharides and polysaccharides, transport and anchor the product to the cell surface. Therefore, the cps genes achieve new applications of diagnostic techniques and the construction of attenuated mutant strains. In this paper, the basic properties of cps genes, as well as their transcription regulation, coding proteins and biological functions, regulation mechanism of capsular polysaccharide biosynthesis and serotyping, and application of construction of mutant strains are analyzed and discussed. We looked forward to providing a theoretical reference on the new functions research and innovative applications of S. agalactiae cps genes.%无乳链球菌是一种人畜鱼共患的重要病原菌,菌体表面的荚膜多糖是公认的毒力因子,其合成过程受荚膜多糖合成基因的调控.荚膜多糖合成基因存在于荚膜多糖操纵子中,参与无乳链球菌荚膜多糖的合成启动、寡糖和多糖的聚合以及外输并锚定于菌体表面,在新型诊断技术和减毒突变株的构建方面取得良好的应用.本文首次就cps基因的基本属性、转录调节、编码蛋白及其生物功能、对荚膜多糖合成的调控机理、在血清分型和突变株构建的应用这六个方面进行深入分析和讨论,以期为GBS cps基因的新功能研究和创新应用提供理论参考.

  20. Synthesis and preliminary evaluation of 9-(4-[18F]fluoro-3-hydroxymethylbutyl) guanine ([18F]FHBG) in HSV1-tk gene transduced hepatoma cell

    International Nuclear Information System (INIS)

    The HSV1-tk reporter gene system is the most widely used system because of its advantage that direct monitoring is possible without the introduction of a separate reporter gene in case of HSV1-tk suicide gene therapy. In this study, we investigate the usefulness of the reporter probe (substrate), 9-(4-[18F]fluoro-3-hydroxymethylbutyl) guanine ([18F]FHBG) for non-invasive reporter gene imaging using PET in HSV1-tk expressing hepatoma model. Radiolabeled FHBG was prepared in 8 steps from a commercially available triester. The labeling reaction was carried out by NCA nucleophilic substitution with K[18F]/K2.2.2. in acetonitrile using N2-monomethoxytrityl-9-[4-(tosly)-3-monomethoxytritylmethylbutl] guanine as a precursor, followed by deprotection with 1 N HCI. Preliminary biological properties of the probe were evaluated with MCA cells and MCA-tk cells transduced with HSV1-tk reporter gene. In vitro uptake and release-out studies of [18F]FHBG were performed, and was analyzed correlation between [18F]FHBG uptake ratio according to increasing numeric count of MCA-tk cells and degree of gene expression. MicroPET scan image was obtained with MCA and MCA-tk tumor beating Balb/c-nude mouse model. [18F]FHBG was purified by reverse phase semi-HPLC system and collected at around 16-18 min. Radiochemical yield was about 20-25% (corrected for decay), radiochemical purity was > 95% and specific activity was around > 55.5 GBq/ μ mol. Specific accumulation of [18F]FHBG was observed in HSV1-tk gene transduced MCA-tk cells but not MCA cells, and consecutive 1 hour release-out results showed more than 86% of uptaked [18F]FHBG was retained inside of cells. The uptake of [18F]FHBG was showed a highly significant linear correlation (R2 = 0.995) with increasing percentage of MCA-tk numeric cell count. In microPET scan images, remarkable difference of accumulation was observed for the two type of tumors. [18F]FHBG appears to be a useful as non-invasive PET imaging substrate in HSV1-tk

  1. The secretion, synthesis, and metabolism of cortisol and its downstream genes in the H-P-I axis of rare minnows (Gobiocypris rarus) are disrupted by acute waterborne cadmium exposure.

    Science.gov (United States)

    Liu, Xiao-Hong; Xie, Bi-Wen; Wang, Zhi-Jian; Jin, Li; Zhang, Yao-Guang

    2016-01-01

    The H (hypothalamic)-P (pituitary)-I (interrenal) axis plays a critical role in the fish stress response and is regulated by several factors. Cadmium (Cd) is one of the most toxic heavy metals in the world, but its effects on the H-P-I axis of teleosts are largely unknown. Using rare minnow (Gobiocypris rarus) as an experimental animal, we found that Cd only disrupted the secretion and synthesis of cortisol. Neither hormones at the H or P level nor the expressions of their receptor genes (corticotropin-releasing hormone receptor (CRHR) and melanocortin receptor 2 (MC2R)) were affected. Steroidogenic acute regulator (StAR), CYP11A1 and CYP11B1, which encode the key enzymes in the cortisol synthesis pathway, were significantly up-regulated in the kidney (including the head kidney). The level of 11β-HSD2, which is required for the conversion of cortisol to cortisone, was increased in the kidney, intestine, brain, and hepatopancreas, whereas the expression of 11β-HSD1, which encodes the reverse conversion enzyme, was increased in the gill, kidney and almost unchanged in other tissues. The enzyme activity concentration of 11β-HSD2 was increased in the kidney as well. The level of glucocorticoid receptor (GR) decreased in the intestine, gill and muscle, and the key GR regulator FK506 binding protein5 (FKBP5) was up-regulated in the GR-decreased tissues, whereas the level of nuclear receptor co-repressor 1 (NCoR1), another GR regulator remained almost unchanged. Thus, GR, FKBP5 and 11β-HSD2 may be involved in Cd-induced cortisol disruption. PMID:27033032

  2. Multiple phenotypic changes in mice after knockout of the B3gnt5 gene, encoding Lc3 synthase--a key enzyme in lacto-neolacto ganglioside synthesis

    Directory of Open Access Journals (Sweden)

    McLendon Roger E

    2010-11-01

    Full Text Available Abstract Background Ganglioside biosynthesis occurs through a multi-enzymatic pathway which at the lactosylceramide step is branched into several biosynthetic series. Lc3 synthase utilizes a variety of galactose-terminated glycolipids as acceptors by establishing a glycosidic bond in the beta-1,3-linkage to GlcNaAc to extend the lacto- and neolacto-series gangliosides. In order to examine the lacto-series ganglioside functions in mice, we used gene knockout technology to generate Lc3 synthase gene B3gnt5-deficient mice by two different strategies and compared the phenotypes of the two null mouse groups with each other and with their wild-type counterparts. Results B3gnt5 gene knockout mutant mice appeared normal in the embryonic stage and, if they survived delivery, remained normal during early life. However, about 9% developed early-stage growth retardation, 11% died postnatally in less than 2 months, and adults tended to die in 5-15 months, demonstrating splenomegaly and notably enlarged lymph nodes. Without lacto-neolacto series gangliosides, both homozygous and heterozygous mice gradually displayed fur loss or obesity, and breeding mice demonstrated reproductive defects. Furthermore, B3gnt5 gene knockout disrupted the functional integrity of B cells, as manifested by a decrease in B-cell numbers in the spleen, germinal center disappearance, and less efficiency to proliferate in hybridoma fusion. Conclusions These novel results demonstrate unequivocally that lacto-neolacto series gangliosides are essential to multiple physiological functions, especially the control of reproductive output, and spleen B-cell abnormality. We also report the generation of anti-IgG response against the lacto-series gangliosides 3'-isoLM1 and 3',6'-isoLD1.

  3. Synthesis, and Characterization, and Evaluation of Cellular Effects of the FOL-PEG-g-PEI-GAL Nanoparticles as a Potential Non-Viral Vector for Gene Delivery

    OpenAIRE

    S. Ghiamkazemi; Amanzadeh, A.; Dinarvand, R.; M Rafiee-Tehrani; Amini, M.

    2010-01-01

    In this manuscript, we synthesized the potential non viral vector for gene delivery with proper transfection efficiency and low cytotoxicity. Polyethylenimine (PEI) is a well-known cationic polymer which has high positive surface charge for condensing plasmid DNA. However; it is highly cytotoxic in many cell lines because of the high surface charge, non-biodegradability and non-biocompatibility. To enhance PEI biodegradability, the graft copolymer “PEG-g-PEI” was synthesized. To t...

  4. Synthesis of Human Insulin Gene and Transformation into Tremella fuciformis%人胰岛素基因的人工合成及转化银耳的研究

    Institute of Scientific and Technical Information of China (English)

    谢宝贵; 卢启泉; 饶永斌; 孙淑静; 郑金贵

    2007-01-01

    用PCR法合成人胰岛素基因,克隆到pUC18载体上,经测序证实其碱基序列与人胰岛素基因的序列完全一致.本实验还构建了银耳表达载体,由限制性内切酶介导(Restriction enzyme-mediated DNA Integration, REMI)转化银耳芽孢.随机挑取21个抗性菌落,转管繁殖2代后检测其GUS活性,实验结果:18个菌株阳性,3个菌株阴性.从这21个菌株中选取10个菌株,提取染色体DNA,用人胰岛素基因、GUS基因和Tnos序列的特异引物进行PCR,结果表明,这10菌株都能扩增出相应长度的特异片段,证明了它们是人胰岛素基因转化子.%A human insulin gene was synthesized using PCR-based methodology, cloned into plasmid pUC18, and the sequence of the inserted fragment was confirmed. A Tremella fuciformis expression vector was constructed and used to transform T. fuciformis yeast-like conidia employing Restriction Enzyme-Mediated DNA Integration (REMI). A total of 21 antibiotic resistant colonies were selected at random of which 18 tested positive for β-glucuronidase (GUS) activity. Genomic DNA was extracted from 10 of the 21 positive transformants and PCR performed using genomic DNA as the template and specific primers for amplifying the human insulin gene, the GUS gene and the Tnos sequence. PCR data showed that the expected fragment was amplified from all 10 isolates demonstrating that they were true human insulin gene transformants.

  5. Biocatalytic synthesis of flavones and hydroxyl-small molecules by recombinant Escherichia coli cells expressing the cyanobacterial CYP110E1 gene

    OpenAIRE

    Makino Takuya; Otomatsu Toshihiko; Shindo Kazutoshi; Kitamura Emi; Sandmann Gerhard; Harada Hisashi; Misawa Norihiko

    2012-01-01

    Abstract Background Cyanobacteria possess several cytochrome P450s, but very little is known about their catalytic functions. CYP110 genes unique to cyanaobacteria are widely distributed in heterocyst-forming cyanobacteria including nitrogen-fixing genera Nostoc and Anabaena. We screened the biocatalytic functions of all P450s from three cyanobacterial strains of genus Nostoc or Anabaena using a series of small molecules that contain flavonoids, sesquiterpenes, low-molecular-weight drugs, and...

  6. The Expression of the Related Fatty Acid Synthesis Key Enzyme Genes in Bovine Somatic Cell%牛乳腺脂肪合成关键酶基因在乳汁体细胞中的表达研究

    Institute of Scientific and Technical Information of China (English)

    谢佳喜; 朱河水; 杨国宇; 李宏基; 郭豫杰; 汪新建; 王月影

    2011-01-01

    为了阐明乳脂合成的影响因素及其内在分子机理,为反刍动物原料乳的优化,特别是为乳脂肪的营养调控和遗传改良提供理论依据.本试验以奶牛初乳、常乳和末乳中的乳汁体细胞为研究对象,以看家基因GAPDH为内参,对初乳、常乳和末乳中LPL、CD36、VLDLR、ACSS2、ACSL1、FABP3、ACC、FASN、SCD、ADFP、XDH和BTN1A1 mRNA进行半定量RT-PCR分析.结果发现,LPL、CD36、VLDLR、ACSS2、ACSL1、FABP3、SCD、ADFP、XDH和BTN1Al mRNA在初乳、常乳和末乳中均有表达,而ACC和FASN mRNA只在初乳中表达,常乳和末乳中均不表达;半定量结果表明,与初乳相比,常乳和末乳中LPL、CD36、VLDLR、ACSS2、ACSL1、FABP3、SCD、ADFP、XDH和BTN1Al mRNA转录水平显著降低(P<0.05),且常乳与末乳间差异不显著(P>0.05).研究结果提示初乳期乳腺脂肪合成能力明显高于常乳和末乳期乳腺,且脂肪合成关键酶基因的表达与细胞内脂转运和代谢的生理变化有关.%To clarify the molecular mechanism of milk fat synthesis, optimizing ruminant raw milk, particularly providing theory basis for nutrition regulation and heredity improving for milk fat. Somatic cells in colostrum milk, mature milk and involution milk were selected and housekeeping gene GAPDH was selected as reference, the semi-quantitive RT-PCR was used to analyze the expression of LPL, CD36, VLDLR, ACSS2, ACSLl, FABP3, ACC, FASN, SCD, AD-FP, XDH and BTN1A1 mRNA in milk. The results showed that the genes LPL, CD36, VLDLR, ACSS2, ACSLl, FABP3, ACC, FASN, SCD, ADFP, XDH and BTNV1A1 mRNA expressed in the colostrum milk, mature milk and involution milk. However the genes ACC and FASN were not detected in mature milk and involution milk. The relative quantitive results showed that the expression level of LPL, CD36, VLDLR, ACSS2, ACSLl, FABP3, ACC, FASN, SCD, ADFP, XDH and BTN1A1 mRNA in mature milk and involution milk were significantly decreased(P0. 05) between mature

  7. Biocatalytic synthesis of flavones and hydroxyl-small molecules by recombinant Escherichia coli cells expressing the cyanobacterial CYP110E1 gene

    Directory of Open Access Journals (Sweden)

    Makino Takuya

    2012-07-01

    Full Text Available Abstract Background Cyanobacteria possess several cytochrome P450s, but very little is known about their catalytic functions. CYP110 genes unique to cyanaobacteria are widely distributed in heterocyst-forming cyanobacteria including nitrogen-fixing genera Nostoc and Anabaena. We screened the biocatalytic functions of all P450s from three cyanobacterial strains of genus Nostoc or Anabaena using a series of small molecules that contain flavonoids, sesquiterpenes, low-molecular-weight drugs, and other aromatic compounds. Results Escherichia coli cells carrying each P450 gene that was inserted into the pRED vector, containing the RhFRed reductase domain sequence from Rhodococcus sp. NCIMB 9784 P450RhF (CYP116B2, were co-cultured with substrates and products were identified when bioconversion reactions proceeded. Consequently, CYP110E1 of Nostoc sp. strain PCC 7120, located in close proximity to the first branch point in the phylogenetic tree of the CYP110 family, was found to be promiscuous for the substrate range mediating the biotransformation of various small molecules. Naringenin and (hydroxyl flavanones were respectively converted to apigenin and (hydroxyl flavones, by functioning as a flavone synthase. Such an activity is reported for the first time in prokaryotic P450s. Additionally, CYP110E1 biotransformed the notable sesquiterpene zerumbone, anti-inflammatory drugs ibuprofen and flurbiprofen (methylester forms, and some aryl compounds such as 1-methoxy and 1-ethoxy naphthalene to produce hydroxylated compounds that are difficult to synthesize chemically, including novel compounds. Conclusion We elucidated that the CYP110E1 gene, C-terminally fused to the P450RhF RhFRed reductase domain sequence, is functionally expressed in E. coli to synthesize a robust monooxygenase, which shows promiscuous substrate specificity (affinity for various small molecules, allowing the biosynthesis of not only flavones (from flavanones but also a variety of

  8. Energy transfer from carotenoids to bacteriochlorophylls

    Czech Academy of Sciences Publication Activity Database

    Frank, H.A.; Polívka, Tomáš

    Dordrecht : Springer, 2008 - (Hunter, C.; Daldal, F.; Thurnauer, M.; Beatty, J.), s. 218-230 ISBN 978-1-4020-8814-8. - (Advances in Photosynthesis and Respiration. 28) Institutional research plan: CEZ:AV0Z50510513 Keywords : carotenoids * Energy transfer Subject RIV: BO - Biophysics

  9. Synthesis and biological evaluation of [18F]tetrafluoroborate: a PET imaging agent for thyroid disease and reporter gene imaging of the sodium/iodide symporter

    OpenAIRE

    Jauregui-Osoro, Maite; Sunassee, Kavitha; Weeks, Amanda J.; Berry, David J; Paul, Rowena L.; Cleij, Marcel; Banga, Jasvinder Paul; O’Doherty, Michael J.; Marsden, Paul K.; Clarke, Susan E. M.; Ballinger, James R.; Szanda, Istvan; Cheng, Sheue-yann; Blower, Philip J

    2010-01-01

    Purpose The human sodium/iodide symporter (hNIS) is a well-established target in thyroid disease and reporter gene imaging using gamma emitters 123I-iodide, 131I-iodide and 99mTc-pertechnetate. However, no PET imaging agent is routinely available. The aim of this study was to prepare and evaluate 18F-labelled tetrafluoroborate ([18F]TFB) for PET imaging of hNIS. Methods [18F]TFB was prepared by isotopic exchange of BF4 − with [18F]fluoride in hot hydrochloric acid and purified using an alumin...

  10. Association Between Seed Dormancy and Pericarp Color Is Controlled by a Pleiotropic Gene That Regulates Abscisic Acid and Flavonoid Synthesis in Weedy Red Rice

    OpenAIRE

    Gu, Xing-You; Foley, Michael E.; Horvath, David P.; Anderson, James V.; Feng, Jiuhuan; Zhang, Lihua; Mowry, Chase R.; Ye, Heng; Suttle, Jeffrey C; Kadowaki, Koh-ichi; Chen, Zongxiang

    2011-01-01

    Seed dormancy has been associated with red grain color in cereal crops for a century. The association was linked to qSD7-1/qPC7, a cluster of quantitative trait loci for seed dormancy/pericarp color in weedy red rice. This research delimited qSD7-1/qPC7 to the Os07g11020 or Rc locus encoding a basic helix-loop-helix family transcription factor by intragenic recombinants and provided unambiguous evidence that the association arises from pleiotropy. The pleiotropic gene expressed in early devel...

  11. Dual stage synthesis and crucial role of cytoadherence-linked asexual gene 9 in the surface expression of malaria parasite var proteins

    DEFF Research Database (Denmark)

    Goel, Suchi; Valiyaveettil, Manojkumar; Achur, Rajeshwara N;

    2010-01-01

    Plasmodium falciparum erythrocyte membrane protein 1 (PfEMP1) family members mediate the adherence of parasite-infected red blood cells (IRBCs) to various host receptors. A previous study has shown that the parasite protein, cytoadherence-linked asexual gene 9 (CLAG9), is also essential for IRBC...... within the parasite. Based on these findings, we propose that CLAG9 plays a critical role in the trafficking of PfEMP1s onto the IRBC surface. These results have important implications for the development of therapeutics for cerebral, placental, and other cytoadherence-associated malaria illnesses....

  12. Synthesis and function of Actinomyces naeslundii T14V type 1 fimbriae require the expression of additional fimbria-associated genes.

    OpenAIRE

    Yeung, M K; Ragsdale, P A

    1997-01-01

    The nucleotide sequence of the chromosomal DNA flanking the Actinomyces naeslundii (formerly A. viscosus) T14V type 1 fimbrial structural subunit gene (fimP) was determined. Six open reading frames (ORFs), in the order 5' ORF3, ORF2, ORF1,fimP, ORF4, ORF5, ORF6 3', were identified. ORF1 encoded a protein of 408 amino acid residues (Mr = 39,270) and had significant sequence homology with the A. naeslundii T14V type 1 and A. naeslundii WVU45 type 2 fimbrial structural subunits. An in-frame fusi...

  13. The evolution of the protein synthesis system. I - A model of a primitive protein synthesis system

    Science.gov (United States)

    Mizutani, H.; Ponnamperuma, C.

    1977-01-01

    A model is developed to describe the evolution of the protein synthesis system. The model is comprised of two independent autocatalytic systems, one including one gene (A-gene) and two activated amino acid polymerases (O and A-polymerases), and the other including the addition of another gene (N-gene) and a nucleotide polymerase. Simulation results have suggested that even a small enzymic activity and polymerase specificity could lead the system to the most accurate protein synthesis, as far as permitted by transitions to systems with higher accuracy.

  14. Amphiphilic star block copolymers as gene carrier Part I: Synthesis via ATRP using calix[4]resorcinarene-based initiators and characterization

    Energy Technology Data Exchange (ETDEWEB)

    Zheng, Anna; Xue, Yan; Wei, Dafu [Shanghai Key Laboratory of Advanced Polymeric Materials, Key Laboratory for Ultrafine Materials of Ministry of Education, School of Materials Science and Engineering, East China University of Science and Technology, Shanghai 200237 (China); Guan, Yong, E-mail: yguan@ecust.edu.cn [Shanghai Key Laboratory of Advanced Polymeric Materials, Key Laboratory for Ultrafine Materials of Ministry of Education, School of Materials Science and Engineering, East China University of Science and Technology, Shanghai 200237 (China); Xiao, Huining [Department of Chemical Engineering, University of New Brunswick, Fredericton, New Brunswick, Canada E3B 5A3 (Canada)

    2013-01-01

    In this work, a cationic star polymer [poly(2-dimethylamino)ethyl methacrylate (PDMAEMA)] was prepared via atom transfer radical polymerization (ATRP), using brominated calix[4]resorcinarene as an initiator. Hydrophobic moieties, methyl methacrylate (MMA) and butyl acrylate (BA), were further incorporated via 'one-pot' method. Well-defined eight-armed star block copolymers bearing hydrophilic blocks inside and hydrophobic blocks outside were synthesized. The molecular weight, particle size, electrophoretic mobility and apparent charge density were determined by gel permeation chromatography (GPC), dynamic light scattering (DLS), phase analysis light scattering (PALS) and colloidal titration, respectively. The zeta potentials and apparent charge densities of the products exhibited the characteristics of polyelectrolyte. The incorporation of hydrophobic moieties generated electrostatic screening effect. The as-synthesized amphiphilic star copolymer is promising as a thermo-sensitive gene carrier for gene therapy. Highlights: Black-Right-Pointing-Pointer Amphiphilic cationic star block copolymers with well-controlled structures were prepared via ATRP. Black-Right-Pointing-Pointer The molecular structures and properties of the initiator and copolymers were systematically characterized. Black-Right-Pointing-Pointer The products exhibited the positive charged character, and hydrophobic moieties generated electrostatic screening effect.

  15. Detection and Cloning of a Gene Involved in Zwitermicin A Synthesis from Plant Growth Promoting Rhizobacteria of Bacillus sp CR64

    Directory of Open Access Journals (Sweden)

    Aris Tri Wahyudi

    2015-11-01

    Full Text Available Utilization of soil bacteria as biocontrol agent is becoming popular due to its valuable and effective mechanisms to suppress plant pathogenic microbes. We have previously isolated Bacillus sp, designated as Bacillus sp CR64, which exhibited effective plant growth promoting and antifungal activities. In this study, CR64 was examined in inhibiting the growth of Rhizoctonia solani, the causing agent of root rot disease. Partial sequence analysis of 16S rRNA gene revealed that this isolate similar with Bacillus cereus (94%. Furthermore, a gene designated zmaR was detected by means of specific amplification of DNA fragment approximately 950 bp. This fragment was then cloned onto pCRII-TOPO (3.9 kb and sequenced using DNA sequencer ABI PRISM 310. Sequence analysis revealed that it had highest homology with the ZmaR protein (89% identity; 90% similarity of B. thuringiensis serovar kurstaki (AAF82729.2. Alignment analysis with other ZmaR sequences from other antibiotic-producing Bacilli exhibited an almost fully conserved region within ZmaR sequences.Key words : PGPR, Bacillus sp CR64, Zwitermicin A, Cloning, Antifungal.

  16. Cloning and Experssion of Key Enzyme Gene APT1 for ATP Synthesis in Actinomucor elegans%ATP合成关键酶基因APT1在雅致放射毛霉中的克隆表达

    Institute of Scientific and Technical Information of China (English)

    朱家荣; 杨善岩; 陈丽芬; 杨光辉

    2012-01-01

    为构建遗传稳定的ATP高产工程菌,利用PCR技术扩增酿酒酵母ATP合成关键酶基因APT1,并将其克隆至质粒pCB1004-Pgpd的相应位点,得到有强启动子Pgpd驱动的APT1基因超表达质粒pCB1004-Pgpd-APT1。在PEG-CaCl2介导下,超表达质粒转化雅致放射毛霉原生质体,获得ATP高产工程菌。其ATP产量及摩尔转化率比出发菌株提高44.04%。%To construct the engineering strain with genetic stability which can efficient to synthesize ATP,the key enzyme gene APT1 for ATP synthesis which from Saccharomyces cerevisiae was cloned by PCR.The sequence was cloned to corresponding site of pCB1004-Pgpd.Then the super expression plasmid pCB1004-Pgpd-APT1 which controlled by strong promoter Pgpd was obtained.PEG and CaCl2 mediated protoplast transformation of Actinomucor elegans with super expression plasmid was performed and ATP-overproducing transformants were obtained.The ATP yield and Moore conversion were increased 44.04% in comparison with those from original strain.

