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Sample records for branching enzyme isoform

  1. Temperature Stress at Grain Filling Stage Mediates Expression of Three Isoform Genes Encoding Starch Branching Enzymes in Rice Endosperm

    Institute of Scientific and Technical Information of China (English)

    WEI Ke-su; CHENG Fang-min; ZHANG Qi-fang; LIU Kui-gang

    2009-01-01

    An early-maturity indica rice variety Zhefu 49, whose grain quality and starch structure are sensitive to environmental temperature, was subjected to different temperatures (32oC for high temperature and 22oC for optimum temperature) at the grain filling stage in plant growth chambers, and the different expressions of three isoform genes (SBEI, SBEIII and SBEIV) encoding starch branching enzyme (SBE) in the endosperms were studied by the real-time fluorescence quantitative PCR (FQ-PCR) method. Effects of high temperature on the SBE expression in developing rice endosperms were isoform-dependent. High temperature significantly down-regulated the expressions of SBEI and SBEIII, while up-regulated the expression of SBEIV. Compared with SBEIV and SBEIII, the expression of SBEI gene in Zhefu 49 rice endosperms was more sensitive to temperature variation at the grain filling stage. This study indicates that changes in weather/climate conditions especially temperature stress influence rice grain formation and its quality as evidenced by isoform expression.

  2. Starch Granule Re-Structuring by Starch Branching Enzyme and Glucan Water Dikinase Modulation Affects Caryopsis Physiology and Metabolism

    OpenAIRE

    Shaik, Shahnoor S.; Obata, Toshihiro; Hebelstrup, Kim H.; Schwahn, Kevin; Fernie, Alisdair R.; Mateiu, Ramona Valentina; Blennow, Andreas

    2016-01-01

    Starch is of fundamental importance for plant development and reproduction and its optimized molecular assembly is potentially necessary for correct starch metabolism. Re-structuring of starch granules in-planta can therefore potentially affect plant metabolism. Modulation of granule micro-structure was achieved by decreasing starch branching and increasing starch-bound phosphate content in the barley caryopsis starch by RNAi suppression of all three Starch Branching Enzyme (SBE) isoforms or ...

  3. AN ENZYME LINKED IMMUNOSORBENT ASSAY FOR THE HO-1 ISOFORM OF HEME OXYGENASE

    Science.gov (United States)

    AN ENZYME LINKED IMMUNOSORBENT ASSAY FOR THE HO-1 ISOFORM OF HEME OXYGENASE Heme oxygenase (HO) occurs in biological tissues as two major isoforms HO-1 and HO-2. HO-1 is inducible by many treatments, particularly oxidative stress-related conditions such as depletion of gl...

  4. Characterization and Expression Analysis of Starch Branching Enzymes in Sweet Potato

    Institute of Scientific and Technical Information of China (English)

    QIN Hua; ZHOU Shuang; ZHANG Yi-zheng

    2013-01-01

    Spatial and temporal expression patterns of Sbe1 and Sbe2 that encode starch branching enzyme (SBE) I and II, respectively, in sweet potato (Ipomoea batatas L.) were analyzed. Expression of both genes in Escherichia coli indicate that both genes encoded active SBE. Analysis with real-time quantitative polymerase chain reaction technique indicates that IbSbe1 mRNA was expressed at very low levels in leaves but was the predominant isoform in tuberous root while the reverse case was found for IbSbe2. The expression pattern of IbSbe1, closely resembles that of AGPase S, a gene coding for one of the subunits of ADP-glucose pyrophosphorylase, which is the key regulatory enzyme in the starch biosynthetic pathway. Western analysis detected at least two isoforms of SBE I in tuberous roots, those two isoforms showed adverse expression patterns with the development of the tuberous roots. Expression of the two IbSbe genes exhibited a diurnal rhythm during a 12-h cycle when fed a continuous solution of sucrose. Abscisic acid (ABA) was aother potent inducer of IbSbe expression, but bypassed the semidian oscillator.

  5. Starch Granule Re-Structuring by Starch Branching Enzyme and Glucan Water Dikinase Modulation Affects Caryopsis Physiology and Metabolism

    DEFF Research Database (Denmark)

    Shaik, Shahnoor S.; Obata, Toshihiro; Hebelstrup, Kim H;

    2016-01-01

    -structure was achieved by decreasing starch branching and increasing starch-bound phosphate content in the barley caryopsis starch by RNAi suppression of all three Starch Branching Enzyme (SBE) isoforms or overexpression of potato Glucan Water Dikinase (GWD). The resulting lines displayed Amylose-Only (AO) and Hyper...... high levels of some important stress-related metabolites and potentially protective metabolites, possibly to elude deleterious effects. Investigations on starch molecular structure revealed significant increase in starch phosphate and amylose content in HP and AO respectively with obvious differences...

  6. The activity and isoforms of NADP-malic enzyme in Nicotiana benthamiana plants under biotic stress

    Czech Academy of Sciences Publication Activity Database

    Doubnerová, V.; Jirásková, A.; Janošková, M.; Müller, Karel; Baťková, Petra; Synková, Helena; Čeřovská, Noemi; Ryšlavá, H.

    2007-01-01

    Roč. 26, č. 4 (2007), s. 281-289. ISSN 0231-5882 Institutional research plan: CEZ:AV0Z50380511 Keywords : NADP * malic enzyme isoforms * Nicotiana benthamiana Subject RIV: EF - Botanics Impact factor: 1.286, year: 2007 http://www.gpb.sav.sk/2007-4.htm

  7. Properties of the glucan branching enzyme of the hyperthermophilic bacterium Aquifex aeolicus

    OpenAIRE

    van der Maarel, MJEC; de Vos, A.; Sanders, P.; Dijkhuizen, L.

    2003-01-01

    Glucan branching enzymes are responsible for the synthesis of alpha(1-->6) glycosidic bonds in glycogen and amylopectin. The glucan branching enzyme of the hyperthermophile Aquifex aeolicus is the most thermoactive and thermostable glucan branching enzyme described. The gene encoding this glucan branching enzyme was overexpressed in E. coli and purified using gamma-cyclodextrin affinity chromatography. Subsequently, the enzyme was subjected to a biochemical characterization. The optimum tempe...

  8. Highly branched dextrin prepared from high-amylose maize starch using waxy rice branching enzyme (WRBE).

    Science.gov (United States)

    Tian, Yaoqi; Chen, Huangli; Zhang, Xiwen; Zhan, Jinling; Jin, Zhengyu; Wang, Jinpeng

    2016-07-15

    Branching enzyme (BE, EC 2.4.1.18) was isolated from the developing waxy rice endosperm and used to prepare a highly branched dextrin based on high-amylose maize starch (HAMS) as a substrate. The molecular mass of the starch initially degraded quickly from 2.5×10(7) to 4.1×10(5)Da, and then stabilized, with a minimal increase during the BE treatment. The resultant branched dextrin had a narrow size distribution, with a mean molecular weight of 5.1×10(5)Da and a polydispersity index (PI) of 1.567. The results of high-performance anion exchange chromatography indicated that the degree of polymerization (DP) of the branched chains ranged from 3 to 27; approximately 75.26% of these chains were short (DP<10). These findings suggest that the isolated BE can cleave long chains into oligosaccharides, subsequently transferring oligosaccharides into highly branched dextrins with a narrow size distribution and short side chains. PMID:26948647

  9. LOCALIZATION OF BRANCHING ENZYME IN POTATO-TUBER CELLS WITH THE USE OF IMMUNOELECTRON MICROSCOPY

    NARCIS (Netherlands)

    KRAM, AM; OOSTERGETEL, GT; VANBRUGGEN, EFJ

    1993-01-01

    Potato branching enzyme, a key enzyme in the biosynthesis of starch, was localized in amyloplasts in starch-storage cells of potato (Solanum tuberosum L) with the use of immunogold electron microscopy. Branching enzyme was found in the amyloplast stroma, concentrated at the interface of the stroma a

  10. Mutagenesis of Genes for Starch Debranching Enzyme Isoforms in Pea by Zinc-Finger Endonucleases

    International Nuclear Information System (INIS)

    Starch debranching enzymes in plants are divided into two groups based on their ability to hydrolyze different substrates. The first group, pullulanases, hydrolyze α-1,6-glucosidic linkages in substrates such as pullulan, amylopectin and glycogen. The second group of debranching enzymes, isoamylases, hydrolyze glycogen and amylopectin and are not active on pullulan. Three isoforms of isoamylase and a pullulanase have been isolated from a cDNA library of Pisum sativum. These isoamylases have been characterized following their heterologous expression in E. coli. Based on the DNA sequence that encodes these debranching enzymes, a specific mutagenesis targeting these enzymes will be attempted. The technique involves the homologous recombination of DNA mediated by zinc-finger endonucleases. Vectors will be constructed to include a fragment that will modify these genes. Using this technique, it is hoped that null mutants for each enzyme will be created and the exact role of these enzymes for the synthesis and degradation of starch in plants will be elucidated. (author)

  11. Revealing the functions of the transketolase enzyme isoforms in Rhodopseudomonas palustris using a systems biology approach.

    Directory of Open Access Journals (Sweden)

    Chia-Wei Hu

    Full Text Available BACKGROUND: Rhodopseudomonas palustris (R. palustris is a purple non-sulfur anoxygenic phototrophic bacterium that belongs to the class of proteobacteria. It is capable of absorbing atmospheric carbon dioxide and converting it to biomass via the process of photosynthesis and the Calvin-Benson-Bassham (CBB cycle. Transketolase is a key enzyme involved in the CBB cycle. Here, we reveal the functions of transketolase isoforms I and II in R. palustris using a systems biology approach. METHODOLOGY/PRINCIPAL FINDINGS: By measuring growth ability, we found that transketolase could enhance the autotrophic growth and biomass production of R. palustris. Microarray and real-time quantitative PCR revealed that transketolase isoforms I and II were involved in different carbon metabolic pathways. In addition, immunogold staining demonstrated that the two transketolase isoforms had different spatial localizations: transketolase I was primarily associated with the intracytoplasmic membrane (ICM but transketolase II was mostly distributed in the cytoplasm. Comparative proteomic analysis and network construction of transketolase over-expression and negative control (NC strains revealed that protein folding, transcriptional regulation, amino acid transport and CBB cycle-associated carbon metabolism were enriched in the transketolase I over-expressed strain. In contrast, ATP synthesis, carbohydrate transport, glycolysis-associated carbon metabolism and CBB cycle-associated carbon metabolism were enriched in the transketolase II over-expressed strain. Furthermore, ATP synthesis assays showed a significant increase in ATP synthesis in the transketolase II over-expressed strain. A PEPCK activity assay showed that PEPCK activity was higher in transketolase over-expressed strains than in the negative control strain. CONCLUSIONS/SIGNIFICANCE: Taken together, our results indicate that the two isoforms of transketolase in R. palustris could affect photoautotrophic growth

  12. Expression of Escherichia coli glycogen branching enzyme in an Arabidopsis mutant devoid of endogenous starch branching enzymes induces the synthesis of starch-like polyglucans.

    Science.gov (United States)

    Boyer, Laura; Roussel, Xavier; Courseaux, Adeline; Ndjindji, Ofilia M; Lancelon-Pin, Christine; Putaux, Jean-Luc; Tetlow, Ian J; Emes, Michael J; Pontoire, Bruno; D' Hulst, Christophe; Wattebled, Fabrice

    2016-07-01

    Starch synthesis requires several enzymatic activities including branching enzymes (BEs) responsible for the formation of α(1 → 6) linkages. Distribution and number of these linkages are further controlled by debranching enzymes that cleave some of them, rendering the polyglucan water-insoluble and semi-crystalline. Although the activity of BEs and debranching enzymes is mandatory to sustain normal starch synthesis, the relative importance of each in the establishment of the plant storage polyglucan (i.e. water insolubility, crystallinity and presence of amylose) is still debated. Here, we have substituted the activity of BEs in Arabidopsis with that of the Escherichia coli glycogen BE (GlgB). The latter is the BE counterpart in the metabolism of glycogen, a highly branched water-soluble and amorphous storage polyglucan. GlgB was expressed in the be2 be3 double mutant of Arabidopsis, which is devoid of BE activity and consequently free of starch. The synthesis of a water-insoluble, partly crystalline, amylose-containing starch-like polyglucan was restored in GlgB-expressing plants, suggesting that BEs' origin only has a limited impact on establishing essential characteristics of starch. Moreover, the balance between branching and debranching is crucial for the synthesis of starch, as an excess of branching activity results in the formation of highly branched, water-soluble, poorly crystalline polyglucan. PMID:26715025

  13. Repression of both isoforms of disproportionating enzyme leads to higher malto-oligosaccharide content and reduced growth in potato

    DEFF Research Database (Denmark)

    Mogensen, Henrik Lütken; Lloyd, James Richard; Glaring, Mikkel A.;

    2010-01-01

    Two glucanotransferases, disproportionating enzyme 1 (StDPE1) and disproportionating enzyme 2 (StDPE2), were repressed using RNA interference technology in potato, leading to plants repressed in either isoform individually, or both simultaneously. This is the first detailed report of their combined...... the fusion proteins, supporting a cytosolic role of the StDPE2 enzyme in leaf starch metabolism, as has been observed for Arabidopsis DPE2. It is concluded that StDPE1 and StDPE2 have individual essential roles in starch metabolism in potato and consequently repression of these disables regulation of...

  14. Studies on starch structure and the differential properties of starch branching enzymes

    OpenAIRE

    Andersson, Lena

    2001-01-01

    Starch is a staple food in human and animal diets, but also a raw material widely used for industrial purposes. By genetical modification of starch-synthesising enzymes in crop plants, starch yields could be increased and novel starches with particular qualities could be produced for industrial use. However, the process of starch biosynthesis and its regulation is still not completely understood. One of the major groups of enzymes in starch biosynthesis is the starch branching enzymes (SBEs),...

  15. Synergistic amylomaltase and branching enzyme catalysis to suppress cassava starch digestibility.

    Science.gov (United States)

    Sorndech, Waraporn; Meier, Sebastian; Jansson, Anita M; Sagnelli, Domenico; Hindsgaul, Ole; Tongta, Sunanta; Blennow, Andreas

    2015-11-01

    Starch provides our main dietary caloric intake and over-consumption of starch-containing foods results in escalating life-style disease including diabetes. By increasing the content of α-1,6 branch points in starch, digestibility by human amylolytic enzymes is expected to be retarded. Aiming at generating a soluble and slowly digestible starch by increasing the content and changing the relative positioning of the branch points in the starch molecules, we treated cassava starch with amylomaltase (AM) and branching enzyme (BE). We performed a detailed molecular analysis of the products including amylopectin chain length distribution, content of α-1,6 glucosidic linkages, absolute molecular weight distribution and digestibility. Step-by-step enzyme catalysis was the most efficient treatment, and it generated branch structures even more extreme than those of glycogen. All AM- and BE-treated samples showed increased resistance to degradation by porcine pancreatic α-amylase and glucoamylase as compared to cassava starch. The amylolytic products showed chain lengths and branching patterns similar to the products obtained from glycogen. Our data demonstrate that combinatorial enzyme catalysis provides a strategy to generate potential novel soluble α-glucan ingredients with low dietary digestibility assets. PMID:26256365

  16. Starch Granule Re-Structuring by Starch Branching Enzyme and Glucan Water Dikinase Modulation Affects Caryopsis Physiology and Metabolism.

    Science.gov (United States)

    Shaik, Shahnoor S; Obata, Toshihiro; Hebelstrup, Kim H; Schwahn, Kevin; Fernie, Alisdair R; Mateiu, Ramona V; Blennow, Andreas

    2016-01-01

    Starch is of fundamental importance for plant development and reproduction and its optimized molecular assembly is potentially necessary for correct starch metabolism. Re-structuring of starch granules in-planta can therefore potentially affect plant metabolism. Modulation of granule micro-structure was achieved by decreasing starch branching and increasing starch-bound phosphate content in the barley caryopsis starch by RNAi suppression of all three Starch Branching Enzyme (SBE) isoforms or overexpression of potato Glucan Water Dikinase (GWD). The resulting lines displayed Amylose-Only (AO) and Hyper-Phosphorylated (HP) starch chemotypes, respectively. We studied the influence of these alterations on primary metabolism, grain composition, starch structural features and starch granule morphology over caryopsis development at 10, 20 and 30 days after pollination (DAP) and at grain maturity. While HP showed relatively little effect, AO showed significant reduction in starch accumulation with re-direction to protein and β-glucan (BG) accumulation. Metabolite profiling indicated significantly higher sugar accumulation in AO, with re-partitioning of carbon to accumulate amino acids, and interestingly it also had high levels of some important stress-related metabolites and potentially protective metabolites, possibly to elude deleterious effects. Investigations on starch molecular structure revealed significant increase in starch phosphate and amylose content in HP and AO respectively with obvious differences in starch granule morphology at maturity. The results demonstrate that decreasing the storage starch branching resulted in metabolic adjustments and re-directions, tuning to evade deleterious effects on caryopsis physiology and plant performance while only little effect was evident by increasing starch-bound phosphate as a result of overexpressing GWD. PMID:26891365

  17. Early-branching Gut Fungi Possess A Large, And Comprehensive Array Of Biomass-Degrading Enzymes

    Energy Technology Data Exchange (ETDEWEB)

    Solomon, Kevin V.; Haitjema, Charles; Henske, John K.; Gilmore, Sean P.; Borges-Rivera, Diego; Lipzen, Anna; Brewer, Heather M.; Purvine, Samuel O.; Wright, Aaron T.; Theodorou, Michael K.; Grigoriev, Igor V.; Regev, Aviv; Thompson, Dawn; O' Malley, Michelle A.

    2016-03-11

    The fungal kingdom is the source of almost all industrial enzymes in use for lignocellulose bioprocessing. Its more primitive members, however, remain relatively unexploited. We developed a systems-level approach that integrates RNA-Seq, proteomics, phenotype and biochemical studies of relatively unexplored early-branching free-living fungi. Anaerobic gut fungi isolated from herbivores produce a large array of biomass-degrading enzymes that synergistically degrade crude, unpretreated plant biomass, and are competitive with optimized commercial preparations from Aspergillus and Trichoderma. Compared to these model platforms, gut fungal enzymes are unbiased in substrate preference due to a wealth of xylan-degrading enzymes. These enzymes are universally catabolite repressed, and are further regulated by a rich landscape of noncoding regulatory RNAs. Furthermore, we identified several promising sequence divergent enzyme candidates for lignocellulosic bioprocessing.

  18. The characterization of modified starch branching enzymes: toward the control of starch chain-length distributions.

    Directory of Open Access Journals (Sweden)

    Cheng Li

    Full Text Available Starch is a complex branched glucose polymer whose branch molecular weight distribution (the chain-length distribution, CLD influences nutritionally important properties such as digestion rate. Chain-stopping in starch biosynthesis is by starch branching enzyme (SBE. Site-directed mutagenesis was used to modify SBEIIa from Zea mays (mSBEIIa to produce mutants, each differing in a single conserved amino-acid residue. Products at different times from in vitro branching were debranched and the time evolution of the CLD measured by size-exclusion chromatography. The results confirm that Tyr352, Glu513, and Ser349 are important for mSBEIIa activity while Arg456 is important for determining the position at which the linear glucan is cut. The mutant mSBEIIa enzymes have different activities and suggest the length of the transferred chain can be varied by mutation. The work shows analysis of the molecular weight distribution can yield information regarding the enzyme branching sites useful for development of plants yielding starch with improved functionality.

  19. Modification of potato starch composition by introduction and expression of bacterial branching enzyme genes.

    OpenAIRE

    Kortstee, A.J.

    1997-01-01

    Starch consists of two major components; amylose and amylopectin. Amylose is synthesized by the enzyme Granule-Bound Starch Syntase (GBSS) and consists of essentially linear chains of α-1,4 linked glucose residues. Amylopectin is synthesized by the combined activity of the enzymes Soluble Starch Synthase (SSS) and Branching enzyme (BE) and consists of linear α-1,4 linked glucosidic chains with α-1,6 linked branchpoints. The amount and fine structure of each of the components determine the sta...

  20. Changes of Proline Content,Activity,and Active Isoforms of Antioxidative Enzymes in Two Alfalfa Cultivars Under Salt Stress

    Institute of Scientific and Technical Information of China (English)

    WANG Xiao-shan; HAN Jian-guo

    2009-01-01

    The plants of two alfalfa(Medicago sativa L.)cultivars differing in salt tolerance were subjected to three salt treatments,70,140,and 210 mM NaCl for 7 days.Root,shoot,and leaf growths were inhibited by increased salt treatments in both cultivars,and at 140 and 210 mM salt treatments,Zhongmu 1 had significantly higher root,shoot,and leaf dry weights per plant than Defi.The malondialdehyde(MDA)accumulation in Defi was considerably greater than in Zhongmu 1,indicating a higher degree of lipid peroxidation at 140 and 210 mM salt treatments.The changes in the activity and active isoforms of antioxidant enzymes such as superoxide dismutase(SOD,EC 1.15.1.1),catalase(CAT,EC 1.11.1.6),peroxidase(POD,EC 1.11.1.7),and ascorbate peroxidase(APOX,EC 1.11.1.11),accumulation of free proline,and rate of lipid peroxidation in leaves of two alfalfa cultivars were also investigated.After stress,the activity and active isoforms of antioxidative enzymes were altered and the extent of alteration varied between the cultivar Deft and Zhongmu 1.The proline accumulation in Defi was considerably greater than in Zhongmu 1 at 210 mM salt treatment.This indicated that proline accumulation may be the result,instead of the cause,of salt tolerance.

  1. Mutagenesis of genes for starch debranching enzyme isoforms in pea by means of zinc-finger endonucleases

    International Nuclear Information System (INIS)

    Starch debranching enzymes in plants are divided into two groups based on their ability to hydrolyse different substrates. The first group, pullulanases, hydrolyses α-1,6-glucosidic linkages in substrates such as pullulan, amylopectin and glycogen. The second group of debranching enzymes, isoamylases, hydrolyse glycogen and amylopectin and are not active on pullulan. Three isoforms of isoamylase and a pullulanase have been isolated from cDNA library of Pisum sativum. These isoamylases have been characterised based on the their heterologous expression in E coli. Based on the DNA sequence that encodes these debranching enzyme, a specific mutagenesis targeting at these DNA will be attempted. The method that will be employed are based on the techniques developed by Wright et al. (2005). This technique involves the homologous recombination of DNA that is mediated by zinc-finger endonucleases. Vectors will be constructed to include a fragment that will modify these genes. Microinjection technique will be used to insert these vectors into pollen which then will be fertilized. Using this technique, it is hoped that null mutant for each enzyme will be created and the exact role of these enzymes for the synthesis and degradation of starch in plants will be elucidate. (author)

  2. Enzyme-instructed self-assembly of taxol promotes axonal branching

    Science.gov (United States)

    Mei, Bin; Miao, Qingqing; Tang, Anming; Liang, Gaolin

    2015-09-01

    Axonal branching is important for vertebrate neuron signaling. Taxol has a biphasic effect on axonal branching (i.e., high concentration inhibits axonal growth but low concentration restores it). To the best of our knowledge, low concentration of taxol to promote axonal branching has not been reported yet. Herein, we rationally designed a taxol derivative Fmoc-Phe-Phe-Lys(taxol)-Tyr(H2PO4)-OH (1) which could be subjected to alkaline phosphatase (ALP)-catalyzed self-assembly to form taxol nanofibers. We found that, at 10 μM, 1 has a microtubule (MT) condensation effect similar to that of taxol on mammalian cells but with more chronic toxicity than taxol on the cells. At a low concentration of 10 nM, 1 not only promoted neurite elongation as taxol did but also promoted axonal branching which was not achieved by using taxol. We propose that self-assembly of 1 along the MTs prohibited their lateral contacts and thus promoted axonal branching. Our strategy of enzyme-instructed self-assembly (EISA) of a taxol derivative provides a new tool for scientists to study the morphology of neurons, as well as their behaviours.Axonal branching is important for vertebrate neuron signaling. Taxol has a biphasic effect on axonal branching (i.e., high concentration inhibits axonal growth but low concentration restores it). To the best of our knowledge, low concentration of taxol to promote axonal branching has not been reported yet. Herein, we rationally designed a taxol derivative Fmoc-Phe-Phe-Lys(taxol)-Tyr(H2PO4)-OH (1) which could be subjected to alkaline phosphatase (ALP)-catalyzed self-assembly to form taxol nanofibers. We found that, at 10 μM, 1 has a microtubule (MT) condensation effect similar to that of taxol on mammalian cells but with more chronic toxicity than taxol on the cells. At a low concentration of 10 nM, 1 not only promoted neurite elongation as taxol did but also promoted axonal branching which was not achieved by using taxol. We propose that self-assembly of 1

  3. Molecular Genetic Analysis of Glucan Branching Enzymes from Plants and Bacteria in Arabidopsis Reveals Marked Differences in Their Functions and Capacity to Mediate Starch Granule Formation.

    Science.gov (United States)

    Lu, Kuan-Jen; Streb, Sebastian; Meier, Florence; Pfister, Barbara; Zeeman, Samuel C

    2015-11-01

    The major component of starch is the branched glucan amylopectin, the branching pattern of which is one of the key factors determining its ability to form semicrystalline starch granules. Here, we investigated the functions of different branching enzyme (BE) types by expressing proteins from maize (Zea mays BE2a), potato (Solanum tuberosum BE1), and Escherichia coli (glycogen BE [EcGLGB]) in Arabidopsis (Arabidopsis thaliana) mutant plants that are deficient in their endogenous BEs and therefore, cannot make starch. The expression of each of these three BE types restored starch biosynthesis to differing degrees. Full complementation was achieved using the class II BE ZmBE2a, which is most similar to the two endogenous Arabidopsis isoforms. Expression of the class I BE from potato, StBE1, resulted in partial complementation and high amylose starch. Expression of the glycogen BE EcGLGB restored only minimal amounts of starch production, which had unusual chain length distribution, branch point distribution, and granule morphology. Nevertheless, each type of BE together with the starch synthases and debranching enyzmes were able to create crystallization-competent amylopectin polymers. These data add to the knowledge of how the properties of the BE influence the final composition of starch and fine structure of amylopectin. PMID:26358415

  4. Circadian oscillation of starch branching enzyme gene expression in the sorghum endosperm

    Energy Technology Data Exchange (ETDEWEB)

    Mutisya, J.; Sun, C.; Jansson, C.

    2009-08-31

    Expression of the three SBE genes, encoding starch branching enzymes, in the sorghum endosperm exhibited a diurnal rhythm during a 24-h cycle. Remarkably, the oscillation in SBE expression was maintained in cultured spikes after a 48-h dark treatment, also when fed a continuous solution of sucrose or abscisic acid. Our findings suggest that the rhythmicity in SBE expression in the endosperm is independent of cues from the photosynthetic source and that the oscillator resides within the endosperm itself.

  5. Enzyme-instructed self-assembly of taxol promotes axonal branching.

    Science.gov (United States)

    Mei, Bin; Miao, Qingqing; Tang, Anming; Liang, Gaolin

    2015-10-14

    Axonal branching is important for vertebrate neuron signaling. Taxol has a biphasic effect on axonal branching (i.e., high concentration inhibits axonal growth but low concentration restores it). To the best of our knowledge, low concentration of taxol to promote axonal branching has not been reported yet. Herein, we rationally designed a taxol derivative Fmoc-Phe-Phe-Lys(taxol)-Tyr(H2PO4)-OH (1) which could be subjected to alkaline phosphatase (ALP)-catalyzed self-assembly to form taxol nanofibers. We found that, at 10 μM, 1 has a microtubule (MT) condensation effect similar to that of taxol on mammalian cells but with more chronic toxicity than taxol on the cells. At a low concentration of 10 nM, 1 not only promoted neurite elongation as taxol did but also promoted axonal branching which was not achieved by using taxol. We propose that self-assembly of 1 along the MTs prohibited their lateral contacts and thus promoted axonal branching. Our strategy of enzyme-instructed self-assembly (EISA) of a taxol derivative provides a new tool for scientists to study the morphology of neurons, as well as their behaviours. PMID:26359218

  6. GABA-shunt enzymes activity in GH3 cells with reduced level of PMCA2 or PMCA3 isoform

    Energy Technology Data Exchange (ETDEWEB)

    Kowalski, Antoni, E-mail: antoni.kowalski@umed.lodz.pl [Department of Molecular Neurochemistry, Medical University of Lodz, 6/8 Mazowiecka Str., 92-215 Lodz (Poland); Zylinska, Ludmila, E-mail: ludmila.zylinska@umed.lodz.pl [Department of Molecular Neurochemistry, Medical University of Lodz, 6/8 Mazowiecka Str., 92-215 Lodz (Poland); Boczek, Tomasz, E-mail: tomasz.boczek@umed.lodz.pl [Department of Molecular Neurochemistry, Medical University of Lodz, 6/8 Mazowiecka Str., 92-215 Lodz (Poland); Rebas, Elzbieta, E-mail: elzbieta.rebas@umed.lodz.pl [Department of Molecular Neurochemistry, Medical University of Lodz, 6/8 Mazowiecka Str., 92-215 Lodz (Poland)

    2011-08-12

    Highlights: {yields} Suppression of PMCA2 or PMCA3 slows down proliferation of GH3 cells. {yields} PMCA2 suppression lowers the activity of GABA-shunt enzymes. {yields} PMCA3 suppression increases the expression of glutamate decarboxylase 65. {yields} PMCA2 and PMCA3 function appears to be linked to regulation of GABA metabolism. -- Abstract: GABA ({gamma}-aminobutyric acid) is important neurotransmitter and regulator of endocrine functions. Its metabolism involves three enzymes: glutamate decarboxylase (GAD65 and GAD67), GABA aminotransferase (GABA-T) and succinic semialdehyde dehydrogenase (SSADH). As many cellular processes GABA turnover can depend on calcium homeostasis, which is maintained by plasma membrane calcium ATPases (PMCAs). In excitable cells PMCA2 and PMCA3 isoforms are particularly important. In this study we focused on GABA-metabolizing enzymes expression and activity in rat anterior pituitary GH3 cells with suppressed expression of PMCA2 or PMCA3. We observed that PMCA3-reduced cells have increased GAD65 expression. Suppression of PMCA2 caused a decrease in total GAD and GABA-T activity. These results indicate that PMCA2 and PMCA3 presence may be an important regulatory factor in GABA metabolism. Results suggest that PMCA2 and PMCA3 function is rather related to regulation of GABA synthesis and degradation than supplying cells with metabolites, which can be potentially energetic source.

  7. GABA-shunt enzymes activity in GH3 cells with reduced level of PMCA2 or PMCA3 isoform

    International Nuclear Information System (INIS)

    Highlights: → Suppression of PMCA2 or PMCA3 slows down proliferation of GH3 cells. → PMCA2 suppression lowers the activity of GABA-shunt enzymes. → PMCA3 suppression increases the expression of glutamate decarboxylase 65. → PMCA2 and PMCA3 function appears to be linked to regulation of GABA metabolism. -- Abstract: GABA (γ-aminobutyric acid) is important neurotransmitter and regulator of endocrine functions. Its metabolism involves three enzymes: glutamate decarboxylase (GAD65 and GAD67), GABA aminotransferase (GABA-T) and succinic semialdehyde dehydrogenase (SSADH). As many cellular processes GABA turnover can depend on calcium homeostasis, which is maintained by plasma membrane calcium ATPases (PMCAs). In excitable cells PMCA2 and PMCA3 isoforms are particularly important. In this study we focused on GABA-metabolizing enzymes expression and activity in rat anterior pituitary GH3 cells with suppressed expression of PMCA2 or PMCA3. We observed that PMCA3-reduced cells have increased GAD65 expression. Suppression of PMCA2 caused a decrease in total GAD and GABA-T activity. These results indicate that PMCA2 and PMCA3 presence may be an important regulatory factor in GABA metabolism. Results suggest that PMCA2 and PMCA3 function is rather related to regulation of GABA synthesis and degradation than supplying cells with metabolites, which can be potentially energetic source.

  8. Phosphorylated alpha(1 leads to 4) glucans as substrate for potato starch-branching enzyme I

    International Nuclear Information System (INIS)

    The possible involvement of potato (Solanum tuberosum L.) starch-branching enzyme I (PSBE-I) in the in vivo synthesis of phosphorylated amylopectin was investigated in in vitro experiments with isolated PSBE-I using 33P-labeled phosphorylated and 3H end-labeled nonphosphorylated alpha(1 leads to 4) glucans as the substrates. From these radiolabeled substrates PSBE-I was shown to catalyze the formation of dual-labeled (3H/33P) phosphorylated branched polysaccharides with an average degree of polymerization of 80 to 85. The relatively high molecular mass indicated that the product was the result of multiple chain-transfer reactions. The presence of alpha(1 leads to 6) branch points was documented by isoamylase treatment and anion-exchange chromatography. Although the initial steps of the in vivo mechanism responsible for phosphorylation of potato starch remains elusive, the present study demonstrates that the enzyme machinery available in potato has the ability to incorporate phosphorylated alpha(1 leads to 4) glucans into neutral polysaccharides in an interchain catalytic reaction. Potato mini tubers synthesized phosphorylated starch from exogenously supplied 33PO4(3-) and [U-14C]Glc at rates 4 times higher than those previously obtained using tubers from fully grown potato plants. This system was more reproducible compared with soil-grown tubers and was therefore used for preparation of 33P-labeled phosphorylated alpha(1 leads to 4) glucan chains

  9. Improved yields of cyclic nigerosylnigerose from starch by pretreatment with a thermostable branching enzyme.

    Science.gov (United States)

    Aga, Hajime; Okamoto, Iwao; Taniguchi, Mituki; Kawashima, Akira; Abe, Hiroko; Chaen, Hiroto; Fukuda, Shigeharu

    2010-04-01

    Cyclic nigerosylnigerose (CNN) is produced enzymatically from starch by the combined action of 6-alpha-glucosyltransferase and 3-alpha-isomaltosyltransferase. In our previous study, alpha-1,6-branching chains found in the structure of amylopectin and glycogen were shown to be favorable for CNN formation by the two enzymes. Therefore, we examined whether the introduction of alpha-1,6-branch points into starch using the action of branching enzyme (BE) could improve the yield of CNN from starch. Thermostable BE from Geobacillus stearothermophilus TC-91 was prepared as a purified recombinant protein. Pretreatment of amylose with BE considerably increased the CNN yield from 5% to 38%. When BE acted on tapioca starch, the CNN yield was elevated from 47% to 60%. Conversely, BE treatment of waxy corn starch containing very little amylose resulted in a negligible increase in CNN yield. In addition, BE exerted a beneficial effect when starch with a lower degree of hydrolysis was used as a substrate. The present results indicate that the addition of alpha-1,6-glucosidic linkages to starch using BE is an effective strategy to improve the yield of CNN from starch. PMID:20226381

  10. Single-enzyme kinetics with branched pathways: exact theory and series expansion

    CERN Document Server

    Garai, Ashok

    2014-01-01

    The progress of the successive rounds of catalytic conversion of substrates into product(s) by a single enzyme is characterized by the distribution of turnover times. Establishing the most general form of dependence of this distribution on the substrate concentration [S] is one of the fundamental challenges in single molecule enzymology. The distribution of the times of dwell of a molecular motor at the successive positions on its track is an analogous quantity. We derive approximate series expansions for the [ATP]-dependence of the first two moments of the dwell time distributions of motors that catalyze hydrolysis of ATP to draw input energy. Comparison between our results for motors with branched pathways and the corresponding expressions reported earlier for linear enzymatic pathways provides deep insight into the effects of the branches. Such insight is likely to help in discovering the most general form of [S]-dependence of these fundamental distributions.

  11. Direct evidence for the inactivation of branched-chain oxo-acid dehydrogenase by enzyme phosphorylation

    International Nuclear Information System (INIS)

    The branched-chain 2-oxo-acid dehydrogenase (BCOAD) from mitochondria of several different rat tissues is inactivated by ATP and can be reactivated by incubation in Mg2+-containing buffers. Work carried out on the system from skeletal muscle mitochondria has shown that inactivation requires the cleavage of the γ-phosphate group of ATP and that modification is covalent. The non-metabolized ATP analog, p[NH]ppA, can block the inhibitory effect of ATP when added prior to ATP addition, but cannot reverse the inhibition of the inactivated dehydrogenase. These and other data raise the possibility that BCOAD may be regulated by enzyme phosphorylation. This hypothesis is supported by the finding that various procedures which separate the enzyme from its mitochondrial environment (e.g. detergent treatment, ammonium sulfate precipitation and freeze-thawing) do not alter the degree of inhibition induced by ATP in the mitochondrial preincubation. These experiments suggested the feasibility of labelling the enzyme with 32P and purifying it. (Auth.)

  12. Concerted suppression of all starch branching enzyme genes in barley produces amylose-only starch granules

    DEFF Research Database (Denmark)

    Carciofi, Massimiliano; Blennow, Per Gunnar Andreas; Jensen, Susanne Langgård;

    2012-01-01

    Background Starch is stored in higher plants as granules composed of semi-crystalline amylopectin and amorphous amylose. Starch granules provide energy for the plant during dark periods and for germination of seeds and tubers. Dietary starch is also a highly glycemic carbohydrate being degraded to...... glucose and rapidly absorbed in the small intestine. But a portion of dietary starch, termed "resistant starch" (RS) escapes digestion and reaches the large intestine, where it is fermented by colonic bacteria producing short chain fatty acids (SCFA) which are linked to several health benefits. The RS is...... preferentially derived from amylose, which can be increased by suppressing amylopectin synthesis by silencing of starch branching enzymes (SBEs). However all the previous works attempting the production of high RS crops resulted in only partly increased amylose-content and/or significant yield loss. Results In...

  13. Characterization of the reactions of starch branching enzymes from rice endosperm.

    Science.gov (United States)

    Nakamura, Yasunori; Utsumi, Yoshinori; Sawada, Takayuki; Aihara, Satomi; Utsumi, Chikako; Yoshida, Mayumi; Kitamura, Shinichi

    2010-05-01

    To our knowledge the present paper shows for the first time the kinetic parameters of all the three starch branching enzyme (BE) isozymes, BEI, BEIIa and BEIIb, from rice with both amylopectin and synthetic amylose as glucan substrate. The activities of these BE isozymes with a linear glucan amylose decreased with a decrease in the molar size of amylose, and no activities of BEIIa and BEIIb were found when the degree of polymerization (DP) of amylose was lower than at least 80, whereas BEI had an activity with amylose of a DP higher than approximately 50. Detailed analyses of debranched products from BE reactions revealed the distinct chain length preferences of the individual BE isozymes. BEIIb almost exclusively transferred chains of DP7 and DP6 while BEIIa formed a wide range of short chains of DP6 to around DP15 from outer chains of amylopectin and amylose. On the other hand, BEI formed a variety of short chains and intermediate chains of a DP branched glucan while BEIIa or BEIIb could only scarcely or could not attack inner chains, respectively. The comprehensive in vitro studies revealed different enzymatic characteristics of the three BE isozymes and give a new insight into the distinct roles of individual BE isozymes in amylopectin biosynthesis in the endosperm. Based on these results, the functional distinction and interaction of BE isozymes during amylopectin biosynthesis in cereal endosperm is discussed. PMID:20305271

  14. Heterologous expression and characterization of glycogen branching enzyme from Synechocystis sp. PCC6803.

    Science.gov (United States)

    Lee, Byung-Hoo; Yoo, Young-Hee; Ryu, Je-Hoon; Kim, Tae-Jip; Yoo, Sang-Ho

    2008-08-01

    A gene (sll0158) putatively encoding a glycogen branching enzyme (GBE, E.C. 2.4.1.18) was cloned from Synechocystis sp. PCC6803, and the recombinant protein expressed and characterized. The PCR-amplified putative GBE gene was ligated into a pET-21a plasmid vector harboring a T7 promoter, and the recombinant DNA transformed into a host cell, E. coli BL21(DE3). The IPTG-induced enzymes were then extracted and purified using Ni-NTA affinity chromatography. The putative GBE gene was found to be composed of 2,310 nucleotides and encoded 770 amino acids, corresponding to approx. 90.7 kDa, as confirmed by SDS-PAGE and MALDI-TOF-MS analyses. The optimal conditions for GBE activity were investigated by measuring the absorbance change in iodine affinity, and shown to be pH 8.0 and 30 degrees in a 50 mM glycine-NaOH buffer. The action pattern of the GBE on amylose, an alpha-(1,4)-linked linear glucan, was analyzed using high-performance anion-exchange chromatography (HPAEC) after isoamylolysis. As a result, the GBE displayed alpha-glucosyl transferring activity by cleaving the alpha-(1,4)-linkages and transferring the cleaved maltoglycosyl moiety to form new alpha-(1,6)- branch linkages. A time-course study of the GBE reaction was carried out with biosynthetic amylose (BSAM; Mp approximately = 8,000), and the changes in the branch-chain length distribution were evaluated. When increasing the reaction time up to 48 h, the weight- and number-average DP (DPw and DPn) decreased from 19.6 to 8.7 and from 17.6 to 7.8, respectively. The molecular size (Mp, peak Mw approximately = 2.45-2.75 x 10(5)) of the GBE-reacted product from BSAM reached the size of amylose (AM) in botanical starch, yet the product was highly soluble and stable in water, unlike AM molecules. Thus, GBE-generated products can provide new food and non-food applications, owing to their unique physical properties. PMID:18756098

  15. Retrogradation behavior of corn starch treated with 1,4-α-glucan branching enzyme.

    Science.gov (United States)

    Li, Wenwen; Li, Caiming; Gu, Zhengbiao; Qiu, Yijing; Cheng, Li; Hong, Yan; Li, Zhaofeng

    2016-07-15

    The retrogradation behavior of corn starch treated with 1,4-α-glucan branching enzyme (GBE) was investigated using rheometry, pulsed nuclear magnetic resonance (PNMR), differential scanning calorimetry (DSC) and Fourier transform infrared spectroscopy (FTIR). Dynamic time sweep analysis confirmed that the storage modulus (G') of corn starch stored at 4 °C decreased with increasing GBE treatment time. PNMR analysis demonstrated that the transverse relaxation times (T2) of corn starches treated with GBE were higher than that of control during the storage at 4 °C. DSC results demonstrated that the retrogradation enthalpy (ΔHr) of corn starch was reduced by 22.3% after GBE treatment for 10h. Avrami equation analysis showed that GBE treatment reduced the rate of starch retrogradation. FTIR analysis revealed that GBE treatment led to a decrease in hydrogen bonds within the starch. Overall, these results demonstrate that both short- and long-term retrogradation of corn starch were retarded by GBE treatment. PMID:26948619

  16. Enzyme- and pH-Sensitive Branched Polymer-Doxorubicin Conjugate-Based Nanoscale Drug Delivery System for Cancer Therapy.

    Science.gov (United States)

    Wei, Xiaoli; Luo, Qiang; Sun, Ling; Li, Xue; Zhu, Hongyan; Guan, Pujun; Wu, Min; Luo, Kui; Gong, Qiyong

    2016-05-11

    Owing to their dendritic architectural features, branched copolymers have been investigated as drug delivery systems. In this paper, an enzyme- and pH-sensitive branched poly[N-(2-hydroxypropyl)methacrylamide] (polyHPMA) copolymer-doxorubicin (DOX) conjugate possessing a molecular weight (MW) of 165 kDa was designed and prepared via a one-pot reaction and drug conjugation. This conjugate's potential as a smart, nanoscale drug delivery system (NDDS) is also investigated. The branched conjugate was capable of forming nanoparticles with a negative surface charge. The self-assembled nanoparticles were 102 nm in diameter as measured by dynamic light scattering (DLS) and 95 nm in diameter via scanning electron microscopy, respectively. The nanoparticles were degraded to low-MW products (23∼25 kDa) in the presence of papain or cathepsin B, and the degradation was monitored via DLS and size-exclusion chromatography. The nanoparticles demonstrated pH-sensitive drug release, as the DOX was attached to the branched copolymer via a hydrazone bond. In comparison to free DOX, the conjugate-based nanoparticles exhibited greater accumulation in breast tumors, resulting in enhanced antitumor therapeutic indexes. Furthermore, widespread dissemination of the conjugate among breast tumor cells was confirmed by immunohistochemical assay. Finally, no obvious systemic toxicities were observed in vivo in normal mice. Thus, the branched HPMA copolymer-DOX conjugate may be employed as a safe and efficient pH- and enzyme-responsive NDDS for cancer therapy. PMID:27102364

  17. Synthesis of Hyperbranched Glycoconjugates by the Combined Action of Potato Phosphorylase and Glycogen Branching Enzyme from Deinococcus geothermalis

    Directory of Open Access Journals (Sweden)

    Katja Loos

    2012-02-01

    Full Text Available Potato phosphorylase is able to synthesize linear polyglucans from maltoheptaose primers. By coupling maltoheptaose to butane diamine, tris(2-aminoethylamine and amine functionalized amine functionalized poly ethyleneglycol (PEG, new primer molecules became available. The resulting di-, tri- and macro-primers were incubated with potato phosphorylase and glycogen branching enzyme from Deinococcus geothermalis. Due to the action of both enzymes, hyperbranched polyglucan arms were grown from the maltoheptaose derivatives with a maximum degree of branching of 11%. The size of the synthesized hyperbranched polyglucans could be controlled by the ratio monomer over primer. About 60%–80% of the monomers were incorporated in the glycoconjugates. The resulting hyperbranched glycoconjugates were subjected to Dynamic Light Scattering (DLS measurements in order to determine the hydrodynamic radius and it became obvious that the structures formed agglomerates in the range of 14–32 nm.

  18. The different large subunit isoforms of Arabidopsis thaliana ADP-glucose pyrophosphorylase confer distinct kinetic and regulatory properties to the heterotetrameric enzyme.

    Science.gov (United States)

    Crevillén, Pedro; Ballicora, Miguel A; Mérida, Angel; Preiss, Jack; Romero, José M

    2003-08-01

    ADP-glucose pyrophosphorylase catalyzes the first and limiting step in starch biosynthesis and is allosterically regulated by the levels of 3-phosphoglycerate and phosphate in plants. ADP-glucose pyrophosphorylases from plants are heterotetramers composed of two types of subunits (small and large). In this study, the six Arabidopsis thaliana genes coding for ADP-glucose pyrophosphorylase isoforms (two small and four large subunits) have been cloned and expressed in an Escherichia coli mutant deficient in ADP-glucose pyrophosphorylase activity. The co-expression of the small subunit APS1 with the different Arabidopsis large subunits (APL1, APL2, APL3, and APL4) resulted in heterotetramers with different regulatory and kinetic properties. Heterotetramers composed of APS1 and APL1 showed the highest sensitivity to the allosteric effectors as well as the highest apparent affinity for the substrates (glucose-1-phosphate and ATP), whereas heterotetramers formed by APS1 and APL2 showed the lower response to allosteric effectors and the lower affinity for the substrates. No activity was detected for the second gene coding for a small subunit isoform (APS2) annotated in the Arabidopsis genome. This lack of activity is possibly due to the absence of essential amino acids involved in catalysis and/or in the binding of glucose-1-phosphate and 3-phosphoglycerate. Kinetic and regulatory properties of the different heterotetramers, together with sequence analysis has allowed us to make a distinction between sink and source enzymes, because the combination of different large subunits would provide a high plasticity to ADP-glucose pyrophosphorylase activity and regulation. This is the first experimental data concerning the role that all the ADP-glucose pyrophosphorylase isoforms play in a single plant species. This phenomenon could have an important role in vivo, because different large subunits would confer distinct regulatory properties to ADP-glucose pyrophosphorylase according

  19. Enzymes for N-Glycan Branching and Their Genetic and Nongenetic Regulation in Cancer

    Science.gov (United States)

    Kizuka, Yasuhiko; Taniguchi, Naoyuki

    2016-01-01

    N-glycan, a fundamental and versatile protein modification in mammals, plays critical roles in various physiological and pathological events including cancer progression. The formation of N-glycan branches catalyzed by specific N-acetylglucosaminyltransferases [GnT-III, GnT-IVs, GnT-V, GnT-IX (Vb)] and a fucosyltransferase, Fut8, provides functionally diverse N-glycosylated proteins. Aberrations of these branches are often found in cancer cells and are profoundly involved in cancer growth, invasion and metastasis. In this review, we focus on the GlcNAc and fucose branches of N-glycans and describe how their expression is dysregulated in cancer by genetic and nongenetic mechanisms including epigenetics and nucleotide sugar metabolisms. We also survey the roles that these N-glycans play in cancer progression and therapeutics. Finally, we discuss possible applications of our knowledge on basic glycobiology to the development of medicine and biomarkers for cancer therapy. PMID:27136596

  20. Enzymes for N-Glycan Branching and Their Genetic and Nongenetic Regulation in Cancer

    Directory of Open Access Journals (Sweden)

    Yasuhiko Kizuka

    2016-04-01

    Full Text Available N-glycan, a fundamental and versatile protein modification in mammals, plays critical roles in various physiological and pathological events including cancer progression. The formation of N-glycan branches catalyzed by specific N-acetylglucosaminyltransferases [GnT-III, GnT-IVs, GnT-V, GnT-IX (Vb] and a fucosyltransferase, Fut8, provides functionally diverse N-glycosylated proteins. Aberrations of these branches are often found in cancer cells and are profoundly involved in cancer growth, invasion and metastasis. In this review, we focus on the GlcNAc and fucose branches of N-glycans and describe how their expression is dysregulated in cancer by genetic and nongenetic mechanisms including epigenetics and nucleotide sugar metabolisms. We also survey the roles that these N-glycans play in cancer progression and therapeutics. Finally, we discuss possible applications of our knowledge on basic glycobiology to the development of medicine and biomarkers for cancer therapy.

  1. Enzymes for N-Glycan Branching and Their Genetic and Nongenetic Regulation in Cancer.

    Science.gov (United States)

    Kizuka, Yasuhiko; Taniguchi, Naoyuki

    2016-01-01

    N-glycan, a fundamental and versatile protein modification in mammals, plays critical roles in various physiological and pathological events including cancer progression. The formation of N-glycan branches catalyzed by specific N-acetylglucosaminyltransferases [GnT-III, GnT-IVs, GnT-V, GnT-IX (Vb)] and a fucosyltransferase, Fut8, provides functionally diverse N-glycosylated proteins. Aberrations of these branches are often found in cancer cells and are profoundly involved in cancer growth, invasion and metastasis. In this review, we focus on the GlcNAc and fucose branches of N-glycans and describe how their expression is dysregulated in cancer by genetic and nongenetic mechanisms including epigenetics and nucleotide sugar metabolisms. We also survey the roles that these N-glycans play in cancer progression and therapeutics. Finally, we discuss possible applications of our knowledge on basic glycobiology to the development of medicine and biomarkers for cancer therapy. PMID:27136596

  2. Cloning and inactivation of a branched-chain-amino-acid aminotransferase gene from Staphylococcus carnosus and characterization of the enzyme

    DEFF Research Database (Denmark)

    Madsen, Søren M; Beck, Hans Christian; Ravn, Peter; Vrang, Astrid; Hansen, Anne Maria; Israelsen, Hans

    2002-01-01

    Staphylococcus carnosus and Staphylococcus xylosus are widely used as aroma producers in the manufacture of dried fermented sausages. Catabolism of branched-chain amino acids (BCAAs) by these strains contributes to aroma formation by production of methyl-branched aldehydes and carboxy acids. The...... first step in the catabolism is most likely a transamination reaction catalyzed by BCAA aminotransferases (IlvE proteins). In this study, we cloned the ilvE gene from S. carnosus by using degenerate oligonucleotides and PCR. We found that the deduced amino acid sequence was 80% identical to that of the...... corresponding enzyme in Staphylococcus aureus and that the ilvE gene was constitutively expressed as a monocistronic transcript. To study the influence of ilvE on BCAA catabolism, we constructed an ilvE deletion mutant by gene replacement. The IlvE protein from S. carnosus was shown mainly to catalyze the...

  3. Modification of potato starch composition by introduction and expression of bacterial branching enzyme genes.

    NARCIS (Netherlands)

    Kortstee, A.J.

    1997-01-01

    Starch consists of two major components; amylose and amylopectin. Amylose is synthesized by the enzyme Granule-Bound Starch Syntase (GBSS) and consists of essentially linear chains of α-1,4 linked glucose residues. Amylopectin is synthesized by the combined activity of the enzymes Soluble Starch Syn

  4. Cloning and expression of the liver and muscle isoforms of ovine carnitine palmitoyltransferase 1: residues within the N-terminus of the muscle isoform influence the kinetic properties of the enzyme.

    Science.gov (United States)

    Price, Nigel T; Jackson, Vicky N; van der Leij, Feike R; Cameron, Jacqueline M; Travers, Maureen T; Bartelds, Beatrijs; Huijkman, Nicolette C; Zammit, Victor A

    2003-01-01

    The nucleotide sequence data reported will appear in DDBJ, EMBL, GenBank(R) and GSDB Nucleotide Sequence Databases; the sequences of ovine CPT1A and CPT1B cDNAs have the accession numbers Y18387 and AJ272435 respectively and the partial adipose tissue and liver CPT1A clones have the accession numbers Y18830 and Y18829 respectively. Fatty acid and ketone body metabolism differ considerably between monogastric and ruminant species. The regulation of the key enzymes involved may differ accordingly. Carnitine palmitoyltransferase 1 (CPT 1) is the key locus for the control of long-chain fatty acid beta-oxidation and liver ketogenesis. Previously we showed that CPT 1 kinetics in sheep and rat liver mitochondria differ. We cloned cDNAs for both isoforms [liver- (L-) and muscle- (M-)] of ovine CPT 1 in order to elucidate the structural features of these proteins and their genes ( CPT1A and CPT1B ). Their deduced amino acid sequences show a high degree of conservation compared with orthologues from other mammalian species, with the notable exception of the N-terminus of ovine M-CPT 1. These differences were also present in bovine M-CPT 1, whose N-terminal sequence we determined. In addition, the 5'-end of the sheep CPT1B cDNA suggested a different promoter architecture when compared with previously characterized CPT1B genes. Northern blotting revealed differences in tissue distribution for both CPT1A and CPT1B transcripts compared with other species. In particular, ovine CPT1B mRNA was less tissue restricted, and the predominant transcript in the pancreas was CPT1B. Expression in yeast allowed kinetic characterization of the two native enzymes, and of a chimaera in which the distinctive N-terminal segment of ovine M-CPT 1 was replaced with that from rat M-CPT 1. The ovine N-terminal segment influences the kinetics of the enzyme for both its substrates, such that the K (m) for palmitoyl-CoA is decreased and that for carnitine is increased for the chimaera, relative to the

  5. Metabolic gene clusters encoding the enzymes of two branches of the 3-oxoadipate pathway in the pathogenic yeast Candida albicans.

    Science.gov (United States)

    Gérecová, Gabriela; Neboháčová, Martina; Zeman, Igor; Pryszcz, Leszek P; Tomáška, Ľubomír; Gabaldón, Toni; Nosek, Jozef

    2015-05-01

    The pathogenic yeast Candida albicans utilizes hydroxyderivatives of benzene via the catechol and hydroxyhydroquinone branches of the 3-oxoadipate pathway. The genetic basis and evolutionary origin of this catabolic pathway in yeasts are unknown. In this study, we identified C. albicans genes encoding the enzymes involved in the degradation of hydroxybenzenes. We found that the genes coding for core components of the 3-oxoadipate pathway are arranged into two metabolic gene clusters. Our results demonstrate that C. albicans cells cultivated in media containing hydroxybenzene substrates highly induce the transcription of these genes as well as the corresponding enzymatic activities. We also found that C. albicans cells assimilating hydroxybenzenes cope with the oxidative stress by upregulation of cellular antioxidant systems such as alternative oxidase and catalase. Moreover, we investigated the evolution of the enzymes encoded by these clusters and found that most of them share a particularly sparse phylogenetic distribution among Saccharomycotina, which is likely to have been caused by extensive gene loss. We exploited this fact to find co-evolving proteins that are suitable candidates for the missing enzymes of the pathway. PMID:25743787

  6. Key enzymes of the protocatechuate branch of the β-ketoadipate pathway for aromatic degradation in Corynebacterium glutamicum

    Institute of Scientific and Technical Information of China (English)

    SHEN Xihui; LIU Shuangjiang

    2005-01-01

    Although the protocatechuate branch of the β-ketoadipate pathway in Gram bacteria has been well studied, this branch is less understood in Gram+ bacteria. In this study,Corynebacterium glutamicum was cultivated with protocatechuate, p-cresol, vanillate and 4-hydroxybenzoate as sole carbon and energy sources for growth. Enzymatic assays indicated that growing cells on these aromatic compounds exhibited protocatechuate 3,4-dioxygenase activities. Data-mining of the genome of this bacterium revealed that the genetic locus ncg12314-ncg12315 encoded a putative protocatechuate 3,4-dioxygenase. The genes,ncg12314 and ncg12315, were amplified by PCR technique and were cloned into plasmid (pET21aP34D). Recombinant Escherichia coli strain harboring this plasmid actively expressed protocatechuate 3,4-dioxygenase activity. Further, when this locus was disrupted in C. glutamicum, the ability to degrade and assimilate protocatechuate, p-cresol, vanillate or 4-hydroxybenzoate was lost and protocatechuate 3,4-dioxygenase activity was disappeared. The ability to grow with these aromatic compounds and protocatechuate 3,4-dioxygenase activity of C.glutamicum mutant could be restored by gene complementation. Thus, it is clear that the key enzyme for ring-cleavage, protocatechuate 3,4-dioxygenase, was encoded by ncg12314 and ncg12315. The additional genes involved in the protocatechuate branch of the β-ketoadipate pathway were identified by mining the genome data publically available in the GenBank. The functional identification of genes and their unique organization in C. glutamicum provided new insight into the genetic diversity of aromatic compound degradation.

  7. Characterization of a novel debranching enzyme from Nostoc punctiforme possessing a high specificity for long branched chains

    International Nuclear Information System (INIS)

    A novel debranching enzyme from Nostoc punctiforme PCC73102 (NPDE) exhibits hydrolysis activity toward both α-(1,6)- and α-(1,4)-glucosidic linkages. The action patterns of NPDE revealed that branched chains are released first, and the resulting maltooligosaccharides are then hydrolyzed. Analysis of the reaction with maltooligosaccharide substrates labeled with 14C-glucose at the reducing end shows that NPDE specifically liberates glucose from the reducing end. Kinetic analyses showed that the hydrolytic activity of NPDE is greatly affected by the length of the substrate. The catalytic efficiency of NPDE increased considerably upon using substrates that can occupy at least eight glycone subsites such as maltononaose and maltooctaosyl-α-(1,6)-β-cyclodextrin. These results imply that NPDE has a unique subsite structure consisting of -8 to +1 subsites. Given its unique subsite structure, side chains shorter than maltooctaose in amylopectin were resistant to hydrolysis by NPDE, and the population of longer side chains was reduced.

  8. Synthesis of branched polysaccharides with tunable degree of branching

    NARCIS (Netherlands)

    Ciric, Jelena; Loos, Katja

    2013-01-01

    An in vitro enzyme-catalyzed tandem reaction using the enzymes phosphorylase b from rabbit muscle and Deinococcus geothermalis glycogen branching enzyme (Dg GBE) to obtain branched polyglucans with tunable degree of branching (2% divided by 13%) is presented. The tunable degree of branching is obtai

  9. Enzyme

    Science.gov (United States)

    Enzymes are complex proteins that cause a specific chemical change in all parts of the body. For ... use them. Blood clotting is another example of enzymes at work. Enzymes are needed for all body ...

  10. Mouse white adipocytes and 3T3-L1 cells display an anomalous pattern of carnitine palmitoyltransferase (CPT) I isoform expression during differentiation. Inter-tissue and inter-species expression of CPT I and CPT II enzymes.

    Science.gov (United States)

    Brown, N F; Hill, J K; Esser, V; Kirkland, J L; Corkey, B E; Foster, D W; McGarry, J D

    1997-01-01

    The outer mitochondrial membrane enzyme carnitine palmitoyltransferase I (CPT I) represents the initial and regulated step in the beta-oxidation of fatty acids. It exists in at least two isoforms, denoted L (liver) and M (muscle) types, with very different kinetic properties and sensitivities to malonyl-CoA. Here we have examined the relative expression of the CPT I isoforms in two different models of adipocyte differentiation and in a number of rat tissues. Adipocytes from mice, hamsters and humans were also evaluated. Primary monolayer cultures of undifferentiated rat preadipocytes expressed solely L-CPT I, but significant levels of M-CPT I emerged after only 3 days of differentiation in vitro; in the mature cell M-CPT I predominated. In sharp contrast, the murine 3T3-L1 preadipocyte expressed essentially exclusively L-CPT I, both in the undifferentiated state and throughout the differentiation process in vitro. This was also true of the mature mouse white fat cell. Fully developed adipocytes from the hamster and human behaved similarly to those of the rat. Thus the mouse white fat cell differs fundamentally from those of the other species examined in terms of tis choice of a key regulatory enzyme in fatty acid metabolism. In contrast, brown adipose tissue from all three rodents displayed the same isoform profiles, each expressing overwhelmingly M-CPT I. Northern blot analysis of other rat tissues established L-CPT I as the dominant isoform not only in liver but also in kidney, lung, ovary, spleen, brain, intestine and pancreatic islets. In addition to its primacy in skeletal muscle, heart and fat, M-CPT I was also found to dominate the testis. The same inter-tissue isoform pattern (with the exception of white fat) was found in the mouse. Taken together, the data bring to light an intriguing divergence between white adipocytes of the mouse and other mammalian species. They also raise a cautionary note that should be considered in the choice of animal model used

  11. Purification, molecular cloning, and expression of 2-hydroxyphytanoyl- CoA lyase, a peroxisomal thiamine pyrophosphate-dependent enzyme that catalyzes the carbon-carbon bond cleavage during à-oxidation of 3- methyl-branched fatty acids

    CERN Document Server

    Foulon, V; Croes, K; Waelkens, E

    1999-01-01

    Purification, molecular cloning, and expression of 2-hydroxyphytanoyl- CoA lyase, a peroxisomal thiamine pyrophosphate-dependent enzyme that catalyzes the carbon-carbon bond cleavage during à-oxidation of 3- methyl-branched fatty acids

  12. Crystal Structure of Full-length Mycobacterium tuberculosis H37Rv Glycogen Branching Enzyme; Insights of N-Terminal [beta]-Sandwich in Sustrate Specifity and Enzymatic Activity

    Energy Technology Data Exchange (ETDEWEB)

    Pal, Kuntal; Kumar, Shiva; Sharma, Shikha; Garg, Saurabh Kumar; Alam, Mohammad Suhail; Xu, H. Eric; Agrawal, Pushpa; Swaminathan, Kunchithapadam (NU Sinapore); (Van Andel); (IMT-India)

    2010-07-13

    The open reading frame Rv1326c of Mycobacterium tuberculosis (Mtb) H37Rv encodes for an {alpha}-1,4-glucan branching enzyme (MtbGlgB, EC 2.4.1.18, Uniprot entry Q10625). This enzyme belongs to glycoside hydrolase (GH) family 13 and catalyzes the branching of a linear glucose chain during glycogenesis by cleaving a 1 {yields} 4 bond and making a new 1 {yields} 6 bond. Here, we show the crystal structure of full-length MtbGlgB (MtbGlgBWT) at 2.33-{angstrom} resolution. MtbGlgBWT contains four domains: N1 {beta}-sandwich, N2 {beta}-sandwich, a central ({beta}/{alpha}){sub 8} domain that houses the catalytic site, and a C-terminal {beta}-sandwich. We have assayed the amylase activity with amylose and starch as substrates and the glycogen branching activity using amylose as a substrate for MtbGlgBWT and the N1 domain-deleted (the first 108 residues deleted) Mtb{Delta}108GlgB protein. The N1 {beta}-sandwich, which is formed by the first 105 amino acids and superimposes well with the N2 {beta}-sandwich, is shown to have an influence in substrate binding in the amylase assay. Also, we have checked and shown that several GH13 family inhibitors are ineffective against MtbGlgBWT and Mtb{Delta}108GlgB. We propose a two-step reaction mechanism, for the amylase activity (1 {yields} 4 bond breakage) and isomerization (1 {yields} 6 bond formation), which occurs in the same catalytic pocket. The structural and functional properties of MtbGlgB and Mtb{Delta}108GlgB are compared with those of the N-terminal 112-amino acid-deleted Escherichia coli GlgB (EC{Delta}112GlgB).