  17. Organic synthesis

    International Nuclear Information System (INIS)

    The 1988 progress report of the Organic Synthesis Chemistry laboratory (Polytechnic School, France), is presented. The laboratory activities are centered on the chemistry of natural products, which have a biological activity and on the development of new reactions, useful in the organic synthesis. The research works involve the following domains: the natural products chemistry which are applied in pharmacology, the plants and insects chemistry, the organic synthesis, the radical chemistry new reactions and the bio-organic physicochemistry. The published papers, the congress communications and the thesis are listed

  18. 白菜硫代葡萄糖甙合成中MAM的进化分析%Evolutionary Study ofMAM Genes Which Function in the Process of Glucosinolates Synthesis in Brassica rapa

    Institute of Scientific and Technical Information of China (English)

    刘锦; 张冀芳; 梁建丽; 程锋; 武剑; 王晓武

    2014-01-01

    Objective]The objective of this study is to explore the duplication and evolution of Brassica rapaMAM which are key genes in the process of glucosinolates synthesis. This study will provide a theoretical basis for futureMAM research.[Method]The synthenic relationship ofMAMgenes amongArabidopsis thaliana,Schrenkiella parvulaandBrassica rapa were analyzed, according toB. rapa datebase BARD, conserved motif ofMAM genes were predicted by online software MEME, the expression of MAMhomologous genes in 93 materials ofBrassica rapa was analyzed by software R, and the phylogenetic tree was constructed by software MEGA 5.05.[Result]The results showed that theMAMhomologous genes were located in three subgenomes ofB. rapa. Amongst sevenMAM genes, five geneswhich are synthenic orthologs toA. thaliana mainly originated from whole genome triplication. The remaining two genes may derive from translocation. Motif analysis result indicated that there were many differences in both ends of protein sequences in BrMAMhomologous genes, compared withAtMAM genes. Expression data showed that the expression level of these homologous genes was variable,Bra021947andBra018524 expressed little in several materials ofB. rapa, the expression ofBra029356could be detected in all materials. In the phylogenetic tree,BrMAMhomologous genes andAtMAM genes were located in different branches, in addition toBra018524.[Conclusion]These results indicated that theB. rapa MAM genes had an independent evolution after the divergence ofB. rapaandA. thaliana.%目的探索硫代葡萄糖甙合成中关键基因-甲基硫代烷烃基苹果酸合成酶(methylthioalkylmalate, MAM)基因在白菜基因组中的进化情况,为研究白菜MAM的功能提供理论指导。方法根据白菜数据库BARD信息分析拟南芥、盐芥和白菜MAM的共线性关系,利用MEME在线软件预测MAM序列的高度保守基序(motif),通过R软件分析93份白菜材料MAM的表达模式,并利用MEGA5.05软

  19. Synthesis and evaluation of a C-6 alkylated pyrimidine derivative for the in vivo imaging of HSV1-TK gene expression

    International Nuclear Information System (INIS)

    Introduction: We report on the synthesis, radiolabeling, in vitro and in vivo characterization of N-Me-[18F]FHBT (6-(3-[18F]fluoro-2-(hydroxymethyl)propyl)-1,5-dimethylpyrimidin-2,4 (1H,3H)-dione), a C-6-substituted N-1-methylated pyrimidine derivative as a reporter probe for imaging herpes simplex virus type 1 thymidine kinase (HSV1-TK) expression. Methods: N-Me-[18F]FHBT was synthesized via a standard nucleophilic substitution reaction followed by acidic cleavage of the methoxytrityl protecting group. Cell uptake was studied in vitro with control HEK293 (human embryonic kidney cells) and HEK293 cells stably transfected with nonmutant HSV1-tk (HEK293TK+ cells). Positron emission tomography (PET) imaging and biodistribution studies of N-Me-[18F]FHBT or [18F]FHBG were performed in nude mice bearing xenografts of HEK293 control and TK+ cells. Results: N-Me-[18F]FHBT was obtained in a two-step reaction in an overall maximal radiochemical yield (decay-corrected) of 5% and a radiochemical purity >96%. The tracer uptake in HSV1-TK containing HEK293TK+ cells was 14.5 times (at 30 min) and 55.4 times (at 240 min) higher than in control HEK293 cells. In mice, N-Me-[18F]FHBT and [18F]FHBG accumulated significantly and exhibited similar radioactivity levels in the HEK293TK+ xenografts; however, standardized uptake values ratios between HEK293TK+ and HEK293 control xenografts were higher for [18F]FHBG than for N-Me-[18F]FHBT. Both tracers showed high gall bladder and abdominal activity. Conclusion: The biological evaluations demonstrated the feasibility of using N-methylated C-6-substituted pyrimidine derivative N-Me-[18F]FHBT as a PET radiotracer for monitoring HSV1-TK expression in vivo.

  20. Silencing of the violaxanthin de-epoxidase gene in the diatom Phaeodactylum tricornutum reduces diatoxanthin synthesis and non-photochemical quenching.

    Directory of Open Access Journals (Sweden)

    Johann Lavaud

    Full Text Available Diatoms are a major group of primary producers ubiquitous in all aquatic ecosystems. To protect themselves from photooxidative damage in a fluctuating light climate potentially punctuated with regular excess light exposures, diatoms have developed several photoprotective mechanisms. The xanthophyll cycle (XC dependent non-photochemical chlorophyll fluorescence quenching (NPQ is one of the most important photoprotective processes that rapidly regulate photosynthesis in diatoms. NPQ depends on the conversion of diadinoxanthin (DD into diatoxanthin (DT by the violaxanthin de-epoxidase (VDE, also called DD de-epoxidase (DDE. To study the role of DDE in controlling NPQ, we generated transformants of P. tricornutum in which the gene (Vde/Dde encoding for DDE was silenced. RNA interference was induced by genetic transformation of the cells with plasmids containing either short (198 bp or long (523 bp antisense (AS fragments or, alternatively, with a plasmid mediating the expression of a self-complementary hairpin-like construct (inverted repeat, IR. The silencing approaches generated diatom transformants with a phenotype clearly distinguishable from wildtype (WT cells, i.e. a lower degree as well as slower kinetics of both DD de-epoxidation and NPQ induction. Real-time PCR based quantification of Dde transcripts revealed differences in transcript levels between AS transformants and WT cells but also between AS and IR transformants, suggesting the possible presence of two different gene silencing mediating mechanisms. This was confirmed by the differential effect of the light intensity on the respective silencing efficiency of both types of transformants. The characterization of the transformants strengthened some of the specific features of the XC and NPQ and confirmed the most recent mechanistic model of the DT/NPQ relationship in diatoms.

  1. Synthesis, and Characterization, and Evaluation of Cellular Effects of the FOL-PEG-g-PEI-GAL Nanoparticles as a Potential Non-Viral Vector for Gene Delivery

    Directory of Open Access Journals (Sweden)

    S. Ghiamkazemi

    2010-01-01

    Full Text Available In this manuscript, we synthesized the potential non viral vector for gene delivery with proper transfection efficiency and low cytotoxicity. Polyethylenimine (PEI is a well-known cationic polymer which has high positive surface charge for condensing plasmid DNA. However; it is highly cytotoxic in many cell lines because of the high surface charge, non-biodegradability and non-biocompatibility. To enhance PEI biodegradability, the graft copolymer “PEG-g-PEI” was synthesized. To target cancer liver cells, two targeting ligands folic acid and galactose (lactobionic acid which are over expressed on human hepatocyte carcinoma were attached to graft copolymer and “FOL-PEG-g-PEI-GAL” copolymer was synthesized. Composition of this grafted copolymer was characterized using 1H-NMR and FTIR spectra. The molecular weight and zeta potential of this copolymer was compared to PEI. The particle size and zeta potential of FOL-PEG-g-PEI-GAL/DNA complexes at various N/P ratio were measured using dynamic light scattering (DLS. Cytotoxicity of the copolymer was also studied in cultured HepG2 human hepatoblastoma cell line. The FOL-PEG-g-PEI-GAL/DNA complexes at various N/P ratios exhibited no cytotoxicity in HepG2 cell line compared to PEI 25K as a control. The novel copolymer showed enhanced biodegradability in physiological conditions in compared with PEI and targeted cultured HepG2 cells. More importantly, significant transfection efficiency was exhibited in cancer liver cells. Together, our results showed that “FOL-PEG-g-PEI-GAL” nanoparticals could be considered as a useful non-viral vector for targeted gene delivery.

  2. Human Pigmentation, Cutaneous Vitamin D Synthesis and Evolution: Variants of Genes (SNPs) Involved in Skin Pigmentation Are Associated with 25(OH)D Serum Concentration.

    Science.gov (United States)

    Rossberg, Willi; Saternus, Roman; Wagenpfeil, Stefan; Kleber, Marcus; März, Winfried; Reichrath, Sandra; Vogt, Thomas; Reichrath, Jörg

    2016-03-01

    Vitamin D deficiency is common and associated with higher risk for and unfavourable outcome of many diseases. Limited data exist on genetic determinants of serum 25(OH)D concentration. In a cohort of the LURIC study (n=2974, median 25(OH)D concentration 15.5 ng/ml), we tested the hypothesis that variants (SNPs, n=244) of several genes (n=15) involved in different aspects of skin pigmentation, including melanosomal biogenesis (ATP7A, DTNBP1, BLOC1S5, PLDN, PMEL), melanosomal transport within melanocytes (RAB27A, MYO5A, MLPH); or various melanocyte signaling pathways (MC1R, MITF, PAX3, SOX10, DKK1, RACK1, CNR1) are predictive of serum 25(OH)D levels. Eleven SNPs located in 6 genes were associated (p<0.05) with low or high serum 25(OH)D levels, 3 out of these 11 SNPs reached the aimed significance level after correction for multiple comparisons (FDR). In the linear regression model adjusted for sex, body mass index (BMI), year of birth and month of blood sample rs7565264 (MLPH), rs10932949 (PAX3), and rs9328451 (BLOC1S5) showed a significant association with 25(OH)D. The combined impact on variation of 25(OH)D serum levels (coefficient of determination (R(2))) for the 11 SNPs was 1.6% and for the 3 SNPs after FDR 0.3%. In Cox Regression we identified rs2292881 (MLPH) as having a significant association (advantage) with overall survival. Kaplan-Meier analysis did not show any significant impact of individual SNPs on overall survival. In conclusion, these results shed new light on the role of sunlight, skin pigmentation and vitamin D for human evolution. PMID:26977047

  3. Synthesis, and Characterization, and Evaluation of Cellular Effects of the FOL-PEG-g-PEI-GAL Nanoparticles as a Potential Non-Viral Vector for Gene Delivery

    International Nuclear Information System (INIS)

    In this manuscript, we synthesized the potential non viral vector for gene delivery with proper transfection efficiency and low cytotoxicity. Polyethylenimine (PEI) is a well-known cationic polymer which has high positive surface charge for condensing plasmid DNA. However; it is highly cytotoxic in many cell lines because of the high surface charge, non-biodegradability and non-biocompatibility. To enhance PEI biodegradability, the graft copolymer PEG-g-PEI was synthesized. To target cancer liver cells, two targeting ligands folic acid and galactose (lactobionic acid) which are over expressed on human hepatocyte carcinoma were attached to graft copolymer and FOL-PEG-g-PEI-GAL copolymer was synthesized. Composition of this grafted copolymer was characterized using 1H-NMR and FTIR spectra. The molecular weight and zeta potential of this copolymer was compared to PEI. The particle size and zeta potential of FOL-PEG-g-PEI-GAL/DNA complexes at various N/P ratio were measured using dynamic light scattering (DLS). Cytotoxicity of the copolymer was also studied in cultured HepG2 human hepatoblastoma cell line. The FOL-PEG-g-PEI-GAL/DNA complexes at various N/P ratios exhibited no cytotoxicity in HepG2 cell line compared to PEI 25K as a control. The novel copolymer showed enhanced biodegradability in physiological conditions in compared with PEI and targeted cultured HepG2 cells. More importantly, significant transfection efficiency was exhibited in cancer liver cells. Together, our results showed that FOL-PEG-g-PEI-GAL nanoparticles could be considered as a useful non-viral vector for targeted gene delivery.

  4. Effect of nutrition on plasma lipid profile and mRNA levels of ovarian genes involved in steroid hormone synthesis in Hu sheep during luteal phase.

    Science.gov (United States)

    Ying, S J; Xiao, S H; Wang, C L; Zhong, B S; Zhang, G M; Wang, Z Y; He, D Y; Ding, X L; Xing, H J; Wang, F

    2013-11-01

    Ovarian steroid hormones regulate follicular growth and atresia. This study aims to determine whether key ovarian sterol-regulatory genes are differentially expressed in Hu sheep under different short-term nutritional regimens. Estrus was synchronized using intravaginal progestagen sponges. The ewes were assigned randomly to 3 groups. On d 6 to 12 of their estrous cycle, the control (CON) group received a maintenance diet (1.0×M), the supplemented (SUP) group received 1.5×M, and the restricted (R) group received 0.5×M. On d 7 to 12, blood samples were taken. The sheep were slaughtered at the end of the treatment, and their organs and ovaries were collected. The plasma concentrations of urea (P2.5 mm. Follicle size affected the mRNA expression of very low density lipoprotein receptor (VLDLR), estrogen receptor 2 (ESR2), FSH receptor (FSHR), CYP17A1, and CYP19A1 (Pgrowth may involve responses to disrupted reproductive hormone concentrations and influenced the intrafollicular expression of CYP17A1, CYP19A1, and ESR1. This result may be due to increased plasma urea and lipid concentrations. PMID:24045481

  5. Synthesis and biological evaluation of [18F]tetrafluoroborate: a PET imaging agent for thyroid disease and reporter gene imaging of the sodium/iodide symporter

    International Nuclear Information System (INIS)

    The human sodium/iodide symporter (hNIS) is a well-established target in thyroid disease and reporter gene imaging using gamma emitters 123I-iodide, 131I-iodide and 99mTc-pertechnetate. However, no PET imaging agent is routinely available. The aim of this study was to prepare and evaluate 18F-labelled tetrafluoroborate ([18F]TFB) for PET imaging of hNIS. [18F]TFB was prepared by isotopic exchange of BF4- with [18F]fluoride in hot hydrochloric acid and purified using an alumina column. Its identity, purity and stability in serum were determined by HPLC, thin-layer chromatography (TLC) and mass spectrometry. Its interaction with NIS was assessed in vitro using FRTL-5 rat thyroid cells, with and without stimulation by thyroid-stimulating hormone (TSH), in the presence and absence of perchlorate. Biodistribution and PET imaging studies were performed using BALB/c mice, with and without perchlorate inhibition. [18F]TFB was readily prepared with specific activity of 10 GBq/mg. It showed rapid accumulation in FRTL-5 cells that was stimulated by TSH and inhibited by perchlorate, and rapid specific accumulation in vivo in thyroid (SUV = 72 after 1 h) and stomach that was inhibited 95% by perchlorate. [18F]TFB is an easily prepared PET imaging agent for rodent NIS and should be evaluated for hNIS PET imaging in humans. (orig.)

  6. Inhibitory effect of novobiocin on ribonucleic acid synthesis during germination of Bacillus subtilis spores.

    OpenAIRE

    Matsuda, M; Kameyama, T

    1980-01-01

    Novobiocin inhibited ribonculeic acid synthesis during germination of Bacillus subtilis spores. Transcription of certain kinds of genes probably required a preceding conformational change in deoxyribonucleic acid.

  7. Base substitution mutations in uridinediphosphate-dependent glycosyltransferase 76G1 gene of Stevia rebaudiana causes the low levels of rebaudioside A: mutations in UGT76G1, a key gene of steviol glycosides synthesis.

    Science.gov (United States)

    Yang, Yong-Heng; Huang, Su-Zhen; Han, Yu-Lin; Yuan, Hai-Yan; Gu, Chun-Sun; Zhao, Yan-Hai

    2014-07-01

    Steviol glycosides, extracted from the leaves of Stevia rebaudiana (Bert) Bertoni, are calorie-free sugar substitute of natural origin with intensely sweet (Boileau et al., 2012). Stevioside and rebaudioside A are the two main kinds of the diterpenic glycosides. We analyzed the concentration of stevioside and rebaudioside A in Stevia leaves of about 500 samples (hybrid progenies) and discovered a mutation plant "Z05" with very low levels of rebaudioside A. Because UGT76G1, a uridinediphosphate-dependent glycosyltransferases, is responsible for the conversion from stevioside to rebaudioside A (Richman et al., 2005), so mutation identification was done by sequencing the candidate gene, UGT76G1. In this study molecular analysis of two strains revealed a heterozygotic nonsense mutation of c.389T > G (p.L121X) in UGT76G1. Meanwhile, we found some amino acid substitutions significant change the protein structure. And the difference of enzyme activity between two strains proved the lack of functionality of UGT76G1 of the mutation "Z05". So the nonsense mutation and amino acid substitution mutation resulted in the low levels of rebaudioside A. PMID:24811677

  8. Synthesis and biological evaluation of [{sup 18}F]tetrafluoroborate: a PET imaging agent for thyroid disease and reporter gene imaging of the sodium/iodide symporter

    Energy Technology Data Exchange (ETDEWEB)

    Jauregui-Osoro, Maite; Sunassee, Kavitha; Weeks, Amanda J.; Berry, David J.; Paul, Rowena L.; Cleij, Marcel; O' Doherty, Michael J.; Marsden, Paul K.; Szanda, Istvan; Blower, Philip J. [King' s College London, Division of Imaging Sciences, London (United Kingdom); Banga, Jasvinder Paul [King' s College London, Division of Cell and Gene Based Therapy, London (United Kingdom); Clarke, Susan E.M.; Ballinger, James R. [Guy' s and St Thomas' NHS Trust, Department of Nuclear Medicine, London (United Kingdom); Cheng, Sheue-Yann [National Cancer Institute, Laboratory of Molecular Biology, Bethesda (United States)

    2010-11-15

    The human sodium/iodide symporter (hNIS) is a well-established target in thyroid disease and reporter gene imaging using gamma emitters {sup 123}I-iodide, {sup 131}I-iodide and {sup 99m}Tc-pertechnetate. However, no PET imaging agent is routinely available. The aim of this study was to prepare and evaluate {sup 18}F-labelled tetrafluoroborate ([{sup 18}F]TFB) for PET imaging of hNIS. [{sup 18}F]TFB was prepared by isotopic exchange of BF{sub 4} {sup -} with [{sup 18}F]fluoride in hot hydrochloric acid and purified using an alumina column. Its identity, purity and stability in serum were determined by HPLC, thin-layer chromatography (TLC) and mass spectrometry. Its interaction with NIS was assessed in vitro using FRTL-5 rat thyroid cells, with and without stimulation by thyroid-stimulating hormone (TSH), in the presence and absence of perchlorate. Biodistribution and PET imaging studies were performed using BALB/c mice, with and without perchlorate inhibition. [{sup 18}F]TFB was readily prepared with specific activity of 10 GBq/mg. It showed rapid accumulation in FRTL-5 cells that was stimulated by TSH and inhibited by perchlorate, and rapid specific accumulation in vivo in thyroid (SUV = 72 after 1 h) and stomach that was inhibited 95% by perchlorate. [{sup 18}F]TFB is an easily prepared PET imaging agent for rodent NIS and should be evaluated for hNIS PET imaging in humans. (orig.)

  9. Design,synthesis and expression of TRAIL genes%TRAIL胞外区编码基因的设计、人工合成及其表达

    Institute of Scientific and Technical Information of China (English)

    张英莉; 林陈水

    2011-01-01

    According to the preferential codons of E. coli, a DNA fraction encoding the extracellular domain of TRAIL(114~281aa)was designed. Splicing primers were synthesized and ligased, and the target gene was obtained. The DNA fraction was amplified by PCR and coloned into the T vector. After sequenced, the fraction was added to a sequence encoding enterokinase recognised site (EKsite) at 5', terminal, and then recombined to the intracellular fusing expressional T-Vector. The recombinant expression plasmid pDsbA-6his-EKsite-TRAIL was constructed and transformed into E. coli BL21 (DE3). The recombinant strains were expressed under 0.4 mmol/L IPTG induction at 33 ℃. The whole cell was analyzed by SDS-PAGE, and the result showed that there was a band at 44 KD on the SDS-PAGE with an expected molecular weight.%根据大肠杆菌密码子偏爱性要求,设计编码 TRAIL 胞外段(114~281氨基酸)的DNA序列,分段合成拼接引物并连接,得到目的基因,PCR 扩增后克隆于T载体中,经测序证实后,PCR 法在目的基因的5',末端引入肠激酶识别位点 EKsite 的编码序列,重纽于胞内融合表达型T栽体中,构建成pDsbA-6His-EKsite-TRAIL 胞内融合表达质粒,重组质粒转化表达宿主E.coli BL21(DE3).在33℃条件下,添加终浓度为0.4 mmol/L 的 IPTG 诱导表达,全菌 SDS-PAGE 分析结果表明:所构建的工程菌能表达44 kD左右的 TRAIL 融合蛋白,与理论值相符.

  10. Genetic Synthesis of Periodic Protein Materials

    OpenAIRE

    Fournier, M J; Creel, H. S.; Krejchi, M. T.; Mason, T L; Tirrell, D A; McGrath, K. P.; Atkins, E. D. T.

    1991-01-01

    Genetic engineering offers a novel approach to the development of advanced polymeric materials, in particular protein-based materials. Biological synthesis provides levels of control of polymer chain architecture that cannot yet be attained by current methods of chemical synthesis. In addition to employing naturally occurring genetic templates artificial genes can be designed to encode completely new materials with customized properties. In the present paper we: 1) review th...

  11. Prebiotic RNA Synthesis by Montmorillonite Catalysis

    OpenAIRE

    2014-01-01

    This review summarizes our recent findings on the role of mineral salts in prebiotic RNA synthesis, which is catalyzed by montmorillonite clay minerals. The clay minerals not only catalyze the synthesis of RNA but also facilitate homochiral selection. Preliminary data of these findings have been presented at the “Horizontal Gene Transfer and the Last Universal Common Ancestor (LUCA)” conference at the Open University, Milton Keynes, UK, 5–6 September 2013. The objective of this meeting was to...

  12. Regulation of phospholipid synthesis in yeast

    OpenAIRE

    Carman, George M.; Han, Gil-Soo

    2009-01-01

    Phospholipid synthesis in the yeast Saccharomyces cerevisiae is a complex process that involves regulation by both genetic and biochemical mechanisms. The activity levels of phospholipid synthesis enzymes are controlled by gene expression (e.g., transcription) and by factors (lipids, water-soluble phospholipid precursors and products, and covalent modification of phosphorylation) that modulate catalysis. Phosphatidic acid, whose levels are controlled by the biochemical regulation of key phosp...

  13. Sterol synthesis in diverse bacteria

    Directory of Open Access Journals (Sweden)

    Jeremy H Wei

    2016-06-01

    Full Text Available Sterols are essential components of eukaryotic cells whose biosynthesis and function has been studied extensively. Sterols are also recognized as the diagenetic precursors of steranes preserved in sedimentary rocks where they can function as geological proxies for eukaryotic organisms and/or aerobic metabolisms and environments. However, production of these lipids is not restricted to the eukaryotic domain as a few bacterial species also synthesize sterols. Phylogenomic studies have identified genes encoding homologs of sterol biosynthesis proteins in the genomes of several additional species, indicating that sterol production may be more widespread in the bacterial domain than previously thought. Although the occurrence of sterol synthesis genes in a genome indicates the potential for sterol production, it provides neither conclusive evidence of sterol synthesis nor information about the composition and abundance of basic and modified sterols that are actually being produced. Here, we coupled bioinformatics with lipid analyses to investigate the scope of bacterial sterol production. We identified oxidosqualene cyclase (Osc, which catalyzes the initial cyclization of oxidosqualene to the basic sterol structure, in 34 bacterial genomes from 5 phyla (Bacteroidetes, Cyanobacteria, Planctomycetes, Proteobacteria and Verrucomicrobia and in 176 metagenomes. Our data indicate that bacterial sterol synthesis likely occurs in diverse organisms and environments and also provides evidence that there are as yet uncultured groups of bacterial sterol producers. Phylogenetic analysis of bacterial and eukaryotic Osc sequences confirmed a complex evolutionary history of sterol synthesis in this domain. Finally, we characterized the lipids produced by Osc-containing bacteria and found that we could generally predict the ability to synthesize sterols. However, predicting the final modified sterol based on our current knowledge of sterol synthesis was difficult

  14. Organic Synthesis

    OpenAIRE

    Romea, Pedro

    2014-01-01

    Organic Synthesis is a one-semester course of the fourth year of the Chemistry Degree at the Universitat de Barcelona. This course covers the most important transformations in Organic Chemistry, including a short introduction to the Retrosynthetic Analysis. The aim is to provide a solid knowledge of the main reactions and their mechanism, which could later be improved during Master studies.

  15. Cellulose Synthesis in Agrobacterium tumefaciens

    Energy Technology Data Exchange (ETDEWEB)

    Alan R. White; Ann G. Matthysse

    2004-07-31

    We have cloned the celC gene and its homologue from E. coli, yhjM, in an expression vector and expressed the both genes in E. coli; we have determined that the YhjM protein is able to complement in vitro cellulose synthesis by extracts of A. tumefaciens celC mutants, we have purified the YhjM protein product and are currently examining its enzymatic activity; we have examined whole cell extracts of CelC and various other cellulose mutants and wild type bacteria for the presence of cellulose oligomers and cellulose; we have examined the ability of extracts of wild type and cellulose mutants including CelC to incorporate UDP-14C-glucose into cellulose and into water-soluble, ethanol-insoluble oligosaccharides; we have made mutants which synthesize greater amounts of cellulose than the wild type; and we have examined the role of cellulose in the formation of biofilms by A. tumefaciens. In addition we have examined the ability of a putative cellulose synthase gene from the tunicate Ciona savignyi to complement an A. tumefaciens celA mutant. The greatest difference between our knowledge of bacterial cellulose synthesis when we started this project and current knowledge is that in 1999 when we wrote the original grant very few bacteria were known to synthesize cellulose and genes involved in this synthesis were sequenced only from Acetobacter species, A. tumefaciens and Rhizobium leguminosarum. Currently many bacteria are known to synthesize cellulose and genes that may be involved have been sequenced from more than 10 species of bacteria. This additional information has raised the possibility of attempting to use genes from one bacterium to complement mutants in another bacterium. This will enable us to examine the question of which genes are responsible for the three dimensional structure of cellulose (since this differs among bacterial species) and also to examine the interactions between the various proteins required for cellulose synthesis. We have carried out one

  16. Genetics Home Reference: congenital bile acid synthesis defect type 2

    Science.gov (United States)

    ... DEFECT, CONGENITAL, 2 Sources for This Page Clayton PT. Disorders of bile acid synthesis. J Inherit Metab ... J, Duran M, Overmars H, Scambler PJ, Clayton PT. Mutations in SRD5B1 (AKR1D1), the gene encoding delta( ...

  17. The research and application of TPO's gene

    International Nuclear Information System (INIS)

    Thyro-peroxidase (TPO) is a glycosylated protein bound to the apical plasma membrane of thyrocytes. It is the key enzyme in the synthesis of thyroid hormones. Its gene structure and transcriptional regulation have been deeply studied. The author reviews the development of TPO's gene structure, function, transcriptional regulation, the relationship between TPO with thyroid diseases and radioactive iodide therapy

  18. Inactivation of Mg Chelatase during Transition from Anaerobic to Aerobic Growth in Rhodobacter capsulatus

    OpenAIRE

    Willows, Robert D; Lake, Vanessa; Roberts, Thomas Hugh; Beale, Samuel I.

    2003-01-01

    The facultative photosynthetic bacterium Rhodobacter capsulatus can adapt from an anaerobic photosynthetic mode of growth to aerobic heterotrophic metabolism. As this adaptation occurs, the cells must rapidly halt bacteriochlorophyll synthesis to prevent phototoxic tetrapyrroles from accumulating, while still allowing heme synthesis to continue. A likely control point is Mg chelatase, the enzyme that diverts protoporphyrin IX from heme biosynthesis toward the bacteriochlorophyll biosynthetic ...

  19. Visual gene developer: a fully programmable bioinformatics software for synthetic gene optimization

    Directory of Open Access Journals (Sweden)

    McDonald Karen

    2011-08-01

    Full Text Available Abstract Background Direct gene synthesis is becoming more popular owing to decreases in gene synthesis pricing. Compared with using natural genes, gene synthesis provides a good opportunity to optimize gene sequence for specific applications. In order to facilitate gene optimization, we have developed a stand-alone software called Visual Gene Developer. Results The software not only provides general functions for gene analysis and optimization along with an interactive user-friendly interface, but also includes unique features such as programming capability, dedicated mRNA secondary structure prediction, artificial neural network modeling, network & multi-threaded computing, and user-accessible programming modules. The software allows a user to analyze and optimize a sequence using main menu functions or specialized module windows. Alternatively, gene optimization can be initiated by designing a gene construct and configuring an optimization strategy. A user can choose several predefined or user-defined algorithms to design a complicated strategy. The software provides expandable functionality as platform software supporting module development using popular script languages such as VBScript and JScript in the software programming environment. Conclusion Visual Gene Developer is useful for both researchers who want to quickly analyze and optimize genes, and those who are interested in developing and testing new algorithms in bioinformatics. The software is available for free download at http://www.visualgenedeveloper.net.

  20. Effect of UVB radiation on melanocytes and melanin synthesis-related gene expression%UVB 辐射对黑色素细胞黑素合成及其相关基因表达的影响

    Institute of Scientific and Technical Information of China (English)

    吴姗姗; 许欢欢; 朱丹丹; 居博伟; 闫瑶; 杨建华

    2016-01-01

    -2 in cells.Conclusion UVB radiation has a positive effect on the regulating genes expression which are related to the melanin synthesis pathway and the melanin significantly increased in the cells,which provide more information for constructing pig-mentation models .

  1. 植物花青素生物合成途径相关基因研究进展及其基因工程修饰%Advances in research of genes involved in anthocyanin biological synthesis in plant and the genetic modification of the pathway

    Institute of Scientific and Technical Information of China (English)

    赵德勇

    2012-01-01

    This paper reviews the advances in research of synthetic genes and regulator genes involved in the anthocyanin biological synthesis process as well as in genetic engineering in regulating the anthocyanin biological synthesis. Anthocyanin biological synthesis process of plants belongs to the secondary metabolic pathway, regulates the expression of key enzymes involved in the pathway, and could hence lead to a reduced or increased yield of target compound. Genetic improvement of plants may be realized through modifying the secondary metabolic process. Anthocyanin accumulation helps the plants to act against the UV Further study on the defense molecular mechanism of the anthocyanin facilitates b with resistance to diseases and adversities. radiation, insects and fungi. reeding of new plant cultivars%对植物花青素生物合成及调控基因的研究进展、基因工程在调控花青素合成途径中的应用进行了综述。植物花青素生物合成属次生代谢途径,对该途径关键酶基因的调控可降低或提高目标化合物的产量,可通过调控植物次生代谢的方式对植物进行遗传改良。对植物通过积累花青素来适应紫外线辐射、防卫害虫及真菌侵害的分子机制进行研究,有助于培育抗病、抗逆的植物新品种。

  2. Analysis of Gene Expression Profile in Lung Adenosquamous Carcinoma Using cDNA Microarray

    Institute of Scientific and Technical Information of China (English)

    YANG Fei; YANG Jiong; JIANG Man; YE Bo; ZHANG Yu-xia; CHEN Hong-lei; XIA Dong; LIU Ming-qiu

    2004-01-01

    Gene expression profile of the lung adenosquamous carcinoma was characterized by using cDNA microarray chip containing 4 096 human genes. Among target genes, 508 differentially expressed genes were identified in adenosquamous carcinoma of the lung, 232 genes were overexpressed and 276 genes were underexpressed. Among them, 92 genes are cell signals transduction genes, 34 genes are proto-oncogenes and tumor suppressor genes or cell cycle related genes or cell apoptosis related genes, 29 genes are cell skeleton genes, 28 genes are DNA synthesis, repair and recombination genes, 12 genes are DNA binding and transcription genes. These genes may be associated with the occurence and development of adenosquamous carinome of the lung.

  3. Nucleolar dominance and ribosomal RNA gene silencing

    OpenAIRE

    Tucker, Sarah; Vitins, Alexa; Pikaard, Craig S.

    2010-01-01

    Nucleolar dominance is an epigenetic phenomenon that occurs in genetic hybrids and describes the expression of 45S rRNA genes inherited from one progenitor due to the silencing of the other progenitor’s rRNA genes. Nucleolar dominance is a manifestation of rRNA gene dosage control, which also occurs in non-hybrids, regulating the number of active rRNA genes according to the cellular demand for ribosomes and protein synthesis. Ribosomal RNA gene silencing involves changes in DNA methylation an...

  4. Evaluation of flux synthesis algorithms

    International Nuclear Information System (INIS)

    The flux synthesis algorithm which is the best fit to the numerical solution of the multigroup diffusion equations, was determined. Three different types of synthesis were studied: 1) discontinuous synthesis 2) continuous synthesis 3) pseudo-continuous synthesis. A matrix and a differential formulation were developed for the first two types of synthesis. For pseudo-continuous synthesis only the matrix formulation was used. Some tests were performed and the results allowed us to establish the following order of efficiency for the algorithms: 1) continuous synthesis (matrix formulation) 2) continuous synthesis (differential formulation) 3) pseudo-continuous synthesis 4) discontinuous synthesis (matrix formulation) 5) discontinuous synthesis (differential formulation). (Author)

  5. The Mutational Spectrum of Holoprosencephaly-Associated Changes within the SHH Gene in Humans Predicts Loss-of-Function Through Either Key Structural Alterations of the Ligand or Its Altered Synthesis

    OpenAIRE

    Roessler, Erich; El-Jaick, Kenia B.; Dubourg, Christèle; Vélez, Jorge I.; Solomon, Benjamin D.; Pineda-Álvarez, Daniel E.; Lacbawan, Felicitas; Zhou, Nan; Ouspenskaia, Maia; Paulussen, Aimée; Smeets, Hubert J.; Hehr, Ute; Bendavid, Claude; Bale, Sherri; Odent, Sylvie

    2009-01-01

    Mutations within either the SHH gene or its related pathway components are the most common, and best understood, pathogenetic changes observed in holoprosencephaly patients; this fact is consistent with the essential functions of this gene during forebrain development and patterning. Here we summarize the nature and types of deleterious sequence alterations among over one hundred distinct mutations in the SHH gene (64 novel mutations) and compare these to over a dozen mutations in disease-rel...

  6. Gene co-expression network analysis in Rhodobacter capsulatus and application to comparative expression analysis of Rhodobacter sphaeroides

    Energy Technology Data Exchange (ETDEWEB)

    Pena-Castillo, Lourdes; Mercer, Ryan; Gurinovich, Anastasia; Callister, Stephen J.; Wright, Aaron T.; Westbye, Alexander; Beatty, J. T.; Lang, Andrew S.

    2014-08-28

    annotation. We identified R. capsulatus modules enriched with genes for ribosomal proteins, porphyrin and bacteriochlorophyll anabolism, and biosynthesis of secondary metabolites to be preserved in R. sphaeroides whereas modules related to RcGTA production and signalling showed lack of preservation in R. sphaeroides. In addition, we demonstrated that network statistics may also be applied within-species to identify congruence between mRNA expression and protein abundance data for which simple correlation measurements have previously had mixed results.

  7. Expression data on liver metabolic pathway genes and proteins

    OpenAIRE

    Mooli Raja Gopal Reddy; Chodisetti Pavan Kumar; Malleswarapu Mahesh; Manchiryala Sravan Kumar; Jeyakumar, Shanmugam M

    2016-01-01

    Here, we present the expression data on various metabolic pathways of liver with special emphasize on lipid and carbohydrate metabolism and long chain polyunsaturated fatty acid (PUFA) synthesis, both at gene and protein levels. The data were obtained to understand the effect of vitamin A deficiency on the expression status (both gene and protein levels) of some of the key factors involved in lipogenesis, fatty acid oxidation, triglyceride secretion, long chain PUFA, resolvin D1 synthesis, gl...

  8. Plastid protein synthesis is required for plant development in tobacco

    OpenAIRE

    Ahlert, Daniela; Ruf, Stephanie; Bock, Ralph

    2003-01-01

    Chloroplasts fulfill important functions in cellular metabolism. The majority of plastid genome-encoded genes is involved in either photosynthesis or chloroplast gene expression. Whether or not plastid genes also can determine extraplastidic functions has remained controversial. We demonstrate here an essential role of plastid protein synthesis in tobacco leaf development. By using chloroplast transformation, we have developed an experimental system that produces recombination-based knockouts...

  9. Muscle Biological Characteristics of Differentially Expressed Genes in Wujin and Landrace Pigs

    Institute of Scientific and Technical Information of China (English)

    XU Hong; HUANG Ying; LI Wei-zhen; YANG Ming-hua; GE Chang-rong; ZHANG Xi; LI Liu-an; GAO Shi-zheng; ZHAO Su-mei

    2014-01-01

    The biological chemistry would be responsible for the meat quality. This study tried to investigate the transcript expression proifle and explain the characteristics of differentially expressed genes between the Wujin and Landrace pigs. The results showed that 526 differentially expressed genes were found by comparing the transcript expression proifle of muscle tissue between Wujin and Landrace pigs. Among them, 335 genes showed up-regulations and 191 genes showed down-regulations in Wujin pigs compared with the Landrace pigs. Gene ontology (GO) analysis indicated that the differentially expressed genes were clustered into three groups involving in protein synthesis, energy metabolism and immune response. Kyoto encyclopedia of genes and genomes (KEGG) pathway analysis found that these differentially expressed genes participated in protein synthesis metabolism, energy metabolism and immune response pathway. The Database for Annotation, Visualization and Integrated Discovery (DAVID) analysis of protein function and protein domains function also conifrmed that differentially expressed genes belonged to protein synthesis, energy metabolism and immune response. Genes related protein synthesis metabolism pathway in Landrace was higher than in Wujin pigs. However, differentially expressed genes related energy metabolism and immune response was up-regulated in Wujin pigs compared with Landrace pigs. Quantitative real-time RT-PCR on selected genes was used to conifrm the results from the microarray. These suggested that the genes related to protein synthesis, energy metabolism and immune response would contribute to the growth performance, meat quality as well as anti-disease capacity.

  10. 奶牛乳糖合成及泌乳相关基因和细胞信号通路的研究进展%Research Progress on Genes and Signal Pathways Associated with Lactose Synthesis and Lactation in Dairy Cows

    Institute of Scientific and Technical Information of China (English)

    李文清; 王加启; 南雪梅; 孙鹏

    2012-01-01

    Lactose is a special component of milk and is closely related to milk yield. The factors of influencing lactose synthesis are extensive. The paper reviews the related genes and cell signaling pathways, including glucose transporters, intracel-lular catalytic lactose synthesis enzymes, hormones (insulin, prolactin, growth hormone, leptin, glucocorticoid ), cell factors (IGF-Ⅰ ,IGF-Ⅱ ,EGF) and its main cell signal transduction pathway.%乳糖是乳中的特有成分,与产奶量密切相关.影响牛奶中乳糖合成的因素很多,作者主要综述与其相关基因,包括乳腺葡萄糖转运蛋白、细胞内催化乳糖合成的酶类、与泌乳和乳糖合成相关的激素(胰岛素、催乳素、生长激素、糖皮质激素、瘦素)、细胞因子(IGF-Ⅰ、IGF-Ⅱ、EGF)及其主要的信号传导通路的研究进展.

  11. Overall Synthesis

    International Nuclear Information System (INIS)

    AMIGO is an OECD/NEA international project on the topic of 'Approaches and Methods for Integrating Geological Information in the Safety Case'. The term safety case here refers to the post-closure safety case for a geological repository for long-lived radioactive waste, and is defined as a synthesis of evidence, analyses and arguments that quantify and substantiate a claim that the repository is safe. Geological or geo-scientific information includes the various types of geophysical, hydrogeological, geochemical and structural information that can contribute to the safety case. The safety case is generally updated periodically throughout the step-wise process of repository siting, planning, construction, operation, as well as prior to closure, and becomes more rigorous over time, as increasing amounts of geological and other data become available, until, for a well-chosen site and design, a point is reached at which the safety case is adequate for repository licensing. AMIGO is structured as a series of workshops. This document summarises the first workshop of the AMIGO series, held at Yverdon-les-Bains, Switzerland, on 3-5 June 2003, a-nd-hosted The focus of the workshop was on 'building confidence (in analyses and arguments that support the safety case) using multiple lines of evidence', but other themes within the overall scope of AMIGO were also discussed, such as the integration of the work of geo-scientists and safety assessors. The main themes addressed by the workshop, which include the topics covered by the Working Group Sessions, can be stated as follows: the role of the geosphere in disposal concepts; the ways in which geological information is used by waste management programmes, and the way in which usage changes as a programme progresses; the synthesis of wide ranging geo-scientific information into a consistent site description or conceptual model; the development of arguments for the long-term safety of disposal systems; the use of multiple lines of

  12. Co-ordinate control of synthesis of mitochondrial and non-mitochondrial hemoproteins: a binding site for the HAP1 (CYP1) protein in the UAS region of the yeast catalase T gene (CTT1).

    OpenAIRE

    Winkler, H.; Adam, G.; Mattes, E; Schanz, M.; Hartig, A; Ruis, H

    1988-01-01

    Control of expression of the Saccharomyces cerevisiae CTT1 (catalase T) gene by the HAP1 (CYP1) gene, a mediator of heme control of mitochondrial cytochromes, was studied. Expression of a CTT1-lacZ fusion in a hap1 mutant showed that the CTT1 promoter is under HAP1 control. As demonstrated by a gel retardation assay, the HAP1 protein binds to a heme control region of the CTT1 gene. This binding in vitro is stimulated by hemin. The HAP1-binding sequence was localized by using DNA fragments spa...

  13. Aerobic Anoxygenic Phototrophic Bacteria

    OpenAIRE

    Yurkov, Vladimir V.; Beatty, J. Thomas

    1998-01-01

    The aerobic anoxygenic phototrophic bacteria are a relatively recently discovered bacterial group. Although taxonomically and phylogenetically heterogeneous, these bacteria share the following distinguishing features: the presence of bacteriochlorophyll a incorporated into reaction center and light-harvesting complexes, low levels of the photosynthetic unit in cells, an abundance of carotenoids, a strong inhibition by light of bacteriochlorophyll synthesis, and the inability to grow photosynt...

  14. Mechanistic approaches to the study of evolution: the functional synthesis

    OpenAIRE

    Dean, Antony M; Joseph W Thornton

    2007-01-01

    An emerging synthesis of evolutionary biology and experimental molecular biology is providing much stronger and deeper inferences about the dynamics and mechanisms of evolution than were possible in the past. The new approach combines statistical analyses of gene sequences with manipulative molecular experiments to reveal how ancient mutations altered biochemical processes and produced novel phenotypes. This functional synthesis has set the stage for major advances in our understanding of fun...

  15. Genes, stress, and depression.

    Science.gov (United States)

    Wurtman, Richard J

    2005-05-01

    A relationship between genetic makeup and susceptibility to major depressive disorder (MDD) has long been suspected on the basis of family and twin studies. A metaanalysis of reports on the basis of twin studies has estimated MDD's degree of heritability to be 0.33 (confidence interval, 0.26-0.39). Among families exhibiting an increased prevalence of MDD, risk of developing the illness was enhanced in members exposed to a highly stressful environment. Aberrant genes can predispose to depression in a number of ways, for example, by diminishing production of growth factors that act during brain development. An aberrant gene could also increase or decrease a neurotransmitter's release into synapses, its actions, or its duration of activity. The gene products of greatest interest at present are those involved in the synthesis and actions of serotonin; among them, the serotonin-uptake protein localized within the terminals and dendrites of serotonin-releasing neurons. It has been found that the Vmax of platelet serotonin uptake is low in some patients with MDD; also, Vmax is highly correlated in twins. Antidepressant drugs such as the selective serotonin reuptake inhibitors act on this uptake protein. The specific genetic locus causing serotonin uptake to be lower in some patients with major depression involves a polymorphic region (5-HTTLPR) in the promoter region of the gene for the uptake protein. The gene itself exists as several alleles, the short "S" allele and the long "L" allele. The S variant is associated with less, and the L variant with more, of the uptake protein. The effect of stressful life events on depressive symptoms in young adults was found to be significantly stronger among SS or SL subjects than among LL subjects. Neuroimaging studies showed that people with the SS or SL alleles exhibited a greater activation of the amygdala in response to fearful stimuli than those with LL. It has been reported recently that mutations in the gene that controls

  16. 灵芝甾醇14α-脱甲基酶基因的克隆及超量表达对三萜合成的影响%Cloning of a sterol 14α-demethylase gene and the effects of over-expression of the gene on biological synthesis of triterpenes in Ganoderma lucidum

    Institute of Scientific and Technical Information of China (English)

    方星; 师亮; 徐颖洁; 赵明文

    2011-01-01

    Ganoderma lucidum has been used for centuries to cure various human diseases in our country, and triterpenoids are the most important pharmacologically active constituents of the fungus. Sterol 14a-demethylase (CYP51) is one of the key enzymes involved in the biological synthesis processes of triterpenes. Degenerate primers were designed according to conservative sites of protein sequences from related species and a specific DNA fragment was obtained, then full length of Gl-cyp51 was obtained using traditional methods. Genomic DNA was 1,981bp and cDNA was 1,635bp. The ORF encoded a 544-amino acid polypeptide with a theoretical pI of 6.36 and a theoretical molecular mass of 61.99kDa. The Gl-cyp51 complete cDNA was ligated to the plasmid pG1-GPD. By successful Agrobacterium tumefaciens mediated transformation to G. lucidum, we realized Gl-cyp51 over expression transforments. We found that the transcript level of Gl-cyp51 was over expressed and triterpenes production was mcrased. Further more, the transcript level of genes (Gl-aact, Gl-hmgr and Gl-ls) involved in the biosynthesis of triterpenes were also increased.%灵芝Ganoderma lucidum是我国传统的药用真菌,三萜类物质是灵芝的主要生物活性成分,甾醇14α-脱甲基酶是三萜合成途径中的关键酶.根据已报道其他物种甾醇14α-脱甲基酶的氨基酸保守序列设计简并引物,获得灵芝甾醇14α-脱甲基酶特异基因片段,并进一步获得灵芝甾醇14α-脱甲基酶基因的全长DNA和cDNA序列.其中DNA序列长1,981bp,cDNA序列长1,635bp.结构基因编码蛋白包含544个氨基酸,分子量为61.99kDa,等电点为6.36.将甾醇14α-脱甲基酶基因的cDNA序列克隆剑灵芝超量表达载体pGl-GPD中,利用农杆菌介导的转化法实现了甾醇14α-脱甲基酶基因在灵芝内的超量表达.转化子的甾醇14α-脱甲基酶基因在转录水平表达量增加,三萜含量增加.进一步研究发现,三萜合成途径的关键酶基因Gl-aact

  17. Synthesis Organic Compound

    OpenAIRE

    Rasyid, Herlina; Firdaus; Hariani, Nunuk

    2015-01-01

    Abstract. Synthesis of metil ??-(p-hidroksifenil)akrilic from ??-(p-hidroksifenil)akrilat acid and methanol using Dean Stark Trap method had been done. Synthesis of ths compound intended to form the starting material in the subsequent synthesis of amide???s compound through the formation of ester compound. This synthesis using H2SO4 catalyst and Dean Stark Trap method, some of benzena which is added to remove the water that resulting from the reaction. Synthesis of this compound be held at re...

  18. Substructural controller synthesis

    Science.gov (United States)

    Su, Tzu-Jeng; Craig, Roy R., Jr.

    1989-01-01

    A decentralized design procedure which combines substructural synthesis, model reduction, decentralized controller design, subcontroller synthesis, and controller reduction is proposed for the control design of flexible structures. The structure to be controlled is decomposed into several substructures, which are modeled by component mode synthesis methods. For each substructure, a subcontroller is designed by using the linear quadratic optimal control theory. Then, a controller synthesis scheme called Substructural Controller Synthesis (SCS) is used to assemble the subcontrollers into a system controller, which is to be used to control the whole structure.

  19. Transcription of interferon-stimulated genes is induced by adenovirus particles but is suppressed by E1A gene products

    Energy Technology Data Exchange (ETDEWEB)

    Reich, N.; Pine, R.; Levy, D.; Darnell, J.E. Jr.

    1988-01-01

    Interferon treatment of cell cultures results in the rapid transcriptional induction of a specific set of genes. In this paper the authors explore the effect of cellular infection by several adenoviruses, both wild type and mutant, on the expression of these genes. Infection with adenovirus induces the transcription of the interferon-stimulated genes in the absence of any protein synthesis. In fact, the inhibition of protein synthesis during a wild-type infection produces enhanced stimulation of transcription of these genes. Experiments with viral mutants indicate the ability to specifically suppress this transcription maps to the E1A gene. In addition, the E1A gene products are capable of suppressing the specific transcriptional induction of interferon-stimulated promoters during cotransfection experiments and therefore presumably during viral infection. The dual effect of adenovirus on the expression of interferon-stimulated genes may represent an example of action and evolutionary reaction between virus and host.

  20. Protein degradation and protein synthesis in long-term memory formation

    OpenAIRE

    Jarome, Timothy J.; Helmstetter, Fred J.

    2014-01-01

    Long-term memory (LTM) formation requires transient changes in the activity of intracellular signaling cascades that are thought to regulate new gene transcription and de novo protein synthesis in the brain. Consistent with this, protein synthesis inhibitors impair LTM for a variety of behavioral tasks when infused into the brain around the time of training or following memory retrieval, suggesting that protein synthesis is a critical step in LTM storage in the brain. However, evidence sugg...

  1. The mutational spectrum of holoprosencephaly-associated changes within the SHH gene in humans predicts loss-of-function through either key structural alterations of the ligand or its altered synthesis.