  13. Repression of a novel isoform of disproportionating enzyme (stDPE2) in potato leads to inhibition of starch degradation in leaves but not tubers stored at low temperature

    DEFF Research Database (Denmark)

    Lloyd, J.R.; Blennow, A.; Burhenne, K.;

    2004-01-01

    A potato (Solanum tuberosum) cDNA encoding an isoform of disproportionating enzyme (stDPE2) was identified in a functional screen in Escherichia coli. The stDPE2 protein was demonstrated to be present in chloroplasts and to accumulate at times of active starch degradation in potato leaves and...... tubers. Transgenic potato plants were made in which its presence was almost completely eliminated. It could be demonstrated that starch degradation was repressed in leaves of the transgenic plants but that cold-induced sweetening was not affected in tubers stored at 4degreesC. No evidence could be found...... homogeneity from potato tuber extracts and could be demonstrated to transfer glucose from maltose to oyster glycogen....

  14. Glycogen synthase isoforms in Synechocystis sp. PCC6803: identification of different roles to produce glycogen by targeted mutagenesis.

    Directory of Open Access Journals (Sweden)

    Sang-Ho Yoo

    Full Text Available Synechocystis sp. PCC6803 belongs to cyanobacteria which carry out photosynthesis and has recently become of interest due to the evolutionary link between bacteria and plant species. Similar to other bacteria, the primary carbohydrate storage source of Synechocystis sp. PCC6803 is glycogen. While most bacteria are not known to have any isoforms of glycogen synthase, analysis of the genomic DNA sequence of Synechocystis sp. PCC6803 predicts that this strain encodes two isoforms of glycogen synthase (GS for synthesizing glycogen structure. To examine the functions of the putative GS genes, each gene (sll1393 or sll0945 was disrupted by double cross-over homologous recombination. Zymogram analysis of the two GS disruption mutants allowed the identification of a protein band corresponding to each GS isoform. Results showed that two GS isoforms (GSI and GSII are present in Synechocystis sp. PCC6803, and both are involved in glycogen biosynthesis with different elongation properties: GSI is processive and GSII is distributive. Total GS activities in the mutant strains were not affected and were compensated by the remaining isoform. Analysis of the branch-structure of glycogen revealed that the sll1393- mutant (GSI- produced glycogen containing more intermediate-length chains (DP 8-18 at the expense of shorter and longer chains compared with the wild-type strain. The sll0945- mutant (GSII- produced glycogen similar to the wild-type, with only a slightly higher proportion of short chains (DP 4-11. The current study suggests that GS isoforms in Synechocystis sp. PCC6803 have different elongation specificities in the biosynthesis of glycogen, combined with ADP-glucose pyrophosphorylase and glycogen branching enzyme.

  15. Roles of the N- and C-terminal domains of carnitine palmitoyltransferase I isoforms in malonyl-CoA sensitivity of the enzymes: insights from expression of chimaeric proteins and mutation of conserved histidine residues.

    Science.gov (United States)

    Swanson, S T; Foster, D W; McGarry, J D; Brown, N F

    1998-01-01

    The mitochondrial outer membrane enzyme carnitine palmitoyltransferase I (CPT I) plays a major role in the regulation of fatty acid entry into the mitochondrial matrix for beta-oxidation by virtue of its inhibition by malonyl-CoA. Two isoforms of CPT I, the liver type (L) and muscle type (M), have been identified, the latter being 100 times more sensitive to malonyl-CoA and having a much higher Km for the substrate carnitine. Here we have examined the roles of different regions of the CPT I molecules in their response to malonyl-CoA, etomoxir (an irreversible inhibitor) and carnitine. To this end, we analysed the properties of engineered rat CPT I constructs in which (a) the N-terminal domain of L-CPT I was deleted, (b) the N-terminal domains of L- and M-CPT I were switched, or (c) each of three conserved histidine residues located towards the N-terminus in L-CPT I was mutated. Several novel points emerged: (1) whereas the N-terminal domain is critical for a normal malonyl-CoA response, it does not itself account for the widely disparate sensitivities of the liver and muscle enzymes to the inhibitor; (2) His-5 and/or His-140 probably play a direct role in the malonyl-CoA response, but His-133 does not; (3) the truncated, chimaeric and point- mutant variants of the enzyme all bound the covalent, active-site- directed ligand, etomoxir; and (4) only the most radical alteration of L-CPT I, i.e. deletion of the N-terminal 82 residues, affected the response to carnitine. We conclude that the N-terminal domain of CPT I plays an essential, but permissive, role in the inhibition of the enzyme by malonyl-CoA. By contrast, the larger C-terminal region dictates the degree of sensitivity to malonyl-CoA, as well as the response to carnitine; it is also sufficient for etomoxir binding. Additionally, further weight is added to the notion that one or more histidine residues may be involved in the CPT I-malonyl-CoA interaction. PMID:9794789

  16. A putative gene sbe3-rs for resistant starch mutated from SBE3 for starch branching enzyme in rice (Oryza sativa L..

    Directory of Open Access Journals (Sweden)

    Ruifang Yang

    Full Text Available Foods high in resistant starch (RS are beneficial to prevent various diseases including diabetes, colon cancers, diarrhea and chronic renal or hepatic diseases. Elevated RS in rice is important for public health since rice is a staple food for half of the world population. A japonica mutant 'Jiangtangdao 1' (RS = 11.67% was crossed with an indica cultivar 'Miyang 23' (RS = 0.41%. The mutant sbe3-rs that explained 60.4% of RS variation was mapped between RM6611 and RM13366 on chromosome 2 (LOD = 36 using 178 F(2 plants genotyped with 106 genome-wide polymorphic SSR markers. Using 656 plants from four F(3:4 families, sbe3-rs was fine mapped to a 573.3 Kb region between InDel 2 and InDel 6 using one STS, five SSRs and seven InDel markers. SBE3 which codes for starch branching enzyme was identified as a candidate gene within the putative region. Nine pairs of primers covering 22 exons were designed to sequence genomic DNA of the wild type for SBE3 and the mutant for sbe3-rs comparatively. Sequence analysis identified a missense mutation site where Leu-599 of the wild was changed to Pro-599 of the mutant in the SBE3 coding region. Because the point mutation resulted in the loss of a restriction enzyme site, sbe3-rs was not digested by a CAPS marker for SpeI site while SBE3 was. Co-segregation of the digestion pattern with RS content among 178 F(2 plants further supported sbe3-rs responsible for RS in rice. As a result, the CAPS marker could be used in marker-assisted breeding to develop rice cultivars with elevated RS which is otherwise difficult to accurately assess in crops. Transgenic technology should be employed for a definitive conclusion of the sbe3-rs.

  17. Activities of the enzymes of the Ehrlich pathway and formation of branched-chain alcohols in Saccharomyces cerevisiae and Candida utilis grown in continuous culture on valine or ammonium as sole nitrogen source.

    Science.gov (United States)

    Derrick, S; Large, P J

    1993-11-01

    Valine aminotransferase, a key enzyme in both biosynthesis and breakdown of branched-chain amino acids, showed consistently higher activity in Candida utilis grown in continuous culture than in Saccharomyces cerevisiae, while pyruvate decarboxylase and alcohol dehydrogenase, the other two enzymes of the Ehrlich pathway of branched-chain alcohol formation, were lower in activity. By spheroplast lysis, it was shown that valine aminotransferase followed the distribution of pyruvate decarboxylase in being located in the cytosol. Replacement of ammonium as nitrogen source by valine during conditions of carbon or nitrogen limitation caused increased specific activities of these three enzymes in S. cerevisiae, but (with one exception) decreased those of C. utilis. Of the metabolites accumulating in the culture medium, little or no ethanol or branched-chain alcohols were present during carbon-limited growth of either organism, but the change to nitrogen limitation resulted in increases in concentration of 20- to 100-fold in pyruvate, acetate and non-pyruvate keto acids as well as the accumulation of branched-chain alcohols in both organisms, and of ethanol, ethyl acetate and glycerol in S. cerevisiae. When valine was the limiting nitrogen source, there was an increase in non-pyruvate keto acids and a 10- to 16-fold increase in 2-methylpropanol. Total branched-chain alcohols formed under nitrogen limitation were 2-fold higher in S. cerevisiae than in C. utilis, irrespective of nitrogen source. Accumulation of branched-chain alcohols, ethanol, acetate and glycerol was also observed during carbon-limited growth of S. cerevisiae with valine as nitrogen source at dilution rates above the critical rate for transition to respirofermentative growth. Less than 70% of the valine carbon metabolized during growth of S. cerevisiae and only 15% of that used during growth of C. utilis was recovered in identified metabolic products. Even allowing for losses by volatilization during

  18. Starch phosphorylation in potato tubers is influenced by allelic variation in the genes encoding glucan water dikinase, starch branching enzymes I and II, and starch synthase III

    Directory of Open Access Journals (Sweden)

    Margaret Ann Carpenter

    2015-03-01

    Full Text Available Starch phosphorylation is an important aspect of plant metabolism due to its role in starch degradation. Moreover, the degree of phosphorylation of starch determines its physicochemical properties and is therefore relevant for industrial uses of starch. Currently, starch is chemically phosphorylated to increase viscosity and paste stability. Potato cultivars with elevated starch phosphorylation would make this process unnecessary, thereby bestowing economic and environmental benefits. Starch phosphorylation is a complex trait which has been previously shown by antisense gene repression to be influenced by a number of genes including those involved in starch synthesis and degradation. We have used an association mapping approach to discover genetic markers associated with the degree of starch phosphorylation. A diverse collection of 193 potato lines was grown in replicated field trials, and the levels of starch phosphorylation at the C6 and C3 positions of the glucosyl residues were determined by mass spectrometry of hydrolyzed starch from tubers. In addition, the potato lines were genotyped by amplicon sequencing and microsatellite analysis, focusing on candidate genes known to be involved in starch synthesis. As potato is an autotetraploid, genotyping included determination of allele dosage. Significant associations (p<0.001 were found with SNPs in the glucan water dikinase (GWD, starch branching enzyme I (SBEI and the starch synthase III (SSIII genes, and with a SSR allele in the SBEII gene. SNPs in the GWD gene were associated with C6 phosphorylation, whereas polymorphisms in the SBEI and SBEII genes were associated with both C6 and C3 phosphorylation and the SNP in the SSIII gene was associated with C3 phosphorylation. These allelic variants have potential as genetic markers for starch phosphorylation in potato.

  19. Structure Function Relationships of ADP-Glucose Pyrophosphorylase and Branching Enzyme: Manipulation of Their Genes for Alteration of Starch Quanlity and Quantity

    Energy Technology Data Exchange (ETDEWEB)

    Jack Preiss

    2006-02-16

    Conversion of the Potato tuber ADP-glucose Pyrophopshorylase Regulatory Subunit into a Catalytic Subunit. ADP-glucose synthesis, a rate-limiting reaction in starch synthesis, is catalyzed by ADP-glucose pyrophosphorylase (ADPGlc PPase). The enzyme in plants is allosterically activated by 3-phosphoglycerate (3PGA) and inhibited by inorganic phosphate (Pi) and is composed of two subunits as a heterotetramer, a2b2. Subunit a is the catalytic subunit and subunit b is designated as the regulatory subunit.The b subunit increases the affinty of the activator for the catalytic subunit. Recent results have shown that the subunits are derived from the same ancestor subunit as the regulatory subunit can be converted to a catalytically subunit via mutation of just two amino acids. Lys44 and Thr54 in the large subunit from potato tuber were converted to the homologous catalytic subunit residues, Arg33 and Lys43. The activity of the large subunit mutants cannot be readily tested with a co-expressed wild-type small (catalytic) subunit because of the intrinsic activity of the latter. We co-expressed the regulatory-subunit mutants with SmallD145N, an inactive S subunit in which the catalytic Asp145 was mutated. The activity of the small (catalytic) subunit was reduced more than three orders of magnitude. Coexpression of the L subunit double mutant LargeK44R/T54K with SmallD145N generated an enzyme with considerable activity, 10% and 18% of the wildtype enzyme, in the ADP-glucose synthetic and pyrophosphorolytic direction, respectively. Replacement of those two residues in the small subunit by the homologous amino acids in the L subunits (mutations R33K and K43T) decreased the activity one and two orders of magnitude. The wild-type enzyme and SmallD145NLargeK44R/T54K had very similar kinetic properties indicating that the substrate site has been conserved. The fact that only two mutations in the L subunit restored enzyme activity is very strong evidence that the large subunit is

  20. Differential role of human choline kinase α and β enzymes in lipid metabolism: Implications in cancer onset and treatment

    OpenAIRE

    Gallego Ortega, David; Ramírez de Molina, Ana; Ramos, Maria Angeles; Valdés Mora, Fátima; Barderas, Maria Gonzalez; Sarmentero Estrada, Jacinto; Lacal, Juan Carlos

    2009-01-01

    Background The Kennedy pathway generates phosphocoline and phosphoethanolamine through its two branches. Choline Kinase (ChoK) is the first enzyme of the Kennedy branch of synthesis of 1phosphocholine, the major component of the plasma membrane. ChoK family of proteins is composed by ChoKα and ChoKβ isoforms, the first one with two different variants of splicing. Recently ChoKα has been implicated in the carcinogenic process, since it is over-expressed in a variety of human cancers. Howev...

  1. DNA signals at isoform promoters.

    Science.gov (United States)

    Dai, Zhiming; Xiong, Yuanyan; Dai, Xianhua

    2016-01-01

    Transcriptional heterogeneity is extensive in the genome, and most genes express variable transcript isoforms. However, whether variable transcript isoforms of one gene are regulated by common promoter elements remain to be elucidated. Here, we investigated whether isoform promoters of one gene have separated DNA signals for transcription and translation initiation. We found that TATA box and nucleosome-disfavored DNA sequences are prevalent in distinct transcript isoform promoters of one gene. These DNA signals are conserved among species. Transcript isoform has a RNA-determined unstructured region around its start site. We found that these DNA/RNA features facilitate isoform transcription and translation. These results suggest a DNA-encoded mechanism by which transcript isoform is generated. PMID:27353836

  2. Development of isoform-specific sensors of polypeptide GalNAc-transferase activity

    DEFF Research Database (Denmark)

    Song, Lina; Bachert, Collin; Schjoldager, Katrine T; Clausen, Henrik; Linstedt, Adam D

    2014-01-01

    Humans express up to 20 isoforms of GalNAc-transferase (herein T1-T20) that localize to the Golgi apparatus and initiate O-glycosylation. Regulation of this enzyme family affects a vast array of proteins transiting the secretory pathway and diseases arise upon misregulation of specific isoforms...

  3. Structure, function and regulation of the enzymes in the starch biosynthetic pathway.

    Energy Technology Data Exchange (ETDEWEB)

    Geiger, Jim

    2013-11-30

    Starch is the major reserve polysaccharide in nature and accounts for the majority of the caloric intact of humans. It is also gaining importance as a renewable and biodegradable industrial material. There is burgeoning interest in increasing the amount and altering the properties of the plant starches by plant genetic modification. A rational approach to this effort will require a detailed, atomic-level understanding of the enzymatic processes that produce the starch granule. The starch granule is a complex particle made up of alternating layers of crystalline and amorphous lamellae. It consists of two types of polymer, amylose, a polymer of relatively long chains of α-1,4-linked glucans that contain virtually no branches, and amylopectin, which is highly branched and contains much shorter chains. This complex structure is synthesized by the coordinate activities of the starch synthases (SS), which elongate the polysaccharide chain by addition of glucose units via α-1,4 linkages using ADP- glucose as a donor, and branching enzymes (BE), which branch the polysaccharide chain by cleavage of α−1,4 linkages and subsequent re-attachment via α−1,6 linkages. Several isoforms of both starch synthase (SS) and branching enzyme (BE) are found in plants, including SSI, SSII, SSIII and granule- bound SS (GBSS), and SBEI, SBEIIa and SBEIIb. These isoforms have different activities and substrate and product specificities and play different roles in creating the granule and determining the properties of the resulting starch. The overarching goal of this proposal is to begin to understand the regulation and specificities of these enzymes at the atomic level. High-resolution X-ray structures of these enzymes bound to substrates and products will be determined to visualize the molecular interactions responsible for the properties of the enzymes. Hypotheses regarding these issues will then be tested using mutagenesis and enzyme assays. To date, we have determined the

  4. Branching out

    CERN Document Server

    Biggins, J D

    2010-01-01

    Results on the behaviour of the rightmost particle in the $n$th generation in the branching random walk are reviewed and the phenomenon of anomalous spreading speeds, noticed recently in related deterministic models, is considered. The relationship between such results and certain coupled reaction-diffusion equations is indicated.

  5. Differential roles of PML isoforms

    Directory of Open Access Journals (Sweden)

    MouniraKChelbi-Alix

    2013-05-01

    Full Text Available The tumor suppressor promyelocytic leukemia protein (PML is fused to the retinoic acid receptor alpha in patients suffering from acute promyelocytic leukemia (APL. Treatment of APL patients with arsenic trioxide (As2O3 reverses the disease phenotype by a process involving the degradation of the fusion protein via its PML moiety. Several PML isoforms are generated from a single PML gene by alternative splicing. They share the same N-terminal region containing the RBCC/TRIM motif but differ in their C-terminal sequences. Recent studies of all the PML isoforms reveal the specific functions of each. Here, we review the nomenclature and structural organization of the PML isoforms in order to clarify the various designations and classifications found in different databases. The functions of the PML isoforms and their differential roles in antiviral defense also are reviewed. Finally, the key players involved in the degradation of the PML isoforms in response to As2O3 or other inducers are discussed.

  6. Quarternary structure and enzymological properties of the different hormone-sensitive lipase (HSL) isoforms

    DEFF Research Database (Denmark)

    Krintel, Christian; Klint, Cecilia; Lindvall, Håkan; Mörgelin, Matthias; Holm, Cecilia

    2010-01-01

    Hormone-sensitive lipase (HSL) is a key enzyme in the mobilization of energy in the form of fatty acids from intracellular stores of neutral lipids. The enzyme has been shown to exist in different isoforms with different molecular masses (84 kDa, 89 kDa and 117 kDa) expressed in a tissue...

  7. Quarternary structure and enzymological properties of the different hormone-sensitive lipase (HSL isoforms.

    Directory of Open Access Journals (Sweden)

    Christian Krintel

    Full Text Available BACKGROUND: Hormone-sensitive lipase (HSL is a key enzyme in the mobilization of energy in the form of fatty acids from intracellular stores of neutral lipids. The enzyme has been shown to exist in different isoforms with different molecular masses (84 kDa, 89 kDa and 117 kDa expressed in a tissue-dependent manner, where the predominant 84 kDa form in adipocytes is the most extensively studied. METHODOLOGY/PRINCIPAL FINDINGS: In this study we employed negative stain electron microscopy (EM to analyze the quarternary structure of the different HSL isoforms. The results show that all three isoforms adopt a head-to-head homodimeric organization, where each monomer contains two structural domains. We also used enzymatic assays to show that despite the variation in the size of the N-terminal domain all three isoforms exhibit similar enzymological properties with regard to psychrotolerance and protein kinase A (PKA-mediated phosphorylation and activation. CONCLUSIONS/SIGNIFICANCE: We present the first data on the quaternary structure and domain organization of the three HSL isoforms. We conclude that despite large differences in the size of the N-terminal, non-catalytic domain all three HSL isoforms exhibit the same three-dimensional architecture. Furthermore, the three HSL isoforms are very similar with regard to two unique enzymological characteristics of HSL, i.e., cold adaptation and PKA-mediated activation.

  8. Opposing effects of fructokinase C and A isoforms on fructose-induced metabolic syndrome in mice

    OpenAIRE

    Ishimoto, Takuji; Lanaspa, Miguel A.; MyPhuong T Le; Garcia, Gabriela E.; Diggle, Christine P; MacLean, Paul S.; Jackman, Matthew R.; Asipu, Aruna; Roncal-Jimenez, Carlos A.; Kosugi, Tomoki; Rivard, Christopher J.; Maruyama, Shoichi; Rodriguez-Iturbe, Bernardo; Sánchez-Lozada, Laura G.; Bonthron, David T.

    2012-01-01

    Fructose intake from added sugars correlates with the epidemic rise in obesity, metabolic syndrome, and nonalcoholic fatty liver disease. Fructose intake also causes features of metabolic syndrome in laboratory animals and humans. The first enzyme in fructose metabolism is fructokinase, which exists as two isoforms, A and C. Here we show that fructose-induced metabolic syndrome is prevented in mice lacking both isoforms but is exacerbated in mice lacking fructokinase A. Fructokinase C is expr...

  9. A novel FADS1 isoform potentiates FADS2-mediated production of eicosanoid precursor fatty acids

    OpenAIRE

    Park, Woo Jung; Kothapalli, Kumar S. D.; Reardon, Holly T; Lawrence, Peter; Qian, Shu-Bing; Brenna, J. Thomas

    2012-01-01

    The fatty acid desaturase (FADS) genes code for the rate-limiting enzymes required for the biosynthesis of long-chain polyunsaturated fatty acids (LCPUFA). Here we report discovery and function of a novel FADS1 splice variant. FADS1 alternative transcript 1 (FADS1AT1) enhances desaturation of FADS2, leading to increased production of eicosanoid precursors, the first case of an isoform modulating the enzymatic activity encoded by another gene. Multiple protein isoforms were detected in primate...

  10. Expression of phosphoinositide-specific phospholipase C isoforms in native endothelial cells.

    Directory of Open Access Journals (Sweden)

    Delphine M Béziau

    Full Text Available Phospholipase C (PLC comprises a superfamily of enzymes that play a key role in a wide array of intracellular signalling pathways, including protein kinase C and intracellular calcium. Thirteen different mammalian PLC isoforms have been identified and classified into 6 families (PLC-β, γ, δ, ε, ζ and η based on their biochemical properties. Although the expression of PLC isoforms is tissue-specific, concomitant expression of different PLC has been reported, suggesting that PLC family is involved in multiple cellular functions. Despite their critical role, the PLC isoforms expressed in native endothelial cells (ECs remains undetermined. A conventional PCR approach was initially used to elucidate the mRNA expression pattern of PLC isoforms in 3 distinct murine vascular beds: mesenteric (MA, pulmonary (PA and middle cerebral arteries (MCA. mRNA encoding for most PLC isoforms was detected in MA, MCA and PA with the exception of η2 and β2 (only expressed in PA, δ4 (only expressed in MCA, η1 (expressed in all but MA and ζ (not detected in any vascular beds tested. The endothelial-specific PLC expression was then sought in freshly isolated ECs. Interestingly, the PLC expression profile appears to differ across the investigated arterial beds. While mRNA for 8 of the 13 PLC isoforms was detected in ECs from MA, two additional PLC isoforms were detected in ECs from PA and MCA. Co-expression of multiple PLC isoforms in ECs suggests an elaborate network of signalling pathways: PLC isoforms may contribute to the complexity or diversity of signalling by their selective localization in cellular microdomains. However in situ immunofluorescence revealed a homogeneous distribution for all PLC isoforms probed (β3, γ2 and δ1 in intact endothelium. Although PLC isoforms play a crucial role in endothelial signal transduction, subcellular localization alone does not appear to be sufficient to determine the role of PLC in the signalling microdomains found

  11. Molecular modeling study on tunnel behavior in different histone deacetylase isoforms.

    Directory of Open Access Journals (Sweden)

    Sundarapandian Thangapandian

    Full Text Available Histone deacetylases (HDACs have emerged as effective therapeutic targets in the treatment of various diseases including cancers as these enzymes directly involved in the epigenetic regulation of genes. However the development of isoform-selective HDAC inhibitors has been a challenge till date since all HDAC enzymes possess conserved tunnel-like active site. In this study, using molecular dynamics simulation we have analyzed the behavior of tunnels present in HDAC8, 10, and 11 enzymes of class I, II, and IV, respectively. We have identified the equivalent tunnel forming amino acids in these three isoforms and found that they are very much conserved with subtle differences to be utilized in selective inhibitor development. One amino acid, methionine of HDAC8, among six tunnel forming residues is different in isoforms of other classes (glutamic acid (E in HDAC10 and leucine (L in HDAC 11 based on which mutations were introduced in HDAC11, the less studied HDAC isoform, to observe the effects of this change. The HDAC8-like (L268M mutation in the tunnel forming residues has almost maintained the deep and narrow tunnel as present in HDAC8 whereas HDAC10-like (L268E mutation has changed the tunnel wider and shallow as observed in HDAC10. These results explained the importance of the single change in the tunnel formation in different isoforms. The observations from this study can be utilized in the development of isoform-selective HDAC inhibitors.

  12. Inference of Isoforms from Short Sequence Reads

    Science.gov (United States)

    Feng, Jianxing; Li, Wei; Jiang, Tao

    Due to alternative splicing events in eukaryotic species, the identification of mRNA isoforms (or splicing variants) is a difficult problem. Traditional experimental methods for this purpose are time consuming and cost ineffective. The emerging RNA-Seq technology provides a possible effective method to address this problem. Although the advantages of RNA-Seq over traditional methods in transcriptome analysis have been confirmed by many studies, the inference of isoforms from millions of short sequence reads (e.g., Illumina/Solexa reads) has remained computationally challenging. In this work, we propose a method to calculate the expression levels of isoforms and infer isoforms from short RNA-Seq reads using exon-intron boundary, transcription start site (TSS) and poly-A site (PAS) information. We first formulate the relationship among exons, isoforms, and single-end reads as a convex quadratic program, and then use an efficient algorithm (called IsoInfer) to search for isoforms. IsoInfer can calculate the expression levels of isoforms accurately if all the isoforms are known and infer novel isoforms from scratch. Our experimental tests on known mouse isoforms with both simulated expression levels and reads demonstrate that IsoInfer is able to calculate the expression levels of isoforms with an accuracy comparable to the state-of-the-art statistical method and a 60 times faster speed. Moreover, our tests on both simulated and real reads show that it achieves a good precision and sensitivity in inferring isoforms when given accurate exon-intron boundary, TSS and PAS information, especially for isoforms whose expression levels are significantly high.

  13. Crystal structures of a halophilic archaeal malate synthase from Haloferax volcanii and comparisons with isoforms A and G

    Directory of Open Access Journals (Sweden)

    Thomas Geoffrey C

    2011-05-01

    Full Text Available Abstract Background Malate synthase, one of the two enzymes unique to the glyoxylate cycle, is found in all three domains of life, and is crucial to the utilization of two-carbon compounds for net biosynthetic pathways such as gluconeogenesis. In addition to the main isoforms A and G, so named because of their differential expression in E. coli grown on either acetate or glycolate respectively, a third distinct isoform has been identified. These three isoforms differ considerably in size and sequence conservation. The A isoform (MSA comprises ~530 residues, the G isoform (MSG is ~730 residues, and this third isoform (MSH-halophilic is ~430 residues in length. Both isoforms A and G have been structurally characterized in detail, but no structures have been reported for the H isoform which has been found thus far only in members of the halophilic Archaea. Results We have solved the structure of a malate synthase H (MSH isoform member from Haloferax volcanii in complex with glyoxylate at 2.51 Å resolution, and also as a ternary complex with acetyl-coenzyme A and pyruvate at 1.95 Å. Like the A and G isoforms, MSH is based on a β8/α8 (TIM barrel. Unlike previously solved malate synthase structures which are all monomeric, this enzyme is found in the native state as a trimer/hexamer equilibrium. Compared to isoforms A and G, MSH displays deletion of an N-terminal domain and a smaller deletion at the C-terminus. The MSH active site is closely superimposable with those of MSA and MSG, with the ternary complex indicating a nucleophilic attack on pyruvate by the enolate intermediate of acetyl-coenzyme A. Conclusions The reported structures of MSH from Haloferax volcanii allow a detailed analysis and comparison with previously solved structures of isoforms A and G. These structural comparisons provide insight into evolutionary relationships among these isoforms, and also indicate that despite the size and sequence variation, and the truncated C

  14. Glutamate dehydrogenase isoforms with N-terminal (His)6- or FLAG-tag retain their kinetic properties and cellular localization

    DEFF Research Database (Denmark)

    Pajęcka, Kamilla; Nielsen, Camilla Wendel; Hauge, Anne;

    2014-01-01

    Glutamate dehydrogenase (GDH) is a crucial enzyme on the crossroads of amino acid and energy metabolism and it is operating in all domains of life. According to current knowledge GDH is present only in one functional isoform in most animals, including mice. In addition to this housekeeping enzyme...

  15. Activity of the acyl-CoA synthetase ACSL6 isoforms: role of the fatty acid Gate-domains

    Directory of Open Access Journals (Sweden)

    Siliakus Melvin

    2010-04-01

    Full Text Available Abstract Background Activation of fatty acids by acyl-CoA synthetase enzymes is required for de novo lipid synthesis, fatty acid catabolism, and remodeling of biological membranes. Human long-chain acyl-CoA synthetase member 6, ASCL6, is a form present in the plasma membrane of cells. Splicing events affecting the amino-terminus and alternative motifs near the ATP-binding site generate different isoforms of ACSL6. Results Isoforms with different fatty acid Gate-domain motifs have different activity and the form lacking this domain, isoform 3, showed no detectable activity. Enzymes truncated of the first 40 residues generate acyl-CoAs at a faster rate than the full-length protein. The gating residue, which prevents entry of the fatty acid substrate unless one molecule of ATP has already accessed the catalytic site, was identified as a tyrosine for isoform 1 and a phenylalanine for isoform 2 at position 319. All isoforms, with or without a fatty acid Gate-domain, as well as recombinant protein truncated of the N-terminus, can interact to form enzymatic complexes with identical or different isoforms. Conclusion The alternative fatty acid Gate-domain motifs are essential determinants for the activity of the human ACSL6 isoforms, which appear to act as homodimeric enzyme as well as in complex with other spliced forms. These findings provide evidence that the diversity of these enzyme species could produce the variety of acyl-CoA synthetase activities that are necessary to generate and repair the hundreds of lipid species present in membranes.

  16. Role of acyl carrier protein isoforms in plant lipid metabolism: Progress report

    Energy Technology Data Exchange (ETDEWEB)

    Ohlrogge, J.B.

    1989-01-01

    Previous research from my lab has revealed that several higher plant species have multiple isoforms of acyl carrier protein (ACP) and therefore this trait appears highly conserved among higher plants. This level of conservation suggests that the existence of ACP isoforms is not merely the results of neutral gene duplications. We have developed techniques to examine a wider range of species. Acyl carrier proteins can be labelled very specifically and to high specific activity using H-palmitate and the E. coli enzyme acyl-ACP synthetase. Isoforms were then resolved by western blotting and native PAGE of H-palmitate labelled ACP's. Multiple isoforms of ACP were observed the leaf tissue of the monocots Avena sativa and Hordeum vulgare and dicots including Arabidopsis thallina, Cuphea wrightii, and Brassica napus. Lower vascular plants including the cycad, Dioon edule, Ginkgo biloba, the gymnosperm Pinus, the fern Anernia phyllitidis and Psilotum nudum, the most primitive known extant vascular plant, were also found to have multiple ACP isoforms as were the nonvascular liverwort, Marchantia and moss, Polytrichum. Therefore, the development of ACP isoforms occurred early in evolution. However, the uniellular alge Chlamydomonas and Dunaliella and the photosynthetic cyanobacteria Synechocystis and Agmnellum have only a single elecrophotetic form of ACP. Thus, multiple forms of ACP do not occur in all photosynthetic organisms but may be associated with multicellular plants.

  17. Branched polynomial covering maps

    DEFF Research Database (Denmark)

    Hansen, Vagn Lundsgaard

    1999-01-01

    A Weierstrass polynomial with multiple roots in certain points leads to a branched covering map. With this as the guiding example, we formally define and study the notion of a branched polynomial covering map. We shall prove that many finite covering maps are polynomial outside a discrete branch...... set. Particular studies are made of branched polynomial covering maps arising from Riemann surfaces and from knots in the 3-sphere....

  18. Branched polynomial covering maps

    DEFF Research Database (Denmark)

    Hansen, Vagn Lundsgaard

    2002-01-01

    A Weierstrass polynomial with multiple roots in certain points leads to a branched covering map. With this as the guiding example, we formally define and study the notion of a branched polynomial covering map. We shall prove that many finite covering maps are polynomial outside a discrete branch...... set. Particular studies are made of branched polynomial covering maps arising from Riemann surfaces and from knots in the 3-sphere. (C) 2001 Elsevier Science B.V. All rights reserved....

  19. [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

  20. Study of the physiological function of carnitine palmitoyltransferase 1C enzyme

    OpenAIRE

    Carrasco Rodríguez, Patricia

    2012-01-01

    Carnitine palmitoyl transferase 1 (CPT1) enzymes catalyze the conversion of long-chain acyl-CoA to acyl-carnitines, thus facilitating the entry of long-chain fatty acids to the mitochondria, where they undergo β-oxidation. There are three isoforms: the liver isoform CPT1A (Esser, V. 1993), the muscle isoform CPT1B (Yamazaki, N. 1995) and the brain-specific isoform CPT1C (Price, N. 2002). CPT1A and CPT1B are localized in the outer mitochondrial membrane and are rate-limiting enzymes in fatty-a...

  1. Bank branches in supermarkets

    OpenAIRE

    Lawrence J. Radecki; John Wenninger; Daniel Orlow

    1996-01-01

    The largest U.S. commercial banks are restructuring their retail operations to reduce the cost disadvantage resulting from a stagnant deposit base and stiffer competition. As part of this effort, some banks are opening "supermarket," or "in-store," branches: a new type of banking office within a large retail outlet. An alternative to the traditional bank office, the supermarket branch enables banks to improve the efficiency of the branch network and offer greater convenience to customers.

  2. FSH isoform pattern in classic galactosemia

    OpenAIRE

    Gubbels, Cynthia S.; Thomas, Chris M.G.; Wodzig, Will K. W. H.; Olthaar, André J.; Jaeken, Jaak; Sweep, Fred C. G. J.; Rubio-Gozalbo, M. Estela

    2010-01-01

    Female classic galactosemia patients suffer from primary ovarian insufficiency (POI). The cause for this long-term complication is not fully understood. One of the proposed mechanisms is that hypoglycosylation of complex molecules, a known secondary phenomenon of galactosemia, leads to FSH dysfunction. An earlier study showed less acidic isoforms of FSH in serum samples of two classic galactosemia patients compared to controls, indicating hypoglycosylation. In this study, FSH isoform patterns...

  3. Isoform Specificity of Protein Kinase Cs in Synaptic Plasticity

    Science.gov (United States)

    Sossin, Wayne S.

    2007-01-01

    Protein kinase Cs (PKCs) are implicated in many forms of synaptic plasticity. However, the specific isoform(s) of PKC that underlie(s) these events are often not known. We have used "Aplysia" as a model system in order to investigate the isoform specificity of PKC actions due to the presence of fewer isoforms and a large number of documented…

  4. PKC Isoform Expression in Modeled Microgravity

    Science.gov (United States)

    Risin, Diana; Sundaresan, Alamelu; Pellis, Neal R.; Dawson, David L. (Technical Monitor)

    1999-01-01

    Our previous studies showed that modeled (MMG) and true (USA Space Shuttle Missions STS-54 and STS-56) microgravity (MG) inhibit human lymphocyte locomotion, Modeled MG also suppressed polyclonal and antigen-specific lymphocyte activation. Activation of PKC by phorbol myristate acetate (PMA) restored the microgravity-inhibited lymphocyte locomotion as well as activation by phytohaemagglutinin (PHA), whereas calcium ionophore (ionomycin) was unable to restore these functions. Based on these results we hypothesized that MG-induced changes in lymphocyte functions are caused by a fundamental defect in signal transduction mechanism. This defect may be localized either at the PKC level or upstream of PKC, most likely, at the cell membrane level. In this study we examined the expression of PKC isoforms alpha, epsilon and delta in PBMC cultured in rotating wall vessel bioreactor, developed at NASA JSC, which models microgravity by sustaining cells in continuous free fall. The assessment of the isoforms was performed by FACS analysis following cell permeabilization. A decrease in the expression of isoforms epsilon and delta, but not isoform a, was observed in PBMC cultured in microgravity conditions. These data suggest that MMG might selectively affect the expression of Ca2+ independent isoforms of PKC Molecular analysis confirm selective suppression of Ca2+ independent isoforms of PKC.

  5. Purification and characterization of soluble (cytosolic) and bound (cell wall) isoforms of invertases in barley (Hordeum vulgare) elongating stem tissue

    Science.gov (United States)

    Karuppiah, N.; Vadlamudi, B.; Kaufman, P. B.

    1989-01-01

    Three different isoforms of invertases have been detected in the developing internodes of barley (Hordeum vulgare). Based on substrate specificities, the isoforms have been identified to be invertases (beta-fructosidases EC 3.2.1.26). The soluble (cytosolic) invertase isoform can be purified to apparent homogeneity by diethylaminoethyl cellulose, Concanavalin-A Sepharose, organo-mercurial Sepharose, and Sephacryl S-300 chromatography. A bound (cell wall) invertase isoform can be released by 1 molar salt and purified further by the same procedures as above except omitting the organo-mercurial Sepharose affinity chromatography step. A third isoform of invertase, which is apparently tightly associated with the cell wall, cannot be isolated yet. The soluble and bound invertase isoforms were purified by factors of 60- and 7-fold, respectively. The native enzymes have an apparent molecular weight of 120 kilodaltons as estimated by gel filtration. They have been identified to be dimers under denaturing and nondenaturing conditions. The soluble enzyme has a pH optimum of 5.5, Km of 12 millimolar, and a Vmax of 80 micromole per minute per milligram of protein compared with cell wall isozyme which has a pH optimum of 4.5, Km of millimolar, and a Vmax of 9 micromole per minute per milligram of protein.

  6. Modeled microgravity-induced protein kinase C isoform expression in human lymphocytes

    Science.gov (United States)

    Sundaresan, A.; Risin, D.; Pellis, N. R.

    2004-01-01

    In long-term space travel, the crew is exposed to microgravity and radiation that invoke potential hazards to the immune system. T cell activation is a critical step in the immune response. Receptor-mediated signaling is inhibited in both microgravity and modeled microgravity (MMG) as reflected by diminished DNA synthesis in peripheral blood lymphocytes and their locomotion through gelled type I collagen. Direct activation of protein kinase C (PKC) bypassing cell surface events using the phorbol ester PMA rescues MMG-inhibited lymphocyte activation and locomotion, whereas the calcium ionophore ionomycin had no rescue effect. Thus calcium-independent PKC isoforms may be affected in MMG-induced locomotion inhibition and rescue. Both calcium-dependent isoforms and calcium-independent PKC isoforms were investigated to assess their expression in lymphocytes in 1 g and MMG culture. Human lymphocytes were cultured and harvested at 24, 48, 72, and 96 h, and serial samples were assessed for locomotion by using type I collagen and expression of PKC isoforms. Expression of PKC-alpha, -delta, and -epsilon was assessed by RT-PCR, flow cytometry, and immunoblotting. Results indicated that PKC isoforms delta and epsilon were downregulated by >50% at the transcriptional and translational levels in MMG-cultured lymphocytes compared with 1-g controls. Events upstream of PKC, such as phosphorylation of phospholipase Cgamma in MMG, revealed accumulation of inactive enzyme. Depressed calcium-independent PKC isoforms may be a consequence of an upstream lesion in the signal transduction pathway. The differential response among calcium-dependent and calcium-independent isoforms may actually result from MMG intrusion events earlier than PKC, but after ligand-receptor interaction.

  7. Regulation of three isoforms of SOD gene by environmental stresses in citrus red mite, Panonychus citri.

    Science.gov (United States)

    Feng, Ying-Cai; Liao, Chong-Yu; Xia, Wen-Kai; Jiang, Xuan-Zhao; Shang, Feng; Yuan, Guo-Rui; Wang, Jin-Jun

    2015-09-01

    Superoxide dismutase (SOD) is a family of enzymes with multiple isoforms that possess antioxidative abilities in response to environmental stresses. Panonychus citri is one of the most important pest mites and has a global distribution. In this study, three distinct isoforms of SOD were cloned from P. citri and identified as cytoplasmic Cu-ZnSOD (PcSOD1), extracellular Cu-ZnSOD (PcSOD2), and mitochondrial MnSOD (PcSOD3). mRNA expression level analysis showed that all three isoforms were up-regulated significantly after exposure to the acaricide abamectin and to UV-B ultraviolet irradiation. In particular, PcSOD3 was up-regulated under almost all environmental stresses tested. The fold change of PcSOD3 expression was significantly higher than those of the two Cu-ZnSOD isoforms. Taken together, the results indicate that abamectin and UV-B can induce transcripts of all three SOD isoforms in P. citri. Furthermore, PcSOD3 seems to play a more important role in P. citri tolerance to oxidative stress. PMID:26063404

  8. [Preparation and properties of isocitrate lyase isoforms from the cotyledons of Glycine max L].

    Science.gov (United States)

    Eprintsev, A T; Diachenko, E V; Lykova, T V; Kuen, C T H; Popov, V N

    2010-01-01

    A four-stage purification procedure including ammonium sulfate precipitation and ion exchange chromatography on DEAE cellulose has been elaborated for isolation of isocitrate lyase (EC 4.1.3.1) isoforms from the cotyledons of soybean Glycine max L. Electrophoretically homogeneous preparations of two forms of the enzyme with specific activity of 5.28 and 5.81 U/mg protein have been obtained. Comparison of physicochemical, kinetic, and regulation characteristics of the isoforms obtained revealed fundamental differences between them. Thus, the isoform that migrated quickly in PAAG had a much lower affinity to isocitrate (K(M) - 50 microM) than the slowly migrating form (K(M) - 16 microM). It has been shown that the conservation of activity of the isoforms obtained depends on the presence of divalent cations (Mn2+ and Mg2+) in the medium. It is suggested to use the isoforms of isocitrate lyase isolated from soybeans for the development of biosensors for biochemical and kinetic assays. PMID:20198926

  9. Crystallization and identification of the glycosylated moieties of two isoforms of the main allergen Hev b 2 and preliminary X-ray analysis of two polymorphs of isoform II

    International Nuclear Information System (INIS)

    Crystallization of important glycoenzymes involved in IgE-mediated latex allergy. Latex from Hevea brasiliensis contains several allergenic proteins that are involved in type I allergy. One of them is Hev b 2, which is a β-1,3-glucanase enzyme that exists in different isoforms with variable glycosylation content. Two glucanase isoforms were isolated from trees of the GV-42 clone by gel filtration, affinity and ion-exchange chromatography. Isoform I had a carbohydrate content of about 20%, with N-linked N-acetyl-glucosamine, N-acetyl-galactosamine, fucose and galactose residues as the main sugars, while isoform II showed 6% carbohydrate content constisting of N-acetyl-glucosamine, fucose, mannose and xylose. Both isoforms were crystallized by the hanging-drop vapour-diffusion method. Isoform I crystals were grown using 0.2 M trisodium citrate dihydrate, 0.1 M Na HEPES pH 7.5 and 20%(v/v) 2-propanol, but these crystals were not appropriate for data collection. Isoform II crystals were obtained under two conditions and X-ray diffraction data were collected from both. In the first condition (0.2 M trisodium citrate, 0.1 M sodium cacodylate pH 6.5, 30% 2-propanol), crystals belonging to the tetragonal space group P41 with unit-cell parameters a = b = 150.17, c = 77.41 Å were obtained. In the second condition [0.2 M ammonium acetate, 0.1 M trisodium citrate dihydrate pH 5.6, 30%(w/v) polyethylene glycol 4000] the isoform II crystals belonged to the monoclinic space group P21, with unit-cell parameters a = 85.08, b = 89.67, c = 101.80 Å, β = 113.6°. Preliminary analysis suggests that there are four molecules of isoform II in both asymmetric units

  10. Crystallization and identification of the glycosylated moieties of two isoforms of the main allergen Hev b 2 and preliminary X-ray analysis of two polymorphs of isoform II

    Energy Technology Data Exchange (ETDEWEB)

    Fuentes-Silva, D. [Instituto de Química, Universidad Nacional Autónoma de México, Circuito Exterior s/n, Cuidad Universitaria, Coyoacán, México, DF 04510 (Mexico); Mendoza-Hernández, G. [Facultad de Medicina, Universidad Nacional Autónoma de México, Circuito Exterior s/n, Cuidad Universitaria, Coyoacán, México, DF 04510 (Mexico); Stojanoff, V. [Brookhaven National Laboratory, National Synchrotron Light Source, Upton, NY (United States); Palomares, L. A. [Instituto de Biotecnología, Universidad Nacional Autónoma de México, Circuito Exterior s/n, Cuidad Universitaria, Coyoacán, México, DF 04510 (Mexico); Zenteno, E. [Facultad de Medicina, Universidad Nacional Autónoma de México, Circuito Exterior s/n, Cuidad Universitaria, Coyoacán, México, DF 04510 (Mexico); Torres-Larios, A. [Instituto de Fisiología Celular, Universidad Nacional Autónoma de México, Circuito Exterior s/n, Cuidad Universitaria, Coyoacán, México, DF 04510 (Mexico); Rodríguez-Romero, A., E-mail: adela@servidor.unam.mx [Instituto de Química, Universidad Nacional Autónoma de México, Circuito Exterior s/n, Cuidad Universitaria, Coyoacán, México, DF 04510 (Mexico)

    2007-09-01

    Crystallization of important glycoenzymes involved in IgE-mediated latex allergy. Latex from Hevea brasiliensis contains several allergenic proteins that are involved in type I allergy. One of them is Hev b 2, which is a β-1,3-glucanase enzyme that exists in different isoforms with variable glycosylation content. Two glucanase isoforms were isolated from trees of the GV-42 clone by gel filtration, affinity and ion-exchange chromatography. Isoform I had a carbohydrate content of about 20%, with N-linked N-acetyl-glucosamine, N-acetyl-galactosamine, fucose and galactose residues as the main sugars, while isoform II showed 6% carbohydrate content constisting of N-acetyl-glucosamine, fucose, mannose and xylose. Both isoforms were crystallized by the hanging-drop vapour-diffusion method. Isoform I crystals were grown using 0.2 M trisodium citrate dihydrate, 0.1 M Na HEPES pH 7.5 and 20%(v/v) 2-propanol, but these crystals were not appropriate for data collection. Isoform II crystals were obtained under two conditions and X-ray diffraction data were collected from both. In the first condition (0.2 M trisodium citrate, 0.1 M sodium cacodylate pH 6.5, 30% 2-propanol), crystals belonging to the tetragonal space group P4{sub 1} with unit-cell parameters a = b = 150.17, c = 77.41 Å were obtained. In the second condition [0.2 M ammonium acetate, 0.1 M trisodium citrate dihydrate pH 5.6, 30%(w/v) polyethylene glycol 4000] the isoform II crystals belonged to the monoclinic space group P2{sub 1}, with unit-cell parameters a = 85.08, b = 89.67, c = 101.80 Å, β = 113.6°. Preliminary analysis suggests that there are four molecules of isoform II in both asymmetric units.

  11. Crystallization and Identification of the Glycosylated Moieties of Two Isoforms of the Main Allergen Hev b 2 and Preliminary X-ray Analysis of Two Polymorphs of Isoform ll

    Energy Technology Data Exchange (ETDEWEB)

    Fuentes-Silva,D.; Mendoza-Hernandez, G.; Stojanoff, V.; Palomares, L.; Zenteno, E.; Torres-Larios, A.; Rodriguez-Romero, A.

    2007-01-01

    Latex from Hevea brasiliensis contains several allergenic proteins that are involved in type I allergy. One of them is Hev b 2, which is a {beta}-1,3-glucanase enzyme that exists in different isoforms with variable glycosylation content. Two glucanase isoforms were isolated from trees of the GV-42 clone by gel filtration, affinity and ion-exchange chromatography. Isoform I had a carbohydrate content of about 20%, with N-linked N-acetyl-glucosamine, N-acetyl-galactosamine, fucose and galactose residues as the main sugars, while isoform II showed 6% carbohydrate content consisting of N-acetyl-glucosamine, fucose, mannose and xylose. Both isoforms were crystallized by the hanging-drop vapor-diffusion method. Isoform I crystals were grown using 0.2 M trisodium citrate dihydrate, 0.1 M Na HEPES pH 7.5 and 20%(v/v) 2-propanol, but these crystals were not appropriate for data collection. Isoform II crystals were obtained under two conditions and X-ray diffraction data were collected from both. In the first condition (0.2 M trisodium citrate, 0.1 M sodium cacodylate pH 6.5, 30% 2-propanol), crystals belonging to the tetragonal space group P4{sub 1} with unit-cell parameters a = b = 150.17, c = 77.41 {angstrom} were obtained. In the second condition [0.2 M ammonium acetate, 0.1 M trisodium citrate dihydrate pH 5.6, 30%(w/v) polyethylene glycol 4000] the isoform II crystals belonged to the monoclinic space group P2{sub 1}, with unit-cell parameters a = 85.08, b = 89.67, c = 101.80 {angstrom}, {beta}= 113.6{sup o}. Preliminary analysis suggests that there are four molecules of isoform II in both asymmetric units.

  12. Identification of a novel CoA synthase isoform, which is primarily expressed in Brain

    International Nuclear Information System (INIS)

    CoA and its derivatives Acetyl-CoA and Acyl-CoA are important players in cellular metabolism and signal transduction. CoA synthase is a bifunctional enzyme which mediates the final stages of CoA biosynthesis. In previous studies, we have reported molecular cloning, biochemical characterization, and subcellular localization of CoA synthase (CoASy). Here, we describe the existence of a novel CoA synthase isoform, which is the product of alternative splicing and possesses a 29aa extension at the N-terminus. We termed it CoASy β and originally identified CoA synthase, CoASy α. The transcript specific for CoASy β was identified by electronic screening and by RT-PCR analysis of various rat tissues. The existence of this novel isoform was further confirmed by immunoblot analysis with antibodies directed to the N-terminal peptide of CoASy β. In contrast to CoASy α, which shows ubiquitous expression, CoASy β is primarily expressed in Brain. Using confocal microscopy, we demonstrated that both isoforms are localized on mitochondria. The N-terminal extension does not affect the activity of CoA synthase, but possesses a proline-rich sequence which can bring the enzyme into complexes with signalling proteins containing SH3 or WW domains. The role of this novel isoform in CoA biosynthesis, especially in Brain, requires further elucidation

  13. Absolute Quantification of Endogenous Ras Isoform Abundance.

    Directory of Open Access Journals (Sweden)

    Craig J Mageean

    Full Text Available Ras proteins are important signalling hubs situated near the top of networks controlling cell proliferation, differentiation and survival. Three almost identical isoforms, HRAS, KRAS and NRAS, are ubiquitously expressed yet have differing biological and oncogenic properties. In order to help understand the relative biological contributions of each isoform we have optimised a quantitative proteomics method for accurately measuring Ras isoform protein copy number per cell. The use of isotopic protein standards together with selected reaction monitoring for diagnostic peptides is sensitive, robust and suitable for application to sub-milligram quantities of lysates. We find that in a panel of isogenic SW48 colorectal cancer cells, endogenous Ras proteins are highly abundant with ≥260,000 total Ras protein copies per cell and the rank order of isoform abundance is KRAS>NRAS≥HRAS. A subset of oncogenic KRAS mutants exhibit increased total cellular Ras abundance and altered the ratio of mutant versus wild type KRAS protein. These data and methodology are significant because Ras protein copy number is required to parameterise models of signalling networks and informs interpretation of isoform-specific Ras functional data.

  14. Deubiquitylating enzymes and disease

    OpenAIRE

    Baker Rohan T; Taylor Matthew C; Singhal Shweta

    2008-01-01

    Abstract Deubiquitylating enzymes (DUBs) can hydrolyze a peptide, amide, ester or thiolester bond at the C-terminus of UBIQ (ubiquitin), including the post-translationally formed branched peptide bonds in mono- or multi-ubiquitylated conjugates. DUBs thus have the potential to regulate any UBIQ-mediated cellular process, the two best characterized being proteolysis and protein trafficking. Mammals contain some 80–90 DUBs in five different subfamilies, only a handful of which have been charac...

  15. Materials Test Branch

    Science.gov (United States)

    Gordon, Gail

    2012-01-01

    The Materials Test Branch resides at Marshall Space Flight Center's Materials and Processing laboratory and has a long history of supporting NASA programs from Mercury to the recently retired Space Shuttle. The Materials Test Branch supports its customers by supplying materials testing expertise in a wide range of applications. The Materials Test Branch is divided into three Teams, The Chemistry Team, The Tribology Team and the Mechanical Test Team. Our mission and goal is to provide world-class engineering excellence in materials testing with a special emphasis on customer service.

  16. 马铃薯遗传转化体系的优化及玉米淀粉分支酶基因SBEⅡb的导入%Optimization of genetic transformation system of potato and introduction of maize starch branching enzyme gene SBEⅡb into potatoFAN

    Institute of Scientific and Technical Information of China (English)

    范亚丽; 阮颖; 刘春林

    2012-01-01

    The highly effective potato transformation systems were established and optimized using tube potato plant-lets as transformation materials in this study. The maize starch branching enzyme gene(SBE Ⅱ b)was transformed into potato using Agrobacterium tumefaciens-mediated transformation method. 762 stem sections were infec ted by Agrobaterium-mediated tumefacien method. 4 transgenic plants were obtained by PCR measurement, implying that SEE Ⅱ b gene had been integrated into potato genome. It was also found that GUS gene could normal ly be expressed in stem sections and potato microtubers of the trasgenetic potato plants.%以马铃薯脱毒试管苗茎段为转化受体材料,建立并优化了农杆菌介导的马铃薯遗传转化体系.通过农杆菌介导法将玉米淀粉分支酶基因(Starch branching enzyme b,SBEⅡb)的过表达载体转化马铃薯,接种762个茎段,共获得35株抗性植株.经PCR检测获得了4株转基因阳性植株;对转基因植株进一步进行GUS活性组织化学染色,发现转基因植株的茎段与试管薯均被染上蓝色,表明外源SBEⅡb基因已整合到马铃薯基因组,且正常表达.

  17. Ikaros isoforms:The saga continues

    Institute of Scientific and Technical Information of China (English)

    Laura; A; Perez-Casellas; Aleksandar; Savic; Sinisa; Dovat

    2011-01-01

    Through alternate splicing,the Ikaros gene produces multiple proteins.Ikaros is essential for normal hematopoiesis and possesses tumor suppressor activity.Ikaros isoforms interact to form dimers and potentially multimeric complexes.Diverse Ikaros complexes produced by the presence of different Ikaros isoforms are hypothesized to confer distinct functions.Small dominantnegative Ikaros isoforms have been shown to inhibit the tumor suppressor activity of full-length Ikaros.Here,we describe how Ikaros activity is regulated by the coordinated expression of the largest Ikaros isoforms IK-1 and IK-H.Although IK-1 is described as full-length Ikaros,IK-H is the longest Ikaros isoform.IK-H,which includes residues coded by exon 3B (60 bp that lie between exons 3 and 4),is abundant in human but not murine hematopoietic cells.Specific residues that lie within the 20 amino acids encoded by exon 3B give IK-H DNA-binding characteristics that are distinct from those of IK-1.Moreover,IK-H can potentiate or inhibit the ability of IK-1 to bind DNA.IK-H binds to the regulatory regions of genes that are upregulated by Ikaros,but not genes that are repressed by Ikaros.Although IK-1 localizes to pericentromeric heterochromatin,IK-H can be found in both pericentromeric and non-pericentromeric locations.Anti-silencing activity of gamma satellite DNA has been shown to depend on the binding of IK-H,but not other Ikaros isoforms.The unique features of IK-H,its influence on Ikaros activity,and the lack of IK-H expression in mice suggest that Ikaros function in humans may be more complex and possibly distinct from that in mice.

  18. Branching processes in biology

    CERN Document Server

    Kimmel, Marek

    2015-01-01

    This book provides a theoretical background of branching processes and discusses their biological applications. Branching processes are a well-developed and powerful set of tools in the field of applied probability. The range of applications considered includes molecular biology, cellular biology, human evolution and medicine. The branching processes discussed include Galton-Watson, Markov, Bellman-Harris, Multitype, and General Processes. As an aid to understanding specific examples, two introductory chapters, and two glossaries are included that provide background material in mathematics and in biology. The book will be of interest to scientists who work in quantitative modeling of biological systems, particularly probabilists, mathematical biologists, biostatisticians, cell biologists, molecular biologists, and bioinformaticians. The authors are a mathematician and cell biologist who have collaborated for more than a decade in the field of branching processes in biology for this new edition. This second ex...

  19. The Olive Branch Awards.

    Science.gov (United States)

    Harnack, William

    1984-01-01

    The first annual Olive Branch Awards, sponsored by the Writers' and Publishers Alliance and the Editors' Organizing Committee, were given to ten magazines, out of 60 that submitted entries. Winning entries are described briefly. (IM)

  20. Functional demonstrations of starch binding domains present in Ostreococcus tauri starch synthases isoforms

    OpenAIRE

    Barchiesi, Julieta; Hedin, Nicolás; Gomez-Casati, Diego F.; Miguel A Ballicora; Busi, María V.

    2015-01-01

    Background Starch-binding domains are key modules present in several enzymes involved in polysaccharide metabolism. These non-catalytic modules have already been described as essential for starch-binding and the catalytic activity of starch synthase III from the higher plant Arabidopsis thaliana. In Ostreococcus tauri, a unicellular green alga of the Prasinophyceae family, there are three SSIII isoforms, known as Ostta SSIII-A, SSIII-B and SSIII-C. Results In this work, using in silico and in...

  1. Koenigs function and branching processes

    OpenAIRE

    Tchikilev, O. G.

    2001-01-01

    An explicit solution of time-homogeneous pure birth branching processes is described. It gives alternative extensions for the negative binomial distribution (branching processes with immigration) and for the Furry-Yule distribution (branching processes without immigration).

  2. EFSA Panel on Food Contact Materials, Enzymes, Flavourings and Processing Aids (CEF); Scientific Opinion on Flavouring Group Evaluation 3, Revision 2 (FGE.03Rev2): Acetals of branched- and straight-chain aliphatic saturated primary alcohols and branched- and straight-chain saturated or unsaturated

    DEFF Research Database (Denmark)

    Larsen, John Christian; Nørby, Karin Kristiane; Beltoft, Vibe Meister;

    The Panel on Food Contact Materials, Enzymes, Flavourings and Processing Aids of the European Food Safety Authority was requested to evaluate one flavouring substance, acetaldehyde ethyl isopropyl acetal [FL-no: 06.137], structurally related to the 58 flavouring substances in the Flavouring Group...... structure-activity relationships, intake from current uses, toxicological threshold of concern, and available data on metabolism and toxicity. The Panel concluded as for the other already evaluated substances that the substance [FL-no: 06.137] do not give rise to safety concern at its level of dietary...

  3. Different characteristics and nucleotide binding properties of inosine monophosphate dehydrogenase (IMPDH isoforms.

    Directory of Open Access Journals (Sweden)

    Elaine C Thomas

    Full Text Available We recently reported that Inosine Monophosphate Dehydrogenase (IMPDH, a rate-limiting enzyme in de novo guanine nucleotide biosynthesis, clustered into macrostructures in response to decreased nucleotide levels and that there were differences between the IMPDH isoforms, IMPDH1 and IMPDH2. We hypothesised that the Bateman domains, which are present in both isoforms and serve as energy-sensing/allosteric modules in unrelated proteins, would contribute to isoform-specific differences and that mutations situated in and around this domain in IMPDH1 which give rise to retinitis pigmentosa (RP would compromise regulation. We employed immuno-electron microscopy to investigate the ultrastructure of IMPDH macrostructures and live-cell imaging to follow clustering of an IMPDH2-GFP chimera in real-time. Using a series of IMPDH1/IMPDH2 chimera we demonstrated that the propensity to cluster was conferred by the N-terminal 244 amino acids, which includes the Bateman domain. A protease protection assay suggested isoform-specific purine nucleotide binding characteristics, with ATP protecting IMPDH1 and AMP protecting IMPDH2, via a mechanism involving conformational changes upon nucleotide binding to the Bateman domain without affecting IMPDH catalytic activity. ATP binding to IMPDH1 was confirmed in a nucleotide binding assay. The RP-causing mutation, R224P, abolished ATP binding and nucleotide protection and this correlated with an altered propensity to cluster. Collectively these data demonstrate that (i the isoforms are differentially regulated by AMP and ATP by a mechanism involving the Bateman domain, (ii communication occurs between the Bateman and catalytic domains and (iii the RP-causing mutations compromise such regulation. These findings support the idea that the IMPDH isoforms are subject to distinct regulation and that regulatory defects contribute to human disease.