    Science.gov (United States)

    Roessler, Erich; El-Jaick, Kenia B; Dubourg, Christèle; Vélez, Jorge I; Solomon, Benjamin D; Pineda-Alvarez, Daniel E; Lacbawan, Felicitas; Zhou, Nan; Ouspenskaia, Maia; Paulussen, Aimée; Smeets, Hubert J; Hehr, Ute; Bendavid, Claude; Bale, Sherri; Odent, Sylvie; David, Véronique; Muenke, Maximilian

    2009-10-01

    Mutations within either the SHH gene or its related pathway components are the most common, and best understood, pathogenetic changes observed in holoprosencephaly patients; this fact is consistent with the essential functions of this gene during forebrain development and patterning. Here we summarize the nature and types of deleterious sequence alterations among over one hundred distinct mutations in the SHH gene (64 novel mutations) and compare these to over a dozen mutations in disease-related Hedgehog family members IHH and DHH. This combined structural analysis suggests that dysfunction of Hedgehog signaling in human forebrain development can occur through truncations or major structural changes to the signaling domain, SHH-N, as well as due to defects in the processing of the mature ligand from its pre-pro-precursor or defective post-translation bi-lipid modifications with palmitate and cholesterol. PMID:19603532

  2. Survey On Speech Synthesis

    Directory of Open Access Journals (Sweden)

    A. Indumathi

    2012-12-01

    Full Text Available The primary goal of this paper is to provide an overview of existing Text-To-Speech (TTS Techniques by highlighting its usage and advantage. First Generation Techniques includes Formant Synthesis and Articulatory Synthesis. Formant Synthesis works by using individually controllable formant filters, which can be set to produce accurate estimations of the vocal-track transfer function. Articulatory Synthesis produces speech by direct modeling of Human articulator behavior. Second Generation Techniques incorporates Concatenative synthesis and Sinusoidal synthesis. Concatenative synthesis generates speech output by concatenating the segments of recorded speech. Generally, Concatenative synthesis generates the natural sounding synthesized speech. Sinusoidal Synthesis use a harmonic model and decompose each frame into a set of harmonics of an estimated fundamental frequency. The model parameters are the amplitudes and periods of the harmonics. With these, the value of the fundamental can be changed while keeping the same basic spectral..In adding, Third Generation includes Hidden Markov Model (HMM and Unit Selection Synthesis.HMM trains the parameter module and produce high quality Speech. Finally, Unit Selection operates by selecting the best sequence of units from a large speech database which matches the specification.

  3. Gene therapy

    Institute of Scientific and Technical Information of China (English)

    2005-01-01

    2005147 CNHK200-hA-a gene-viral therapeutic system and its antitumor effect on lung cancer. WANG Wei-guo(王伟国),et al. Viral & Gene Ther Center, Eastern Hepatobilli Surg Instit 2nd Milit Univ, Shanghai 200438. Chin J Oncol,2005:27(2):69-72. Objective: To develop a novel vector system, which combines the advantages of the gene therapy,

  4. Synthesis and preliminary evaluation of 9-(4-[{sup 18}F]fluoro-3-hydroxymethylbutyl) guanine ([{sup 18}F]FHBG) in HSV1-tk gene transduced hepatoma cell

    Energy Technology Data Exchange (ETDEWEB)

    Moon, Byung Seok; Lee, Tae Sup [Korea Institute of Radiological and Medical Sciences, Seoul (Korea, Republic of); Lee, Myoung Keun [Yonsei University, Wonju (Korea, Republic of)] (and others)

    2006-08-15

    The HSV1-tk reporter gene system is the most widely used system because of its advantage that direct monitoring is possible without the introduction of a separate reporter gene in case of HSV1-tk suicide gene therapy. In this study, we investigate the usefulness of the reporter probe (substrate), 9-(4-[{sup 18}F]fluoro-3-hydroxymethylbutyl) guanine ([{sup 18}F]FHBG) for non-invasive reporter gene imaging using PET in HSV1-tk expressing hepatoma model. Radiolabeled FHBG was prepared in 8 steps from a commercially available triester. The labeling reaction was carried out by NCA nucleophilic substitution with K[{sup 18}F]/K2.2.2. in acetonitrile using N2-monomethoxytrityl-9-[4-(tosly)-3-monomethoxytritylmethylbutl] guanine as a precursor, followed by deprotection with 1 N HCI. Preliminary biological properties of the probe were evaluated with MCA cells and MCA-tk cells transduced with HSV1-tk reporter gene. In vitro uptake and release-out studies of [{sup 18}F]FHBG were performed, and was analyzed correlation between [{sup 18}F]FHBG uptake ratio according to increasing numeric count of MCA-tk cells and degree of gene expression. MicroPET scan image was obtained with MCA and MCA-tk tumor beating Balb/c-nude mouse model. [{sup 18}F]FHBG was purified by reverse phase semi-HPLC system and collected at around 16-18 min. Radiochemical yield was about 20-25% (corrected for decay), radiochemical purity was > 95% and specific activity was around > 55.5 GBq/ {mu} mol. Specific accumulation of [{sup 18}F]FHBG was observed in HSV1-tk gene transduced MCA-tk cells but not MCA cells, and consecutive 1 hour release-out results showed more than 86% of uptaked [{sup 18}F]FHBG was retained inside of cells. The uptake of [{sup 18}F]FHBG was showed a highly significant linear correlation (R{sup 2} = 0.995) with increasing percentage of MCA-tk numeric cell count. In microPET scan images, remarkable difference of accumulation was observed for the two type of tumors. [{sup 18}F]FHBG appears

  5. On the bursting of gene products

    CERN Document Server

    Yvinec, Romain

    2011-01-01

    In this article we demonstrate that the so-called bursting production of molecular species during gene expression may be an artifact caused by low time resolution in experimental data collection and not an actual burst in production. We reach this conclusion through an analysis of a two-stage and binary model for gene expression, and demonstrate that in the limit when mRNA degradation is much faster than protein degradation they are equivalent. The negative binomial distribution is shown to be a limiting case of the binary model for fast "on to off" state transitions and high values of the ratio between protein synthesis and degradation rates. The gene products population increases by unity but multiple times in a time interval orders of magnitude smaller than protein half-life or the precision of the experimental apparatus employed in its detection. This rare-and-fast one-by-one protein synthesis has been interpreted as bursting.

  6. Organization of genes for tetrapyrrole biosynthesis in gram--positive bacteria.

    Science.gov (United States)

    Johansson, P; Hederstedt, L

    1999-03-01

    Clusters of genes encoding enzymes for tetrapyrrole biosynthesis were cloned from Bacillus sphaericus, Bacillus stearothermophilus, Brevibacillus brevis and Paenibacillus macerans. The sequences of all hemX genes found, and of a 6.3 kbp hem gene cluster from P. macerans, were determined. The structure of the hem gene clusters was compared to that of other Gram-positive bacteria. The Bacillus and Brevibacillus species have a conserved organization of the genes hemAXCDBL, required for biosynthesis of uroporphyrinogen III (UroIII) from glutamyl-tRNA. In P. macerans, the hem genes for UroIII synthesis are also closely linked but their organization is different: there is no hemX gene and the gene cluster also contains genes, cysG8 and cysG(A)-hemD, encoding the enzymes required for synthesis of sirohaem from UroIII. Bacillus subtilis contains genes for three proteins, NasF, YInD and YInF, with sequence similarity to Escherichia coli CysG, which is a multi-functional protein catalysing sirohaem synthesis from UroIII. It is shown that YInF is required for sirohaem synthesis and probably catalyses the precorrin-2 to sirohaem conversion. YInD probably catalyses precorrin-2 synthesis from UroIII and NasF seems to be specific for nitrite reduction. PMID:10217486

  7. Gene therapy.

    OpenAIRE

    Mota Biosca, Anna

    1992-01-01

    Applications of gene therapy have been evaluated in virtually every oral tissue, and many of these have proved successful at least in animal models. While gene therapy will not be used routinely in the next decade, practitioners of oral medicine should be aware of the potential of this novel type of treatment that doubtless will benefit many patients with oral diseases.

  8. Trichoderma genes

    Science.gov (United States)

    Foreman, Pamela; Goedegebuur, Frits; Van Solingen, Pieter; Ward, Michael

    2012-06-19

    Described herein are novel gene sequences isolated from Trichoderma reesei. Two genes encoding proteins comprising a cellulose binding domain, one encoding an arabionfuranosidase and one encoding an acetylxylanesterase are described. The sequences, CIP1 and CIP2, contain a cellulose binding domain. These proteins are especially useful in the textile and detergent industry and in pulp and paper industry.

  9. Origin of laccase gene structural diversity in edible mushrooms

    OpenAIRE

    Billette, Christophe; Gibard, Thierry; Foulongne Oriol, Marie; Savoie, Jean-Michel

    2011-01-01

    Laccase genes have been found in fungi, plants, insects and bacteria. In Basidiomycetes, the number of laccase genes ranges from 0 to 17. The role of these genes is not well known. It seems to be important in fungal interaction, development, melanine synthesis, human and plant pathogenesis, [ectomycorrhizal association and nutrition of the fungi]. Their role as ligninmodifying enzymes is controversial. Laccase phylogeny already published is not congruent with species phylogeny. Phylogeny of g...

  10. The Role of Nuclear Bodies in Gene Expression and Disease

    OpenAIRE

    Marie Morimoto; Boerkoel, Cornelius F.

    2013-01-01

    This review summarizes the current understanding of the role of nuclear bodies in regulating gene expression. The compartmentalization of cellular processes, such as ribosome biogenesis, RNA processing, cellular response to stress, transcription, modification and assembly of spliceosomal snRNPs, histone gene synthesis and nuclear RNA retention, has significant implications for gene regulation. These functional nuclear domains include the nucleolus, nuclear speckle, nuclear stress body, transc...

  11. Pancreatic expression of human insulin gene in transgenic mice.

    OpenAIRE

    Bucchini, D; Ripoche, M A; Stinnakre, M G; Desbois, P; Lorès, P; Monthioux, E; Absil, J; Lepesant, J A; Pictet, R; Jami, J

    1986-01-01

    We have investigated the possibility of obtaining integration and expression of a native human gene in transgenic mice. An 11-kilobase (kb) human chromosomal DNA fragment including the insulin gene (1430 base pairs) was microinjected into fertilized mouse eggs. This fragment was present in the genomic DNA of several developing animals. One transgenic mouse and its progeny were analyzed for expression of the foreign gene. Synthesis and release of human insulin was revealed by detection of the ...

  12. The genetic basis of evolutionary change in gene expression levels

    OpenAIRE

    Emerson, J. J.; Li, Wen-Hsiung

    2010-01-01

    The regulation of gene expression is an important determinant of organismal phenotype and evolution. However, the widespread recognition of this fact occurred long after the synthesis of evolution and genetics. Here, we give a brief sketch of thoughts regarding gene regulation in the history of evolution and genetics. We then review the development of genome-wide studies of gene regulatory variation in the context of the location and mode of action of the causative genetic changes. In particu...

  13. First step toward a virus-derived vector for gene cloning and expression in spiroplasmas, organisms which read UGA as a tryptophan codon: synthesis of chloramphenicol acetyltransferase in Spiroplasma citri.

    OpenAIRE

    Stamburski, C; Renaudin, J; Bove, J M

    1991-01-01

    Spiroplasmas are wall-less procaryotes in which the UGA codon serves not as a stop signal but as a code for the amino acid tryptophan. Spiroplasma genes that contain UGA codons thus cannot be studied in the usual Escherichia coli cloning and expression systems. Although this problem can be circumvented by using UGA-suppressor strains of E. coli, spiroplasmas themselves would provide a more efficient cloning and expression host. We have now successfully employed the replicative form (RF) of a ...

  14. Prebiotic RNA Synthesis by Montmorillonite Catalysis

    Directory of Open Access Journals (Sweden)

    Sohan Jheeta

    2014-08-01

    Full Text Available This review summarizes our recent findings on the role of mineral salts in prebiotic RNA synthesis, which is catalyzed by montmorillonite clay minerals. The clay minerals not only catalyze the synthesis of RNA but also facilitate homochiral selection. Preliminary data of these findings have been presented at the “Horizontal Gene Transfer and the Last Universal Common Ancestor (LUCA” conference at the Open University, Milton Keynes, UK, 5–6 September 2013. The objective of this meeting was to recognize the significance of RNA in LUCA. We believe that the prebiotic RNA synthesis from its monomers must have been a simple process. As a first step, it may have required activation of the 5'-end of the mononucleotide with a leaving group, e.g., imidazole in our model reaction (Figure 1. Wide ranges of activating groups are produced from HCN under plausible prebiotic Earth conditions. The final step is clay mineral catalysis in the presence of mineral salts to facilitate selective production of functional RNA. Both the clay minerals and mineral salts would have been abundant on early Earth. We have demonstrated that while montmorillonite (pH 7 produced only dimers from its monomers in water, addition of sodium chloride (1 M enhanced the chain length multifold, as detected by HPLC. The effect of monovalent cations on RNA synthesis was of the following order: Li+ > Na+ > K+. A similar effect was observed with the anions, enhancing catalysis in the following order: Cl− > Br− > I−. The montmorillonite-catalyzed RNA synthesis was not affected by hydrophobic or hydrophilic interactions. We thus show that prebiotic synthesis of RNA from its monomers was a simple process requiring only clay minerals and a small amount of salt.

  15. Prebiotic RNA Synthesis by Montmorillonite Catalysis

    Science.gov (United States)

    Jheeta, Sohan; Joshi, Prakash C.

    2014-08-01

    This review summarizes our recent findings on the role of mineral salts in prebiotic RNA synthesis, which is catalyzed by montmorillonite clay minerals. The clay minerals not only catalyze the synthesis of RNA but also facilitate homochiral selection. Preliminary data of these findings have been presented at the "Horizontal Gene Transfer and the Last Universal Common Ancestor (LUCA)" conference at the Open University, Milton Keynes, UK, 5-6 September 2013. The objective of this meeting was to recognize the significance of RNA in LUCA. We believe that the prebiotic RNA synthesis from its monomers must have been a simple process. As a first step, it may have required activation of the 5'-end of the mononucleotide with a leaving group, e.g., imidazole in our model reaction (Figure 1). Wide ranges of activating groups are produced from HCN under plausible prebiotic Earth conditions. The final step is clay mineral catalysis in the presence of mineral salts to facilitate selective production of functional RNA. Both the clay minerals and mineral salts would have been abundant on early Earth. We have demonstrated that while montmorillonite (pH 7) produced only dimers from its monomers in water, addition of sodium chloride (1 M) enhanced the chain length multifold, as detected by HPLC. The effect of monovalent cations on RNA synthesis was of the following order: Li+ > Na+ > K+. A similar effect was observed with the anions, enhancing catalysis in the following order: Cl- > Br- > I-. The montmorillonite-catalyzed RNA synthesis was not affected by hydrophobic or hydrophilic interactions. We thus show that prebiotic synthesis of RNA from its monomers was a simple process requiring only clay minerals and a small amount of salt.

  16. Protein Synthesis Initiation Factors: Phosphorylation and Regulation

    Energy Technology Data Exchange (ETDEWEB)

    Karen S. Browning

    2009-06-15

    The initiation of the synthesis of proteins is a fundamental process shared by all living organisms. Each organism has both shared and unique mechanisms for regulation of this vital process. Higher plants provide for a major amount of fixation of carbon from the environment and turn this carbon into food and fuel sources for our use. However, we have very little understanding of how plants regulate the synthesis of the proteins necessary for these metabolic processes. The research carried out during the grant period sought to address some of these unknowns in the regulation of protein synthesis initiation. Our first goal was to determine if phosphorylation plays a significant role in plant initiation of protein synthesis. The role of phosphorylation, although well documented in mammalian protein synthesis regulation, is not well studied in plants. We showed that several of the factors necessary for the initiation of protein synthesis were targets of plant casein kinase and showed differential phosphorylation by the plant specific isoforms of this kinase. In addition, we identified and confirmed the phosphorylation sites in five of the plant initiation factors. Further, we showed that phosphorylation of one of these factors, eIF5, affected the ability of the factor to participate in the initiation process. Our second goal was to develop a method to make initiation factor 3 (eIF3) using recombinant methods. To date, we successfully cloned and expressed 13/13 subunits of wheat eIF3 in E. coli using de novo gene construction methods. The final step in this process is to place the subunits into three different plasmid operons for co-expression. Successful completion of expression of eIF3 will be an invaluable tool to the plant translation community.

  17. Diversity-Oriented Organic Synthesis

    Institute of Scientific and Technical Information of China (English)

    J. Wu

    2005-01-01

    @@ 1Introduction A goal of chemical genetics is to find small molecules that modulate the individual functions of gene products with high potency and high specificity[1,2]. Natural products and natural product-derived compounds provide many of the most striking examples, particularly in terms of their specificity. It seems unlikely that natural products alone will provide the hypothetical "complete" set of small molecules that would allow the functions of all proteins, as well as their individual domains, to be determined. For chemistry to have its maximal effect on biology, efficient methods based on diversity-oriented organic synthesis for discovering this set of small molecules are in great demand(See Fig. 1).

  18. Synthesis of Pandamarilactone-1

    OpenAIRE

    Seah, Kang Yee; Macnaughton, Sarah J.; Dallimore, Jonathan W. P.; Robertson, Jeremy

    2014-01-01

    The first total synthesis of pandamarilactone-1, an alkaloid of Pandanus amaryllifolius, is reported. The nine-step synthesis features furan oxidation with singlet oxygen and then spiro-N,O-acetalization and elimination to generate the natural product and further Pandanus alkaloids, pandamarilactonines A–D.

  19. Total Synthesis of (-)-Cardiopetaline.

    Science.gov (United States)

    Nishiyama, Yoshitake; Yokoshima, Satoshi; Fukuyama, Tohru

    2016-05-20

    The total synthesis of (-)-cardiopetaline, an aconitine-type natural product, has been accomplished. Our synthesis involved a Wagner-Meerwein rearrangement of a sulfonyloxirane that enabled, in a single step, the construction of the bicyclo[3.2.1] system in the aconitine skeleton and effective introduction of oxygen functional groups at the appropriate positions. PMID:27166640

  20. Synthesis of oligonucleotide phosphorodithioates

    DEFF Research Database (Denmark)

    Beaton, G.; Brill, W. K D; Grandas, A.;

    1991-01-01

    The synthesis of DNA containing sulfur at the two nonbonding internucleotide valencies is reported. Several different routes using either tervalent or pentavalent mononucleotide synthons are described. © 1991.......The synthesis of DNA containing sulfur at the two nonbonding internucleotide valencies is reported. Several different routes using either tervalent or pentavalent mononucleotide synthons are described. © 1991....

  1. Synthesis of Mechanisms

    DEFF Research Database (Denmark)

    Hansen, John Michael

    1999-01-01

    These notes describe an automated procedure for analysis and synthesis of mechanisms. The analysis method is based on the body coordinate formulation, and the synthesis is based on applying optimization methods, used to minimize the difference between an actual and a desired behaviour...

  2. Synthesis of Isoiminosugars

    DEFF Research Database (Denmark)

    Hyldtoft, Lene; Godskesen, Michael Anders; Lundt, Inge

    1998-01-01

    A short synthesis of isoiminosugars have been developed. Bromolactones are diastereoselectively alkylated at C-2 followed by ring closure to the corresponding lactams. Reduction of these then gives isoiminosugars......A short synthesis of isoiminosugars have been developed. Bromolactones are diastereoselectively alkylated at C-2 followed by ring closure to the corresponding lactams. Reduction of these then gives isoiminosugars...

  3. Homeobox Genes in the Rodent Pineal Gland

    DEFF Research Database (Denmark)

    Rath, Martin Fredensborg; Rohde, Kristian; Klein, David C;

    2013-01-01

    The pineal gland is a neuroendocrine gland responsible for nocturnal synthesis of melatonin. During early development of the rodent pineal gland from the roof of the diencephalon, homeobox genes of the orthodenticle homeobox (Otx)- and paired box (Pax)-families are expressed and are essential for...... normal pineal development consistent with the well-established role that homeobox genes play in developmental processes. However, the pineal gland appears to be unusual because strong homeobox gene expression persists in the pineal gland of the adult brain. Accordingly, in addition to developmental...... functions, homeobox genes appear to be key regulators in postnatal phenotype maintenance in this tissue. In this paper, we review ontogenetic and phylogenetic aspects of pineal development and recent progress in understanding the involvement of homebox genes in rodent pineal development and adult function...

  4. [Language gene].

    Science.gov (United States)

    Takahashi, Hiroshi

    2006-11-01

    The human capacity for acquiring speech and language must derive, at least in part, from the genome. Recent advance in the field of molecular genetics finally discovered 'Language Gene'. Disruption of FOXP2 gene, the firstly identified 'language gene' causes severe speech and language disorder. To elucidate the anatomical basis of language processing in the brain, we examined the expression pattern of FOXP2/Foxp2 genes in the monkey and rat brains through development. We found the preferential expression of FOXP2/Foxp2 in the striosomal compartment of the developing striatum. Thus, we suggest the striatum, particularly striosomal system may participate in neural information processing for language and speech. Our suggestion is consistent with the declarative/ procedural model of language proposed by Ullman (1997, 2001), which the procedural memory-dependent mental grammar is rooted in the basal ganglia and the frontal cortex, and the declarative memory-dependent mental lexicon is rooted in the temporal lobe. PMID:17432197

  5. Gene Silencing

    Czech Academy of Sciences Publication Activity Database

    Kertbundit, Sunee; Juříček, Miloslav; Hall, T.C.

    Dordrecht : Springer, 2010 - (Jain, S.; Brar, D.), s. 631-652 ISBN 978-90-481-2966-9 Institutional research plan: CEZ:AV0Z50380511 Keywords : Gene Silencing * RISC complex Subject RIV: EB - Genetics ; Molecular Biology

  6. Synthesis of zeolite membranes

    Institute of Scientific and Technical Information of China (English)

    JIANG Haiyang; ZHANG Baoquan; Y. S. Lin; LI Yongdan

    2004-01-01

    Zeolite membranes offer great application potentials in membrane separation and/or reaction due to their excellent separation performance and catalytic ability. Up to present, various synthesis methods of zeolite membranes have been developed, including embedded method,in-situ hydrothermal synthesis method, and secondary growth method etc. Compared with the in-situ hydrothermal synthesis method, the secondary growth method possesses a variety of advantages such as easier operation, higher controllability in crystal orientation, microstructure and film thickness, leading to much better reproducibility. This review provides a concise summary and analysis of various synthesis methods reported in the literature. In particular, the secondary growth method was discussed in detail in terms of crystal orientation, defects and crystal grain layers. Some critical issues were also highlighted, which were conducive to the improvement in the synthesis technology of zeolite membranes.

  7. VHDL for logic synthesis

    CERN Document Server

    Rushton, Andrew

    2011-01-01

    Many engineers encountering VHDL (very high speed integrated circuits hardware description language) for the first time can feel overwhelmed by it. This book bridges the gap between the VHDL language and the hardware that results from logic synthesis with clear organisation, progressing from the basics of combinational logic, types, and operators; through special structures such as tristate buses, register banks and memories, to advanced themes such as developing your own packages, writing test benches and using the full range of synthesis types. This third edition has been substantially rewritten to include the new VHDL-2008 features that enable synthesis of fixed-point and floating-point hardware. Extensively updated throughout to reflect modern logic synthesis usage, it also contains a complete case study to demonstrate the updated features. Features to this edition include: * a common VHDL subset which will work across a range of different synthesis systems, targeting a very wide range of technologies...

  8. A grass-based diet favours muscle n-3 long-chain PUFA deposition without modifying gene expression of proteins involved in their synthesis or uptake in Charolais steers.

    Science.gov (United States)

    Cherfaoui, M; Durand, D; Bonnet, M; Bernard, L; Bauchart, D; Ortigues-Marty, I; Gruffat, D

    2013-11-01

    N-3 long-chain polyunsaturated fatty acids (n-3 LCPUFA) are subject of growing interest as they are of particular relevance for meat quality and human health. However, their content in the muscles of cattle is generally low probably as the complex result of their biosynthesis from dietary n-3 PUFA in the muscle and/or in other tissues/organs and of their subsequent uptake by the muscle. In view of this, this study aimed at understanding whether the changes in the muscle n-3 LCPUFA content, depending on the diet (maize silage v. grass) or the muscle type (Rectus abdominis, RA v. Semitendinosus, ST) in 12 Charolais steers, were related to variations in the gene expression of proteins involved in n-3 LCPUFA biosynthesis or cellular uptake. Tissue fatty acid composition was analysed by gas-liquid chromatography and mRNA abundance of proteins by quantitative real-time PCR. The grass-based diet resulted in a 2.3-fold (P Charolais steers did not seem to be mediated by the gene expression regulation of proteins involved in the biosynthesis or uptake of these fatty acids. PMID:23916277

  9. Regulation of expression of a soybean storage protein subunit gene. Progress report

    International Nuclear Information System (INIS)

    We have found that the methionine repression of the β-subunit gene expression is not due to degradation of the β-subunit but is due to an effect on synthesis of the β-subunit. The effect of methionine on the synthesis of the β-is due to an inhibition of β-subunit mRNA synthesis. 3 references, 1 figure

  10. Experiment and quantitative modeling of cell-free gene expression dynamics

    OpenAIRE

    Stögbauer, Tobias Roland

    2012-01-01

    Genexpression that is the cellular synthesis of proteins is comprised of the sub-steps tran- scription (mRNA synthesis based on the DNA master), translation (protein synthesis based on the mRNA) and protein folding. Owing to the large number of interactions between individual components this process is very complex in vivo and therefore mathematical modeling is extremely laborious. By means of simpli�ed in vitro model systems individual aspects of cellular gene expression can be studied...