  4. Glutamic acid decarboxylase isoform distribution in transgenic mouse septum: an anti-GFP immunofluorescence study.

    Science.gov (United States)

    Verimli, Ural; Sehirli, Umit S

    2016-09-01

    The septum is a basal forebrain region located between the lateral ventricles in rodents. It consists of lateral and medial divisions. Medial septal projections regulate hippocampal theta rhythm whereas lateral septal projections are involved in processes such as affective functions, memory formation, and behavioral responses. Gamma-aminobutyric acidergic neurons of the septal region possess the 65 and 67 isoforms of the enzyme glutamic acid decarboxylase. Although data on the glutamic acid decarboxylase isoform distribution in the septal region generally appears to indicate glutamic acid decarboxylase 67 dominance, different studies have given inconsistent results in this regard. The aim of this study was therefore to obtain information on the distributions of both of these glutamic acid decarboxylase isoforms in the septal region in transgenic mice. Two animal groups of glutamic acid decarboxylase-green fluorescent protein knock-in transgenic mice were utilized in the experiment. Brain sections from the region were taken for anti-green fluorescent protein immunohistochemistry in order to obtain estimated quantitative data on the number of gamma-aminobutyric acidergic neurons. Following the immunohistochemical procedures, the mean numbers of labeled cells in the lateral and medial septal nuclei were obtained for the two isoform groups. Statistical analysis yielded significant results which indicated that the 65 isoform of glutamic acid decarboxylase predominates in both lateral and medial septal nuclei (unpaired two-tailed t-test p first to reveal the dominance of glutamic acid decarboxylase isoform 65 in the septal region in glutamic acid decarboxylase-green fluorescent protein transgenic mice. PMID:26643381

  5. Regulation of carnitine palmitoyltransferase I (CPT-Iα) gene expression by the peroxisome proliferator activated receptor gamma coactivator (PGC-1) isoforms

    OpenAIRE

    Sadana, Prabodh; Zhang, Yi; Song, Shulan; Cook, George A.; Elam, Marshall B.; Park, Edwards A.

    2006-01-01

    The peroxisome proliferator-activated receptor gamma coactivators (PGC-1) have important roles in mitochondrial biogenesis and metabolic control in a variety of tissues. There are multiple isoforms of PGC-1 including PGC-1α and PGC-1β. Both the PGC-1α and β isoforms promote mitochondrial biogenesis and fatty acid oxidation, but only PGC-1α stimulates gluconeogenesis in the liver. Carnitine palmitoyltransferase I (CPT-I) is a key enzyme regulating mitochondrial fatty acid oxidation. In these s...

  6. Kunstige Enzymer

    DEFF Research Database (Denmark)

    Bols, Mikael; Bjerre, Jeannette; Marinescu, Lavinia

    2007-01-01

    Enzymer har en enestående evne til at accelerere kemiske processer. Der forskes målrettet i at optimere enzymer baseret på cyclodextrin.......Enzymer har en enestående evne til at accelerere kemiske processer. Der forskes målrettet i at optimere enzymer baseret på cyclodextrin....

  7. p53 isoforms change p53 paradigm

    OpenAIRE

    Bourdon, JC

    2014-01-01

    Although p53 defines cellular responses to cancer treatment it is not clear how p53 can be used to control cell fate outcome. Data demonstrate that so-called p53 does not exist as a single protein, but is in fact a group of p53 protein isoforms whose expression can be manipulated to control the cellular response to treatment.

  8. New isoforms of rat Aquaporin-4

    DEFF Research Database (Denmark)

    Moe, Svein Erik; Sorbo, Jan Gunnar; Søgaard, Rikke;

    2008-01-01

    Aquaporin-4 (AQP4) is a brain aquaporin implicated in the pathophysiology of numerous clinical conditions including brain edema. Here we show that rat AQP4 has six cDNA isoforms, formed by alternative splicing. These are named AQP4a-f, where AQP4a and AQP4c correspond to the two classical M1 and M...

  9. Fragrance Release from the Surface of Branched Poly (Amide S

    Directory of Open Access Journals (Sweden)

    T. Youngs

    2005-01-01

    Full Text Available Enzymes are powerful tools in organic synthesis that are able to catalyse a wide variety of selective chemical transformations under mild and environmentally friendly conditions. Enzymes such as the lipases have also found applications in the synthesis and degradation of polymeric materials. However, the use of these natural catalysts in the synthesis and the post-synthetic modification of dendrimers and hyperbranched molecules is an application of chemistry yet to be explored extensively. In this study the use of two hydrolytic enzymes, a lipase from Candida cylindracea and a cutinase from Fusarium solani pisii, were investigated in the selective cleavage of ester groups situated on the peripheral layer of two families of branched polyamides. These branched polyamides were conjugated to simple fragrances citronellol and L-menthol via ester linkages. Hydrolysis of the ester linkage between the fragrances and the branched polyamide support was carried out in aqueous buffered systems at slightly basic pH values under the optimum operative conditions for the enzymes used. These preliminary qualitative investigations revealed that partial cleavage of the ester functionalities from the branched polyamide support had occurred. However, the ability of the enzymes to interact with the substrates decreased considerably as the branching density, the rigidity of the structure and the bulkiness of the polyamide-fragrance conjugates increased.

  10. Integrated genomic and functional analyses of histone demethylases identify oncogenic KDM2A isoform in breast cancer.

    Science.gov (United States)

    Liu, Hui; Liu, Lanxin; Holowatyj, Andreana; Jiang, Yuanyuan; Yang, Zeng-Quan

    2016-05-01

    Histone lysine demethylases (KDMs) comprise a large class of enzymes that catalyze site-specific demethylation of lysine residues on histones and other proteins. They play critical roles in controlling transcription, chromatin architecture, and cellular differentiation. However, the genomic landscape and clinical significance of KDMs in breast cancer remain poorly characterized. Here, we conducted a meta-analysis of 24 KDMs in breast cancer and identified associations among recurrent copy number alterations, gene expression, breast cancer subtypes, and clinical outcome. Two KDMs, KDM2A and KDM5B, had the highest frequency of genetic amplification and overexpression. Furthermore, among the 24 KDM genes, KDM2A had the highest correlation between copy number and mRNA expression, and high mRNA levels of KDM2A were significantly associated with shorter survival of breast cancer patients. KDM2A has two isoforms: the long isoform is comprised of a JmjC domain, CXXC-zinc finger, PHD zinc finger, F-box, and the AMN1 protein domain; whereas the short isoform of KDM2A lacks the N-terminal JmjC domain but contains all other motifs. Detailed characterization of KDM2A in breast cancer revealed that the short isoform of KDM2A is more abundant than the long isoform at DNA, mRNA, and protein levels in a subset of breast cancers. Furthermore, our data indicate that the short isoform of KDM2A has oncogenic potential and functions as an oncogenic isoform in a subset of breast cancers. Taken together, our findings suggest that amplification and overexpression of the KDM2A short isoform is critical in breast cancer progression. © 2015 Wiley Periodicals, Inc. PMID:26207617

  11. Isolation and characterization of cytosolic alanine aminotransferase isoforms from starved rat liver.

    Science.gov (United States)

    Vedavathi, M; Girish, K S; Kumar, M Karuna

    2004-12-01

    Alanine is the most effective precursor for gluconeogenesis among amino acids and the initial reaction is catalyzed by alanine aminotransferases (AlaATs). It is a less extensively studied enzyme under starvation and known to that the enzyme activity increases in liver under starvation. The present study describes the purification and characterization of two isoforms of alanine aminotransferases from starved male rat liver under starvation. The molecular mass of isoforms was found to be 17.7 and 112.2 kDa with isoelectric points of 4.2 and 5.3 respectively for AlaAT I and AlaAT II. Both the enzymes showed narrow substrate specificity for L-alanine with different Km for alanine and 2-oxoglutarate. Both the enzymes were glycoprotein in nature. Inhibition, modification and spectroscopic studies showed that both PLP and free-SH groups are directly involved in the enzymatic catalysis. PLP activated both the enzymes with a Km 0.057 mM and 0.2 mM for AlaAT I and II respectively. PMID:15663181

  12. Right bundle branch block

    DEFF Research Database (Denmark)

    Bussink, Barbara E; Holst, Anders Gaarsdal; Jespersen, Lasse;

    2013-01-01

    AimsTo determine the prevalence, predictors of newly acquired, and the prognostic value of right bundle branch block (RBBB) and incomplete RBBB (IRBBB) on a resting 12-lead electrocardiogram in men and women from the general population.Methods and resultsWe followed 18 441 participants included in...... men vs. 0.5%/2.3% in women, P <0.001). Significant predictors of newly acquired RBBB were male gender, increasing age, high systolic blood pressure, and presence of IRBBB, whereas predictors of newly acquired IRBBB were male gender, increasing age, and low BMI. Right bundle branch block was associated...... with significantly increased all-cause and cardiovascular mortality in both genders with age-adjusted hazard ratios (HR) of 1.31 [95% confidence interval (CI), 1.11-1.54] and 1.87 (95% CI, 1.48-2.36) in the gender pooled analysis with little attenuation after multiple adjustment. Right bundle branch...

  13. Heterogeneity of serum gelatinases MMP-2 and MMP-9 isoforms and charge variants.

    Science.gov (United States)

    Rossano, Rocco; Larocca, Marilena; Riviello, Lea; Coniglio, Maria Gabriella; Vandooren, Jennifer; Liuzzi, Grazia Maria; Opdenakker, Ghislain; Riccio, Paolo

    2014-02-01

    The matrix metalloproteinases (MMPs) gelatinase A (MMP-2) and gelatinase B (MMP-9) are mediators of brain injury in multiple sclerosis (MS) and valuable biomarkers of disease activity. We applied bidimensional zymography (2-DZ) as an extension of classic monodimensional zymography (1-DZ) to analyse the complete pattern of isoforms and post-translational modifications of both MMP-9 and MMP-2 present in the sera of MS patients. The enzymes were separated on the basis of their isoelectric points (pI) and apparent molecular weights (Mw) and identified both by comparison with standard enzyme preparations and by Western blot analysis. Two MMP-2 isoforms, and at least three different isoforms and two different states of organization of MMP-9 (the multimeric MMP-9 and the N-GAL-MMP-9 complex) were observed. In addition, 2-DZ revealed for the first time that all MMP-9 and MMP-2 isoforms actually exist in the form of charge variants: four or five variants in the NGAL complex, more charge variants in the case of MMP-9; and five to seven charge variants for MMP-2. Charge variants were also observed in recombinant enzymes and, after concentration, also in sera from healthy individuals. Sialylation (MMP-9) and phosphorylation (MMP-2) contributed to molecular heterogeneity. The detection of charge variants of MMP-9 and MMP-2 in MS serum samples illustrates the power of 2-DZ and demonstrates that in previous studies MMP mixtures, rather than single molecules, were analysed. These observations open perspectives for better diagnosis and prognosis of many diseases and need to be critically interpreted when applying other methods for MS and other diseases. PMID:24616914

  14. Purification and catalytic properties of polygalacturonase isoforms from ripe avocado (Persea americana) fruit mesocarp.

    Science.gov (United States)

    Wakabayashi, Kazuyuki; Huber, Donald J.

    2001-10-01

    Endo-polygalacturonase (PG; EC 3.2.1.15) was recovered from the cell walls of avocado mesocarp (Persea americana Mill cv. Lula) tissue and purified by sequential ion exchange and gel permeation chromatography. Two isoforms (S-I and S-II) were recovered, exhibiting molecular masses of about 41 kD on size exclusion media and about 48 (S-I) and 46 (S-II) kDa as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Both isoforms exhibited maximum activity at pH 6.0 against polygalacturonic acid (PGA) and hydrolyzed PGA of about 180 kDa to polymers of about 4 kDa. The catalytic activity of the 48-kDa isoform against PGA was slightly higher than that of the 46-kDa isoform. The purified PGs catalyzed significant molecular mass downshifts in the polyuronides of pre-ripe avocados; however, the capacity of the enzymes to solubilize polyuronides from cell walls of pre-ripe fruit was limited. PMID:12060298

  15. Over-expression in Escherichia coli and characterization of two recombinant isoforms of human FAD synthetase

    International Nuclear Information System (INIS)

    FAD synthetase (FADS) (EC 2.7.7.2) is a key enzyme in the metabolic pathway that converts riboflavin into the redox cofactor FAD. Two hypothetical human FADSs, which are the products of FLAD1 gene, were over-expressed in Escherichia coli and identified by ESI-MS/MS. Isoform 1 was over-expressed as a T7-tagged protein which had a molecular mass of 63 kDa on SDS-PAGE. Isoform 2 was over-expressed as a 6-His-tagged fusion protein, carrying an extra 84 amino acids at the N-terminal with an apparent molecular mass of 60 kDa on SDS-PAGE. It was purified near to homogeneity from the soluble cell fraction by one-step affinity chromatography. Both isoforms possessed FADS activity and had a strict requirement for MgCl2, as demonstrated using both spectrophotometric and chromatographic methods. The purified recombinant isoform 2 showed a specific activity of 6.8 ± 1.3 nmol of FAD synthesized/min/mg protein and exhibited a K M value for FMN of 1.5 ± 0.3 μM. This is First report on characterization of human FADS, and First cloning and over-expression of FADS from an organism higher than yeast

  16. Activity of selected hydrolytic enzymes in Allium sativum L. anthers.

    Science.gov (United States)

    Winiarczyk, Krystyna; Gębura, Joanna

    2016-05-01

    The aim of the study was to determine enzymatic activity in sterile Allium sativum anthers in the final stages of male gametophyte development (the stages of tetrads and free microspores). The analysed enzymes were shown to occur in the form of numerous isoforms. In the tetrad stage, esterase activity was predominant, which was manifested by the greater number of isoforms of the enzyme. In turn, in the microspore stage, higher numbers of isoforms of acid phosphatases and proteases were detected. The development of sterile pollen grains in garlic is associated with a high level of protease and acid phosphatase activity and lower level of esterase activities in the anther locule. Probably this is the first description of the enzymes activity (ACPH, EST, PRO) in the consecutives stages of cell wall formation which is considered to be one of the causes of male sterility in flowering plant. PMID:26901781

  17. A Crucial Role in Fertility for the Oyster Angiotensin-Converting Enzyme Orthologue CgACE

    OpenAIRE

    Riviere, Guillaume; Fellous, Alexandre; Franco, Alban; Bernay, Benoit; Favrel, Pascal

    2011-01-01

    Angiotensin-converting enzyme (ACE) is a highly conserved metallopeptidase. In mammals, the somatic isoform governs blood pressure whereas the germinal isoform (tACE) is required for fertility. In Ecdysozoans, ACE-like enzymes are implicated in reproduction. Despite ACE orthologues being present from bacteria to humans, their function(s) remain(s) unknown in distant organisms such as Lophotrochozoans. In silico analysis of an oyster (Crassostrea gigas) EST library suggested the presence of an...

  18. Tracheobronchial Branching Anomalies

    Energy Technology Data Exchange (ETDEWEB)

    Hong, Min Ji; Kim, Young Tong; Jou, Sung Shick [Soonchunhyang University, Cheonan Hospital, Cheonan (Korea, Republic of); Park, A Young [Soonchunhyang University College of Medicine, Asan (Korea, Republic of)

    2010-04-15

    There are various congenital anomalies with respect to the number, length, diameter, and location of tracheobronchial branching patterns. The tracheobronchial anomalies are classified into two groups. The first one, anomalies of division, includes tracheal bronchus, cardiac bronchus, tracheal diverticulum, pulmonary isomerism, and minor variations. The second one, dysmorphic lung, includes lung agenesis-hypoplasia complex and lobar agenesis-aplasia complex

  19. Characterization of protein kinase C and its isoforms in human T lymphocytes.

    Science.gov (United States)

    Beyers, A D; Hanekom, C; Rheeder, A; Strachan, A F; Wooten, M W; Nel, A E

    1988-11-15

    Protein kinase C (PKC) regulates numerous T cell functions and is present in abundance in normal human T cells and certain T cell lines. Although crude Triton X-100 soluble material obtained from T cell pellets contains minimal PKC activity, DEAE chromatography revealed that 12 to 37% of cellular PKC was membrane associated, probably due to removal of an inhibitor through column chromatography. As in other tissues, PKC from lymphoid tissue was phospholipid and Ca2+ dependent and diolein reduced the Ca2+ requirements for enzyme activity. Hydroxylapatite chromatography revealed that T cells possess two major peaks of PKC activity. Although, the enzyme in these peaks had similar m.w. and identical iso-electric mobility, the proteins differed with respect to their autophosphorylation sites and immunoreactivity toward an isoform specific antibody. Furthermore, differences in their activities in the presence of phospholipid, diolein, and limiting amounts of Ca2+ imply that these isoforms may be differentially activated. We discuss optimal conditions for activation of PKC and its isoforms for study of T lymphocyte cellular function. PMID:3263426

  20. Differential water permeability and regulation of three aquaporin 4 isoforms

    DEFF Research Database (Denmark)

    Fenton, Robert A.; Moeller, Hanne B; Zelenina, Marina;

    2010-01-01

    Aquaporin 4 (AQP4) is expressed in the perivascular glial endfeet and is an important pathway for water during formation and resolution of brain edema. In this study, we examined the functional properties and relative unit water permeability of three functional isoforms of AQP4 expressed...... in the brain (M1, M23, Mz). The M23 isoform gave rise to square arrays when expressed in Xenopus laevis oocytes. The relative unit water permeability differed significantly between the isoforms in the order of M1 > Mz > M23. None of the three isoforms were permeable to small osmolytes nor were they affected...... by changes in external K(+) concentration. Upon protein kinase C (PKC) activation, oocytes expressing the three isoforms demonstrated rapid reduction of water permeability, which correlated with AQP4 internalization. The M23 isoform was more sensitive to PKC regulation than the longer isoforms...

  1. Lipoprotein lipase isoelectric point isoforms in humans

    DEFF Research Database (Denmark)

    Badia-Villanueva, M.; Carulla, P.; Carrascal, M.;

    2014-01-01

    characterization of these forms was carried out by 2DE combined with Western blotting and mass spectrometry (MALDI-TOF/MS and LC-MS/MS). Further studies are needed to discover their molecular origin, the pattern of pI isoforms in human tissues, their possible physiological functions and possible modifications of......-heparin plasma (PHP), LPL consists of a pattern of more than 8 forms of the same apparent molecular weight, but different isoelectric point (pI). In the present study we describe, for the first time, the existence of at least nine LPL pI isoforms in human PHP, with apparent pI between 6.8 and 8.6. Separation and...

  2. EASI—enrichment of alternatively spliced isoforms

    OpenAIRE

    Julian P Venables; Burn, John

    2006-01-01

    Alternative splicing produces more than one protein from the majority of genes and the rarer forms can have dominant functions. Instability of alternative transcripts can also hinder the study of regulation of gene expression by alternative splicing. To investigate the true extent of alternative splicing we have developed a simple method of enriching alternatively spliced isoforms (EASI) from PCRs using beads charged with Thermus aquaticus single-stranded DNA-binding protein (T.Aq ssb). This ...

  3. Functional studies of sodium pump isoforms

    DEFF Research Database (Denmark)

    Clausen, Michael Jakob

    The Na+,K+-ATPase is an essential ion pump found in all animal cells. It uses the energy from ATP hydrolysis to export three Na+ and import two K+, both against their chemical gradients and for Na+ also against the electrical potential. Mammals require four Na+,K+-ATPase isoforms that each have...... synthesized cohorts of pumps from the Golgi apparatus to the plasma membrane....

  4. Production of ligninolytic enzymes by cultures of white rot fungi.

    Science.gov (United States)

    Górska, Ewa B; Jankiewicz, Urszula; Dobrzyński, Jakub; Agnieszka Gałązka; Sitarek, Monika; Gozdowski, Dariusz; Russel, Stefan; Kowalczyk, Paweł

    2014-01-01

    Some Basidiomycota were chosen for studies of key ligninases synthesis (25°C, 30 days) in modified medium (shaken or not cultures) with added wheat straw. Liquid Czapek medium with straw yielded a higher amount of laccase than peroxidase, ground straw induced enzyme worse than chopped straw. With peroxidase the reverse dependencies were observed. Laccase of Lentinus edodes synthesized two enzyme isoforms (ca 30 and 16 kDa). In T. versicolor culture active laccase protein with highest molecular mass ca 65 kDa was found. P. sajor-caju yielded three different peroxidase isoforms. Ligninase biosynthesis depended on strain, straw fragmentation extent, culture method and growth medium. PMID:25804067

  5. Deep Proteomics of Mouse Skeletal Muscle Enables Quantitation of Protein Isoforms, Metabolic Pathways, and Transcription Factors*

    Science.gov (United States)

    Deshmukh, Atul S.; Murgia, Marta; Nagaraj, Nagarjuna; Treebak, Jonas T.; Cox, Jürgen; Mann, Matthias

    2015-01-01

    Skeletal muscle constitutes 40% of individual body mass and plays vital roles in locomotion and whole-body metabolism. Proteomics of skeletal muscle is challenging because of highly abundant contractile proteins that interfere with detection of regulatory proteins. Using a state-of-the art MS workflow and a strategy to map identifications from the C2C12 cell line model to tissues, we identified a total of 10,218 proteins, including skeletal muscle specific transcription factors like myod1 and myogenin and circadian clock proteins. We obtain absolute abundances for proteins expressed in a muscle cell line and skeletal muscle, which should serve as a valuable resource. Quantitation of protein isoforms of glucose uptake signaling pathways and in glucose and lipid metabolic pathways provides a detailed metabolic map of the cell line compared with tissue. This revealed unexpectedly complex regulation of AMP-activated protein kinase and insulin signaling in muscle tissue at the level of enzyme isoforms. PMID:25616865

  6. Tau leptonic branching ratios

    CERN Document Server

    Buskulic, Damir; De Bonis, I; Décamp, D; Ghez, P; Goy, C; Lees, J P; Lucotte, A; Minard, M N; Odier, P; Pietrzyk, B; Ariztizabal, F; Chmeissani, M; Crespo, J M; Efthymiopoulos, I; Fernández, E; Fernández-Bosman, M; Gaitan, V; Garrido, L; Martínez, M; Orteu, S; Pacheco, A; Padilla, C; Palla, Fabrizio; Pascual, A; Perlas, J A; Sánchez, F; Teubert, F; Colaleo, A; Creanza, D; De Palma, M; Farilla, A; Gelao, G; Girone, M; Iaselli, Giuseppe; Maggi, G; Maggi, M; Marinelli, N; Natali, S; Nuzzo, S; Ranieri, A; Raso, G; Romano, F; Ruggieri, F; Selvaggi, G; Silvestris, L; Tempesta, P; Zito, G; Huang, X; Lin, J; Ouyang, Q; Wang, T; Xie, Y; Xu, R; Xue, S; Zhang, J; Zhang, L; Zhao, W; Bonvicini, G; Cattaneo, M; Comas, P; Coyle, P; Drevermann, H; Engelhardt, A; Forty, Roger W; Frank, M; Hagelberg, R; Harvey, J; Jacobsen, R; Janot, P; Jost, B; Kneringer, E; Knobloch, J; Lehraus, Ivan; Markou, C; Martin, E B; Mato, P; Minten, Adolf G; Miquel, R; Oest, T; Palazzi, P; Pater, J R; Pusztaszeri, J F; Ranjard, F; Rensing, P E; Rolandi, Luigi; Schlatter, W D; Schmelling, M; Schneider, O; Tejessy, W; Tomalin, I R; Venturi, A; Wachsmuth, H W; Wiedenmann, W; Wildish, T; Witzeling, W; Wotschack, J; Ajaltouni, Ziad J; Bardadin-Otwinowska, Maria; Barrès, A; Boyer, C; Falvard, A; Gay, P; Guicheney, C; Henrard, P; Jousset, J; Michel, B; Monteil, S; Montret, J C; Pallin, D; Perret, P; Podlyski, F; Proriol, J; Rossignol, J M; Saadi, F; Fearnley, Tom; Hansen, J B; Hansen, J D; Hansen, J R; Hansen, P H; Nilsson, B S; Kyriakis, A; Simopoulou, Errietta; Siotis, I; Vayaki, Anna; Zachariadou, K; Blondel, A; Bonneaud, G R; Brient, J C; Bourdon, P; Passalacqua, L; Rougé, A; Rumpf, M; Tanaka, R; Valassi, Andrea; Verderi, M; Videau, H L; Candlin, D J; Parsons, M I; Focardi, E; Parrini, G; Corden, M; Delfino, M C; Georgiopoulos, C H; Jaffe, D E; Antonelli, A; Bencivenni, G; Bologna, G; Bossi, F; Campana, P; Capon, G; Chiarella, V; Felici, G; Laurelli, P; Mannocchi, G; Murtas, F; Murtas, G P; Pepé-Altarelli, M; Dorris, S J; Halley, A W; ten Have, I; Knowles, I G; Lynch, J G; Morton, W T; O'Shea, V; Raine, C; Reeves, P; Scarr, J M; Smith, K; Smith, M G; Thompson, A S; Thomson, F; Thorn, S; Turnbull, R M; Becker, U; Braun, O; Geweniger, C; Graefe, G; Hanke, P; Hepp, V; Kluge, E E; Putzer, A; Rensch, B; Schmidt, M; Sommer, J; Stenzel, H; Tittel, K; Werner, S; Wunsch, M; Beuselinck, R; Binnie, David M; Cameron, W; Colling, D J; Dornan, Peter J; Konstantinidis, N P; Moneta, L; Moutoussi, A; Nash, J; San Martin, G; Sedgbeer, J K; Stacey, A M; Dissertori, G; Girtler, P; Kuhn, D; Rudolph, G; Bowdery, C K; Brodbeck, T J; Colrain, P; Crawford, G; Finch, A J; Foster, F; Hughes, G; Sloan, Terence; Whelan, E P; Williams, M I; Galla, A; Greene, A M; Kleinknecht, K; Quast, G; Raab, J; Renk, B; Sander, H G; Wanke, R; Van Gemmeren, P; Zeitnitz, C; Aubert, Jean-Jacques; Bencheikh, A M; Benchouk, C; Bonissent, A; Bujosa, G; Calvet, D; Carr, J; Diaconu, C A; Etienne, F; Thulasidas, M; Nicod, D; Payre, P; Rousseau, D; Talby, M; Abt, I; Assmann, R W; Bauer, C; Blum, Walter; Brown, D; Dietl, H; Dydak, Friedrich; Ganis, G; Gotzhein, C; Jakobs, K; Kroha, H; Lütjens, G; Lutz, Gerhard; Männer, W; Moser, H G; Richter, R H; Rosado-Schlosser, A; Schael, S; Settles, Ronald; Seywerd, H C J; Saint-Denis, R; Wolf, G; Alemany, R; Boucrot, J; Callot, O; Cordier, A; Courault, F; Davier, M; Duflot, L; Grivaz, J F; Heusse, P; Jacquet, M; Kim, D W; Le Diberder, F R; Lefrançois, J; Lutz, A M; Musolino, G; Nikolic, I A; Park, H J; Park, I C; Schune, M H; Simion, S; Veillet, J J; Videau, I; Abbaneo, D; Azzurri, P; Bagliesi, G; Batignani, G; Bettarini, S; Bozzi, C; Calderini, G; Carpinelli, M; Ciocci, M A; Ciulli, V; Dell'Orso, R; Fantechi, R; Ferrante, I; Foà, L; Forti, F; Giassi, A; Giorgi, M A; Gregorio, A; Ligabue, F; Lusiani, A; Marrocchesi, P S; Messineo, A; Rizzo, G; Sanguinetti, G; Sciabà, A; Spagnolo, P; Steinberger, Jack; Tenchini, Roberto; Tonelli, G; Triggiani, G; Vannini, C; Verdini, P G; Walsh, J; Betteridge, A P; Blair, G A; Bryant, L M; Cerutti, F; Gao, Y; Green, M G; Johnson, D L; Medcalf, T; Mir, L M; Perrodo, P; Strong, J A; Bertin, V; Botterill, David R; Clifft, R W; Edgecock, T R; Haywood, S; Edwards, M; Maley, P; Norton, P R; Thompson, J C; Bloch-Devaux, B; Colas, P; Emery, S; Kozanecki, Witold; Lançon, E; Lemaire, M C; Locci, E; Marx, B; Pérez, P; Rander, J; Renardy, J F; Roussarie, A; Schuller, J P; Schwindling, J; Trabelsi, A; Vallage, B; Johnson, R P; Kim, H Y; Litke, A M; McNeil, M A; Taylor, G; Beddall, A; Booth, C N; Boswell, R; Cartwright, S L; Combley, F; Dawson, I; Köksal, A; Letho, M; Newton, W M; Rankin, C; Thompson, L F; Böhrer, A; Brandt, S; Cowan, G D; Feigl, E; Grupen, Claus; Lutters, G; Minguet-Rodríguez, J A; Rivera, F; Saraiva, P; Smolik, L; Stephan, F; Apollonio, M; Bosisio, L; Della Marina, R; Giannini, G; Gobbo, B; Ragusa, F; Rothberg, J E; Wasserbaech, S R; Armstrong, S R; Bellantoni, L; Elmer, P; Feng, Z; Ferguson, D P S; Gao, Y S; González, S; Grahl, J; Harton, J L; Hayes, O J; Hu, H; McNamara, P A; Nachtman, J M; Orejudos, W; Pan, Y B; Saadi, Y; Schmitt, M; Scott, I J; Sharma, V; Turk, J; Walsh, A M; Wu Sau Lan; Wu, X; Yamartino, J M; Zheng, M; Zobernig, G

    1996-01-01

    A sample of 62249 \\tau-pair events is selected from data taken with the ALEPH detector in 1991, 1992 and 1993. The measurement of the branching fractions for \\tau decays into electrons and muons is presented with emphasis on the study of systematic effects from selection, particle identification and decay classification. Combined with the most recent ALEPH determination of the \\tau lifetime, these results provide a relative measurement of the leptonic couplings in the weak charged current for transverse W bosons.

  7. Holographic Coulomb branch vevs

    CERN Document Server

    Skenderis, K; Skenderis, Kostas; Taylor, Marika

    2006-01-01

    We compute holographically the vevs of all chiral primary operators for supergravity solutions corresponding to the Coulomb branch of N=4 SYM and find exact agreement with the corresponding field theory computation. Using the dictionary between 10d geometries and field theory developed to extract these vevs, we propose a gravity dual of a half supersymmetric deformation of N=4 SYM by certain irrelevant operators.

  8. Modeling branching in cereals

    OpenAIRE

    Jochem B. Evers; Vos, Jan

    2013-01-01

    Cereals and grasses adapt their structural development to environmental conditions and the resources available. The primary adaptive response is a variable degree of branching, called tillering in cereals. Especially for heterogeneous plant configurations the degree of tillering varies per plant. Functional–structural plant modeling (FSPM) is a modeling approach allowing simulation of the architectural development of individual plants, culminating in the emergent behavior at the canopy level....

  9. Analysis of knockout mutants reveals non-redundant functions of poly(ADP-ribose)polymerase isoforms in Arabidopsis.

    Science.gov (United States)

    Pham, Phuong Anh; Wahl, Vanessa; Tohge, Takayuki; de Souza, Laise Rosado; Zhang, Youjun; Do, Phuc Thi; Olas, Justyna J; Stitt, Mark; Araújo, Wagner L; Fernie, Alisdair R

    2015-11-01

    The enzyme poly(ADP-ribose)polymerase (PARP) has a dual function being involved both in the poly(ADP-ribosyl)ation and being a constituent of the NAD(+) salvage pathway. To date most studies, both in plant and non-plant systems, have focused on the signaling role of PARP in poly(ADP-ribosyl)ation rather than any role that can be ascribed to its metabolic function. In order to address this question we here used a combination of expression, transcript and protein localization studies of all three PARP isoforms of Arabidopsis alongside physiological analysis of the corresponding mutants. Our analyses indicated that whilst all isoforms of PARP were localized to the nucleus they are also present in non-nuclear locations with parp1 and parp3 also localised in the cytosol, and parp2 also present in the mitochondria. We next isolated and characterized insertional knockout mutants of all three isoforms confirming a complete knockout in the full length transcript levels of the target genes as well as a reduced total leaf NAD hydrolase activity in the two isoforms (PARP1, PARP2) that are highly expressed in leaves. Physiological evaluation of the mutant lines revealed that they displayed distinctive metabolic and root growth characteristics albeit unaltered leaf morphology under optimal growth conditions. We therefore conclude that the PARP isoforms play non-redundant non-nuclear metabolic roles and that their function is highly important in rapidly growing tissues such as the shoot apical meristem, roots and seeds. PMID:26428915

  10. Subcellular localization and responses of superoxide dismutase isoforms in local wheat varieties subjected to continuous soil drought.

    Science.gov (United States)

    Huseynova, Irada M; Aliyeva, Durna R; Aliyev, Jalal A

    2014-08-01

    Water is a key factor influencing the yield and quality of crops. One of the parameters of plant biological tolerance to constantly changing environmental conditions is the change of activities and numerous molecular forms of antioxidant enzymes. Two durum (Triticum durum Desf.) wheat varieties contrasting for drought tolerance, such as Barakatli-95 (drought tolerant) and Garagylchyg-2 (drought sensitive) were grown over a wide area in the field. Experiments were carried out to study the effect of soil drought on changes in activities and subcellular localization of superoxide dismutase isoforms. The levels of malondialdehyde, glycine betaine and total proteins were also analyzed. The level of the enzyme activity appeared to depend on the wheat varieties, duration of drought and stages of leaf development. Native polyacrylamide gel electrophoresis (PAGE) revealed the presence of 9 isoenzymes of superoxide dismutase in wheat leaves during drought. Mn-SOD was found in the mitochondrial fractions, Fe-SOD in the chloroplast fraction and Cu/Zn-SOD is localized in all subcellular fractions. Wheat leaves contain three different isoforms of SOD (Mn-, Fe-, Cu/Zn-SOD). Three isoforms of Mn-SOD, one isoform of Fe-SOD and five of Cu/Zn-SOD were observed in wheat leaves using 3 mM KCN and 5 mM H2O2 as selective inhibitors. The expression of Mn-SOD was preferentially enhanced by drought stress. It seems that Mn-SOD isoforms more than SOD ones play a major role in the scavenging of superoxide radicals. The observed data showed that status of antioxidant enzymes such as SOD could provide a meaningful tool for depicting drought tolerance of wheat genotype. PMID:24560039

  11. Epimorphin is a novel regulator of the progesterone receptor isoform-a.

    Science.gov (United States)

    Bascom, Jamie L; Radisky, Derek C; Koh, Eileen; Fata, Jimmie E; Lo, Alvin; Mori, Hidetoshi; Roosta, Neda; Hirai, Yohei; Bissell, Mina J

    2013-09-15

    Epimorphin/syntaxin-2 is a membrane-tethered protein localized extracellularly (Epim) and intracellularly (Stx-2). The extracellular form Epim stimulates morphogenic processes in a range of tissues, including in murine mammary glands where its overexpression in luminal epithelial cells is sufficient to drive hyperplasia and neoplasia. We analyzed WAP-Epim transgenic mice to gain insight into how Epim promotes malignancy. Ectopic overexpression of Epim during postnatal mammary gland development led to early side-branching onset, precocious bud formation, and increased proliferation of mammary epithelial cells. Conversely, peptide-based inhibition of Epim function reduced side branching. Because increased side branching and hyperplasia occurs similarly in mice upon overexpression of the progesterone receptor isoform-a (Pgr-a), we investigated whether Epim exhibits these phenotypes through Pgr modulation. Epim overexpression indeed led to a steep upregulation of both total Pgr mRNA and Pgr-a protein levels. Notably, the Pgr antagonist RU486 abrogated Epim-induced ductal side branching, mammary epithelial cell proliferation, and bud formation. Evaluation of Epim signaling in a three-dimensional ex vivo culture system showed that its action was dependent on binding to its extracellular receptor, integrin-αV, and on matrix metalloproteinase 3 activity downstream of Pgr-a. These findings elucidate a hitherto unknown transcriptional regulator of Pgr-a, and shed light on how overexpression of Epim leads to malignancy. PMID:23867473

  12. Localization and functional characterization of the human NKCC2 isoforms

    DEFF Research Database (Denmark)

    Carota, I; Theilig, F; Oppermann, M;

    2010-01-01

    inhibited by bumetanide than by furosemide. A sequence analysis of the amino acids encoded by exon 4 variants revealed high similarities between human and rodent NKCC2 isoforms, suggesting that differences in ion transport characteristics between species may be related to sequence variations outside the...... isoforms have specific localizations and transport characteristics, as assessed for rabbit, rat and mouse. In the present study, we aimed to address the localization and transport characteristics of the human NKCC2 isoforms. METHODS: RT-PCR, in situ hybridization and uptake studies in Xenopus oocytes were...... performed to characterize human NKCC2 isoforms. RESULTS: All three classical NKCC2 isoforms were detected in the human kidney; in addition, we found splice variants with tandem duplicates of the variable exon 4. Contrary to rodents, in which NKCC2F is the most abundant NKCC2 isoform, NKCC2A was the dominant...

  13. Tumorigenic properties of alternative osteopontin isoforms in mesothelioma

    Energy Technology Data Exchange (ETDEWEB)

    Ivanov, Sergey V., E-mail: Sergey.Ivanov@med.nyu.edu [Thoracic Surgery Laboratory, Cardiothoracic Surgery Department, NYU Langone Medical Center, 462 First Ave., Bellevue Hospital, Room 15N20, NY 10016 (United States); Ivanova, Alla V.; Goparaju, Chandra M.V.; Chen, Yuanbin; Beck, Amanda; Pass, Harvey I. [Thoracic Surgery Laboratory, Cardiothoracic Surgery Department, NYU Langone Medical Center, 462 First Ave., Bellevue Hospital, Room 15N20, NY 10016 (United States)

    2009-05-08

    Osteopontin (SPP1) is an inflammatory cytokine that we previously characterized as a diagnostic marker in patients with asbestos-induced malignant mesothelioma (MM). While SPP1 shows both pro- and anti-tumorigenic biological effects, little is known about the molecular basis of these activities. In this study, we demonstrate that while healthy pleura possesses all three differentially spliced SPP1 isoforms (A-C), in clinical MM specimens isoform A is markedly up-regulated and predominant. To provide a clue to possible functions of the SPP1 isoforms we next performed their functional evaluation via transient expression in MM cell lines. As a result, we report that isoforms A-C demonstrate different activities in cell proliferation, wound closure, and invasion assays. These findings suggest different functions for SPP1 isoforms and underline pro-tumorigenic properties of isoforms A and B.

  14. Starch branching enzymes and their genes in Sorghum

    OpenAIRE

    Mutisya, Joel

    2004-01-01

    Starch is an important raw material both for food and non-food purposes. It is synthesized and stored in source and sink tissues in plants. The starch deposited in amyloplasts of storage tissues possesses several physico-chemical properties, which makes it desirable for diverse applications. For industrial applications, starches high in either amylose or amylopectin are preferred in order to minimize chemical modification. The main purpose in our research is to understand starch synthesis in ...

  15. Method for altering starch using a microbial branching enzyme

    NARCIS (Netherlands)

    van der Maarel, Marc; Palomo Reixach, Marta; Booiman, Thijs; Dijkhuizen, Lubbert

    2010-01-01

    The present invention relates to methods for construction of pharmamers i.e. vaccine components characterized by their multimerization domain and the attached biologically active molecules, and their use in preparation of vaccines that contains the pharmamers alone or in combination with other molec

  16. A novel functional rabbit IL- 7 isoform

    OpenAIRE

    Siewe, Basile T.; Kalis, Susan L.; Esteves, Pedro J; Zhou, Tong; Knight, Katherine L.

    2010-01-01

    IL-7 is required for B cell development in mouse and is a key regulator of T cell development and peripheral T cell homeostasis in mouse and human. Recently, we found that IL-7 is expressed in rabbit bone marrow and in vitro, is required for differentiation of lymphoid progenitors to B and T lineage cells. Herein, we report the identification of a novel rabbit IL-7 isoform, IL-7II. Recombinant IL-7II (rIL-7II) binds lymphocytes via the IL-7R and induces phosphorylation of STAT5. Further, rIL-...

  17. Thermal Energy Conversion Branch

    Science.gov (United States)

    Bielozer, Matthew C.; Schreiber, Jeffrey, G.; Wilson, Scott D.

    2004-01-01

    The Thermal Energy Conversion Branch (5490) leads the way in designing, conducting, and implementing research for the newest thermal systems used in space applications at the NASA Glenn Research Center. Specifically some of the most advanced technologies developed in this branch can be broken down into four main areas: Dynamic Power Systems, Primary Solar Concentrators, Secondary Solar Concentrators, and Thermal Management. Work was performed in the Dynamic Power Systems area, specifically the Stirling Engine subdivision. Today, the main focus of the 5490 branch is free-piston Stirling cycle converters, Brayton cycle nuclear reactors, and heat rejection systems for long duration mission spacecraft. All space exploring devices need electricity to operate. In most space applications, heat energy from radioisotopes is converted to electrical power. The Radioisotope Thermoelectric Generator (RTG) already supplies electricity for missions such as the Cassini Spacecraft. The focus of today's Stirling research at GRC is aimed at creating an engine that can replace the RTG. The primary appeal of the Stirling engine is its high system efficiency. Because it is so efficient, the Stirling engine will significantly reduce the plutonium fuel mission requirements compared to the RTG. Stirling is also being considered for missions such as the lunar/Mars bases and rovers. This project has focused largely on Stirling Engines of all types, particularly the fluidyne liquid piston engine. The fluidyne was developed by Colin D. West. This engine uses the same concepts found in any type of Stirling engine, with the exception of missing mechanical components. All the working components are fluid. One goal was to develop and demonstrate a working Stirling Fluidyne Engine at the 2nd Annual International Energy Conversion Engineering Conference in Providence, Rhode Island.

  18. Combustion Branch Website Development

    Science.gov (United States)

    Bishop, Eric

    2004-01-01

    The NASA combustion branch is a leader in developing and applying combustion science to focused aerospace propulsion systems concepts. It is widely recognized for unique facilities, analytical tools, and personnel. In order to better communicate the outstanding research being done in this Branch to the public and other research organization, a more substantial website was desired. The objective of this project was to build an up-to-date site that reflects current research in a usable and attractive manner. In order to accomplish this, information was requested from all researchers in the Combustion branch, on their professional skills and on the current projects. This information was used to fill in the Personnel and Research sections of the website. A digital camera was used to photograph all personnel and these photographs were included in the personnel section as well. The design of the site was implemented using the latest web standards: xhtml and external css stylesheets. This implementation conforms to the guidelines recommended by the w3c. It also helps to ensure that the web site is accessible by disabled users, and complies with Section 508 Federal legislation (which mandates that all Federal websites be accessible). Graphics for the new site were generated using the gimp (www.gimp.org) an open-source graphics program similar to Adobe Photoshop. Also, all graphics on the site were of a reasonable size (less than 20k, most less than 2k) so that the page would load quickly. Technologies such as Macromedia Flash and Javascript were avoided, as these only function on some clients which have the proper software installed or enabled. The website was tested on different platforms with many different browsers to ensure there were no compatibility issues. The website was tested on windows with MS IE 6, MSIE 5 , Netscape 7, Mozilla and Opera. On a Mac, the site was tested with MS IE 5 , Netscape 7 and Safari.

  19. Androgen receptor isoforms in human and rat prostate

    Institute of Scientific and Technical Information of China (English)

    Shu-JieXIA; Gang-YaoHAO; Xiao-DaTANG

    2000-01-01

    Aim: To investigate the androgen receptor (AR) isoforms and its variability of expression in human and rat prostatic tissues. Methods: Human benign prostatic hyperplasia (BPH) and prostatic cancer tissues were obtained from patients undergoing prostatectomy, and rat ventral prostate was incised 3 days after castration. Forty-one AR-positive BPH specimens, 3 prostatic cancer specimens, and 6 rat prostates were used. After processing at 4℃, the tissues were examined by means of high resolution isoelectric focusing (IEF) technique to determine their AR isoforms. Results:From the prostatic specimens, 3 types of AR isoforms were detected with pI values at 6.5, 6.0, and 5.3. In human BPH tissues, 15/41 (36.6%) specimens showed all the three types of isoforms, while 19/41 (46.3%) showed 2 isoforms at various combinations and 7/41(17.1%), 1 isoform. For the 3 prostatic cancer specimens, one showed 3 isoforms, one, 2 isoforms, and the other failed to show any isoform. All rat prostatic tissues showed 2 isoforms at different combinations. Binding of 3H-dihydrotestosterone (DHT) to the isoforms was inhibited by the addition of 100-fold excess of DHT or testosterone, but not progesterone, oestradiol or diethylstilboestrol. Conclusion: AR isoforms are different in different patients. Although their genesis is not clear, the therapeutic implication of the present observation appears to be interesting, that may help clarifying the individual differences in the response to hormonal therapy.(Asian J Androl 2000 Dec;2:307-310)

  20. The Kinetics of Enzyme Mixtures

    Directory of Open Access Journals (Sweden)

    Simon Brown

    2014-03-01

    Full Text Available Even purified enzyme preparations are often heterogeneous. For example, preparations of aspartate aminotransferase or cytochrome oxidase can consist of several different forms of the enzyme. For this reason we consider how different the kinetics of the reactions catalysed by a mixture of forms of an enzyme must be to provide some indication of the characteristics of the species present. Based on the standard Michaelis-Menten model, we show that if the Michaelis constants (Km of two isoforms differ by a factor of at least 20 the steady-state kinetics can be used to characterise the mixture. However, even if heterogeneity is reflected in the kinetic data, the proportions of the different forms of the enzyme cannot be estimated from the kinetic data alone. Consequently, the heterogeneity of enzyme preparations is rarely reflected in measurements of their steady-state kinetics unless the species present have significantly different kinetic properties. This has two implications: (1 it is difficult, but not impossible, to detect molecular heterogeneity using kinetic data and (2 even when it is possible, a considerable quantity of high quality data is required.

  1. Identification of a novel TDRD7 isoforms

    Directory of Open Access Journals (Sweden)

    Filonenko V. V.

    2011-12-01

    Full Text Available The aim of our study was to investigate the tudor domain-containing protein 7 (TDRD7 subcellular localization, which could be linked to diverse functions of this protein within the cell. Methods. In this study we employed cell imaging technique for detecting TDRD7 subcellular localization, Western blot analysis of HEK293 cell fractions with anti-TDRD7 monoclonal antibodies and bioinformatical search of possible TDRD7 isoforms in Uniprot, Ensemble, UCSC databases. Results. We have observed specific TDRD7-containing structures in cytoplasm as well as in the nucleus in HEK293 cells. The Western blot analysis of subcellular fractions (cytoplasm, mitochondria, nucleus allowed us to detect three lower immunoreactive bands, with the aproximate molecular weight of 130, 110 and 60 kDa (we termed them as TDRD7, TDRD7 and TDRD7 and specific subcellular localization. The bioinformatical analysis of TDRD7 primary structure allowed us to determine two alternative transcripts from TDRD7 gene coding for proteins with calculated molecular weight of 130 and 60 kDa. Conclusion. The presented data demonstrate the existence at protein level of potential TDRD7 isoforms: TDRD7, TDRD7 and TDRD7. The expression profile of these splice variants and their role in cells remains to be elucidated.

  2. Compensatory expression and substrate inducibility of γ-glutamyl transferase GGT2 isoform in Arabidopsis thaliana

    OpenAIRE

    Destro, Tiziana; Prasad, Dinesh; Martignago, Damiano; Lliso Bernet, Ignacio; Trentin, Anna Rita; Renu, Indu Kumari; Ferretti, Massimo; Masi, Antonio

    2010-01-01

    γ-Glutamyl transferases (GGT; EC 2.3.2.2) are glutathione-degrading enzymes that are represented in Arabidopsis thaliana by a small gene family of four members. Two isoforms, GGT1 and GGT2, are apoplastic, sharing broad similarities in their amino acid sequences, but they are differently expressed in the tissues: GGT1 is expressed in roots, leaves, and siliques, while GGT2 was thought to be expressed only in siliques. It is demonstrated here that GGT2 is also expressed in wild-type roots, alb...

  3. Cooperation between two ClpB isoforms enhances the recovery of the recombinant β-galactosidase from inclusion bodies

    International Nuclear Information System (INIS)

    Highlights: ► An important role of synergistic cooperation between the two ClpB isoforms. ► Both ClpB isoforms are associated with IBs of β-galactosidase. ► ClpB is a key chaperone in IB protein release. -- Abstract: Bacterial ClpB is a molecular chaperone that solubilizes and reactivates aggregated proteins in cooperation with the DnaK chaperone system. The mechanism of protein disaggregation mediated by ClpB is linked to translocation of substrates through the central channel within the ring-hexameric structure of ClpB. Two isoforms of ClpB are produced in vivo: the full-length ClpB95 and the truncated ClpB80 (ClpBΔN), which does not contain the N-terminal domain. The functional specificity of the two ClpB isoforms and the biological role of the N-terminal domain are still not fully understood. Recently, it has been demonstrated that ClpB may achieve its full potential as an aggregate-reactivating chaperone through the functional interaction and synergistic cooperation of its two isoforms. It has been found that the most efficient resolubilization and reactivation of stress-aggregated proteins occurred in the presence of both ClpB95 and ClpB80. In this work, we asked if the two ClpB isoforms functionally cooperate in the solubilization and reactivation of proteins from insoluble inclusion bodies (IBs) in Escherichia coli cells. Using the model β-galactosidase fusion protein (VP1LAC), we found that solubilization and reactivation of enzymes entrapped in IBs occurred more efficiently in the presence of ClpB95 with ClpB80 than with either ClpB95 or ClpB80 alone. The two isoforms of ClpB chaperone acting together enhanced the solubility and enzymatic activity of β-galactosidase sequestered into IBs. Both ClpB isoforms were associated with IBs of β-galactosidase, what demonstrates their affinity to this type of aggregates. These results demonstrate a synergistic cooperation between the two isoforms of ClpB chaperone. In addition, no significant recovery of

  4. Cooperation between two ClpB isoforms enhances the recovery of the recombinant {beta}-galactosidase from inclusion bodies

    Energy Technology Data Exchange (ETDEWEB)

    Guenther, Izabela [Department of Biochemistry, University of Gdansk, Wita Stwosza 59, 80-308 Gdansk (Poland); Zolkiewski, Michal [Department of Biochemistry, Kansas State University, Manhattan, KS 66506 (United States); Kedzierska-Mieszkowska, Sabina, E-mail: kedzie@biotech.ug.gda.pl [Department of Biochemistry, University of Gdansk, Wita Stwosza 59, 80-308 Gdansk (Poland)

    2012-10-05

    Highlights: Black-Right-Pointing-Pointer An important role of synergistic cooperation between the two ClpB isoforms. Black-Right-Pointing-Pointer Both ClpB isoforms are associated with IBs of {beta}-galactosidase. Black-Right-Pointing-Pointer ClpB is a key chaperone in IB protein release. -- Abstract: Bacterial ClpB is a molecular chaperone that solubilizes and reactivates aggregated proteins in cooperation with the DnaK chaperone system. The mechanism of protein disaggregation mediated by ClpB is linked to translocation of substrates through the central channel within the ring-hexameric structure of ClpB. Two isoforms of ClpB are produced in vivo: the full-length ClpB95 and the truncated ClpB80 (ClpB{Delta}N), which does not contain the N-terminal domain. The functional specificity of the two ClpB isoforms and the biological role of the N-terminal domain are still not fully understood. Recently, it has been demonstrated that ClpB may achieve its full potential as an aggregate-reactivating chaperone through the functional interaction and synergistic cooperation of its two isoforms. It has been found that the most efficient resolubilization and reactivation of stress-aggregated proteins occurred in the presence of both ClpB95 and ClpB80. In this work, we asked if the two ClpB isoforms functionally cooperate in the solubilization and reactivation of proteins from insoluble inclusion bodies (IBs) in Escherichia coli cells. Using the model {beta}-galactosidase fusion protein (VP1LAC), we found that solubilization and reactivation of enzymes entrapped in IBs occurred more efficiently in the presence of ClpB95 with ClpB80 than with either ClpB95 or ClpB80 alone. The two isoforms of ClpB chaperone acting together enhanced the solubility and enzymatic activity of {beta}-galactosidase sequestered into IBs. Both ClpB isoforms were associated with IBs of {beta}-galactosidase, what demonstrates their affinity to this type of aggregates. These results demonstrate a synergistic

  5. Distinction between human cytochrome P450 (CYP) isoforms and identification of new phosphorylation sites by mass spectrometry

    DEFF Research Database (Denmark)

    Redlich, Gorden; Zanger, Ulrich M; Riedmaier, Stephan;

    2008-01-01

    establish that the isoforms CYP1A2, CYP2A6, CYP2B6, CYP2E1, CYP2C8, CYP2D6, CYP3A4, CYP3A7, and CYP8B1 are phosphorylated in vivo. These results will aid in further investigation of the functional significance of protein phosphorylation for this important group of enzymes....

  6. Peroxiredoxin isoforms are associated with cardiovascular risk factors in type 2 diabetes mellitus

    Energy Technology Data Exchange (ETDEWEB)

    El Eter, E. [Physiology Department, Faculty of Medicine, King Saud University, Riyadh (Saudi Arabia); Cardiovascular Research Group, Faculty of Medicine, King Saud University, Riyadh (Saudi Arabia); Physiology Department, Faculty of Medicine, Alexandria University, Alexandria (Egypt); Al-Masri, A.A. [Physiology Department, Faculty of Medicine, King Saud University, Riyadh (Saudi Arabia); Cardiovascular Research Group, Faculty of Medicine, King Saud University, Riyadh (Saudi Arabia)

    2015-03-03

    The production of oxygen free radicals in type 2 diabetes mellitus contributes to the development of complications, especially the cardiovascular-related ones. Peroxiredoxins (PRDXs) are antioxidant enzymes that combat oxidative stress. The aim of this study was to investigate the associations between the levels of PRDX isoforms (1, 2, 4, and 6) and cardiovascular risk factors in type 2 diabetes mellitus. Fifty-three patients with type 2 diabetes mellitus (28F/25M) and 25 healthy control subjects (7F/18M) were enrolled. We measured the plasma levels of each PRDX isoform and analyzed their correlations with cardiovascular risk factors. The plasma PRDX1, -2, -4, and -6 levels were higher in the diabetic patients than in the healthy control subjects. PRDX2 and -6 levels were negatively correlated with diastolic blood pressure, fasting blood sugar, and hemoglobin A1c. In contrast, PRDX1 levels were positively correlated with low-density lipoprotein and C-reactive protein levels. PRDX4 levels were negatively correlated with triglycerides. In conclusion, PRDX1, -2, -4, and -6 showed differential correlations with a variety of traditional cardiovascular risk factors. These results should encourage further research into the crosstalk between PRDX isoforms and cardiovascular risk factors.

  7. Peroxiredoxin isoforms are associated with cardiovascular risk factors in type 2 diabetes mellitus

    International Nuclear Information System (INIS)

    The production of oxygen free radicals in type 2 diabetes mellitus contributes to the development of complications, especially the cardiovascular-related ones. Peroxiredoxins (PRDXs) are antioxidant enzymes that combat oxidative stress. The aim of this study was to investigate the associations between the levels of PRDX isoforms (1, 2, 4, and 6) and cardiovascular risk factors in type 2 diabetes mellitus. Fifty-three patients with type 2 diabetes mellitus (28F/25M) and 25 healthy control subjects (7F/18M) were enrolled. We measured the plasma levels of each PRDX isoform and analyzed their correlations with cardiovascular risk factors. The plasma PRDX1, -2, -4, and -6 levels were higher in the diabetic patients than in the healthy control subjects. PRDX2 and -6 levels were negatively correlated with diastolic blood pressure, fasting blood sugar, and hemoglobin A1c. In contrast, PRDX1 levels were positively correlated with low-density lipoprotein and C-reactive protein levels. PRDX4 levels were negatively correlated with triglycerides. In conclusion, PRDX1, -2, -4, and -6 showed differential correlations with a variety of traditional cardiovascular risk factors. These results should encourage further research into the crosstalk between PRDX isoforms and cardiovascular risk factors

  8. A novel FADS1 isoform potentiates FADS2-mediated production of eicosanoid precursor fatty acids.

    Science.gov (United States)

    Park, Woo Jung; Kothapalli, Kumar S D; Reardon, Holly T; Lawrence, Peter; Qian, Shu-Bing; Brenna, J Thomas

    2012-08-01

    The fatty acid desaturase (FADS) genes code for the rate-limiting enzymes required for the biosynthesis of long-chain polyunsaturated fatty acids (LCPUFA). Here we report discovery and function of a novel FADS1 splice variant. FADS1 alternative transcript 1 (FADS1AT1) enhances desaturation of FADS2, leading to increased production of eicosanoid precursors, the first case of an isoform modulating the enzymatic activity encoded by another gene. Multiple protein isoforms were detected in primate liver, thymus, and brain. In human neuronal cells, their expression patterns are modulated by differentiation and result in alteration of cellular fatty acids. FADS1, but not FADS1AT1, localizes to endoplasmic reticulum and mitochondria. Ribosomal footprinting demonstrates that all three FADS genes are translated at similar levels. The noncatalytic regulation of FADS2 desaturation by FADS1AT1 is a novel, plausible mechanism by which several phylogenetically conserved FADS isoforms may regulate LCPUFA biosynthesis in a manner specific to tissue, organelle, and developmental stage. PMID:22619218

  9. Changes in the expression of NO synthase isoforms after ozone: the effects of allergen exposure

    Directory of Open Access Journals (Sweden)

    Lee June-Hyuk

    2004-06-01

    Full Text Available Abstract Background The functional role of nitric oxide (NO and various nitric oxide synthase (NOS isoforms in asthma remains unclear. Objective This study investigated the effects of ozone and ovalbumin (OVA exposure on NOS isoforms. Methods The expression of inducible NOS (iNOS, neuronal NOS (nNOS, and endothelial NOS (eNOS in lung tissue was measured. Enhanced pause (Penh was measured as a marker of airway obstruction. Nitrate and nitrite in bronchoalveolar lavage (BAL fluid were measured using a modified Griess reaction. Results The nitrate concentration in BAL fluid from the OVA-sensitized/ozone-exposed/OVA-challenged group was greater than that of the OVA-sensitized/saline-challenged group. Methacholine-induced Penh was increased in the OVA-sensitized/ozone-exposed/OVA-challenged group, with a shift in the dose-response curve to the left, compared with the OVA-sensitized/saline-challenged group. The levels of nNOS and eNOS were increased significantly in the OVA-sensitized/ozone-exposed/OVA-challenged group and the iNOS levels were reduced compared with the OVA-sensitized/saline-challenged group. Conclusion In mice, ozone is associated with increases in lung eNOS and nNOS, and decreases in iNOS. None of these enzymes are further affected by allergens, suggesting that the NOS isoforms play different roles in airway inflammation after ozone exposure.

  10. Path-valued branching processes and nonlocal branching superprocesses

    CERN Document Server

    Li, Zenghu

    2012-01-01

    A family of continuous-state branching processes with immigration are constructed as the solution flow of a stochastic equation system driven by time-space noises. The family can be regarded as an inhomogeneous increasing path-valued branching process with immigration. Two nonlocal branching immigration superprocesses can be defined from the flow. We identify explicitly the branching and immigration mechanisms of those processes. The results provide new perspectives into the tree-valued Markov processes of Aldous and Pitman [Ann. Inst. H. Poincare Probab. Statist. 34 (1998), 637--686] and Abraham and Delmas [Ann. Probab. To appear].

  11. Maritime Branch Analysis

    DEFF Research Database (Denmark)

    Mougaard, Krestine; Neugebauer, Line Maria; Garcia i Mateu, Adrià;

    after-sales service and have therefore embarked on business development activities that tightly combine product and service offerings in their portfolios. Closer customer contact, commoditisation of goods, total cost of ownership, and product liability are just some of the reasons for this transition......The vast majority of countries in the developed world are now dependent on their service sectors for between 70-80% of their gross domestic product. Even companies with decades of expertise in producing manufactured products are experiencing an increased need to understand before-, during- and...... and influencing a move towards intensified service integration into business- and product development activities. The workbook has been created by condensing the transcripts and the insights we have gained from a wealth of studies across the maritime branch, and it serves to provide a unique insight...

  12. Quiver Varieties and Branching

    Directory of Open Access Journals (Sweden)

    Hiraku Nakajima

    2009-01-01

    Full Text Available Braverman and Finkelberg recently proposed the geometric Satake correspondence for the affine Kac-Moody group Gaff [Braverman A., Finkelberg M., arXiv:0711.2083]. They conjecture that intersection cohomology sheaves on the Uhlenbeck compactification of the framed moduli space of Gcpt-instantons on $R^4/Z_r$ correspond to weight spaces of representations of the Langlands dual group $G_{aff}^{vee}$ at level $r$. When $G = SL(l$, the Uhlenbeck compactification is the quiver variety of type $sl(r_{aff}$, and their conjecture follows from the author's earlier result and I. Frenkel's level-rank duality. They further introduce a convolution diagram which conjecturally gives the tensor product multiplicity [Braverman A., Finkelberg M., Private communication, 2008]. In this paper, we develop the theory for the branching in quiver varieties and check this conjecture for $G = SL(l$.

  13. Transcriptional profiles of glutathione-S-Transferase isoforms, Cyp, and AOE genes in atrazine-exposed zebrafish embryos.

    Science.gov (United States)

    Glisic, Branka; Hrubik, Jelena; Fa, Svetlana; Dopudj, Nela; Kovacevic, Radmila; Andric, Nebojsa

    2016-02-01

    Glutathione-S-transferase (GST) superfamily consists of multiple members involved in xenobiotic metabolism. Expressional pattern of the GST isoforms in adult fish has been used as a biomarker of exposure to environmental chemicals. However, GST transcriptional responses vary across organs, thus requiring a cross-tissue examination of multiple mRNAs for GST profiling in an animal after chemical exposure. Zebrafish embryos express all GST isoforms as adult fish and could therefore represent an alternative model for identification of biomarkers of exposure. To evaluate such a possibility, we studied a set of cytosolic and microsomal GST isoform-specific expression profiles in the zebrafish embryos after exposure to atrazine, a widely used herbicide. Expression of the GST isoforms was compared with that of CYP genes involved in the phase I of xenobiotic metabolism and antioxidant enzyme (AOE) genes. Using quantitative real-time PCR, we showed dynamic changes in the expressional pattern of twenty GST isoforms, cyp1a, cyp3a65, ahr2, and four AOEs in early development of zebrafish. Acute (48 and 72 h) exposure of 24 h-old embryos to atrazine, from environmentally relevant (0.005 mg/L) to high (40 mg/L) concentrations, caused a variety of transient, albeit minor changes (GST isoforms, ahr2 and AOE genes response. However, expression of cyp1a and cyp3a65 mRNA was markedly and consistently induced by high doses of atrazine (5 and 40 mg/L). In summary, an analysis of the response of multiple systems in the zebrafish embryos provided a comprehensive understanding of atrazine toxicity and its potential impact on biological processes. PMID:25158112

  14. Spinach pyruvate kinase isoforms: partial purification and regulatory properties

    Energy Technology Data Exchange (ETDEWEB)

    Baysdorfer, C.; Bassham, J.A.