  11. Gene regulation in parthenocarpic tomato fruit.

    Science.gov (United States)

    Martinelli, Federico; Uratsu, Sandra L; Reagan, Russell L; Chen, Ying; Tricoli, David; Fiehn, Oliver; Rocke, David M; Gasser, Charles S; Dandekar, Abhaya M

    2009-01-01

    Parthenocarpy is potentially a desirable trait for many commercially grown fruits if undesirable changes to structure, flavour, or nutrition can be avoided. Parthenocarpic transgenic tomato plants (cv MicroTom) were obtained by the regulation of genes for auxin synthesis (iaaM) or responsiveness (rolB) driven by DefH9 or the INNER NO OUTER (INO) promoter from Arabidopsis thaliana. Fruits at a breaker stage were analysed at a transcriptomic and metabolomic level using microarrays, real-time reverse transcription-polymerase chain reaction (RT-PCR) and a Pegasus III TOF (time of flight) mass spectrometer. Although differences were observed in the shape of fully ripe fruits, no clear correlation could be made between the number of seeds, transgene, and fruit size. Expression of auxin synthesis or responsiveness genes by both of these promoters produced seedless parthenocarpic fruits. Eighty-three percent of the genes measured showed no significant differences in expression due to parthenocarpy. The remaining 17% with significant variation (P auxin in particular), and metabolism of sugars and lipids. Up-regulation of lipid transfer proteins and differential expression of several indole-3-acetic acid (IAA)- and ethylene-associated genes were observed in transgenic parthenocarpic fruits. Despite differences in several fatty acids, amino acids, and other metabolites, the fundamental metabolic profile remains unchanged. This work showed that parthenocarpy with ovule-specific alteration of auxin synthesis or response driven by the INO promoter could be effectively applied where such changes are commercially desirable. PMID:19700496

  12. Studies of globin gene expression in differentiating erythroid cells

    International Nuclear Information System (INIS)

    The author has addressed questions concerning globin gene expression and the loss of protein synthesis in the terminal stages of erythroid development. (1) The hypothesis that the rate of cell division affects the relative synthesis of γ and β globin in erythroid cells was investigated. The effect of hydroxyurea, aminopterin, or low culture temperature on the in vitro growth of erythroid progenitor cells and on the relative synthesis of γ and β globin was measured. No consistent change in γ globin synthesis was detected. (2) The hypothesis that the ratio of γ and β globin synthesis decreases during erythroid maturation because of differential mRNA stability was investigated. The half-lives of γ and β globin mRNAs and γ and β globin protein synthesis were measured in cultured reticulocytes. γ and β globin mRNAs were assayed by solution hybridization and by in vitro translation. Globin synthesis was determined by 3H-leucine incorporation into the γ and β globin chains. γ and β globin mRNAs decay with similar half-lives in cultured reticulocytes. Therefore, the change in the ratio of γ and β globin synthesis during erythroid maturation cannot be explained by differences in mRNA stability and is likely to result from asynchronous transcription of the genes. These data suggest that protein synthesis in maturing reticulocytes is not limited by the quantity of mRNA but by the availability of translation factors. (3) The hypothesis was tested that the initiation factor GEF becomes limiting for protein synthesis during reticulocyte maturation

  13. Role for deoxyribonucleic acid ligase in deoxyribonucleic acid polymerase I-dependent repair synthesis in toluene-treated Escherichia coli

    International Nuclear Information System (INIS)

    In a toluene-treated mutant of Escherichia coli K-12 having a temperature-sensitive, conditionally lethal mutation in the structural gene for deoxyribonucleic acid (DNA) ligase, an extensive DNA repair synthesis occurred in x-irradiated cells at the nonpermissive temperature, 420C. At the permissive temperature, 300C, nearly normal semiconservative synthesis and limited repair synthesis were observed when DNA ligase was activated by the addition of nicotinamide adenine dinucleotide. (auth)

  14. Regulation of toxin synthesis in Clostridium botulinum and Clostridium tetani.

    Science.gov (United States)

    Connan, Chloé; Denève, Cécile; Mazuet, Christelle; Popoff, Michel R

    2013-12-01

    Botulinum and tetanus neurotoxins are structurally and functionally related proteins that are potent inhibitors of neuroexocytosis. Botulinum neurotoxin (BoNT) associates with non-toxic proteins (ANTPs) to form complexes of various sizes, whereas tetanus toxin (TeNT) does not form any complex. The BoNT and ANTP genes are clustered in a DNA segment called the botulinum locus, which has different genomic localization (chromosome, plasmid, phage) in the various Clostridium botulinum types and subtypes. The botulinum locus genes are organized in two polycistronic operons (ntnh-bont and ha/orfX operons) transcribed in opposite orientations. A gene called botR lying between the two operons in C. botulinum type A encodes an alternative sigma factor which regulates positively the synthesis of BoNT and ANTPs at the late exponential growth phase and beginning of the stationary phase. In Clostridium tetani, the gene located immediately upstream of tent encodes a positive regulatory protein, TetR, which is related to BotR. C. botulinum and C. tetani genomes contain several two-component systems and predicted regulatory orphan genes. In C. botulinum type A, four two-component systems have been found that positively or negatively regulate the synthesis of BoNT and ANTPs independently of BotR/A. The synthesis of neurotoxin in Clostridia seems to be under the control of complex network of regulation. PMID:23769754

  15. Total Synthesis of Teixobactin.

    Science.gov (United States)

    Giltrap, Andrew M; Dowman, Luke J; Nagalingam, Gayathri; Ochoa, Jessica L; Linington, Roger G; Britton, Warwick J; Payne, Richard J

    2016-06-01

    The first total synthesis of the cyclic depsipeptide natural product teixobactin is described. Synthesis was achieved by solid-phase peptide synthesis, incorporating the unusual l-allo-enduracididine as a suitably protected synthetic cassette and employing a key on-resin esterification and solution-phase macrolactamization. The synthetic natural product was shown to possess potent antibacterial activity against a range of Gram-positive pathogenic bacteria, including a virulent strain of Mycobacterium tuberculosis and methicillin-resistant Staphylococcus aureus (MRSA). PMID:27191730

  16. Asymmetric synthesis v.4

    CERN Document Server

    Morrison, James

    1984-01-01

    Asymmetric Synthesis, Volume 4: The Chiral Carbon Pool and Chiral Sulfur, Nitrogen, Phosphorus, and Silicon Centers describes the practical methods of obtaining chiral fragments. Divided into five chapters, this book specifically examines initial chiral transmission and extension. The opening chapter describes the so-called chiral carbon pool, the readily available chiral carbon fragments used as building blocks in synthesis. This chapter also provides a list of 375 chiral building blocks, along with their commercial sources, approximate prices, and methods of synthesis. Schemes involving

  17. Gas Phase Nanoparticle Synthesis

    Science.gov (United States)

    Granqvist, Claes; Kish, Laszlo; Marlow, William

    This book deals with gas-phase nanoparticle synthesis and is intended for researchers and research students in nanomaterials science and engineering, condensed matter physics and chemistry, and aerosol science. Gas-phase nanoparticle synthesis is instrumental to nanotechnology - a field in current focus that raises hopes for environmentally benign, resource-lean manufacturing. Nanoparticles can be produced by many physical, chemical, and even biological routes. Gas-phase synthesis is particularly interesting since one can achieve accurate manufacturing control and hence industrial viability.

  18. Synthesis of Chiral Cyclopentenones.

    Science.gov (United States)

    Simeonov, Svilen P; Nunes, João P M; Guerra, Krassimira; Kurteva, Vanya B; Afonso, Carlos A M

    2016-05-25

    The cyclopentenone unit is a very powerful synthon for the synthesis of a variety of bioactive target molecules. This is due to the broad diversity of chemical modifications available for the enone structural motif. In particular, chiral cyclopentenones are important precursors in the asymmetric synthesis of target chiral molecules. This Review provides an overview of reported methods for enantioselective and asymmetric syntheses of cyclopentenones, including chemical and enzymatic resolution, asymmetric synthesis via Pauson-Khand reaction, Nazarov cyclization and organocatalyzed reactions, asymmetric functionalization of the existing cyclopentenone unit, and functionalization of chiral building blocks. PMID:27101336

  19. Complete Genome Sequence of a Klebsiella pneumoniae Isolate with Chromosomally Encoded Carbapenem Resistance and Colibactin Synthesis Loci

    OpenAIRE

    Conlan, Sean; Deming, Clayton; Tsai, Yu-Chih; Lau, Anna F.; Dekker, John P.; Korlach, Jonas; Segre, Julia A

    2014-01-01

    Klebsiella pneumoniae is an important nosocomial pathogen, and multidrug-resistant strains have become a worldwide concern. Here, we report the complete genome of a K. pneumoniae isolate with chromosomally integrated bla KPC genes and a colibactin synthesis locus.

  20. 蛋白质谷氨酰胺酶基因的合成表达及性质研究%Gene Synthesis, Expression and Property Research of Protein-glutaminase

    Institute of Scientific and Technical Information of China (English)

    汪正华; 朱蓓霖; 赵云; 周杰; 吴自荣; 黄静

    2012-01-01

    Protein-glutaminase, a novel protein-deamidating enzyme, which has potential for industrial applications. The gene encoding the protein was synthesized using overlap extension PCR and the mature PG gene was cloned into expression vector pET32a( + ). The recombinant protein was expressed in Escherichia coli BL21 (DE3) as inclusion bodies, and the active PG was obtained after denaturation and renaturation. In order to improve the solubility of PG,the culture was incubated in cold-shocked condition and a chaperone plasmid pTfl6-tig was also cloned into pET32a-matPG/BL21 (DE3). The results showed that low temperature can improve the solubility of PG slightly, but pTfl6-tig was futile. For deamidating activity assay, Cbz-Gln-Gly was used as substrate. The reaction showed that PG can effectively hydrolyzed glutaminyl residues in the Cbz-Gln-Gly and resulting in release of ammonia. The research on enzymatic properties of PG showed that the optimum temperature is 40℃ and the optimal pH is 6. 0. The gene encoding protein-glutaminase was synthesized and expressed successfully. The research provided a new idea for heterologous expression of this particular food-enzyme using genetic engineering.%蛋白质谷氨酰胺酶(Protein-glutaminase,PG)是引起蛋白质脱酰胺作用的一种新型水解酶,在食品工业中具有十分广泛的应用前景.通过重叠延伸PCR法合成了PG全长基因,将其成熟肽序列克隆至原核表达载体pET32a(+)上,转化大肠杆菌BL21(DE3),经IPTG诱导,重组蛋白主要以包涵体形式表达.通过降低诱导温度和共表达分子伴侣两种策略,以期实现重组蛋白的可溶性表达,结果发现低温对可溶性的提高有一定的作用,而分子伴侣影响甚微.收集包涵体经变性、复性后获得活性PG.将PG与底物Cbz-Gln-Gly进行孵育反应,结果表明,PG可有效水解Cbz-Gln-Gly谷氨酰胺残基上的氨酰基,释放出氨;酶学性质的研究表明,此酶的最适反应温度为40

  1. Two-directional synthesis as a tool for diversity-oriented synthesis: Synthesis of alkaloid scaffolds

    OpenAIRE

    Kieron M. G. O’Connell; Monica Díaz-Gavilán; Galloway, Warren R. J. D.; Spring, David R.

    2012-01-01

    Two-directional synthesis represents an ideal strategy for the rapid elaboration of simple starting materials and their subsequent transformation into complex molecular architectures. As such, it is becoming recognised as an enabling technology for diversity-oriented synthesis. Herein, we provide a thorough account of our work combining two-directional synthesis with diversity-oriented synthesis, with particular reference to the synthesis of polycyclic alkaloid scaffolds.

  2. [Application of levansucrase in levan synthesis--a review].

    Science.gov (United States)

    Lu, Juan; Lu, Lili; Xiao, Min

    2014-06-01

    Levan is a fructan mainly linked by beta-(2,6)-glycosidic bonds with some beta-(2,1)-linked branch chains. Some microbial levan exhibit biological activities such as antitumor, antidiabetic and immunostimulating activities. hypolipidemic effect, and function as prebiotics, which has a wide and potential application in the pharmaceutical and food industry. Because of low extraction yields from microbial fermentation and a very complex process for chemical synthesis of levan, enzymatic synthesis of levan has attracted tremendous interest. Levansucrase (EC 2. 4. 1. 10), a beta-propeller protein belonging to the glycoside hydrolase family 68 (GH68) with reaction mechanism of non-Leloir glycosyltransferase, catalyzes the synthesis of levan by transferring the fructosyl group of non-activated sucrose into the fructan chain. The molecular structure and regulation of gene expression of some microbial levansucrases have been elucidated. Meanwhile, the enzymatic synthesis of levan by levansucrase is widely studied. In this review, catalytic mechanism of levansucrase, molecular structure and regulation of gene expression of some microbial levansucrases, and the application of levansucrases in enzymatic synthesis of levan were summarized. PMID:25272807

  3. Synthesis of Acetylhomoagmatine

    Directory of Open Access Journals (Sweden)

    Carmenza Duque

    2006-08-01

    Full Text Available Abstract: The first total synthesis of acetylhomoagmatine, a natural product isolated form the methanolic extracts from the sponge Cliona celata, is performed in four steps in a very high yield.

  4. 2002 Annual report: synthesis

    International Nuclear Information System (INIS)

    This synthesis of the Annual Report 2002 presents information of the main activities on the scope of the radiation protection and nuclear safety of the Nuclear Regulatory Authority (ARN) of the Argentina during 2002

  5. 2000 Annual report: synthesis

    International Nuclear Information System (INIS)

    This synthesis of the Annual Report 2000 present information of the main activities on the scope of the radiation protection and nuclear safety of the Nuclear Regulatory Authority (NRA) of the Argentina during 2000

  6. 2001 Annual report: synthesis

    International Nuclear Information System (INIS)

    This synthesis of the Annual Report 2001 presents information of the main activities on the scope of the radiation protection and nuclear safety of the Nuclear Regulatory Authority (ARN) of the Argentina during 2001

  7. Total synthesis of atropurpuran.

    Science.gov (United States)

    Gong, Jing; Chen, Huan; Liu, Xiao-Yu; Wang, Zhi-Xiu; Nie, Wei; Qin, Yong

    2016-01-01

    Due to their architectural intricacy and biological significance, the synthesis of polycyclic diterpenes and their biogenetically related alkaloids have been the subject of considerable interest over the last few decades, with progress including the impressive synthesis of several elusive targets. Despite tremendous efforts, conquering the unique structural types of this large natural product family remains a long-term challenge. The arcutane diterpenes and related alkaloids, bearing a congested tetracyclo[5.3.3.0(4,9).0(4,12)]tridecane unit, are included in these unsolved enigmas. Here we report a concise approach to the construction of the core structure of these molecules and the first total synthesis of (±)-atropurpuran. Pivotal features of the synthesis include an oxidative dearomatization/intramolecular Diels-Alder cycloaddition cascade, sequential aldol and ketyl-olefin cyclizations to assemble the highly caged framework, and a chemoselective and stereoselective reduction to install the requisite allylic hydroxyl group in the target molecule. PMID:27387707

  8. Catalytic Synthesis Lactobionic Acid

    Directory of Open Access Journals (Sweden)

    V.G. Borodina

    2014-07-01

    Full Text Available Gold nanoparticles are obtained, characterized and deposited on the carrier. Conducted catalytic synthesis of lactobionic acid from lactose. Received lactobionic acid identify on the IR spectrum.

  9. 乙酸钠和β-羟丁酸钠对奶牛乳腺上皮细胞乳脂和乳蛋白合成相关基因表达的影响%Effects of Sodium Acetate and Sodium β-Hydroxybutyrate on ExPressions of Genes Involved in Milk Fat and Protein Synthesis in Bovine Mammary EPithelial Cells

    Institute of Scientific and Technical Information of China (English)

    塔娜; 李红磊; 侯先志; 考桂兰; 高民; 李大彪

    2014-01-01

    本试验旨在研究乙酸钠和β-羟丁酸钠对奶牛乳腺上皮细胞(BMECs)乳脂和乳蛋白合成相关基因表达的影响。试验分2部分,均采用单因子完全随机试验设计。第1部分,单独添加试验,乙酸钠的添加浓度分别为0(对照)、6.00、9.00、12.00和15.00 mmol/ L,β-羟丁酸钠的添加浓度分别为0(对照)、0.80、1.60和2.40 mmol/ L。第2部分,混合添加试验,以单独添加试验得出的乙酸钠和β-羟丁酸钠的适宜浓度,二者之和为总添加浓度,设定3种不同配比,即乙酸钠∶β-羟丁酸钠分别为1∶1、2∶1和4∶1,对照组不添加乙酸钠和β-羟丁酸钠。结果表明:1)与对照组相比,12.00 mmol/ L 乙酸钠能够显著提高 BMECs 乙酰辅酶 A 羧化酶( ACC)、脂肪酸合成酶(FAS)、二酰甘油酰基转移酶( DGAT)、乙酰辅酶 A 合成酶2( ACSS2)、过氧化物酶体增殖物激活受体(PPARG)、κ酪蛋白(CSN3)和雷帕霉素靶蛋白( mTOR)基因的表达量及甘油三酯(TAG)的含量( P <0.05)。2)与对照组相比,2.40 mmol/ L 的β-羟丁酸钠能够显著提高BMECs ACC、FAS、ACSS2、PPARG、mTOR 基因表达量及 TAG 含量(P<0.05)。3)添加不同配比的乙酸钠和β-羟丁酸钠均不同程度地促进了乳脂合成相关基因的表达,乙酸钠∶β-羟丁酸钠为2∶1和4∶1时,BMECs 中 TAG 含量显著高于为1∶1时和对照组( P<0.05)。综合各项指标,以9.60 mmol/ L乙酸钠和4.80 mmol/ L β-羟丁酸钠混合添加对奶牛乳腺上皮细胞乳脂和乳蛋白合成的促进效果较好。%The aim of this study was to determine the effects of sodium acetate and sodium β-hydroxybutyrate on expressions of genes involved in milk fat and protein synthesis in bovine mammary epithelial cells (BMECs). The study was consisted of two parts,and completely random single-factor designs were adopted. Part

  10. Fusions of flagellar operons to lactose genes on a mu lac bacteriophage.

    OpenAIRE

    Komeda, Y

    1982-01-01

    Previous studies have defined 29 genes necessary for synthesis of the Escherichia coli flagellar apparatus. This study analyzed the transcriptional control of flagellar genes, using Mu d (Apr lac) phage to generate flagellar mutants by insertion. These mutants contained operon fusions of flagellar genes to the lac genes of the Mu d phage and allowed the measurement of flagellar operon expression by detection of beta-galactosidase activity. These fusion mutants expressed the enzyme activity co...

  11. Ethics of Chemical Synthesis

    OpenAIRE

    Joachim Schummer

    2001-01-01

    Unlike other branches of science, the scientific products of synthetic chemistry are not only ideas but also new substances that change our material world, for the benefit or harm of living beings. This paper provides for the first time a systematical analysis of moral issues arising from chemical synthesis, based on concepts of responsibility and general morality. Topics include the questioning of moral neutrality of chemical synthesis as an end in itself, chemical weapons research, moral ob...

  12. Characterisation of esterase genes in the genomes of Streptomyces coelicolor A3(2) and Streptomyces avermitilis

    OpenAIRE

    Soror, Sameh

    2007-01-01

    Esterases and lipases are widely used as industrial enzymes and for the synthesis of chiral drugs. Because of their rich secondary metabolism, Streptomyces species offer a relatively untapped source of interesting esterases and lipases. S. coelicolor and S. avermitilis contain 51 genes annotated as esterases and/or lipases. In this study I have cloned 14 different genes encoding for lipolytic enzymes from S. coelicolor (11 genes) and S. avermitilis (four genes). Some of these genes were over-...

  13. Genes, enzymes and regulation of arginine biosynthesis in plants.

    Science.gov (United States)

    Slocum, Robert D

    2005-08-01

    Arabidopsis genes encoding enzymes for each of the eight steps in L-arginine (Arg) synthesis were identified, based upon sequence homologies with orthologs from other organisms. Except for N-acetylglutamate synthase (NAGS; EC 2.3.1.1), which is encoded by two genes, all remaining enzymes are encoded by single genes. Targeting predictions for these enzymes, based upon their deduced sequences, and subcellular fractionation studies, suggest that most enzymes of Arg synthesis reside within the plastid. Synthesis of the L-ornthine (Orn) intermediate in this pathway from L-glutamate occurs as a series of acetylated intermediates, as in most other organisms. An N-acetylornithine:glutamate acetyltransferase (NAOGAcT; EC 2.3.1.35) facilitates recycling of the acetyl moiety during Orn formation (cyclic pathway). A putative N-acetylornithine deacetylase (NAOD; EC 3.5.1.16), which participates in the "linear" pathway for Orn synthesis in some organisms, was also identified. Previous biochemical studies have indicated that allosteric regulation of the first and, especially, the second steps in Orn synthesis (NAGS; N-acetylglutamate kinase (NAGK), EC 2.7.2.8) by the Arg end-product are the major sites of metabolic control of the pathway in organisms using the cyclic pathway. Gene expression profiling for pathway enzymes further suggests that NAGS, NAGK, NAOGAcT and NAOD are coordinately regulated in response to changes in Arg demand during plant growth and development. Synthesis of Arg from Orn is further coordinated with pyrimidine nucleotide synthesis, at the level of allocation of the common carbamoyl-P intermediate. PMID:16122935

  14. Gene regulation in parthenocarpic tomato fruit

    OpenAIRE

    Martinelli, Federico; Uratsu, Sandra L.; Reagan, Russell L.; Chen, Ying; Tricoli, David; Fiehn, Oliver; Rocke, David M.; Gasser, Charles S.; Abhaya M. Dandekar

    2009-01-01

    Parthenocarpy is potentially a desirable trait for many commercially grown fruits if undesirable changes to structure, flavour, or nutrition can be avoided. Parthenocarpic transgenic tomato plants (cv MicroTom) were obtained by the regulation of genes for auxin synthesis (iaaM) or responsiveness (rolB) driven by DefH9 or the INNER NO OUTER (INO) promoter from Arabidopsis thaliana. Fruits at a breaker stage were analysed at a transcriptomic and metabolomic level using microarrays, real-time re...

  15. Magnetic nanoparticles for gene and drug delivery

    Directory of Open Access Journals (Sweden)

    Stuart C McBain

    2008-06-01

    Full Text Available Stuart C McBain, Humphrey HP Yiu, Jon DobsonInstitute of Science and Technology in Medicine, Keele University, Thornburrow Drive, Hartshill, Stoke-on-Trent, Staffordshire, ST4 7QB, U.K.Abstract: Investigations of magnetic micro- and nanoparticles for targeted drug delivery began over 30 years ago. Since that time, major progress has been made in particle design and synthesis techniques, however, very few clinical trials have taken place. Here we review advances in magnetic nanoparticle design, in vitro and animal experiments with magnetic nanoparticle-based drug and gene delivery, and clinical trials of drug targeting.Keywords: magnetic nanoparticles, gene delivery, biotechnology

  16. Autoinducer-mediated regulation of rhamnolipid biosurfactant synthesis in Pseudomonas aeruginosa.

    OpenAIRE

    Ochsner, U A; Reiser, J

    1995-01-01

    The opportunistic human pathogen Pseudomonas aeruginosa produces a variety of virulence factors, including exotoxin A, elastase, alkaline protease, alginate, phospholipases, and extracellular rhamnolipids. The previously characterized rhlABR gene cluster encodes a regulatory protein (RhlR) and a rhamnosyltransferase (RhlAB), both of which are required for rhamnolipid synthesis. Another gene, rhII, has now been identified downstream of the rhlABR gene cluster. The putative RhlI protein shares ...

  17. Reliable reference genes for normalization of gene expression in cucumber grown under different nitrogen nutrition.

    Directory of Open Access Journals (Sweden)

    Anna Warzybok

    Full Text Available In plants, nitrogen is the most important nutritional factor limiting the yield of cultivated crops. Since nitrogen is essential for synthesis of nucleotides, amino acids and proteins, studies on gene expression in plants cultivated under different nitrogen availability require particularly careful selection of suitable reference genes which are not affected by nitrogen limitation. Therefore, the objective of this study was to select the most reliable reference genes for qPCR analysis of target cucumber genes under varying nitrogen source and availability. Among twelve candidate cucumber genes used in this study, five are highly homologous to the commonly used internal controls, whereas seven novel candidates were previously identified through the query of the cucumber genome. The expression of putative reference genes and the target CsNRT1.1 gene was analyzed in roots, stems and leaves of cucumbers grown under nitrogen deprivation, varying nitrate availability or different sources of nitrogen (glutamate, glutamine or NH3. The stability of candidate genes expression significantly varied depending on the tissue type and nitrogen supply. However, in most of the outputs genes encoding CACS, TIP41, F-box protein and EFα proved to be the most suitable for normalization of CsNRT1.1 expression. In addition, our results suggest the inclusion of 3 or 4 references to obtain highly reliable results of target genes expression in all cucumber organs under nitrogen-related stress.

  18. Alkane Biosynthesis Genes in Cyanobacteria and Their Transcriptional Organization

    OpenAIRE

    Stephan eKlähn; Desiree eBaumgartner; Ulrike ePfreundt; Karsten eVoigt; Verena eSchoen; Claudia eSteglich; Hess, Wolfgang R

    2014-01-01

    In cyanobacteria, alkanes are synthesized from a fatty acyl-ACP by two enzymes, acyl-acyl carrier protein reductase (AAR) and aldehyde deformylating oxygenase (ADO). Despite the great interest in the exploitation for biofuel production, nothing is known about the transcriptional organization of their genes or the physiological function of alkane synthesis. The comparison of 115 microarray datasets indicates the relatively constitutive expression of aar and ado genes. The analysis of 181 avail...

  19. 棕色棉类黄酮3′-羟化酶基因(F3′H)的克隆及色素合成途径中相关基因表达特性研究%Cloning of Flavon oid 3'-hydroxylase Gene (GhF3'H) and Expressional Characteristics of Several Genes Associated with Pigment Synthesis in Brown Cotton (Gossypium hirsutum L.) Fibers

    Institute of Scientific and Technical Information of China (English)

    梁明炜; 刘海峰; 陆雪莹; 李艳红; 宋武; 鲁春芳; 肖向文; 李晓波

    2011-01-01

    The purpose of this research is to explore the role of anthocyanidin synthesis pathway in pigmentation of brown cotton fibers. Two full-length cDNAs encoding flavonoid 3' -hydroxylase gene (F3'H) was cloned from XC-5 fibers of 16 days post anthesis (DPA) by rapid amplification of Cdna ends (RACE) and RT-PCR using specific primers based on the Gossypium hirsutum EST sequences (GenBank accession: DT545210, CO071403 and BG447485)from the blast results with the full-length Cdna of F3'H in Vitis vinifera that had already known and were named as GhF3 H-1 and GhF3 'H-2 (GenBank accession: HM598123, HM598124). The length of the two Cdna sequences were 1 761 and 1 892 bp, respectively. They contained an identical opening reading frame (ORF) of 1 533 bp, encoding a protein of 510 amino acids. There were few differences betweenthe two sequences except for the sequence length of the 3' untranslational region (UTR). The temporal and spatial expression patterns of chalcone synthase (CHS), flavonoid-3'?5'-hydroxylase as (F3'5'H) and flavonoid 3'-hy-droxylase (F3'H) genes were examined by the semi-quantitative RT-PCR analysis with the specific primers. The expression analysis indicated that the expressed levels of CHS and F3'5'H gene in white and brown-fiber cotton cultivars during development were relatively high. But the F3 7/ gene was mainly express in the ovule of 1 day p ost anthesis, and were almost not expressed in the fibers and petals. All the results suggest that the production of pigments in brown cotton fibers is related to the biosynthesis pathway of anthocyanins, the principle of pigment formation in colored cotton has been disclosed, and this is of help to improve the colors of naturally colored cotton.%为探讨花色苷途径在彩棉色素形成中的作用及彩棉色素形成规律,本研究根据葡萄(Vitis vinifera 的类黄酮3'-羟化酶(flavonoid 3'-hydroxylase,F3'H)基因全长cDNA序列blast所得棉花(Gossypium hirsutum

  20. Genes and Psoriasis

    Science.gov (United States)

    ... Diet Tips" to find out more! Email * Zipcode Genes and Psoriasis Genes hold the key to understanding ... is responsible for causing psoriatic disease. How do genes work? Genes control everything from height to eye ...

  1. Genes and Hearing Loss

    Science.gov (United States)

    ... Meeting Calendar Find an ENT Doctor Near You Genes and Hearing Loss Genes and Hearing Loss Patient ... mutation may only have dystopia canthorum. How Do Genes Work? Genes are a road map for the ...

  2. A post-processing method for optimizing synthesis strategy for oligonucleotide microarrays

    OpenAIRE

    Ning, Kang; Choi, Kwok Pui; Leong, Hon Wai; Zhang, Louxin

    2005-01-01

    The broad applicability of gene expression profiling to genomic analyses has generated huge demand for mass production of microarrays and hence for improving the cost effectiveness of microarray fabrication. We developed a post-processing method for deriving a good synthesis strategy. In this paper, we assessed all the known efficient methods and our post-processing method for reducing the number of synthesis cycles for manufacturing a DNA-chip of a given set of oligos. Our experimental resul...

  3. Direct Dimethyl Ether Synthesis

    Institute of Scientific and Technical Information of China (English)

    Takashi Ogawa; Norio Inoue; Tutomu Shikada; Yotaro Ohno

    2003-01-01

    Dimethyl ether (DME) is a clean and economical alternative fuel which can be produced from natural gas through synthesis gas. The properties of DME are very similar to those of LP gas. DME can be used for various fields as a fuel such as power generation, transportation, home heating and cooking,etc. It contains no sulfur or nitrogen. It is not corrosive to any metal and not harmful to human body. An innovative process of direct synthesis of DME from synthesis gas has been developed. Newly developed catalyst in a slurry phase reactor gave a high conversion and high selectivity of DME production. One and half year pilot scale plant (5 tons per day) testing, which was supported by METI, had successfully finished with about 400 tons DME production.

  4. Synthesis of organosilicon compounds

    Energy Technology Data Exchange (ETDEWEB)

    Zhao, G.

    1996-12-31

    Silicon-containing polymers have been a focus of synthesis and study in Dr. Barton`s group because of their chemistry and properties which are not offered by other systems or materials. For example, the polymer -[-SiMe{sub 2}C{triple_bond}C-]{sub n}-can be easily processed to films or fibers from melt or solution, and thermally converted to a SiC-containing ceramic in high yield at high temperature. In recent years, carbosilane dendritic polymers have been of great interests in many research groups. However, no synthesis of carbosilane dendrimers with functionalties both inside and outside the dendrimer has been reported. Functionality is very important in the synthesis of preceramic polymers. This thesis will be devoted to exploring several new organosilicon polymer systems.

  5. Stereoselective Synthesis of (+)-Boronolide

    Institute of Scientific and Technical Information of China (English)

    HU,Shou-Gang; HU,Tai-Shan; WU,Yu-Lin

    2004-01-01

    @@ (+)-Boronolide (1) and its deacetylated products have attracted much attention of synthetic chemists due to their diverse biological properties as well as their structural complexities.[1] Many of these reported synthesis involved dehydrogenation of δ-lactone by using benzeneseleninic anhydride or ring-closing olefin metathesis (RCM) to introduce the requisite α,β-unsaturated δ-lactone in boronolide. Here, we report the synthesis of boronolide with diastereoselective propargylation of α-hydroxy aldehyde as the key step and D-gluconolactone as the starting material.

  6. Supercritical Synthesis of Biodiesel

    Directory of Open Access Journals (Sweden)

    Michel Vaultier

    2012-07-01

    Full Text Available The synthesis of biodiesel fuel from lipids (vegetable oils and animal fats has gained in importance as a possible source of renewable non-fossil energy in an attempt to reduce our dependence on petroleum-based fuels. The catalytic processes commonly used for the production of biodiesel fuel present a series of limitations and drawbacks, among them the high energy consumption required for complex purification operations and undesirable side reactions. Supercritical fluid (SCF technologies offer an interesting alternative to conventional processes for preparing biodiesel. This review highlights the advances, advantages, drawbacks and new tendencies involved in the use of supercritical fluids (SCFs for biodiesel synthesis.

  7. Diophantine frequency synthesis.

    Science.gov (United States)

    Sotiriadis, Paul Peter

    2006-11-01

    A methodology for fine-step, fast-hopping, low-spurs phase-locked loop based frequency synthesis is presented. It uses mathematical properties of integer numbers and linear Diophantine equations to overcome the constraining relation between frequency step and phase-comparator frequency that is inherent in conventional phase-locked loop based frequency synthesis. The methodology leads to fine-step, fast-hopping, modular-structured frequency synthesizers with potentially very low spurs, especially in the vicinity of the carrier. The paper focuses on the mathematical principles of the new methodology and the related number theoretic algorithms. PMID:17091835

  8. Effects of β-Hydroxybutyric Acid on Relative Expression Levels of Genes Related to Milk Fat Synthesis in Bovine Mammary Epithelial Cells%β-羟丁酸对奶牛乳腺上皮细胞内乳脂肪合成及其相关基因相对表达量的影响

    Institute of Scientific and Technical Information of China (English)

    常晨城; 齐利枝; 闫素梅; 生冉; 赵艳丽

    2015-01-01

    本试验主要研究了不同浓度的β-羟丁酸( BHBA)对奶牛乳腺上皮细胞( BMECs)活力、甘油三酯( TAG)含量、脂滴形成以及乳脂肪合成相关基因转录水平的影响。将传至第3代的BMECs悬液(1×105个/孔)接种于细胞培养板上,每孔加入含10%胎牛血清( FBS)的DMEM/F12培养液,于37℃的5%二氧化碳(CO2)培养箱培养48 h。再将培养48 h的BMECs随机分配到6个组,各组向培养孔中加入含不同浓度BHBA的DMEM/F12培养液,培养液中的FBS用1 g/L无脂肪酸的牛血清白蛋白( BSA)代替,并使反应体系中BHBA的最终浓度分别为0(对照)、0.58、1.16、2.32、4.64和9.28 mmol/L。置于37℃的5%CO2培养箱继续培养48 h。试验结果显示:随着BHBA浓度的增加,BMECs活力[(相对增殖率( RGR)]呈显著的二次曲线增加(P=0.041),其中BMECs活力以0.58~4.64 mmol/L BHBA组较高,9.28 mmol/L BHBA组较低;低浓度(0.58~2.32 mmol/L)的 BHBA 可促进 BMECs 内脂滴的形成,而较高浓度(4.64~9.28 mmol/L)的BHBA对脂滴形成的促进作用减弱;BHBA与TAG含量及乳脂肪合成相关基因脂肪酸合成酶( FASN )、乙酰辅酶 A 羧化酶α( ACACA )、硬脂酰辅酶 A 去饱和酶( SCD)、脂肪酸结合蛋白3( FABP3)、过氧化物酶体增殖物激活受体γ( PPARG)和分化抗原簇36(CD36)的相对表达量均无显著的一次线性或二次曲线关系(P>0.05)。综上,BHBA 对BMECs活力的促进作用呈显著的二次曲线增加,即BHBA对BMECs活力呈显著浓度依赖关系;BHBA对细胞内乳脂肪的合成有提高的趋势。%This study was conducted to determine the effects ofβ-hydroxybutyric acid ( BHBA) on cell viabili-ty, triacylglycerol ( TAG) content, lipid droplet formation and relative expression levels of genes related to milk fat synthesis in bovine mammary epithelial cells (BMECs). The 3th passage cells were plated 1×105 cells/well in culture plates and DMEM/F12 medium containing 10% fetal bovine serum ( FBS) was added to

  9. Fusion of the promoter region of rRNA operon rrnB to lac Z gene.

    OpenAIRE

    Glaser, G; Kobi, S.; Oppenheim, A B

    1980-01-01

    A Lambda phage was constructed in which the structural gene for beta galactosidase is fused to a DNA segment carrying the ribosomal promoter rrnB of E. coli. In this hybrid operon beta galactosidase synthesis in vitro is repressed by ppGpp. Repression of beta galactosidase synthesis by cAMP is reported.

  10. Genetic association analysis of 13 nuclear-encoded mitochondrial candidate genes with type II diabetes mellitus : the DAMAGE study

    NARCIS (Netherlands)

    Reiling, Erwin; van Vliet-Ostaptchouk, Jana V.; van't Riet, Esther; van Haeften, Timon W.; Arp, Pascal A.; Hansen, Torben; Kremer, Dennis; Groenewoud, Marlous J.; van Hove, Els C.; Romijn, Johannes A.; Smit, Jan W. A.; Nijpels, Giel; Heine, Robert J.; Uitterlinden, Andre G.; Pedersen, Oluf; Slagboom, P. Eline; Maassen, Johannes A.; Hofker, Marten H.; 't Hart, Leen M.; Dekker, Jacqueline M.

    2009-01-01

    Mitochondria play an important role in many processes, like glucose metabolism, fatty acid oxidation and ATP synthesis. In this study, we aimed to identify association of common polymorphisms in nuclear-encoded genes involved in mitochondrial protein synthesis and biogenesis with type II diabetes me

  11. Molecular cloning of the Salmonella typhimurium lep gene in Escherichia coli

    NARCIS (Netherlands)

    van Dijl, J M; van den Bergh, R; Reversma, T; Smith, H; Bron, S; Venema, G

    1990-01-01

    A system is described which enabled the selection of a heterologous lep gene, encoding signal peptidase I, in Escherichia coli. It is based on complementation of an E. coli mutant, in which the synthesis of signal peptidase I can be regulated. With this system the lep gene of Salmonella typhimurium

  12. The QQS orphan gene regulates carbon and nitrogen partitioning across species via NF-YC interactions

    Science.gov (United States)

    The allocation of carbon and nitrogen resources to the synthesis of plant proteins, carbohydrates, and lipids is complex and under the control of many genes; much remains to be understood about this process. QQS (Qua Quine Starch, At3g30720), an orphan gene unique to Arabidopsis thaliana, regulates...

  13. DNA polymerase I-mediated ultraviolet repair synthesis in toluene-treated Escherichia coli

    International Nuclear Information System (INIS)

    DNA synthesis after ultraviolet irradiation is low in wild type toluene-treated cells. The level of repair incorporation is greater in strains deficient in DNA polymerase I. The low level of repair synthesis is attributable to the concerted action of DNA polymerase I and polynucleotide ligase. Repair synthesis is stimulated by blocking ligase activity with the addition of nicotinamide mononucleotide (NMN) or the use of a ligase temperature-sensitive mutant. NMN stimulation is specific for DNA polymerase I-mediated repair synthesis, as it is absent in isogenic strains deficient in the polymerase function or the 5' yields 3' exonuclease function associated with DNA polymerase I. DNA synthesis that is stimulated by NMN is proportional to the ultraviolet exposure at low doses, nonconservative in nature, and is dependent on the uvrA gene product but is independent of the recA gene product. These criteria place this synthesis in the excision repair pathway. The NMN-stimulated repair synthesis requires ATP and is N-ethylmaleimide-resistant. The use of NMN provides a direct means for evaluating the involvement of DNA polymerase I in excision repair

  14. Age-related synthesis of glucocorticoids in thymocytes

    International Nuclear Information System (INIS)

    Glucocorticoids (GCs) are primarily synthesized in the adrenal glands but an ectopic production has also been reported in the brain, the gastrointestinal tract and in thymic epithelial cells (TEC). Here we show that thymocytes express genes encoding for all enzymes required for de novo GC synthesis and produce the hormone as demonstrated by both a GC specific reporter assay and a corticosterone specific ELISA assay. Interestingly, GC synthesis is detectable in cells from young mice (4 weeks) and thereafter increases during aging (14-22 weeks) together with an increased gene expression of the rate-limiting enzymes StAR and CYP11A1. Hormone production occurred at a thymocyte differentiation stage characterized by being double positive for the CD4 and CD8 surface markers but was found to be unrelated to CD69 expression, a marker for thymocytes undergoing positive selection. No GC synthesis was found in resting or anti-CD3 activated CD4 and CD8 positive T cells isolated from the spleen. Thymocyte-derived GC had an anti-proliferative effect on a GR-transfected cell line and induced apoptosis in thymocytes. The age- and differentiation stage-related GC synthesis in thymocytes may play a role in the involution process that the thymus gland undergoes

  15. Glycosyltransferase complexes improve glycolipid synthesis.

    Science.gov (United States)

    Spessott, Waldo; Crespo, Pilar M; Daniotti, José Luis; Maccioni, Hugo J F

    2012-07-30

    The synthesis of gangliosides GM3 and GD3 is carried out by the successive addition of sialic acid residues on lactosylceramide (LacCer) by the Golgi located sialyltransferases Sial-T1 and Sial-T2, respectively. CHO-K1 cells lack Sial-T2 and only express GM3. Here we show that the activity of Sial-T1 was near 2.5-fold higher in homogenates of CHO-K1 cells transfected to express Sial-T2 (CHO-K1(Sial-T2)) than in untransfected cells. The appearance of Sial-T1 enzyme or gene transcription activators or the stabilization of the Sial-T1 protein were discarded as possible causes of the activation. Sial-T2 lacking the catalytic domain failed to promote Sial-T1 activation. Since Gal-T1, Sial-T1 and Sial-T2 form a multienzyme complex, we propose that transformation of formed GM3 into GD3 and GT3 by Sial-T2 in the complex leaves Sial-T1 unoccupied, enabled for new rounds of LacCer utilization, which results in its apparent activation. PMID:22687240

  16. An Approach to Interface Synthesis

    DEFF Research Database (Denmark)

    Madsen, Jan; Hald, Bjarne

    1995-01-01

    Presents a novel interface synthesis approach based on a one-sided interface description. Whereas most other approaches consider interface synthesis as optimizing a channel to existing client/server modules, we consider the interface synthesis as part of the client/server module synthesis (which...... may contain the re-use of existing modules). The interface synthesis approach describes the basic transformations needed to transform the server interface description into an interface description on the client side of the communication medium. The synthesis approach is illustrated through a point......-to-point communication, but is applicable to synthesis of a multiple client/server environment. The interface description is based on a formalization of communication events....

  17. ATP Synthesis in the Extremely Halophilic Bacteria

    Science.gov (United States)

    Hochstein, Lawrence I.; Morrison, David (Technical Monitor)

    1994-01-01

    The proton-translocating ATPases are multimeric enzymes that carry out a multitude of essential functions. Their origin and evolution represent a seminal event in the early evolution of life. Amino acid sequences of the two largest subunits from archaeal ATPases (A-ATPases), vacuolar ATPases (V-ATPases), and FOF1-ATP syntheses (FATPases) suggest these ATPases evolved from an ancestral vacuolar-like ATP syntheses. A necessary consequence of this notion is that the A-ATPases are ATP syntheses. With the possible exception of the A-ATPase from Halobacterium salinarium. no A-ATPase has been demonstrated to synthesize ATP. The evidence for this case is dubious since ATP synthesis occurs only when conditions are distinctively unphysiological. We demonstrated that ATP synthesis in H.saccharovorum is inconsistent with the operation of an A-type ATPase. In order to determine if this phenomenon was unique to H. saccharovorum, ATP synthesis was examined in various extremely halophilic bacteria with the goal of ascertaining if it resembled what occurred in a. saccharovorum, or was consistent with the operation of an A-type ATPase. A-, V-, and F-type ATPases respond singularly to certain inhibitors. Therefore, the effect of these inhibitors on ATP synthesis in several extreme halophiles was determined. Inhibitors that either blocked or collapsed proton-gradients inhibited the steady state synthesis of ATP thus verifying that synthesis took place at the expense of a proton gradient. Azide, an inhibitor of F-ATPases inhibited ATP synthesis. Since the arginine-dependent synthesis of ATP, which occurs by way of substrate-level phosphorylation, was unaffected by azide, it was unlikely that azide acted as an "uncoupler." N -ethylmaleimide and nitrate, which inhibit V- and A-ATPases, either did not inhibit ATP synthesis or resulted in higher steady-state levels of ATP. These results suggest there are two types of proton-motive ATPases in the extreme halophiles (and presumably in other

  18. Rhodoferax-related pufM gene cluster dominates the aerobic anoxygenic phototrophic communities in German freshwater lakes

    Czech Academy of Sciences Publication Activity Database

    Salka, I.; Čuperová, Zuzana; Mašín, Michal; Koblížek, Michal; Grossart, H. P.

    2011-01-01

    Roč. 13, č. 11 (2011), s. 2865-2875. ISSN 1462-2912 R&D Projects: GA ČR GAP501/10/0221 Institutional research plan: CEZ:AV0Z50200510 Keywords : 16S RIBOSOMAL-RNA * GRADIENT GEL - ELECTROPHORESIS * BACTERIOCHLOROPHYLL-A Subject RIV: EE - Microbiology, Virology Impact factor: 5.843, year: 2011

  19. Efficient and Stereoselective Synthesis of C-(β-2'-Deoxynucleosides)

    Institute of Scientific and Technical Information of China (English)

    WU,Qin-Pei; REES,B.Colin

    2004-01-01

    @@ The formation of triplex DNA is, in principle, an effective way of regulating the expression of selected genes.[1]Many cytosine mimics were investigated to form triplex DNA, e.g., 2-amino-(2'-deoxy-β-D-ribofuranosyl)pyridine (9)was proved to have some effectiveness in stabilizing triplex DNA under neutral conditions.[2] Stereoselective synthesis of 9 was achieved starting from 2-deoxyribose.

  20. Biotin Synthesis Begins by Hijacking the Fatty Acid Synthetic Pathway

    OpenAIRE

    Lin, Steven; Hanson, Ryan E.; Cronan, John E.

    2010-01-01

    Although biotin is an essential enzyme cofactor found in all three domains of life, our knowledge of its biosynthesis remains fragmentary. Most of the carbon atoms of biotin are derived from pimelic acid, a seven carbon dicarboxylic acid, but the mechanism whereby Escherichia coli assembles this intermediate remains unknown. Genetic analysis identified only two genes of unknown function required for pimelate synthesis, bioC and bioH. We report in vivo and in vitro evidence that the pimeloyl m...

  1. Fluorochemicals used in food packaging inhibit male sex hormone synthesis

    DEFF Research Database (Denmark)

    Rosenmai, Anna Kjerstine; Nielsen, F. K.; Pedersen, Mikael;

    2013-01-01

    . The aim of this study was to elucidate the effects of six fluorochemicals on sex hormone synthesis and androgen receptor (AR) activation in vitro. Four PAPS and two metabolites, perfluorooctanoic acid (PFOA) and 8:2 fluorotelomer alcohol (8:2 FTOH) were tested. Hormone profiles, including eight steroid....... Overall, these results demonstrate that fluorochemicals present in food packaging materials and their metabolites can affect steroidogenesis through decreased Bzrp and increased CYP19 gene expression leading to lower androgen and higher estrogen levels....

  2. Distributed Priority Synthesis

    Directory of Open Access Journals (Sweden)

    Harald Ruess

    2012-11-01

    Full Text Available Given a set of interacting components with non-deterministic variable update and given safety requirements, the goal of priority synthesis is to restrict, by means of priorities, the set of possible interactions in such a way as to guarantee the given safety conditions for all possible runs. In distributed priority synthesis we are interested in obtaining local sets of priorities, which are deployed in terms of local component controllers sharing intended next moves between components in local neighborhoods only. These possible communication paths between local controllers are specified by means of a communication architecture. We formally define the problem of distributed priority synthesis in terms of a multi-player safety game between players for (angelically selecting the next transition of the components and an environment for (demonically updating uncontrollable variables. We analyze the complexity of the problem, and propose several optimizations including a solution-space exploration based on a diagnosis method using a nested extension of the usual attractor computation in games together with a reduction to corresponding SAT problems. When diagnosis fails, the method proposes potential candidates to guide the exploration. These optimized algorithms for solving distributed priority synthesis problems have been integrated into the VissBIP framework. An experimental validation of this implementation is performed using a range of case studies including scheduling in multicore processors and modular robotics.

  3. Synthesis of new radiotracers

    International Nuclear Information System (INIS)

    The brain's sensibility besides to the rigorous selectivity of changes taking place on brain's barriers leads us to synthesis specifics radiotracers based on diamine ethylene and marked with technetium radioisotope to form a radiotracer able to pass these barriers and diagnose illnesses in an early stage. These radiotracers are tested by a biodistribution on a small animal to be ratified. (Author)

  4. Synthesis of acrylic prepolymer

    International Nuclear Information System (INIS)

    An acrylic prepolymer was synthesized from glycidyl methacrylate (GMA), butyl methacrylate (BMA), methyl methacrylate (MMA) and acrylic acid (AA). Butyl acetate (BAc), benzoyl peroxide (BzO), 4-methoxyphenol (MPh) and triethylamine (TEA) were used as solvent, initiator, inhibitor and catalyst respectively. Observations of the synthesis leading to the formation of acrylic prepolymer are described. (author)

  5. Synthesis de 1-dotriacotanol

    International Nuclear Information System (INIS)

    In order to prepare isotopic labeled long chain aliphatic primary alcohol's, the synthesis overall yielding and chemical purity of 1-dotriacotanol were 41% and 98%, respectively. This procedure is very useful for carbon-14 and tritium labeling at Beta position of saturated fatty alcohol's

  6. The Synthesis of Glycoglycerolipids

    Institute of Scientific and Technical Information of China (English)

    Chun Xia LI; Ying Xia LI; Ling Bo YU; Hua ZHANG; Shi Dong CHU; Hua Shi GUAN

    2003-01-01

    A convenient synthetic route was developed for the synthesis of the novel glycolipids: 1, 2-di-O-acyl-3-O-(2'-acylamide-2'-deoxy-α-D-glucopyranosyl)-sn-glycerols. 10 new compounds of glycolipids with different acyl groups were obtained.

  7. Enzymatic synthesis of vanillin

    NARCIS (Netherlands)

    van den Heuvel, RHH; Fraaije, MW; Laane, C; van Berkel, WJH; Heuvel, Robert H.H. van den; Berkel, Willem J.H. van

    2001-01-01

    Due to increasing interest in natural vanillin, two enzymatic routes for the synthesis of vanillin were developed. The flavoprotein vanillyl alcohol oxidase (VAO) acts on a wide range of phenolic compounds and converts both creosol and vanillylamine to vanillin with high yield. The VAO-mediated conv

  8. Matrix Synthesis and Characterization

    Science.gov (United States)

    1984-01-01

    The role of NASA in the area of composite material synthesis; evaluation techniques; prediction analysis techniques; solvent-resistant tough composite matrix; resistance to paint strippers; acceptable processing temperature and pressure for thermoplastics; and the role of computer modeling and fiber interface improvement were discussed.