    1984-02-01

    Pyruvate kinase from spinach (Spinacea oleracea L.) leaves consists of two isoforms, separable by blue agarose chromatography. Both isoforms share similar pH profiles and substrate and alternate nucleotide K/sub m/ values. In addition, both isoforms are inhibited by oxalate and ATP and activated by AMP. The isoforms differ in their response to three key metabolites; citrate, aspartate, and glutamate. The first isoform is similar to previously reported plant pyruvate kinases in its sensitivity to citrate inhibition. The K/sub i/ for this inhibition is 1.2 millimolar citrate. The second isoform is not affected by citrate but is regulated by aspartate and glutamate. Aspartate is an activator with a K/sub a/ of 0.05 millimolar, and glutamate is an inhibitor with a K/sub i/ of 0.68 millimolar. A pyruvate kinase with these properties has not been previously reported. Based on these considerations, the authors suggest that the activity of the first isoform is regulated by respiratory metabolism. The second isoform, in contrast, may be regulated by the demand for carbon skeletons for use in ammonia assimilation.

  15. Prokaryotic expression and characterization of a pea actin isoform (PEAcl) fused to GFP

    Institute of Scientific and Technical Information of China (English)

    ZHANG Shaobin; REN Dongtao; XU Xiaojing; LIU Guoqin

    2004-01-01

    Actins widely exist in eukaryotic cells and play important roles in many living activities. As there are many kinds of actin isoforms in plant cells, it is difficult to purify each actin isoform in sufficient quantities for analyzing its physicochemical properties. In the present study, a pea(Pisum Sativum L.) actin isoform (PEAc1) fused to His-tag at its amino terminus and GFP (green fluorescent protein) at its Carboxyl terminus were expressed in E. Coli in inclusion bodies. The fusion protein (PEAc1-GFP) was highly purified with the yield of above 2 mg/L culture by dissolving inclusions in 8 mol/L urea, renaturing by dialysis in a gradient of urea, and affinity binding to Ni-resin. The purified mono meric PEAc1-GFP could efficiently bind on Dnase Ⅰ and inhibit the latter's enzyme activity. PEAc1-GFP could polymerize into green fluorescent filamentous structures (F-PEAc1-GFP), which could be labeled by TRITC-phalloidin, a specific agent for observing microfilaments. The PEAc1-GFP polymerization curve was identical with that of chicken skeletal muscle actin. The critical concentration for PEAc1-GFP to polymerize into filaments is 0.24 μmol/L. The F-PEAc1-GFP could stimulate myosin Mg-ATPase activity in a protein concentration dependant manner (about 4 folds at1 mg/mL F-PEAc1-GFP). The results above show that the PEAcl fused to GFP retained the assembly characteristic of actin, indicating that gene fusion, prokaryotic expression,denaturation and renaturation, and affinity chromatography is a useful strategy for obtaining plant actin isoform proteins in a large amount.

  16. Isolation of a human myocardial cytosolic phospholipase A2 isoform. Fast atom bombardment mass spectroscopic and reverse-phase high pressure liquid chromatography identification of choline and ethanolamine glycerophospholipid substrates.

    OpenAIRE

    Hazen, S. L.; Hall, C. R.; Ford, D. A.; Gross, R W

    1993-01-01

    Recent studies have demonstrated the existence of a novel family of calcium-independent plasmalogen-selective phospholipases A2 in canine myocardium that have been implicated as enzymic mediators of ischemic membrane damage. We now report that human myocardium contains two functionally distinct isoforms of cytosolic calcium-independent phospholipase A2. The major cytosolic phospholipase A2 isoform preferentially hydrolyzes plasmalogen substrate, possesses a pH optimum of 7.0, and is chromatog...

  17. Methods and Technologies Branch (MTB)

    Science.gov (United States)

    The Methods and Technologies Branch focuses on methods to address epidemiologic data collection, study design and analysis, and to modify technological approaches to better understand cancer susceptibility.

  18. Can the branching exponent reliably relate the branching indexes?

    Czech Academy of Sciences Publication Activity Database

    Netopilík, Miloš

    2015-01-01

    Roč. 24, č. 2 (2015), s. 80-84. ISSN 1022-1344 R&D Projects: GA ČR(CZ) GA13-02938S Institutional support: RVO:61389013 Keywords : branching exponent * branching indexes * intrinsic viscosity Subject RIV: CD - Macromolecular Chemistry Impact factor: 1.667, year: 2014

  19. Branched-chain 2-keto acid decarboxylases derived from Psychrobacter.

    Science.gov (United States)

    Wei, Jiashi; Timler, Jacobe G; Knutson, Carolann M; Barney, Brett M

    2013-09-01

    The conversion of branched-chain amino acids to branched-chain acids or alcohols is an important aspect of flavor in the food industry and is dependent on the Ehrlich pathway found in certain lactic acid bacteria. A key enzyme in the pathway, the 2-keto acid decarboxylase (KDC), is also of interest in biotechnology applications to produce small branched-chain alcohols that might serve as improved biofuels or other commodity feedstocks. This enzyme has been extensively studied in the model bacterium Lactococcus lactis, but is also found in other bacteria and higher organisms. In this report, distinct homologs of the L. lactis KDC originally annotated as pyruvate decarboxylases from Psychrobacter cryohalolentis K5 and P. arcticus 273-4 were cloned and characterized, confirming a related activity toward specific branched-chain 2-keto acids derived from branched-chain amino acids. Further, KDC activity was confirmed in intact cells and cell-free extracts of P. cryohalolentis K5 grown on both rich and defined media, indicating that the Ehrlich pathway may also be utilized in some psychrotrophs and psychrophiles. A comparison of the similarities and differences in the P. cryohalolentis K5 and P. arcticus 273-4 KDC activities to other bacterial KDCs is presented. PMID:23826991

  20. Expression, purification and enzymatic characterization of the catalytic domains of human tryptophan hydroxylase isoforms

    DEFF Research Database (Denmark)

    Windahl, Michael Skovbo; Boesen, Jane; Karlsen, Pernille Efferbach; Christensen, Hans Erik Mølager

    Tryptophan hydroxylase exists in two isoforms: Isoform 1 catalyses the first and rate-limiting step in the synthesis of serotonin in the peripheral parts of the body while isoform 2 catalyses this step in the brain. The catalytic domains of human tryptophan hydroxylase 1 and 2 have been expressed......, purified and the kinetic properties have been studied and are compared. Substrate inhibition by tryptophan is observed for isoform 1 but not for isoform 2. Large differences are observed in the K m,tetrahydrobiopterin values for the two isoforms, being >10 times larger for isoform 1 compared to isoform 2....

  1. Tau hadronic branching ratios

    CERN Document Server

    Buskulic, Damir; De Bonis, I; Décamp, D; Ghez, P; Goy, C; Lees, J P; Lucotte, A; Minard, M N; Odier, P; Pietrzyk, B; Ariztizabal, F; Chmeissani, M; Crespo, J M; Efthymiopoulos, I; Fernández, E; Fernández-Bosman, M; Gaitan, V; Martínez, M; Orteu, S; Pacheco, A; Padilla, C; Palla, Fabrizio; Pascual, A; Perlas, J A; Sánchez, F; Teubert, F; Colaleo, A; Creanza, D; De Palma, M; Farilla, A; Gelao, G; Girone, M; Iaselli, Giuseppe; Maggi, G; Maggi, M; Marinelli, N; Natali, S; Nuzzo, S; Ranieri, A; Raso, G; Romano, F; Ruggieri, F; Selvaggi, G; Silvestris, L; Tempesta, P; Zito, G; Huang, X; Lin, J; Ouyang, Q; Wang, T; Xie, Y; Xu, R; Xue, S; Zhang, J; Zhang, L; Zhao, W; Bonvicini, G; Cattaneo, M; Comas, P; Coyle, P; Drevermann, H; Engelhardt, A; Forty, Roger W; Frank, M; Hagelberg, R; Harvey, J; Jacobsen, R; Janot, P; Jost, B; Kneringer, E; Knobloch, J; Lehraus, Ivan; Markou, C; Martin, E B; Mato, P; Minten, Adolf G; Miquel, R; Oest, T; Palazzi, P; Pater, J R; Pusztaszeri, J F; Ranjard, F; Rensing, P E; Rolandi, Luigi; Schlatter, W D; Schmelling, M; Schneider, O; Tejessy, W; Tomalin, I R; Venturi, A; Wachsmuth, H W; Wiedenmann, W; Wildish, T; Witzeling, W; Wotschack, J; Ajaltouni, Ziad J; Bardadin-Otwinowska, Maria; Barrès, A; Boyer, C; Falvard, A; Gay, P; Guicheney, C; Henrard, P; Jousset, J; Michel, B; Monteil, S; Pallin, D; Perret, P; Podlyski, F; Proriol, J; Rossignol, J M; Saadi, F; Fearnley, Tom; Hansen, J B; Hansen, J D; Hansen, J R; Hansen, P H; Nilsson, B S; Kyriakis, A; Simopoulou, Errietta; Siotis, I; Vayaki, Anna; Zachariadou, K; Blondel, A; Bonneaud, G R; Brient, J C; Bourdon, P; Passalacqua, L; Rougé, A; Rumpf, M; Tanaka, R; Valassi, Andrea; Verderi, M; Videau, H L; Candlin, D J; Parsons, M I; Focardi, E; Parrini, G; Corden, M; Delfino, M C; Georgiopoulos, C H; Jaffe, D E; Antonelli, A; Bencivenni, G; Bologna, G; Bossi, F; Campana, P; Capon, G; Chiarella, V; Felici, G; Laurelli, P; Mannocchi, G; Murtas, F; Murtas, G P; Pepé-Altarelli, M; Dorris, S J; Halley, A W; ten Have, I; Knowles, I G; Lynch, J G; Morton, W T; O'Shea, V; Raine, C; Reeves, P; Scarr, J M; Smith, K; Smith, M G; Thompson, A S; Thomson, F; Thorn, S; Turnbull, R M; Becker, U; Braun, O; Geweniger, C; Graefe, G; Hanke, P; Hepp, V; Kluge, E E; Putzer, A; Rensch, B; Schmidt, M; Sommer, J; Stenzel, H; Tittel, K; Werner, S; Wunsch, M; Beuselinck, R; Binnie, David M; Cameron, W; Colling, D J; Dornan, Peter J; Konstantinidis, N P; Moneta, L; Moutoussi, A; Nash, J; San Martin, G; Sedgbeer, J K; Stacey, A M; Dissertori, G; Girtler, P; Kuhn, D; Rudolph, G; Bowdery, C K; Brodbeck, T J; Colrain, P; Crawford, G; Finch, A J; Foster, F; Hughes, G; Sloan, Terence; Whelan, E P; Williams, M I; Galla, A; Greene, A M; Kleinknecht, K; Quast, G; Raab, J; Renk, B; Sander, H G; Wanke, R; Van Gemmeren, P; Zeitnitz, C; Aubert, Jean-Jacques; Bencheikh, A M; Benchouk, C; Bonissent, A; Bujosa, G; Calvet, D; Carr, J; Diaconu, C A; Etienne, F; Thulasidas, M; Nicod, D; Payre, P; Rousseau, D; Talby, M; Abt, I; Assmann, R W; Bauer, C; Blum, Walter; Brown, D; Dietl, H; Dydak, Friedrich; Ganis, G; Gotzhein, C; Jakobs, K; Kroha, H; Lütjens, G; Lutz, Gerhard; Männer, W; Moser, H G; Richter, R H; Rosado-Schlosser, A; Schael, S; Settles, Ronald; Seywerd, H C J; Saint-Denis, R; Wolf, G; Alemany, R; Boucrot, J; Callot, O; Cordier, A; Courault, F; Davier, M; Duflot, L; Grivaz, J F; Heusse, P; Jacquet, M; Kim, D W; Le Diberder, F R; Lefrançois, J; Lutz, A M; Musolino, G; Nikolic, I A; Park, H J; Park, I C; Schune, M H; Simion, S; Veillet, J J; Videau, I; Abbaneo, D; Azzurri, P; Bagliesi, G; Batignani, G; Bettarini, S; Bozzi, C; Calderini, G; Carpinelli, M; Ciocci, M A; Ciulli, V; Dell'Orso, R; Fantechi, R; Ferrante, I; Foà, L; Forti, F; Giassi, A; Giorgi, M A; Gregorio, A; Ligabue, F; Lusiani, A; Marrocchesi, P S; Messineo, A; Rizzo, G; Sanguinetti, G; Sciabà, A; Spagnolo, P; Steinberger, Jack; Tenchini, Roberto; Tonelli, G; Triggiani, G; Vannini, C; Verdini, P G; Walsh, J; Betteridge, A P; Blair, G A; Bryant, L M; Cerutti, F; Gao, Y; Green, M G; Johnson, D L; Medcalf, T; Mir, L M; Perrodo, P; Strong, J A; Bertin, V; Botterill, David R; Clifft, R W; Edgecock, T R; Haywood, S; Edwards, M; Maley, P; Norton, P R; Thompson, J C; Bloch-Devaux, B; Colas, P; Emery, S; Kozanecki, Witold; Lançon, E; Lemaire, M C; Locci, E; Marx, B; Pérez, P; Rander, J; Renardy, J F; Roussarie, A; Schuller, J P; Schwindling, J; Trabelsi, A; Vallage, B; Johnson, R P; Kim, H Y; Litke, A M; McNeil, M A; Taylor, G; Beddall, A; Booth, C N; Boswell, R; Cartwright, S L; Combley, F; Dawson, I; Köksal, A; Letho, M; Newton, W M; Rankin, C; Thompson, L F; Böhrer, A; Brandt, S; Cowan, G D; Feigl, E; Grupen, Claus; Lutters, G; Minguet-Rodríguez, J A; Rivera, F; Saraiva, P; Smolik, L; Stephan, F; Apollonio, M; Bosisio, L; Della Marina, R; Giannini, G; Gobbo, B; Ragusa, F; Rothberg, J E; Wasserbaech, S R; Armstrong, S R; Bellantoni, L; Elmer, P; Feng, Z; Ferguson, D P S; Gao, Y S; González, S; Grahl, J; Harton, J L; Hayes, O J; Hu, H; McNamara, P A; Nachtman, J M; Orejudos, W; Pan, Y B; Saadi, Y; Schmitt, M; Scott, I J; Sharma, V; Turk, J; Walsh, A M; Wu Sau Lan; Wu, X; Yamartino, J M; Zheng, M; Zobernig, G

    1996-01-01

    From 64492 selected \\tau-pair events, produced at the Z^0 resonance, the measurement of the tau decays into hadrons from a global analysis using 1991, 1992 and 1993 ALEPH data is presented. Special emphasis is given to the reconstruction of photons and \\pi^0's, and the removal of fake photons. A detailed study of the systematics entering the \\pi^0 reconstruction is also given. A complete and consistent set of tau hadronic branching ratios is presented for 18 exclusive modes. Most measurements are more precise than the present world average. The new level of precision reached allows a stringent test of \\tau-\\mu universality in hadronic decays, g_\\tau/g_\\mu \\ = \\ 1.0013 \\ \\pm \\ 0.0095, and the first measurement of the vector and axial-vector contributions to the non-strange hadronic \\tau decay width: R_{\\tau ,V} \\ = \\ 1.788 \\ \\pm \\ 0.025 and R_{\\tau ,A} \\ = \\ 1.694 \\ \\pm \\ 0.027. The ratio (R_{\\tau ,V} - R_{\\tau ,A}) / (R_{\\tau ,V} + R_{\\tau ,A}), equal to (2.7 \\pm 1.3) \\ \\%, is a measure of the importance of Q...

  2. Salicylate, a catalytic inhibitor of topoisomerase II, inhibits DNA cleavage and is selective for the α isoform.

    Science.gov (United States)

    Bau, Jason T; Kang, Zhili; Austin, Caroline A; Kurz, Ebba U

    2014-02-01

    Topoisomerase II (topo II) is a ubiquitous enzyme that is essential for cell survival through its role in regulating DNA topology and chromatid separation. Topo II can be poisoned by common chemotherapeutics (such as doxorubicin and etoposide), leading to the accumulation of cytotoxic enzyme-linked DNA double-stranded breaks. In contrast, nonbreak-inducing topo II catalytic inhibitors have also been described and have more limited use in clinical chemotherapy. These agents, however, may alter the efficacy of regimens incorporating topo II poisons. We previously identified salicylate, the primary metabolite of aspirin, as a novel catalytic inhibitor of topo II. We have now determined the mechanism by which salicylate inhibits topo II. As catalytic inhibitors can act at a number of steps in the topo II catalytic cycle, we used multiple independent, biochemical approaches to interrogate the catalytic cycle. Furthermore, as mammalian cells express two isoforms of topo II (α and β), we examined whether salicylate was isoform selective. Our results demonstrate that salicylate is unable to intercalate DNA, and does not prevent enzyme-DNA interaction, nor does it promote stabilization of topo IIα in closed clamps on DNA. Although salicylate decreased topo IIα ATPase activity in a dose-dependent noncompetitive manner, this was secondary to salicylate-mediated inhibition of DNA cleavage. Surprisingly, comparison of salicylate's effects using purified human topo IIα and topo IIβ revealed that salicylate selectively inhibits the α isoform. These findings provide a definitive mechanism for salicylate-mediated inhibition of topo IIα and provide support for further studies determining the basis for its isoform selectivity. PMID:24220011

  3. Differential role of human choline kinase alpha and beta enzymes in lipid metabolism: implications in cancer onset and treatment.

    Directory of Open Access Journals (Sweden)

    David Gallego-Ortega

    Full Text Available BACKGROUND: The Kennedy pathway generates phosphocoline and phosphoethanolamine through its two branches. Choline Kinase (ChoK is the first enzyme of the Kennedy branch of synthesis of phosphocholine, the major component of the plasma membrane. ChoK family of proteins is composed by ChoKalpha and ChoKbeta isoforms, the first one with two different variants of splicing. Recently ChoKalpha has been implicated in the carcinogenic process, since it is over-expressed in a variety of human cancers. However, no evidence for a role of ChoKbeta in carcinogenesis has been reported. METHODOLOGY/PRINCIPAL FINDINGS: Here we compare the in vitro and in vivo properties of ChoKalpha1 and ChoKbeta in lipid metabolism, and their potential role in carcinogenesis. Both ChoKalpha1 and ChoKbeta showed choline and ethanolamine kinase activities when assayed in cell extracts, though with different affinity for their substrates. However, they behave differentially when overexpressed in whole cells. Whereas ChoKbeta display an ethanolamine kinase role, ChoKalpha1 present a dual choline/ethanolamine kinase role, suggesting the involvement of each ChoK isoform in distinct biochemical pathways under in vivo conditions. In addition, while overexpression of ChoKalpha1 is oncogenic when overexpressed in HEK293T or MDCK cells, ChoKbeta overexpression is not sufficient to induce in vitro cell transformation nor in vivo tumor growth. Furthermore, a significant upregulation of ChoKalpha1 mRNA levels in a panel of breast and lung cancer cell lines was found, but no changes in ChoKbeta mRNA levels were observed. Finally, MN58b, a previously described potent inhibitor of ChoK with in vivo antitumoral activity, shows more than 20-fold higher efficiency towards ChoKalpha1 than ChoKbeta. CONCLUSION/SIGNIFICANCE: This study represents the first evidence of the distinct metabolic role of ChoKalpha and ChoKbeta isoforms, suggesting different physiological roles and implications in human

  4. Uptake of Branched-Chain Amino Acids by Streptococcus thermophilus

    OpenAIRE

    Akpemado, K. M.; Bracquart, P. A.

    1983-01-01

    The transport of branched-chain amino acids in Streptococcus thermophilus was energy dependent. The metabolic inhibitors of glycolysis and ATPase enzymes were active, but the proton-conducting uncouplers were not. Transport was optimal at temperatures of between 30 and 45°C and at pH 7.0 for the three amino acids leucine, valine, and isoleucine; a second peak existed at pH 5.0 with valine and isoleucine. By competition and kinetics studies, the branched-chain amino acids were found to share a...

  5. Subunit-specific sulphation of oligosaccharides relating to charge-heterogeneity in porcine lutrophin isoforms.

    Science.gov (United States)

    Ujihara, M; Yamamoto, K; Nomura, K; Toyoshima, S; Demura, H; Nakamura, Y; Ohmura, K; Osawa, T

    1992-06-01

    Lutrophin (LH) consists of an array of isoforms with different charges and bioactivities. This study was undertaken to clarify specifically how oligosaccharides of alpha and beta subunits contribute to LH isoform charges. Porcine LH (pLH) was separated into four isoforms by isoelectric focusing (IEF), followed by subunit isolation. Their oligosaccharides were released by hydrazinolysis, labelled by reduction with NaB3H4, and fractionated by HPLC with a Mono Q column into five populations differing in the number of sulphate (S) and sialic acid (N) residues, designated as Neutral, N-1, S-1, S-N and S-2. Oligosaccharides were predominantly sulphated (S-1 and S-2) and infrequently sialylated (N-1 and S-N). Further analysis, including concanavalin A (Con A) affinity chromatography, desialylation, desulphation, sequential exoglycosidase digestion and methylation, clarified the structures of the acidic oligosaccharides. All were of the biantennary complex type. Their two peripheral branches were SO4-4GalNAc beta 1-4Glc-NAc and GalNAc beta 1-4GlcNAc or GlcNAc in S-1, SO4-4GalNAc beta 1-4GlcNAc and Sia alpha 2-6Gal beta 1-4GlcNAc in S-N, and (SO4-4GalNAc beta 1-4GlcNAc)2 in S-2 (where GalNAc is N-acetylgalactosamine and GlcNAc is N-acetylglucosamine). Ten percent of S-1 and of S-N had a bisecting GlcNAc residue. Sulphate residues occurred in nearly the same amount for both subunits; however, the alpha and beta subunits were sulphated differently. S-1 predominated in the alpha subunit, while S-1 and S-2 were major components in the beta subunit.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:1498420

  6. Targeted Mutations in the Na,K-ATPase Alpha 2 Isoform Confer Ouabain Resistance and Result in Abnormal Behavior in Mice

    OpenAIRE

    Schaefer, Tori L.; Lingrel, Jerry B; Moseley, Amy E.; Vorhees, Charles V.; Williams, Michael T.

    2010-01-01

    Sodium and potassium-activated adenosine triphosphatases (Na,K-ATPase) are ubiquitous, participate in osmotic balance and membrane potential, and are composed of α, β, and γ subunits. The α subunit is required for the catalytic and transport properties of the enzyme and contains binding sites for cations, ATP, and digitalis-like compounds including ouabain. There are four known α isoforms; three that are expressed in the CNS in a regional and cell-specific manner. The α2 isoform is most commo...

  7. Enzyme immunoassay

    DEFF Research Database (Denmark)

    Feldt-Rasmussen, B; Dinesen, B; Deckert, M

    1985-01-01

    An enzyme linked immunoadsorbent assay for urinary albumin using commercially available reagents is described. The assay range is 2.5-120 micrograms/l. When samples are analysed in two standard dilutions, the assayable albumin concentration range is 2.5-240 mg/l, covering the clinical range from...

  8. Food Enzymes

    Science.gov (United States)

    McBroom, Rachel; Oliver-Hoyo, Maria T.

    2007-01-01

    Many students view biology and chemistry as two unrelated, separate sciences; how these courses are generally taught in high schools may do little to change that impression. The study of enzymes provide a great opportunity for both biology and chemistry teachers to share with students the interdisciplinary nature of science. This article describes…

  9. p53 Family: Role of Protein Isoforms in Human Cancer

    Directory of Open Access Journals (Sweden)

    Jinxiong Wei

    2012-01-01

    Full Text Available TP53, TP63, and TP73 genes comprise the p53 family. Each gene produces protein isoforms through multiple mechanisms including extensive alternative mRNA splicing. Accumulating evidence shows that these isoforms play a critical role in the regulation of many biological processes in normal cells. Their abnormal expression contributes to tumorigenesis and has a profound effect on tumor response to curative therapy. This paper is an overview of isoform diversity in the p53 family and its role in cancer.

  10. Catabolism of Branched Chain Amino Acids Supports Respiration but Not Volatile Synthesis in Tomato Fruits

    Institute of Scientific and Technical Information of China (English)

    Andrej Kochevenko; Wagner L.Araújo; Gregory S.Maloney; Denise M.Tieman; Phuc Thi Do; Mark G.Taylor; Harry J.Klee; Alisdair R.Fernie

    2012-01-01

    The branched-chain amino acid transaminases (BCATs) have a crucial role in metabolism of the branched-chain amino acids leucine,isoleucine,and valine.These enzymes catalyze the last step of synthesis and the initial step of degradation of these amino acids.Although the biosynthetic pathways of branched chain amino acids in plants have been extensively investigated and a number of genes have been characterized,their catabolism in plants is not yet completely understood.We previously characterized the branched chain amino acid transaminase gene family in tomato,revealing both the subcellular localization and kinetic properties of the enzymes encoded by six genes.Here,we examined possible functions of the enzymes during fruit development.We further characterized transgenic plants differing in the expression of branched chain amino acid transaminases 1 and 3,evaluating the rates of respiration in fruits deficient in BCAT1 and the levels of volatiles in lines overexpressing either BCAT1 or BCAT3.We quantitatively tested,via precursor and isotope feeding experiments,the importance of the branched chain amino acids and their corresponding keto acids in the formation of fruit volatiles.Our results not only demonstrate for the first time the importance of branched chain amino acids in fruit respiration,but also reveal that keto acids,rather than amino acids,are the likely precursors for the branched chain flavor volatiles.

  11. Functional characterization of flax fatty acid desaturase FAD2 and FAD3 isoforms expressed in yeast reveals a broad diversity in activity.

    Science.gov (United States)

    Radovanovic, Natasa; Thambugala, Dinushika; Duguid, Scott; Loewen, Evelyn; Cloutier, Sylvie

    2014-07-01

    With 45 % or more oil content that contains more than 55 % alpha linolenic (LIN) acid, linseed (Linum usitatissimum L.) is one of the richest plant sources of this essential fatty acid. Fatty acid desaturases 2 (FAD2) and 3 (FAD3) are the main enzymes responsible for the Δ12 and Δ15 desaturation in planta. In linseed, the oilseed morphotype of flax, two paralogous copies, and several alleles exist for each gene. Here, we cloned three alleles of FAD2A, four of FAD2B, six of FAD3A, and seven of FAD3B into a pYES vector and transformed all 20 constructs and an empty construct in yeast. The transformants were induced in the presence of oleic (OLE) acid substrate for FAD2 constructs and linoleic (LIO) acid for FAD3. Conversion rates of OLE acid into LIO acid and LIO acid into LIN acid were measured by gas chromatography. Conversion rate of FAD2 exceeded that of FAD3 enzymes with FAD2B having a conversion rate approximately 10 % higher than FAD2A. All FAD2 isoforms were active, but significant differences existed between isoforms of both FAD2 enzymes. Two FAD3A and three FAD3B isoforms were not functional. Some nonfunctional enzymes resulted from the presence of nonsense mutations causing premature stop codons, but FAD3B-C and FAD3B-F seem to be associated with single amino acid changes. The activity of FAD3A-C was more than fivefold greater than the most common isoform FAD3A-A, while FAD3A-F was fourfold greater. Such isoforms could be incorporated into breeding lines to possibly further increase the proportion of LIN acid in linseed. PMID:24522837

  12. Continuous-state branching processes

    CERN Document Server

    Li, Zenghu

    2012-01-01

    These notes were used in a short graduate course on branching processes the author gave in Beijing Normal University. The following main topics are covered: scaling limits of Galton--Watson processes, continuous-state branching processes, extinction probabilities, conditional limit theorems, decompositions of sample paths, martingale problems, stochastic equations, Lamperti's transformations, independent and dependent immigration processes. Some of the results are simplified versions of those in the author's book "Measure-valued branching Markov processes" (Springer, 2011). We hope these simplified results will set out the main ideas in an easy way and lead the reader to a quick access of the subject.

  13. Cadmium-induced Membrane Lipid Peroxidation and Changes in Antioxidant Enzyme Activities and Peroxidase Isoforms in Jerusalem Artichoke Seedlings%镉诱导的菊芋幼苗膜脂过氧化和抗氧化酶活性及过氧化物酶同工酶的改变

    Institute of Scientific and Technical Information of China (English)

    陶毅明; 陈燕珍; 梁杨琳; 徐美燕; 徐祥明

    2007-01-01

    Jerusalem artichoke (Helianthus tuberosus L.)seedlings cultured in sandy media were treated with Hoagland nutrition solution with different concentrations of Cd(NO3)2 from 0 to 400 μmol/L. After 50 days' treatment, Cd accumulation, activities of peroxidase (POD, EC 1.11.1.7), superoxide dismutase (SOD, EC 1.15.1.1), catalase (CAT, EC 1.11.1.6) were measured and electrophoretograms of POD isoenzymes were analyzed. The accumulation of Cd in seedlings increased from Cd 50-100 μmol/L, after which further increases in Cd concentration resulted in only small increases in accumulation of Cd in seedlings. MDA content was markedly higher than control values indicating the enhanced membrane lipid peroxidation in roots and leaves. POD activities in leaf and root extracts increased with an increase of Cd concentration from 0 to 50 and 100 μmol/L and then decreased with further increases to 200 and 400 μmol/L. Under moderate Cd level of 50-200 μmol/L,SOD activities in leaf and root extracts increased whereas with a higher Cd level of 400 μmol/L marked inhibitions in enzyme activities were observed. With increase in Cd concentration marked elevations in CAT activities in leaves and roots were observed. Results of electrophoresis show that the alteration of POD isoenzyme was noticeable to Cd and an additional POD isoenzyme LP10 appeared. It is suggested that POD isoenzyme of Jerusalem artichoke seedlings could be used as bioindicator for soil contamination by Cd.%用含有不同浓度(0~400 μmol/L)Cd(NO3)2的Hoagland营养液处理砂培的菊芋.处理50 d后,测定植物体内镉积累量以及过氧化物酶(POD)、超氧化物歧化酶(SOD)、过氧化氢酶(CAT)活性,并对POD同工酶进行电泳分析.发现在Cd 50~100 μmol/L浓度内,随着镉浓度的升高,菊芋根和叶中镉的积累量显著增加,而随后积累量的增加有所减少.根和叶中MDA含量显著上升,说明镉引起了膜脂过氧化.0~100 μmol/L Cd处理,根和叶中POD活性

  14. Photoreactivating enzymes

    International Nuclear Information System (INIS)

    Photoreactivating enzymes (PRE) also called photolyases (EC 4.1.99.3) catalyze the light 300 to 600 nm)-dependent monomerization of cyclobutyl pyrimidine dimers, formed between adjacent pyrimidines on the same DNA strand, upon exposure to ultraviolet (uv) irradiation (220 to 320 nm). Although much is known about the substrate and product of these unusual enzymes, their identification required the development and synthesis of such fields as photochemistry, biochemistry, and microbiology. Photoreactivation was first known as a biological recovery phenomenon: cells exposed to visible light following uv irradiation showed higher survival than those kept in the dark. Early investigators examined the photoreactivability of an enormous range of cellular damage in both prokaryotes and eukaryotes. This review article discusses the purification and properties of PRE, the kinetics of photoreactivation and the biological role of this repair process

  15. Engineering enzymes

    OpenAIRE

    Dutton, P. Leslie; Moser, Christopher C.