  9. Ascorbate Synthesis Pathway: DUAL ROLE OF ASCORBATE IN BONE HOMEOSTASIS*

    OpenAIRE

    Gabbay, Kenneth H.; Bohren, Kurt M.; Morello, Roy; Bertin, Terry; Liu, Jeff; Vogel, Peter

    2010-01-01

    Using mouse gene knock-out models, we identify aldehyde reductase (EC 1.1.1.2, Akr1a4 (GR)) and aldose reductase (EC 1.1.1.21, Akr1b3 (AR)) as the enzymes responsible for conversion of d-glucuronate to l-gulonate, a key step in the ascorbate (ASC) synthesis pathway in mice. The gene knock-out (KO) mice show that the two enzymes, GR and AR, provide ∼85 and ∼15% of l-gulonate, respectively. GRKO/ARKO double knock-out mice are unable to synthesize ASC (>95% ASC deficit) and develop scurvy. The G...

  10. Design, synthesis, and testing toward a 57-codon genome.

    Science.gov (United States)

    Ostrov, Nili; Landon, Matthieu; Guell, Marc; Kuznetsov, Gleb; Teramoto, Jun; Cervantes, Natalie; Zhou, Minerva; Singh, Kerry; Napolitano, Michael G; Moosburner, Mark; Shrock, Ellen; Pruitt, Benjamin W; Conway, Nicholas; Goodman, Daniel B; Gardner, Cameron L; Tyree, Gary; Gonzales, Alexandra; Wanner, Barry L; Norville, Julie E; Lajoie, Marc J; Church, George M

    2016-08-19

    Recoding--the repurposing of genetic codons--is a powerful strategy for enhancing genomes with functions not commonly found in nature. Here, we report computational design, synthesis, and progress toward assembly of a 3.97-megabase, 57-codon Escherichia coli genome in which all 62,214 instances of seven codons were replaced with synonymous alternatives across all protein-coding genes. We have validated 63% of recoded genes by individually testing 55 segments of 50 kilobases each. We observed that 91% of tested essential genes retained functionality with limited fitness effect. We demonstrate identification and correction of lethal design exceptions, only 13 of which were found in 2229 genes. This work underscores the feasibility of rewriting genomes and establishes a framework for large-scale design, assembly, troubleshooting, and phenotypic analysis of synthetic organisms. PMID:27540174

  11. Intestinal triacylglycerol synthesis in fat absorption and systemic energy metabolism.

    Science.gov (United States)

    Yen, Chi-Liang Eric; Nelson, David W; Yen, Mei-I

    2015-03-01

    The intestine plays a prominent role in the biosynthesis of triacylglycerol (triglyceride; TAG). Digested dietary TAG is repackaged in the intestine to form the hydrophobic core of chylomicrons, which deliver metabolic fuels, essential fatty acids, and other lipid-soluble nutrients to the peripheral tissues. By controlling the flux of dietary fat into the circulation, intestinal TAG synthesis can greatly impact systemic metabolism. Genes encoding many of the enzymes involved in TAG synthesis have been identified. Among TAG synthesis enzymes, acyl-CoA:monoacylglycerol acyltransferase 2 and acyl-CoA:diacylglycerol acyltransferase (DGAT)1 are highly expressed in the intestine. Their physiological functions have been examined in the context of whole organisms using genetically engineered mice and, in the case of DGAT1, specific inhibitors. An emerging theme from recent findings is that limiting the rate of TAG synthesis in the intestine can modulate gut hormone secretion, lipid metabolism, and systemic energy balance. The underlying mechanisms and their implications for humans are yet to be explored. Pharmacological inhibition of TAG hydrolysis in the intestinal lumen has been employed to combat obesity and associated disorders with modest efficacy and unwanted side effects. The therapeutic potential of inhibiting specific enzymes involved in intestinal TAG synthesis warrants further investigation. PMID:25231105

  12. Alterations in gastric mucin synthesis by Helicobacter pylori

    Institute of Scientific and Technical Information of China (English)

    James C, Byrd; Robert S, Bresalier

    2000-01-01

    AIM To determine the role of Helicobacter pylori in altering gastric mucin synthesis and define how thprocess relates to H. pylori-related diseases.METHODS Analyses of human gastric tissues using immunohistochemistry and in situ hybridizatiodocument the role of H. pylori in altering the composition and distribution of gastric mucins.RESULTS These data indicate a decrease in the product of the MUC5 (MUC5AC) gene and aberraexpression of MUC6 in the surface epithelium of H. pylori-infected patients. A normal pattern was restorby H. pylori eradication. Inhibition of mucin synthesis including MUC5AC and MUCl mucins by H. pvlohas been established in vitro using biochemical and Western blot analyses. This effect is not due to inhibitiof glycosylation, but results from inhibition of synthesis of mucin core structures. In vitro experiments usiinhibitors of mucin synthesis indicate that cell surface mucins decrease adhesion of H. pylori to gastepithelial cells.CONCLUSION Inhibition of mucin synthesis by H. pylori in vivo can disrupt the protective mucous layand facilitate bacterial adhesion, which may lead to increased inflammation in thc gastric epithelium.

  13. Adeno-associated virus rep protein synthesis during productive infection

    Energy Technology Data Exchange (ETDEWEB)

    Redemann, B.E.; Mendelson, E.; Carter, B.J.

    1989-02-01

    Adeno-associated virus (AAV) Rep proteins mediate viral DNA replication and can regulate expression from AAV genes. The authors studied the kinetics of synthesis of the four Rep proteins, Rep78, Rep68, Rep52, and Rep40, during infection of human 293 or KB cells with AAV and helper adenovirus by in vivo labeling with (/sup 35/S)methionine, immunoprecipitation, and immunoblotting analyses. Rep78 and Rep52 were readily detected concomitantly with detection of viral monomer duplex DNA replicating about 10 to 12 h after infection, and Rep68 and Rep40 were detected 2 h later. Rep78 and Rep52 were more abundant than Rep68 and Rep40 owing to a higher synthesis rate throughout the infectious cycle. In some experiments, very low levels of Rep78 could be detected as early as 4 h after infection. The synthesis rates of Rep proteins were maximal between 14 and 24 h and then decreased later after infection. Isotopic pulse-chase experiments showed that each of the Rep proteins was synthesized independently and was stable for at least 15 h. A slower-migrating, modified form of Rep78 was identified late after infection. AAV capsid protein synthesis was detected at 10 to 12 h after infection and also exhibited synthesis kinetics similar to those of the Rep proteins. AAV DNA replication showed at least two clearly defined stages. Bulk duplex replicating DNA accumulation began around 10 to 12 h and reached a maximum level at about 20 h when Rep and capsid protein synthesis was maximal. Progeny single-stranded DNA accumulation began about 12 to 13 h, but most of this DNA accumulated after 24 h when Rep and capsid protein synthesis had decreased.

  14. In vitro synthesis of the iron-molybdenum cofactor of nitrogenase.

    OpenAIRE

    Shah, V K; Imperial, J; Ugalde, R A; Ludden, P W; Brill, W J

    1986-01-01

    Molybdate- and ATP-dependent in vitro synthesis of the iron-molybdenum cofactor (FeMo-co) of nitrogenase requires the protein products of at least the nifB, nifN, and nifE genes. Extracts of FeMo-co-negative mutants of Klebsiella pneumoniae and Azotobacter vinelandii with lesions in different genes can be complemented for FeMo-co synthesis. Both K. pneumoniae and A. vinelandii dinitrogenase (component I) deficient in FeMo-co can be activated by FeMo-co synthesized in vitro. Properties of the ...

  15. Viral RNA polymerase scanning and the gymnastics of Sendai virus RNA synthesis

    International Nuclear Information System (INIS)

    mRNA synthesis from nonsegmented negative-strand RNA virus (NNV) genomes is unique in that the genome RNA is embedded in an N protein assembly (the nucleocapsid) and the viral RNA polymerase does not dissociate from the template after release of each mRNA, but rather scans the genome RNA for the next gene-start site. A revised model for NNV RNA synthesis is presented, in which RNA polymerase scanning plays a prominent role. Polymerase scanning of the template is known to occur as the viral transcriptase negotiates gene junctions without falling off the template

  16. Local gene expression in nerve endings.

    Science.gov (United States)

    Crispino, Marianna; Chun, Jong Tai; Cefaliello, Carolina; Perrone Capano, Carla; Giuditta, Antonio

    2014-03-01

    At the Nobel lecture for physiology in 1906, Ramón y Cajal famously stated that "the nerve elements possess reciprocal relationships in contiguity but not in continuity," summing up the neuron doctrine. Sixty years later, by the time the central dogma of molecular biology formulated the axis of genetic information flow from DNA to mRNA, and then to protein, it became obvious that neurons with extensive ramifications and long axons inevitably incur an innate problem: how can the effect of gene expression be extended from the nucleus to the remote and specific sites of the cell periphery? The most straightforward solution would be to deliver soma-produced proteins to the target sites. The influential discovery of axoplasmic flow has supported this scheme of protein supply. Alternatively, mRNAs can be dispatched instead of protein, and translated locally at the strategic target sites. Over the past decades, such a local system of protein synthesis has been demonstrated in dendrites, axons, and presynaptic terminals. Moreover, the local protein synthesis in neurons might even involve intercellular trafficking of molecules. The innovative concept of glia-neuron unit suggests that the local protein synthesis in the axonal and presynaptic domain of mature neurons is sustained by a local supply of RNAs synthesized in the surrounding glial cells and transferred to these domains. Here, we have reviewed some of the evidence indicating the presence of a local system of protein synthesis in axon terminals, and have examined its regulation in various model systems. PMID:23853157

  17. GENE DOPING IN SPORT – PERSPECTIVES AND RISKS

    OpenAIRE

    2014-01-01

    In the past few years considerable progress regarding the knowledge of the human genome map has been achieved. As a result, attempts to use gene therapy in patients’ management are more and more often undertaken. The aim of gene therapy is to replace defective genes in vivo and/or to promote the long-term endogenous synthesis of deficient protein. In vitro studies improve the production of human recombinant proteins, such as insulin (INS), growth hormone (GH), insulin-like growth factor-1 (IG...

  18. Function and evolution of the serotonin-synthetic bas-1 gene and other aromatic amino acid decarboxylase genes in Caenorhabditis

    Directory of Open Access Journals (Sweden)

    Hare Emily E

    2004-08-01

    Full Text Available Abstract Background Aromatic L-amino acid decarboxylase (AADC enzymes catalyze the synthesis of biogenic amines, including the neurotransmitters serotonin and dopamine, throughout the animal kingdom. These neurotransmitters typically perform important functions in both the nervous system and other tissues, as illustrated by the debilitating conditions that arise from their deficiency. Studying the regulation and evolution of AADC genes is therefore desirable to further our understanding of how nervous systems function and evolve. Results In the nematode C. elegans, the bas-1 gene is required for both serotonin and dopamine synthesis, and maps genetically near two AADC-homologous sequences. We show by transformation rescue and sequencing of mutant alleles that bas-1 encodes an AADC enzyme. Expression of a reporter construct in transgenics suggests that the bas-1 gene is expressed, as expected, in identified serotonergic and dopaminergic neurons. The bas-1 gene is one of six AADC-like sequences in the C. elegans genome, including a duplicate that is immediately downstream of the bas-1 gene. Some of the six AADC genes are quite similar to known serotonin- and dopamine-synthetic AADC's from other organisms whereas others are divergent, suggesting previously unidentified functions. In comparing the AADC genes of C. elegans with those of the congeneric C. briggsae, we find only four orthologous AADC genes in C. briggsae. Two C. elegans AADC genes – those most similar to bas-1 – are missing from C. briggsae. Phylogenetic analysis indicates that one or both of these bas-1-like genes were present in the common ancestor of C. elegans and C. briggsae, and were retained in the C. elegans line, but lost in the C. briggsae line. Further analysis of the two bas-1-like genes in C. elegans suggests that they are unlikely to encode functional enzymes, and may be expressed pseudogenes. Conclusions The bas-1 gene of C. elegans encodes a serotonin- and dopamine

  19. Bacterial cellulose synthesis mechanism of facultative anaerobe Enterobacter sp. FY-07.

    Science.gov (United States)

    Ji, Kaihua; Wang, Wei; Zeng, Bing; Chen, Sibin; Zhao, Qianqian; Chen, Yueqing; Li, Guoqiang; Ma, Ting

    2016-01-01

    Enterobacter sp. FY-07 can produce bacterial cellulose (BC) under aerobic and anaerobic conditions. Three potential BC synthesis gene clusters (bcsI, bcsII and bcsIII) of Enterobacter sp. FY-07 have been predicted using genome sequencing and comparative genome analysis, in which bcsIII was confirmed as the main contributor to BC synthesis by gene knockout and functional reconstitution methods. Protein homology, gene arrangement and gene constitution analysis indicated that bcsIII had high identity to the bcsI operon of Enterobacter sp. 638; however, its arrangement and composition were same as those of BC synthesizing operon of G. xylinum ATCC53582 except for the flanking sequences. According to the BC biosynthesizing process, oxygen is not directly involved in the reactions of BC synthesis, however, energy is required to activate intermediate metabolites and synthesize the activator, c-di-GMP. Comparative transcriptome and metabolite quantitative analysis demonstrated that under anaerobic conditions genes involved in the TCA cycle were downregulated, however, genes in the nitrate reduction and gluconeogenesis pathways were upregulated, especially, genes in three pyruvate metabolism pathways. These results suggested that Enterobacter sp. FY-07 could produce energy efficiently under anaerobic conditions to meet the requirement of BC biosynthesis. PMID:26911736

  20. Experimental and computational validation of models of fluorescent and luminescent reporter genes in bacteria

    OpenAIRE

    Pinel Corinne; Ropers Delphine; Ranquet Caroline; de Jong Hidde; Geiselmann Johannes

    2010-01-01

    Abstract Background Fluorescent and luminescent reporter genes have become popular tools for the real-time monitoring of gene expression in living cells. However, mathematical models are necessary for extracting biologically meaningful quantities from the primary data. Results We present a rigorous method for deriving relative protein synthesis rates (mRNA concentrations) and protein concentrations by means of kinetic models of gene expression. We experimentally and computationally validate t...

  1. The Application of Nanoparticles in Gene Therapy and Magnetic Resonance Imaging

    OpenAIRE

    Herranz, Fernando; Almarza, Elena; Rodríguez, Ignacio; Salinas, Beatriz; ROSELL, YAMILKA; Desco, Manuel; BULTE, JEFF W.; Ruiz-Cabello, Jesús

    2011-01-01

    The combination of nanoparticles, gene therapy, and medical imaging has given rise to a new field known as gene theranostics, in which a nanobioconjugate is used to diagnose and treat the disease. The process generally involves binding between a vector carrying the genetic information and a nanoparticle, which provides the signal for imaging. The synthesis of this probe generates a synergic effect, enhancing the efficiency of gene transduction and imaging contrast. We discuss the latest appro...

  2. Homeobox genes in the rodent pineal gland: roles in development and phenotype maintenance

    OpenAIRE

    Rath, Martin F.; Rohde, Kristian; Klein, David C.; Møller, Morten

    2012-01-01

    The pineal gland is a neuroendocrine gland responsible for nocturnal synthesis of melatonin. During early development of the rodent pineal gland from the roof of the diencephalon, homeobox genes of the orthodenticle homeobox (Otx)- and paired box (Pax)-families are expressed and are essential for normal pineal development consistent with the well-established role that homeobox genes play in developmental processes. However, the pineal gland appears to be unusual because strong homeobox gene e...

  3. Osmoregulated periplasmic glucans synthesis gene family of Shigella flexneri

    Science.gov (United States)

    Osmoregulated periplasmic glucans (OPGs) of foodborne enteropathogen Shigella flexneri were characterized. OPGs were composed of 100 percent glucose with 2-linked glucose as the most abundant residue with terminal glucose, 2-linked and 2,6-linked glucose also present in high quantities. Most dominan...

  4. Gene gymnastics

    Science.gov (United States)

    Vijayachandran, Lakshmi S; Thimiri Govinda Raj, Deepak B; Edelweiss, Evelina; Gupta, Kapil; Maier, Josef; Gordeliy, Valentin; Fitzgerald, Daniel J; Berger, Imre

    2013-01-01

    Most essential activities in eukaryotic cells are catalyzed by large multiprotein assemblies containing up to ten or more interlocking subunits. The vast majority of these protein complexes are not easily accessible for high resolution studies aimed at unlocking their mechanisms, due to their low cellular abundance and high heterogeneity. Recombinant overproduction can resolve this bottleneck and baculovirus expression vector systems (BEVS) have emerged as particularly powerful tools for the provision of eukaryotic multiprotein complexes in high quality and quantity. Recently, synthetic biology approaches have begun to make their mark in improving existing BEVS reagents by de novo design of streamlined transfer plasmids and by engineering the baculovirus genome. Here we present OmniBac, comprising new custom designed reagents that further facilitate the integration of heterologous genes into the baculovirus genome for multiprotein expression. Based on comparative genome analysis and data mining, we herein present a blueprint to custom design and engineer the entire baculovirus genome for optimized production properties using a bottom-up synthetic biology approach. PMID:23328086

  5. Transcriptome analysis reveals key differentially expressed genes involved in wheat grain development

    Directory of Open Access Journals (Sweden)

    Yonglong Yu

    2016-04-01

    Full Text Available Wheat seed development is an important physiological process of seed maturation and directly affects wheat yield and quality. In this study, we performed dynamic transcriptome microarray analysis of an elite Chinese bread wheat cultivar (Jimai 20 during grain development using the GeneChip Wheat Genome Array. Grain morphology and scanning electron microscope observations showed that the period of 11–15 days post-anthesis (DPA was a key stage for the synthesis and accumulation of seed starch. Genome-wide transcriptional profiling and significance analysis of microarrays revealed that the period from 11 to 15 DPA was more important than the 15–20 DPA stage for the synthesis and accumulation of nutritive reserves. Series test of cluster analysis of differential genes revealed five statistically significant gene expression profiles. Gene ontology annotation and enrichment analysis gave further information about differentially expressed genes, and MapMan analysis revealed expression changes within functional groups during seed development. Metabolic pathway network analysis showed that major and minor metabolic pathways regulate one another to ensure regular seed development and nutritive reserve accumulation. We performed gene co-expression network analysis to identify genes that play vital roles in seed development and identified several key genes involved in important metabolic pathways. The transcriptional expression of eight key genes involved in starch and protein synthesis and stress defense was further validated by qRT-PCR. Our results provide new insight into the molecular mechanisms of wheat seed development and the determinants of yield and quality.

  6. Transcriptome analysis reveals key differentially expressed genes involved in wheat grain development

    Institute of Scientific and Technical Information of China (English)

    Yonglong Yu; Dong Zhu; Chaoying Ma; Hui Cao; Yaping Wang; Yanhao Xu; Wenying Zhang; Yueming Yan

    2016-01-01

    Wheat seed development is an important physiological process of seed maturation and directly affects wheat yield and quality. In this study, we performed dynamic transcriptome microarray analysis of an elite Chinese bread wheat cultivar (Jimai 20) during grain development using the GeneChip Wheat Genome Array. Grain morphology and scanning electron microscope observations showed that the period of 11–15 days post-anthesis (DPA) was a key stage for the synthesis and accumulation of seed starch. Genome-wide transcriptional profiling and significance analysis of microarrays revealed that the period from 11 to 15 DPA was more important than the 15–20 DPA stage for the synthesis and accumulation of nutritive reserves. Series test of cluster analysis of differential genes revealed five statistically significant gene expression profiles. Gene ontology annotation and enrichment analysis gave further informa-tion about differentially expressed genes, and MapMan analysis revealed expression changes within functional groups during seed development. Metabolic pathway network analysis showed that major and minor metabolic pathways regulate one another to ensure regular seed development and nutritive reserve accumulation. We performed gene co-expression network analysis to identify genes that play vital roles in seed development and identified several key genes involved in important metabolic pathways. The transcriptional expression of eight key genes involved in starch and protein synthesis and stress defense was further validated by qRT-PCR. Our results provide new insight into the molecular mechanisms of wheat seed development and the determinants of yield and quality.

  7. Cloning and characterization of a locus encoding an indolepyruvate decarboxylase involved in indole-3-acetic acid synthesis in Erwinia herbicola.

    OpenAIRE

    Brandl, M. T.; Lindow, S E

    1996-01-01

    Erwinia herbicola 299R synthesizes indole-3-acetic acid (IAA) primarily by the indole-3-pyruvic acid pathway. A gene involved in the biosynthesis of IAA was cloned from strain 299R. This gene (ipdC) conferred the synthesis of indole-3-acetaldehyde and tryptophol upon Escherichia coli DH5 alpha in cultures supplemented with L-tryptophan. The deduced amino acid sequence of the gene product has high similarity to that of the indolepyruvate decarboxylase of Enterobacter cloacae. Regions within py...

  8. Ethics of Chemical Synthesis

    Directory of Open Access Journals (Sweden)

    Joachim Schummer

    2001-10-01

    Full Text Available Unlike other branches of science, the scientific products of synthetic chemistry are not only ideas but also new substances that change our material world, for the benefit or harm of living beings. This paper provides for the first time a systematical analysis of moral issues arising from chemical synthesis, based on concepts of responsibility and general morality. Topics include the questioning of moral neutrality of chemical synthesis as an end in itself, chemical weapons research, moral objections against improving material conditions of life by chemical means, and freedom of research. The paper aims at providing both a sound basis for moral judgements of chemistry in a public discourse and a framework for chemists to reflect on the moral relevance of their activity.

  9. Benzoylurea Chitin Synthesis Inhibitors.

    Science.gov (United States)

    Sun, Ranfeng; Liu, Chunjuan; Zhang, Hao; Wang, Qingmin

    2015-08-12

    Benzoylurea chitin synthesis inhibitors are widely used in integrated pest management (IPM) and insecticide resistance management (IRM) programs due to their low toxicity to mammals and predatory insects. In the past decades, a large number of benzoylurea derivatives have been synthesized, and 15 benzoylurea chitin synthesis inhibitors have been commercialized. This review focuses on the history of commercial benzolyphenylureas (BPUs), synthetic methods, structure-activity relationships (SAR), action mechanism research, environmental behaviors, and ecotoxicology. Furthermore, their disadvantages of high risk to aquatic invertebrates and crustaceans are pointed out. Finally, we propose that the para-substituents at anilide of benzoylphenylureas should be the functional groups, and bipartite model BPU analogues are discussed in an attempt to provide new insight for future development of BPUs. PMID:26168369

  10. Prebiotic synthesis of histidine

    Science.gov (United States)

    Shen, C.; Yang, L.; Miller, S. L.; Oro, J.

    1990-01-01

    The prebiotic formation of histidine (His) has been accomplished experimentally by the reaction of erythrose with formamidine followed by a Strecker synthesis. In the first step of this reaction sequence, the formation of imidazole-4-acetaldehyde took place by the condensation of erythrose and formamidine, two compounds that are known to be formed under prebiotic conditions. In a second step, the imidazole-4-acetaldehyde was converted to His, without isolation of the reaction products by adding HCN and ammonia to the reaction mixture. LC, HPLC, thermospray liquid chromatography-mass spectrometry, and tandem mass spectrometry were used to identify the product, which was obtained in a yield of 3.5% based on the ratio of His/erythrose. This is a new chemical synthesis of one of the basic amino acids which had not been synthesized prebiotically until now.

  11. Synthesis of approximation errors

    Energy Technology Data Exchange (ETDEWEB)

    Bareiss, E.H.; Michel, P.

    1977-07-01

    A method is developed for the synthesis of the error in approximations in the large of regular and irregular functions. The synthesis uses a small class of dimensionless elementary error functions which are weighted by the coefficients of the expansion of the regular part of the function. The question is answered whether a computer can determine the analytical nature of a solution by numerical methods. It is shown that continuous least-squares approximations of irregular functions can be replaced by discrete least-squares approximation and how to select the discrete points. The elementary error functions are used to show how the classical convergence criterions can be markedly improved. There are eight numerical examples included, 30 figures and 74 tables.

  12. 花青素合成转录因子基因在玉米中的表达研究:一种新型基因可视化跟踪表达系统%Expression of foreign transcription genes Bi and Cl on anthocyanin synthesis pathway in maize: A novel expression system of visual tracking for transgene

    Institute of Scientific and Technical Information of China (English)

    宫硖; 杨凤萍; 薛静; 陈绪清; 张立全; 李向龙; 张晓东

    2012-01-01

    Anthocyanin is a kind of water-soluble flavonoids, which make the plants showing red, blue, purple, red purple or other colors. Expression of the metabolic regulation gene causes the accumulation of anthocyanins in plant cells, and makes the plant organs change color, easy to observe. In this paper, using Bi and Cl gene which involved in anthocyanin synthesis pathway as a visual reporter, and glyphosate resistant gene epsp, encoding 5-enolpyruvylshikimate-3 phosphate synthase, as a selection marker, plant expression vector pBAC9009 was constructed. Immature embryos and embryogenic callus of several excellent maize inbred lines were transformed with biolistic bombardment. Undertaking in vitro culture steps of callus induction, glyphosate resistance selection, differentiation and plant regeneration, 75 transgenic plants were regenerated, and 43 ears of corn were harvested. There are 18 T0 lines expressed parts or whole purple on the seedling or the growth stage, 8 ears of these lines with scattered purple seeds. Analysis of PCR, RT-PCR and anthocyanin content showed that exogenous genes were well integrated in the genome of maize and efficiently expressed in the purple leaves and seeds. In conclusion, we successfully constructed a visual tracking system for transgene which make the transformed plants can be easily distinguished from their appearance, only by its color appearance at the early stage. Application of the system can greatly improve efficiency of transgene, and save costs. It is very important for plant transgenic research.%花青素是一种水溶性的黄酮类物质,可使植物呈现出红、蓝、紫和红紫等颜色.外源花青素代谢调控基因的表达可使花青素在植物细胞内积累,使植物体外观上表现出色彩的变化,易于观察,因此可作为报告基因用于植物转基因研究,快速报告细胞、组织、器官或植株是否被转化.本研究用花青素代谢调控基因Bi和Cl作为报告基因,以epsp作为筛

  13. Total synthesis of (-)-spinosyn A.

    Science.gov (United States)

    Mergott, Dustin J; Frank, Scott A; Roush, William R

    2004-08-17

    A convergent, highly stereoselective total synthesis of (-)-spinosyn A (1) is described. Key features of the synthesis include the transannular Diels-Alder reaction of macrocyclic pentaene 11 and the transannular Morita-Baylis-Hillman cyclization of 12 that generates tetracycle 26. The total synthesis of (-)-spinosyn A was completed by a sequence involving the highly beta-selective glycosidation reaction of 13 and glycosyl imidate 30. PMID:15173590

  14. Modulation of gene expression by RNAi.

    Science.gov (United States)

    Wójcik, Cezary; Fabunmi, Rosalind; DeMartino, George N

    2005-01-01

    RNA interference (RNAi) is a form of posttranscriptional gene silencing in which the presence within the cell of double-stranded RNA (dsRNA) leads to the specific degradation of mRNA with a complimentary sequence. RNAi is a natural phenomenon that can be exploited as a powerful tool to study gene function by generating gene "knockdowns" in various cell types. RNAi is mediated by short interfering RNAs (siRNAs), which are generated within cells from long dsRNAs. To avoid generalized toxic effects, mammalian cells are transfected directly with 21-23-bp-long siRNAs generated either by chemical synthesis or obtained by a series of enzymatic reactions. The present chapter deals with siRNA design, synthesis, transfection, and readout of efficiency in a mammalian cell culture system. The general principle is illustrated by the functional knockdown of p97/VCP (valosin-containing protein) in HeLa cells using five different siRNA sequences. PMID:16028696

  15. Microwave Multicomponent Synthesis

    OpenAIRE

    Helmut M Hügel

    2009-01-01

    In the manner that very important research is often performed by multidisciplinary research teams, the applications of multicomponent reactions involving the combination of multiple starting materials with different functional groups leading to the higher efficiency and environmentally friendly construction of multifunctional/complex target molecules is growing in importance. This review will explore the advances and advantages in microwave multicomponent synthesis (MMS) that have been achiev...

  16. Enzymatic synthesis of vanillin

    OpenAIRE

    van den Heuvel, RHH; Fraaije, MW; Laane, C; van Berkel, WJH; Heuvel, Robert H.H. van den; van Berkel, Willem J. H.

    2001-01-01

    Due to increasing interest in natural vanillin, two enzymatic routes for the synthesis of vanillin were developed. The flavoprotein vanillyl alcohol oxidase (VAO) acts on a wide range of phenolic compounds and converts both creosol and vanillylamine to vanillin with high yield. The VAO-mediated conversion of creosol proceeds via a two-step process in which the initially formed vanillyl alcohol is further oxidized to vanillin. Catalysis is limited by the formation of an abortive complex betwee...

  17. Total Synthesis of Millingtonine.

    Science.gov (United States)

    Brown, Patrick D; Lawrence, Andrew L

    2016-07-11

    Millingtonine is a glycosidic alkaloid that exists as a pair of pseudo-enantiomeric diastereomers. Consideration of the likely biosynthetic origins of this unusual natural product has resulted in the development of a seven-step total synthesis. Results from this synthetic work provide evidence in support of a proposed network of biosynthetic pathways that can account for the formation of several phenylethanoid natural products. PMID:27249628

  18. Phase contrast image synthesis

    DEFF Research Database (Denmark)

    Glückstad, J.

    1996-01-01

    A new method is presented for synthesizing arbitrary intensity patterns based on phase contrast imaging. The concept is grounded on an extension of the Zernike phase contrast method into the domain of full range [0; 2 pi] phase modulation. By controlling the average value of the input phase...... function and by choosing appropriate phase retardation at the phase contrast filter, a pure phase to intensity imaging is accomplished. The method presented is also directly applicable in dark field image synthesis....

  19. Stochastic component mode synthesis

    OpenAIRE

    Bah, Mamadou T.; Nair, Prasanth B.; Bhaskar, Atul; Keane, Andy J.

    2003-01-01

    In this paper, a stochastic component mode synthesis method is developed for the dynamic analysis of large-scale structures with parameter uncertainties. The main idea is to represent each component displacement using a subspace spanned by a set of stochastic basis vectors in the same fashion as in stochastic reduced basis methods [1, 2]. These vectors represent however stochastic modes in contrast to the deterministic modes used in conventional substructuring methods [3]. The Craig-Bampton r...

  20. Total synthesis of teixobactin.

    Science.gov (United States)

    Jin, Kang; Sam, Iek Hou; Po, Kathy Hiu Laam; Lin, Du'an; Ghazvini Zadeh, Ebrahim H; Chen, Sheng; Yuan, Yu; Li, Xuechen

    2016-01-01

    To cope with the global bacterial multidrug resistance, scientific communities have devoted significant efforts to develop novel antibiotics, particularly those with new modes of actions. Teixobactin, recently isolated from uncultured bacteria, is considered as a promising first-in-class drug candidate for clinical development. Herein, we report its total synthesis by a highly convergent Ser ligation approach and this strategy allows us to prepare several analogues of the natural product. PMID:27484680

  1. HNAB: synthesis and characterization

    Energy Technology Data Exchange (ETDEWEB)

    O' Keefe, D.M.

    1976-05-01

    The laboratory and pilot-plant scale synthesis of hexanitroazobenzene (HNAB) are detailed. Some of its physical, chemical, and explosive properties are presented and discussed. Among these are included polymorphic forms, thermal behavior, spectra, solubility, compatibility, toxicity, sensitivity to gap test, detonation velocity and pressure, impact sensitivity, and explosive classification. Finally, some alternate syntheses of HNAB and its intermediates are considered; some of these were successful and some were not.

  2. Total synthesis of ochnaflavone

    OpenAIRE

    Monica M. Ndoile; van Heerden, Fanie R

    2013-01-01

    The first total syntheses of ochnaflavone, an asymmetric biflavone consisting of apigenin and luteolin moieties, and the permethyl ether of 2,3,2'',3''-tetrahydroochnaflavone have been achieved. The key steps in the synthesis of ochnaflavone were the formation of a diaryl ether and ring cyclization of an ether-linked dimeric chalcone to assemble the two flavone nuclei. Optimal experimental conditions for the oxidative cyclization to form ochnaflavone were established.

  3. Total synthesis of ochnaflavone

    Directory of Open Access Journals (Sweden)

    Monica M. Ndoile

    2013-07-01

    Full Text Available The first total syntheses of ochnaflavone, an asymmetric biflavone consisting of apigenin and luteolin moieties, and the permethyl ether of 2,3,2'',3''-tetrahydroochnaflavone have been achieved. The key steps in the synthesis of ochnaflavone were the formation of a diaryl ether and ring cyclization of an ether-linked dimeric chalcone to assemble the two flavone nuclei. Optimal experimental conditions for the oxidative cyclization to form ochnaflavone were established.

  4. Towards a new Synthesis

    OpenAIRE

    Foss, Nicolai J.; Klein, Peter G.; Kor, Yasemin Y.; Joseph T. Mahoney

    2006-01-01

    This paper maintains that the consistent application of subjectivism helps to reconcile contemporary entrepreneurship theory with strategic management research in general, and the resource−based view in particular. The paper synthesizes theoretical insights from Austrian economics and Penrose’s (1959) resources approach, arguing that entrepreneurship is inherently subjective and firm specific. This new synthesis describes how entrepreneurship is manifested in teams, and is driven by both hete...

  5. Combinatorial materials synthesis

    OpenAIRE

    Ichiro Takeuchi; Jochen Lauterbach; Michael J. Fasolka

    2005-01-01

    The pace at which major technological changes take place is often dictated by the rate at which new materials are discovered, and the timely arrival of new materials has always played a key role in bringing advances to our society. It is no wonder then that the so-called combinatorial or high-throughput strategy has been embraced by practitioners of materials science in virtually every field. High-throughput experimentation allows simultaneous synthesis and screening of large arrays of differ...

  6. Phase contrast image synthesis

    DEFF Research Database (Denmark)

    Glückstad, J.

    A new method is presented for synthesizing arbitrary intensity patterns based on phase contrast imaging. The concept is grounded on an extension of the Zernike phase contrast method into the domain of full range [0; 2 pi] phase modulation. By controlling the average value of the input phase funct...... function and by choosing appropriate phase retardation at the phase contrast filter, a pure phase to intensity imaging is accomplished. The method presented is also directly applicable in dark field image synthesis....

  7. Adenosine triphosphate inhibits melatonin synthesis in the rat pineal gland.

    Science.gov (United States)

    Souza-Teodoro, Luis Henrique; Dargenio-Garcia, Letícia; Petrilli-Lapa, Camila Lopes; Souza, Ewerton da Silva; Fernandes, Pedro A C M; Markus, Regina P; Ferreira, Zulma S

    2016-03-01

    Adenosine triphosphate (ATP) is released onto the pinealocyte, along with noradrenaline, from sympathetic neurons and triggers P2Y1 receptors that enhance β-adrenergic-induced N-acetylserotonin (NAS) synthesis. Nevertheless, the biotransformation of NAS into melatonin, which occurs due to the subsequent methylation by acetylserotonin O-methyltransferase (ASMT; EC 2.1.1.4), has not yet been evaluated in the presence of purinergic stimulation. We therefore evaluated the effects of purinergic signaling on melatonin synthesis induced by β-adrenergic stimulation. ATP increased NAS levels, but, surprisingly, inhibited melatonin synthesis in an inverse, concentration-dependent manner. Our results demonstrate that enhanced NAS levels, which depend on phospholipase C (PLC) activity (but not the induction of gene transcription), are a post-translational effect. By contrast, melatonin reduction is related to an ASMT inhibition of expression at both the gene transcription and protein levels. These results were independent of nuclear factor-kappa B (NF-kB) translocation. Neither the P2Y1 receptor activation nor the PLC-mediated pathway was involved in the decrease in melatonin, indicating that ATP regulates pineal metabolism through different mechanisms. Taken together, our data demonstrate that purinergic signaling differentially modulates NAS and melatonin synthesis and point to a regulatory role for ATP as a cotransmitter in the control of ASMT, the rate-limiting enzyme in melatonin synthesis. The endogenous production of melatonin regulates defense responses; therefore, understanding the mechanisms involving ASMT regulation might provide novel insights into the development and progression of neurological disorders since melatonin presents anti-inflammatory, neuroprotective, and neurogenic effects. PMID:26732366

  8. Methods of component mode synthesis

    Science.gov (United States)

    Craig, R. R., Jr.

    1977-01-01

    A generalized substructure coupling, or component mode synthesis, procedure is described. Specific methods, applications, and such special topics as damping and experimental verification are surveyed.

  9. Radiochemical synthesis of etomoxir

    International Nuclear Information System (INIS)

    Sodium 2-{6-(4-chlorophenoxy)hexyl}oxirane-2-carboxylate (Etomoxir) inhibits transport of fatty acids via the carnitine shuttle into mitochondria of muscle cells and prevents long chain fatty acids from providing energy through β-oxidation especially for muscle contraction. The objective of this synthesis is to develop a method for radioiodination of Etomoxir in order to explore its potential in diagnostic metabolic studies and molecular imaging. Thus, a method is described for the radiochemical synthesis and purification of ethyl 2-{6-(4-[131I]iodophenoxy)hexyl}oxirane-2-carboxylate (3) and 2-{6-(4-[131I]iodo-phenoxy)hexyl}oxirane-2-carboxylic acid (4). For the synthesis of these new agents, ethyl 2-{6-(4-bromophenoxy)hexyl}oxirane-2-carboxylate (1) and 2-{6-(4-bromophenoxy)hexyl}oxirane-2-carboxylic acid (2) were refluxed with [131I]NaI in the presence of anhydrous acetone at a temperature of 80 oC and 90 oC for a period of 3-4 hours, respectively. The method of radiolabeling, based on the nucleophilic exchange reaction, resulted in a radiochemical yield of 43% and 67% for compounds 3 and 4, respectively. This paper reports on the labeling of etomoxir with radioiodine as 124I labeled etomoxir may be of great importance in molecular imaging.

  10. Chemistry of Ammonothermal Synthesis

    Directory of Open Access Journals (Sweden)

    Theresia M. M. Richter

    2014-02-01

    Full Text Available Ammonothermal synthesis is a method for synthesis and crystal growth suitable for a large range of chemically different materials, such as nitrides (e.g., GaN, AlN, amides (e.g., LiNH2, Zn(NH22, imides (e.g., Th(NH2, ammoniates (e.g., Ga(NH33F3, [Al(NH36]I3 · NH3 and non-nitrogen compounds like hydroxides, hydrogen sulfides and polychalcogenides (e.g., NaOH, LiHS, CaS, Cs2Te5. In particular, large scale production of high quality crystals is possible, due to comparatively simple scalability of the experimental set-up. The ammonothermal method is defined as employing a heterogeneous reaction in ammonia as one homogenous fluid close to or in supercritical state. Three types of milieus may be applied during ammonothermal synthesis: ammonobasic, ammononeutral or ammonoacidic, evoked by the used starting materials and mineralizers, strongly influencing the obtained products. There is little known about the dissolution and materials transport processes or the deposition mechanisms during ammonothermal crystal growth. However, the initial results indicate the possible nature of different intermediate species present in the respective milieus.

  11. Gold Nanoparticle Microwave Synthesis

    Energy Technology Data Exchange (ETDEWEB)

    Krantz, Kelsie E. [Savannah River Site (SRS), Aiken, SC (United States). Savannah River National Lab. (SRNL); Christian, Jonathan H. [Savannah River Site (SRS), Aiken, SC (United States). Savannah River National Lab. (SRNL); Coopersmith, Kaitlin [Savannah River Site (SRS), Aiken, SC (United States). Savannah River National Lab. (SRNL); Washington, II, Aaron L. [Savannah River Site (SRS), Aiken, SC (United States). Savannah River National Lab. (SRNL); Murph, Simona H. [Savannah River Site (SRS), Aiken, SC (United States). Savannah River National Lab. (SRNL)

    2016-07-27

    At the nanometer scale, numerous compounds display different properties than those found in bulk material that can prove useful in areas such as medicinal chemistry. Gold nanoparticles, for example, display promise in newly developed hyperthermia therapies for cancer treatment. Currently, gold nanoparticle synthesis is performed via the hot injection technique which has large variability in final particle size and a longer reaction time. One underdeveloped area by which these particles could be produced is through microwave synthesis. To initiate heating, microwaves agitate polar molecules creating a vibration that gives off the heat energy needed. Previous studies have used microwaves for gold nanoparticle synthesis; however polar solvents were used that partially absorbed incident microwaves, leading to partial thermal heating of the sample rather than taking full advantage of the microwave to solely heat the gold nanoparticle precursors in a non-polar solution. Through this project, microwaves were utilized as the sole heat source, and non-polar solvents were used to explore the effects of microwave heating only as pertains to the precursor material. Our findings show that the use of non-polar solvents allows for more rapid heating as compared to polar solvents, a reduction in reaction time from 10 minutes to 1 minute, maximizes the efficiency of the reaction, and allows for reproducibility in the size/shape of the fabricated nanoparticles.

  12. Integrated methanol synthesis

    International Nuclear Information System (INIS)

    This invention concerns a plant for methanol manufacture from gasified coal, particularly using nuclear power. In order to reduce the cost of the hydrogen circuits, the methanol synthesis is integrated in the coal gasification plant. The coal used is gasified with hydration by means of hydrogen and the crude gas emerging, after cooling and separating the carbon dioxide and hydrogen sulphide, is mixed with the synthetic gas leaving the methane cracking furnace. This mixture is taken to the methanol synthesis and more than 90% is converted into methanol in one pass. The gas mixture remaning after condensation and separation of methanol is decomposed into three fractions in low temperature gas decomposition with a high proportion of unconverted carbon monoxide. The flow of methane is taken to the cracking furnace with steam, the flow of hydrogen is taken to the hydrating coal gasifier, and the flow of carbon monoxide is taken to the methanol synthesis. The heat required for cracking the methane can either be provided by a nuclear reactor or by the coke left after hydrating gasification. (orig./RB)

  13. Principles of gene therapy

    OpenAIRE

    Mammen Biju; Ramakrishnan T; Sudhakar Uma; Vijayalakshmi

    2007-01-01

    Genes are specific sequences of bases that encode instructions to make proteins. When genes are altered so that encoded proteins are unable to carry out their normal functions, genetic disorders can result. Gene therapy is designed to introduce genetic material into cells to compensate for abnormal genes or to make a beneficial protein. This article reviews the fundamentals in gene therapy and its various modes of administration with an insight into the role of gene therapy in Periodontics an...

  14. Recombinant DNA repair genes

    International Nuclear Information System (INIS)

    We have developed a gene transfer system with Chinese hamster ovary (CHO) cells to identify, characterize, and potentially isolate functionally homologous human or CHO genes regulating repair initiation

  15. Stochastic mRNA synthesis in mammalian cells.

    Science.gov (United States)

    Raj, Arjun; Peskin, Charles S; Tranchina, Daniel; Vargas, Diana Y; Tyagi, Sanjay

    2006-10-01

    Individual cells in genetically homogeneous populations have been found to express different numbers of molecules of specific proteins. We investigated the origins of these variations in mammalian cells by counting individual molecules of mRNA produced from a reporter gene that was stably integrated into the cell's genome. We found that there are massive variations in the number of mRNA molecules present in each cell. These variations occur because mRNAs are synthesized in short but intense bursts of transcription beginning when the gene transitions from an inactive to an active state and ending when they transition back to the inactive state. We show that these transitions are intrinsically random and not due to global, extrinsic factors such as the levels of transcriptional activators. Moreover, the gene activation causes burst-like expression of all genes within a wider genomic locus. We further found that bursts are also exhibited in the synthesis of natural genes. The bursts of mRNA expression can be buffered at the protein level by slow protein degradation rates. A stochastic model of gene activation and inactivation was developed to explain the statistical properties of the bursts. The model showed that increasing the level of transcription factors increases the average size of the bursts rather than their frequency. These results demonstrate that gene expression in mammalian cells is subject to large, intrinsically random fluctuations and raise questions about how cells are able to function in the face of such noise. PMID:17048983

  16. Imaging gene expression in gene therapy

    Energy Technology Data Exchange (ETDEWEB)

    Wiebe, Leonard I. [Alberta Univ., Edmonton (Canada). Noujaim Institute for Pharmaceutical Oncology Research

    1997-12-31

    Full text. Gene therapy can be used to introduce new genes, or to supplement the function of indigenous genes. At the present time, however, there is non-invasive test to demonstrate efficacy of the gene transfer and expression processes. It has been postulated that scintigraphic imaging can offer unique information on both the site at which the transferred gene is expressed, and the degree of expression, both of which are critical issue for safety and clinical efficacy. Many current studies are based on `suicide gene therapy` of cancer. Cells modified to express these genes commit metabolic suicide in the presence of an enzyme encoded by the transferred gene and a specifically-convertible pro drug. Pro drug metabolism can lead to selective metabolic trapping, required for scintigraphy. Herpes simplex virus type-1 thymidine kinase (H S V-1 t k{sup +}) has been use for `suicide` in vivo tumor gene therapy. It has been proposed that radiolabelled nucleosides can be used as radiopharmaceuticals to detect H S V-1 t k{sup +} gene expression where the H S V-1 t k{sup +} gene serves a reporter or therapeutic function. Animal gene therapy models have been studied using purine-([{sup 18} F]F H P G; [{sup 18} F]-A C V), and pyrimidine- ([{sup 123}/{sup 131} I]I V R F U; [{sup 124}/{sup 131I}]) antiviral nucleosides. Principles of gene therapy and gene therapy imaging will be reviewed and experimental data for [{sup 123}/{sup 131I}]I V R F U imaging with the H S V-1 t k{sup +} reporter gene will be presented

  17. Imaging gene expression in gene therapy

    International Nuclear Information System (INIS)

    Full text. Gene therapy can be used to introduce new genes, or to supplement the function of indigenous genes. At the present time, however, there is non-invasive test to demonstrate efficacy of the gene transfer and expression processes. It has been postulated that scintigraphic imaging can offer unique information on both the site at which the transferred gene is expressed, and the degree of expression, both of which are critical issue for safety and clinical efficacy. Many current studies are based on 'suicide gene therapy' of cancer. Cells modified to express these genes commit metabolic suicide in the presence of an enzyme encoded by the transferred gene and a specifically-convertible pro drug. Pro drug metabolism can lead to selective metabolic trapping, required for scintigraphy. Herpes simplex virus type-1 thymidine kinase (H S V-1 t k+) has been use for 'suicide' in vivo tumor gene therapy. It has been proposed that radiolabelled nucleosides can be used as radiopharmaceuticals to detect H S V-1 t k+ gene expression where the H S V-1 t k+ gene serves a reporter or therapeutic function. Animal gene therapy models have been studied using purine-([18 F]F H P G; [18 F]-A C V), and pyrimidine- ([123/131 I]I V R F U; [124/131I]) antiviral nucleosides. Principles of gene therapy and gene therapy imaging will be reviewed and experimental data for [123/131I]I V R F U imaging with the H S V-1 t k+ reporter gene will be presented

  18. Hydrothermal organic synthesis experiments

    Science.gov (United States)

    Shock, Everett L.

    1992-01-01

    The serious scientific debate about spontaneous generation which raged for centuries reached a climax in the nineteenth century with the work of Spallanzani, Schwann, Tyndall, and Pasteur. These investigators demonstrated that spontaneous generation from dead organic matter does not occur. Although no aspects of these experiments addressed the issue of whether organic compounds could be synthesized abiotically, the impact of the experiments was great enough to cause many investigators to assume that life and its organic compounds were somehow fundamentally different than inorganic compounds. Meanwhile, other nineteenth-century investigators were showing that organic compounds could indeed be synthesized from inorganic compounds. In 1828 Friedrich Wohler synthesized urea in an attempt to form ammonium cyanate by heating a solution containing ammonia and cyanic acid. This experiment is generally recognized to be the first to bridge the artificial gap between organic and inorganic chemistry, but it also showed the usefulness of heat in organic synthesis. Not only does an increase in temperature enhance the rate of urea synthesis, but Walker and Hambly showed that equilibrium between urea and ammonium cyanate was attainable and reversible at 100 C. Wohler's synthesis of urea, and subsequent syntheses of organic compounds from inorganic compounds over the next several decades dealt serious blows to the 'vital force' concept which held that: (1) organic compounds owe their formation to the action of a special force in living organisms; and (2) forces which determine the behavior of inorganic compounds play no part in living systems. Nevertheless, such progress was overshadowed by Pasteur's refutation of spontaneous generation which nearly extinguished experimental investigations into the origins of life for several decades. Vitalism was dealt a deadly blow in the 1950's with Miller's famous spark-discharge experiments which were undertaken in the framework of the Oparin

  19. Identifying Gene Interaction Enrichment for Gene Expression Data

    OpenAIRE

    Jigang Zhang; Jian Li; Hong-Wen Deng

    2009-01-01

    Gene set analysis allows the inclusion of knowledge from established gene sets, such as gene pathways, and potentially improves the power of detecting differentially expressed genes. However, conventional methods of gene set analysis focus on gene marginal effects in a gene set, and ignore gene interactions which may contribute to complex human diseases. In this study, we propose a method of gene interaction enrichment analysis, which incorporates knowledge of predefined gene sets (e.g. gene ...

  20. Regulation of Plant Immunity through Modulation of Phytoalexin Synthesis

    Directory of Open Access Journals (Sweden)

    Olga V. Zernova

    2014-06-01

    Full Text Available Soybean hairy roots transformed with the resveratrol synthase and resveratrol oxymethyl transferase genes driven by constitutive Arabidopsis actin and CsVMV promoters were characterized. Transformed hairy roots accumulated glycoside conjugates of the stilbenic compound resveratrol and the related compound pterostilbene, which are normally not synthesized by soybean plants. Expression of the non-native stilbenic phytoalexin synthesis in soybean hairy roots increased their resistance to the soybean pathogen Rhizoctonia solani. The expression of the AhRS3 gene resulted in 20% to 50% decreased root necrosis compared to that of untransformed hairy roots. The expression of two genes, the AhRS3 and ROMT, required for pterostilbene synthesis in soybean, resulted in significantly lower root necrosis (ranging from 0% to 7% in transgenic roots than in untransformed hairy roots that had about 84% necrosis. Overexpression of the soybean prenyltransferase (dimethylallyltransferase G4DT gene in soybean hairy roots increased accumulation of the native phytoalexin glyceollin resulting in decreased root necrosis.