    2011-01-01

    Fundamental research into bioinorganic catalysis of the kind presented at this Faraday Discussion has the potential to turn inspiration drawn from impressive natural energy and chemical transformations into artificial catalyst constructions useful to mankind. Creating bio-inspired artificial constructions requires a level of understanding well beyond simple description of structures and mechanisms of natural enzymes. To be useful, such description must be augmented by a practical sense of str...

  16. Expression of novel isoforms of carnitine palmitoyltransferase I (CPT-1) generated by alternative splicing of the CPT-ibeta gene.

    Science.gov (United States)

    Yu, G S; Lu, Y C; Gulick, T

    1998-01-01

    Carnitine palmitoyltransferase I (CPT-I) catalyses the rate-determining step in mitochondrial fatty acid beta-oxidation. The enzyme has two cognate structural genes that are preferentially expressed in liver (alpha) or fat and muscle (beta). We hypothesized the existence of additional isoforms in heart to account for unique kinetic characteristics of enzyme activity in this tissue. Hybridization and PCR screening of a human cardiac cDNA library revealed the expression of two novel CPT-I isoforms generated by alternative splicing of the CPT-Ibeta transcript, in addition to the beta and alpha cDNA species previously described. Ribonuclease protection and reverse transcriptase-mediated PCR assays confirmed the presence of mRNA species of each splicing variant in heart, skeletal muscle and liver, with differing relative concentrations in the tissues. The novel splicing variants omit exons or utilize a cryptic splice donor site within an exon. Deduced polypeptide sequences of the novel enzymes include omissions in the region of putative membrane-spanning and malonyl-CoA regulatory domains compared with the previously described CPT-Is, implying that the encoded enzymes will exhibit unique features with respect to outer mitochondrial membrane topology and response to physiological and pharmacological inhibitors. PMID:9693124

  17. Specific regulation of NRG1 isoform expression by neuronal activity

    OpenAIRE

    Liu, Xihui; Bates, Ryan; Wang, Fay; Su, Nan; Kirov, Sergei A.; Luo, Yuling; Wang, Jian-Zhi; Xiong, Wen-Cheng; Mei, Lin

    2011-01-01

    Neuregulin 1 (NRG1) is a trophic factor that has been implicated in neural development, neurotransmission and synaptic plasticity. NRG1 has multiple isoforms that are generated by usage of different promoters and alternative splicing of a single gene. However, little is known about NRG1 isoform composition profile, whether it changes during development or the underlying mechanisms. We found that each of the six types of NRG1 has a distinct expression pattern in the brain at different ages, re...

  18. Left bundle-branch block

    DEFF Research Database (Denmark)

    Risum, Niels; Strauss, David; Sogaard, Peter;

    2013-01-01

    The relationship between myocardial electrical activation by electrocardiogram (ECG) and mechanical contraction by echocardiography in left bundle-branch block (LBBB) has never been clearly demonstrated. New strict criteria for LBBB based on a fundamental understanding of physiology have recently...

  19. Does Japanese medaka (Oryzias latipes) exhibit a gill Na(+)/K(+)-ATPase isoform switch during salinity change?

    Science.gov (United States)

    Bollinger, Rebecca J; Madsen, Steffen S; Bossus, Maryline C; Tipsmark, Christian K

    2016-05-01

    Some euryhaline teleosts exhibit a switch in gill Na(+)/K(+)-ATPase (Nka) α isoform when moving between fresh water (FW) and seawater (SW). The present study tested the hypothesis that a similar mechanism is present in Japanese medaka and whether salinity affects ouabain, Mg(2+), Na(+) and K(+) affinity of the gill enzyme. Phylogenetic analysis classified six separate medaka Nka α isoforms (α1a, α1b, α1c, α2, α3a and α3b). Medaka acclimated long-term (>30 days) to either FW or SW had similar gill expression of α1c, α2, α3a and α3b, while both α1a and α1b were elevated in SW. Since a potential isoform shift may rely on early changes in transcript abundance, we conducted two short-term (1-3 days) salinity transfer experiments. FW to SW acclimation induced an elevation of α1b and α1a after 1 and 3 days. SW to FW acclimation reduced α1b after 3 days with no other α isoforms affected. To verify that the responses were typical, additional transport proteins were examined. Gill ncc and nhe3 expression were elevated in FW, while cftr and nkcc1a were up-regulated in SW. This is in accordance with putative roles in ion-uptake and secretion. SW-acclimated medaka had higher gill Nka V max and lower apparent K m for Na(+) compared to FW fish, while apparent affinities for K(+), Mg(2+) and ouabain were unchanged. The present study showed that the Japanese medaka does not exhibit a salinity-induced α isoform switch and therefore suggests that Na(+) affinity changes involve altered posttranslational modification or intermolecular interactions. PMID:26920794

  20. A Network of Splice Isoforms for the Mouse.

    Science.gov (United States)

    Li, Hong-Dong; Menon, Rajasree; Eksi, Ridvan; Guerler, Aysam; Zhang, Yang; Omenn, Gilbert S; Guan, Yuanfang

    2016-01-01

    The laboratory mouse is the primary mammalian species used for studying alternative splicing events. Recent studies have generated computational models to predict functions for splice isoforms in the mouse. However, the functional relationship network, describing the probability of splice isoforms participating in the same biological process or pathway, has not yet been studied in the mouse. Here we describe a rich genome-wide resource of mouse networks at the isoform level, which was generated using a unique framework that was originally developed to infer isoform functions. This network was built through integrating heterogeneous genomic and protein data, including RNA-seq, exon array, protein docking and pseudo-amino acid composition. Through simulation and cross-validation studies, we demonstrated the accuracy of the algorithm in predicting isoform-level functional relationships. We showed that this network enables the users to reveal functional differences of the isoforms of the same gene, as illustrated by literature evidence with Anxa6 (annexin a6) as an example. We expect this work will become a useful resource for the mouse genetics community to understand gene functions. The network is publicly available at: http://guanlab.ccmb.med.umich.edu/isoformnetwork. PMID:27079421

  1. Rice PROTEIN l-ISOASPARTYL METHYLTRANSFERASE isoforms differentially accumulate during seed maturation to restrict deleterious isoAsp and reactive oxygen species accumulation and are implicated in seed vigor and longevity.

    Science.gov (United States)

    Petla, Bhanu Prakash; Kamble, Nitin Uttam; Kumar, Meenu; Verma, Pooja; Ghosh, Shraboni; Singh, Ajeet; Rao, Venkateswara; Salvi, Prafull; Kaur, Harmeet; Saxena, Saurabh Chandra; Majee, Manoj

    2016-07-01

    PROTEIN l-ISOASPARTYL O-METHYLTRANSFERASE (PIMT) is a protein-repairing enzyme involved in seed vigor and longevity. However, the regulation of PIMT isoforms during seed development and the mechanism of PIMT-mediated improvement of seed vigor and longevity are largely unknown. In this study in rice (Oryza sativa), we demonstrate the dynamics and correlation of isoaspartyl (isoAsp)-repairing demands and PIMT activity, and their implications, during seed development, germination and aging, through biochemical, molecular and genetic studies. Molecular and biochemical analyses revealed that rice possesses various biochemically active and inactive PIMT isoforms. Transcript and western blot analyses clearly showed the seed development stage and tissue-specific accumulation of active isoforms. Immunolocalization studies revealed distinct isoform expression in embryo and aleurone layers. Further analyses of transgenic lines for each OsPIMT isoform revealed a clear role in the restriction of deleterious isoAsp and age-induced reactive oxygen species (ROS) accumulation to improve seed vigor and longevity. Collectively, our data suggest that a PIMT-mediated, protein repair mechanism is initiated during seed development in rice, with each isoform playing a distinct, yet coordinated, role. Our results also raise the intriguing possibility that PIMT repairs antioxidative enzymes and proteins which restrict ROS accumulation, lipid peroxidation, etc. in seed, particularly during aging, thus contributing to seed vigor and longevity. PMID:26987457

  2. In vitro identification of human cytochrome P450 isoforms involved in the metabolism of Geissoschizine methyl ether, an active component of the traditional Japanese medicine Yokukansan.

    Science.gov (United States)

    Matsumoto, Takashi; Kushida, Hirotaka; Maruyama, Takeshi; Nishimura, Hiroaki; Watanabe, Junko; Maemura, Kazuya; Kase, Yoshio

    2016-01-01

    1. Yokukansan (YKS) is a traditional Japanese medicine also called kampo, which has been used to treat neurosis, insomnia, and night crying and peevishness in children. Geissoschizine methyl ether (GM), a major indole alkaloid found in Uncaria hook, has been identified as a major active component of YKS with psychotropic effects. Recently, GM was reported to have a partial agonistic effect on serotonin 5-HT1A receptors. However, there is little published information on GM metabolism in humans, although several studies reported the blood kinetics of GM in rats and humans. In this study, we investigated the GM metabolic pathways and metabolizing enzymes in humans. 2. Using recombinant human cytochrome P450 (CYP) isoforms and polyclonal antibodies to CYP isoforms, we found that GM was metabolized into hydroxylated, dehydrogenated, hydroxylated+dehydrogenated, demethylated and water adduct forms by some CYP isoforms. 3. The relative activity factors in human liver microsomes were calculated to determine the relative contributions of individual CYP isoforms to GM metabolism in human liver microsomes (HLMs). We identified CYP3A4 as the CYP isoform primarily responsible for GM metabolism in human liver microsomes. 4. These findings provide an important basis for understanding the pharmacokinetics and pharmacodynamics of GM and YKS. PMID:26337900

  3. Evaluation on the nutritional value of protein in genetically modified rice with double antisense starch-branching enzyme gene%转双反义淀粉分支酶基因大米中蛋白质的营养价值评价

    Institute of Scientific and Technical Information of China (English)

    李敏; 胡贻椿; 朴建华; 杨晓光

    2011-01-01

    目的 对转双反义淀粉分支酶基因大米和亲本大米蛋白质的营养价值进行评价.方法 采用通用的非生物学评价方法,即化学评分、氨基酸评分、蛋白质消化率校正后的氨基酸评分、必需氨基酸指数、营养指数、氨基酸比值系数和氨基酸比值系数分,评价大米蛋白质的营养价值.结果 转基因大米和亲本大米的化学评分分别为62,59;氨基酸评分分别为75,62;蛋白质消化率校正后的氨基酸评分分别为65,56;必需氨基酸指数分别为79,80;营养指数分别为11,8;氨基酸比值系数分分别为76.7,67.0;氨基酸比值系数都是l.1.结论 转双反义淀粉分支氨基因大来中抗性淀粉含童显著提高,但并未影响到其自身蛋白质的营养价值.该转基因大米蛋白质的营养价值与亲本大米具有"实质等同性".%Objective To assess the nutritional value of protein in the genetically modified rice with double antisense starch-branching enzyme (SBE) gene and its parental rice.Methods The adopted nutritional value assessment methods, including chemical score (CS), amino acid score (AAS), protein digestibility-corrected amino acid score (PDCAAS), essential amino acid index (EAAI), nutritional index (NI), ratio coefficient of amino acid (RCAA) and score of RCAA ( SRCAA ) was applied to assess the protein nutritional value of the rice.Results The CS value of genetically modified rice and parental rice were 62 and 59, AAS value were 75 and 62, PDCAAS value were 65 and 56,EAAI were 79 and 80, NI were 11 and 8, SRCAA were 76.7 and 67.0, respectively.RCAA were the same.Conclusion The nutritional value of protein in the genetically modified rice with double antisense SBE gene were not greatly changed by the increase of resistant starch content in the rice.So, the nutritional value of protein in the genetically modified rice and its parental rice was substantial equivalent.

  4. Fragrance release from the surface of branched poly (amide)s.

    Science.gov (United States)

    Aulenta, Francesca; Drew, Michael G B; Foster, Alison; Hayes, Wayne; Rannard, Steven; Thornthwaite, David W; Youngs, Tristan G A

    2005-01-01

    Enzymes are powerful tools in organic synthesis that are able to catalyse a wide variety of selective chemical transformations under mild and environmentally friendly conditions. Enzymes such as the lipases have also found applications in the synthesis and degradation of polymeric materials. However, the use of these natural catalysts in the synthesis and the post-synthetic modification of dendrimers and hyperbranched molecules is an application of chemistry yet to be explored extensively. In this study the use of two hydrolytic enzymes, a lipase from Candida cylindracea and a cutinase from Fusarium solani pisii, were investigated in the selective cleavage of ester groups situated on the peripheral layer of two families of branched polyamides. These branched polyamides were conjugated to simple fragrances citronellol and L-menthol via ester linkages. Hydrolysis of the ester linkage between the fragrances and the branched polyamide support was carried out in aqueous buffered systems at slightly basic pH values under the optimum operative conditions for the enzymes used. These preliminary qualitative investigations revealed that partial cleavage of the ester functionalities from the branched polyamide support had occurred. However, the ability of the enzymes to interact with the substrates decreased considerably as the branching density, the rigidity of the structure and the bulkiness of the polyamide-fragrance conjugates increased. PMID:18007278

  5. Characterization of Rabbit CD5 Isoforms

    OpenAIRE

    Pospisil, Richard; Kabat, Juraj; Mage, Rose G.

    2009-01-01

    Previously described polyclonal or monoclonal antibodies (mAb) to rabbit CD5, raised against expressed recombinant protein or peptides, recognize CD5 on most rabbit B cells. The mAb KEN-5 was originally reported to recognize rabbit CD5. However, KEN-5 binds almost exclusively to T cells and only to a minor population of B cells. We show here that by Enzyme-linked Immunosorbent Assay (ELISA), KEN-5 binds to recombinant rabbit CD5. This interaction is partially inhibited by polyclonal goat anti...

  6. Interaction between oblongifolin C and UDP-glucuronosyltransferase isoforms in human liver and intestine microsomes.

    Science.gov (United States)

    Gao, Cui; Shi, Rong; Wang, Tianming; Tan, Hongsheng; Xu, Hongxi; Ma, Yueming

    2015-01-01

    1. Oblongifolin C (OC) is a potential natural anticancer candidate, and its metabolic profile has not yet been established. 2. One major OC glucuronidation metabolite (OCG) has been identified in a pool of human liver microsomes (HLMs). Chemical inhibition experiments suggested that OCG was mainly formed by UGT1A. A screen of recombinant UDP-glucuronosyltransferase isoforms (UGTs) indicated that UGT1A1 primarily mediates OC conjugation, with minor contributions from UGT1A3 and UGT1A8. Enzyme kinetic studies showed that UGT1A1 was the main UGT isoform involved in OCG in HLMs. 3. Further investigation suggested that OC is a broad inhibitor of UGTs. Additionally, OC competitively inhibited UGT1A6 with a Ki value of 3.49 ± 0.57 μM, whereas non-competitively inhibited UGT1A10 with a Ki value of 2.12 ± 0.18 μM. 4. Understanding the interaction between OC and UGTs will greatly contribute to future investigations regarding the inter-individual differences in OC metabolism in clinical trials and potential drug-drug interactions. PMID:25714435

  7. Mitochondrial Sulfide Detoxification Requires a Functional Isoform O-Acetylserine(thiol)lyase C in Arabidopsis thaliana

    Institute of Scientific and Technical Information of China (English)

    Consolación (A)lvarez; Irene García; Luis C.Romero; Cecilia Gotor

    2012-01-01

    In non-cyanogenic species,the main source of cyanide derives from ethylene and camalexin biosyntheses.In mitochondria,cyanide is a potent inhibitor of the cytochrome c oxidase and is metabolized bythe β-cyanoalanine synthase CYS-C1,catalyzing the conversion of cysteine and cyanide to hydrogen sulfide and β-cyanoalanine.The hydrogen sulfide released also inhibits the cytochrome c oxidase and needs to be detoxified by the O-acetylserine(thiol)lyase mitochondrial isoform,OAS-C,which catalyzes the incorporation of sulfide to O-acetylserine to produce cysteine,thus generating a cyclic pathway in the mitochondria.The loss of functional OAS-C isoforms causes phenotypic characteristics very similar to the loss of the CYS-C1 enzyme,showing defects in root hair formation.Genetic complementation with the OAS-C gene rescues the impairment of root hair elongation,restoring the wild-type phenotype.The mitochondria compromise their capacity to properly detoxify cyanide and the resulting sulfide because the latter cannot re-assimilate into cysteine in the oas-c null mutant.Consequently,we observe an accumulation of sulfide and cyanide and of the alternative oxidase,which is unable to prevent the production of reactive oxygen species probably due to the accumulation of both toxic molecules.Our results allow us to suggest that the significance of OAS-C is related to its role in the proper sulfide and cyanide detoxification in mitochondria.

  8. Fluconazole Binding and Sterol Demethylation in Three CYP51 Isoforms Indicate Differences in Active Site Topology

    Energy Technology Data Exchange (ETDEWEB)

    Bellamine, A.; Lepesheva, Galina I.; Waterman, Mike (Vanderbilt)

    2010-11-16

    14{alpha}-Demethylase (CYP51) is a key enzyme in all sterol biosynthetic pathways (animals, fungi, plants, protists, and some bacteria), catalyzing the removal of the C-14 methyl group following cyclization of squalene. Based on mutations found in CYP51 genes from Candida albicans azole-resistant isolates obtained after fluconazole treatment of fungal infections, and using site-directed mutagenesis, we have found that fluconazole binding and substrate metabolism vary among three different CYP51 isoforms: human, fungal, and mycobacterial. In C. albicans, the Y132H mutant from isolates shows no effect on fluconazole binding, whereas the F145L mutant results in a 5-fold increase in its IC{sub 50} for fluconazole, suggesting that F145 (conserved only in fungal 14{alpha}-demethylases) interacts with this azole. In C. albicans, F145L accounts, in part, for the difference in fluconazole sensitivity reported between mammals and fungi, providing a basis for treatment of fungal infections. The C. albicans Y132H and human Y145H CYP51 mutants show essentially no effect on substrate metabolism, but the Mycobacterium tuberculosis F89H CYP51 mutant loses both its substrate binding and metabolism. Because these three residues align in the three isoforms, the results indicate that their active sites contain important structural differences, and further emphasize that fluconazole and substrate binding are uncoupled properties.

  9. Multiple pathways regulate shoot branching

    Directory of Open Access Journals (Sweden)

    Catherine eRameau

    2015-01-01

    Full Text Available Shoot branching patterns result from the spatio-temporal regulation of axillary bud outgrowth. Numerous endogenous, developmental and environmental factors are integrated at the bud and plant levels to determine numbers of growing shoots. Multiple pathways that converge to common integrators are most probably involved. We propose several pathways involving not only the classical hormones auxin, cytokinins and strigolactones, but also other signals with a strong influence on shoot branching such as gibberellins, sugars or molecular actors of plant phase transition. We also deal with recent findings about the molecular mechanisms and the pathway involved in the response to shade as an example of an environmental signal controlling branching. We propose the TCP transcription factor TB1/BRC1 and the polar auxin transport stream in the stem as possible integrators of these pathways. We finally discuss how modeling can help to represent this highly dynamic system by articulating knowledges and hypothesis and calculating the phenotype properties they imply.

  10. Differential regulation of renal phospholipase C isoforms by catecholamines.

    Science.gov (United States)

    Yu, P Y; Asico, L D; Eisner, G M; Jose, P A

    1995-01-01

    Dopamine and D1 agonists and NE all increase phosphatidyl inositol-specific phospholipase C (PLC) activity, but whereas dopamine produces a natriuresis, NE has an antinatriuretic effect. To determine if catecholamines differentially regulate the expression of PLC isoforms, we infused fenoldopam, a D1 agonist, or pramipexole, a D1/D2 agonist, intravenously or infused fenoldopam or NE into the renal artery of anesthetized rats. After 3-4 h of infusion, when the expected natriuresis (fenoldopam or pramipexole) or antinatriuresis (NE) occurred, the kidneys were removed for analysis of PLC isoform protein expression activity. Western blot analysis revealed that in renal cortical membranes, fenoldopam and pramipexole increased expression of PLC beta 1 and decreased expression of PLC gamma 1; PLC delta was unchanged. In the cytosol, pramipexole and fenoldopam increased expression of both PLC beta 1 and PLC gamma 1. No effects were noted in the medulla. A preferential D1 antagonist, SKF 83742, which by itself had no effect, blocked the effects of pramipexole, thus confirming the involvement of the D1 receptor. In contrast, NE also increased PLC beta 1 but did not affect PLC gamma 1 protein expression in membranes. The changes in PLC isoform expression were accompanied by similar changes in PLC isoform activity. These studies demonstrate for the first time differential regulation of PLC isoforms by catecholamines. PMID:7814630

  11. SURVIV for survival analysis of mRNA isoform variation.

    Science.gov (United States)

    Shen, Shihao; Wang, Yuanyuan; Wang, Chengyang; Wu, Ying Nian; Xing, Yi

    2016-01-01

    The rapid accumulation of clinical RNA-seq data sets has provided the opportunity to associate mRNA isoform variations to clinical outcomes. Here we report a statistical method SURVIV (Survival analysis of mRNA Isoform Variation), designed for identifying mRNA isoform variation associated with patient survival time. A unique feature and major strength of SURVIV is that it models the measurement uncertainty of mRNA isoform ratio in RNA-seq data. Simulation studies suggest that SURVIV outperforms the conventional Cox regression survival analysis, especially for data sets with modest sequencing depth. We applied SURVIV to TCGA RNA-seq data of invasive ductal carcinoma as well as five additional cancer types. Alternative splicing-based survival predictors consistently outperform gene expression-based survival predictors, and the integration of clinical, gene expression and alternative splicing profiles leads to the best survival prediction. We anticipate that SURVIV will have broad utilities for analysing diverse types of mRNA isoform variation in large-scale clinical RNA-seq projects. PMID:27279334

  12. Computer simulation of long-chain branching and branching indexes

    Czech Academy of Sciences Publication Activity Database

    Netopilík, Miloš

    Vienna : University of Vienna, 2014. s. 22. [International Conference on Polymer Behaviour /6./. 22.09.2014-26.09.2014, Vienna] R&D Projects: GA ČR(CZ) GA13-02938S Institutional support: RVO:61389013 Keywords : branching indexes * intrinsic viscosity * radius of gyration Subject RIV: CD - Macromolecular Chemistry

  13. Analysis of two Schistosoma mansoni uridine phosphorylases isoforms suggests the emergence of a protein with a non-canonical function.

    Science.gov (United States)

    da Silva Neto, Antônio Marinho; Torini de Souza, Juliana Roberta; Romanello, Larissa; Cassago, Alexandre; Serrão, Vitor Hugo Balasco; DeMarco, Ricardo; Brandão-Neto, José; Garratt, Richard Charles; Pereira, Humberto D'Muniz

    2016-06-01

    Reports of Schistosoma mansoni strains resistant to praziquantel, the only therapeutic strategy available for the treatment of schistosomiasis, have motivated the scientific community towards the search for new possible therapies. Biochemical characterization of the parasite's metabolism is an essential component for the rational development of new therapeutic alternatives. One of the so far uncharacterized enzymes is uridine phosphorylase (UP) (EC 2.4.2.3), for which the parasite genome presents two isoforms (SmUPa and SmUPb) that share 92% sequence identity. In this paper, we present crystal structures for SmUPa and SmUPb in their free states as well as bound to different ligands. This we have complemented by enzyme kinetic characterization and phylogenetic analyses. Both enzymes present an overall fold and active site structure similar to other known UPs. The kinetic analyses showed conclusively that SmUPa is a regular uridine phosphorylase but by contrast SmUPb presented no detectable activity. This is particularly noteworthy given the high level of sequence identity between the two isoforms and is probably the result of the significant differences observed for SmUPb in the vicinity of the active site itself, suggesting that it is not a UP at all. On the other hand, it was not possible to identify an alternative function for SmUPb, although our phylogenetic analyses and expression data suggest that SmUPb is still functional and plays a role in parasite metabolism. The unusual UPb isoform may open up new opportunities for understanding unique features of S. mansoni metabolism. PMID:26898674

  14. Vitamin E Isoforms as Modulators of Lung Inflammation

    Directory of Open Access Journals (Sweden)

    Hiam Abdala-Valencia

    2013-10-01

    Full Text Available Asthma and allergic diseases are complex conditions caused by a combination of genetic and environmental factors. Clinical studies suggest a number of protective dietary factors for asthma, including vitamin E. However, studies of vitamin E in allergy commonly result in seemingly conflicting outcomes. Recent work indicates that allergic inflammation is inhibited by supplementation with the purified natural vitamin E isoform α-tocopherol but elevated by the isoform γ-tocopherol when administered at physiological tissue concentrations. In this review, we discuss opposing regulatory effects of α-tocopherol and γ-tocopherol on allergic lung inflammation in clinical trials and in animal studies. A better understanding of the differential regulation of inflammation by isoforms of vitamin E provides a basis towards the design of clinical studies and diets that would effectively modulate inflammatory pathways in lung disease.

  15. Isoforms of murine and human serum amyloid P component

    DEFF Research Database (Denmark)

    Nybo, Mads; Hackler, R; Kold, B;

    1998-01-01

    affect their number. When the acute-phase response was analysed in three mouse strains, CBA/J and C3H/HeN initially showed seven SAP isoforms in serum and C57BL/6 J three or four. The responses in all three strains peaked at day 2 and were normalized within 14 days. On days 2 and 4, CBA/J and C3H......Isoelectric focusing (IEF) and immunofixation of murine serum amyloid P component (SAP), purified and in serum, showed a distinct and strain-dependent isoform pattern with up to seven bands (pI 5.1-5.7). Neuraminidase treatment caused a shift of the isoforms to more basic pI values, but did not...

  16. Oxygenation properties and isoform diversity of snake hemoglobins

    DEFF Research Database (Denmark)

    Storz, Jay F.; Natarajan, Chandrasekhar; Moriyama, Hideaki;

    2015-01-01

    Available data suggest that snake hemoglobins (Hbs) are characterized by a combination of unusual structural and functional properties relative to the Hbs of other amniote vertebrates, including oxygenation-linked tetramer- dimer dissociation. However, standardized comparative data are lacking for...... snake Hbs, and the Hb isoform composition of snake red blood cells has not been systematically characterized. Here we present the results of an integrated analysis of snake Hbs and the underlying - and -type globin genes to characterize 1) Hb isoform composition of definitive erythrocytes, and 2) the...... oxygenation properties of isolated isoforms as well as composite hemolysates. We used species from three families as subjects for experimental studies of Hb function: South American rattlesnake, Crotalus durissus (Viperidae); Indian python, Python molurus (Pythonidae); and yellow-bellied sea snake, Pelamis...

  17. Laminin isoforms in endothelial and perivascular basement membranes

    Science.gov (United States)

    Yousif, Lema F.; Di Russo, Jacopo; Sorokin, Lydia

    2013-01-01

    Laminins, one of the major functional components of basement membranes, are found underlying endothelium, and encasing pericytes and smooth muscle cells in the vessel wall. Depending on the type of blood vessel (capillary, venule, postcapillary venule, vein or artery) and their maturation state, both the endothelial and mural cell phenotype vary, with associated changes in laminin isoform expression. Laminins containing the α4 and α5 chains are the major isoforms found in the vessel wall, with the added contribution of laminin α2 in larger vessels. We here summarize current data on the precise localization of these laminin isoforms and their receptors in the different layers of the vessel wall, and their potential contribution to vascular homeostasis. PMID:23263631

  18. Stability Mechanisms of Laccase Isoforms using a Modified FoldX Protocol Applicable to Widely Different Proteins

    DEFF Research Database (Denmark)

    Christensen, Niels J.; Kepp, Kasper P.

    2013-01-01

    A recent computational protocol that accurately predicts and rationalizes protein multisite mutant stabilities has been extended to handle widely different isoforms of laccases. We apply the protocol to four isoenzymes of Trametes versicolor laccase (TvL) with variable lengths (498–503 residues) ....... Several sites near Cu, notably 79, 241, and 245, or near substrate, mainly 265, are identified that contribute to stability-function trade-offs, of relevance to the search for new proficient and stable variants of these important industrial enzymes....

  19. Identification and characterization of novel NuMA isoforms

    Energy Technology Data Exchange (ETDEWEB)

    Wu, Jin, E-mail: petersdu2112@hotmail.com [Key Laboratory for Cell Proliferation and Regulation of the Ministry of Education, Beijing Normal University, Beijing (China); Xu, Zhe [Department of Clinical Laboratory Diagnosis, Beijing Tiantan Hospital, Capital Medical University, Beijing (China); Core Laboratory for Clinical Medical Research, Beijing Tiantan Hospital, Capital Medical University, Beijing (China); He, Dacheng [Key Laboratory for Cell Proliferation and Regulation of the Ministry of Education, Beijing Normal University, Beijing (China); Lu, Guanting, E-mail: guantlv@126.com [Beijing DnaLead Science and Technology Co., LTD, Beijing (China)

    2014-11-21

    Highlights: • Seven NuMA isoforms generated by alternative splicing were categorized into 3 groups: long, middle and short. • Both exons 15 and 16 in long NuMA were “hotspot” for alternative splicing. • Lower expression of short NuMA was observed in cancer cells compared with nonneoplastic controls. • Distinct localization pattern of short isoforms indicated different function from that of long and middle NuMA. - Abstract: The large nuclear mitotic apparatus (NuMA) has been investigated for over 30 years with functions related to the formation and maintenance of mitotic spindle poles during mitosis. However, the existence and functions of NuMA isoforms generated by alternative splicing remains unclear. In the present work, we show that at least seven NuMA isoforms (categorized into long, middle and short groups) generated by alternative splicing from a common NuMA mRNA precursor were discovered in HeLa cells and these isoforms differ mainly at the carboxyl terminus and the coiled-coil domains. Two “hotspot” exons with molecular mass of 3366-nt and 42-nt tend to be spliced during alternative splicing in long and middle groups. Furthermore, full-length coding sequences of long and middle NuMA obtained by using fusion PCR were constructed into GFP-tagged vector to illustrate their cellular localization. Long NuMA mainly localized in the nucleus with absence from nucleoli during interphase and translocated to the spindle poles in mitosis. Middle NuMA displayed the similar cell cycle-dependent distribution pattern as long NuMA. However, expression of NuMA short isoforms revealed a distinct subcellular localization. Short NuMA were present in the cytosol during the whole cycle, without colocalization with mitotic apparatus. These results have allowed us tentatively to explore a new research direction for NuMA’s various functions.

  20. Apolipoprotein E isoform-specific effects on lipoprotein receptor processing.

    Science.gov (United States)

    Bachmeier, Corbin; Shackleton, Ben; Ojo, Joseph; Paris, Daniel; Mullan, Michael; Crawford, Fiona

    2014-12-01

    Recent findings indicate an isoform-specific role for apolipoprotein E (apoE) in the elimination of beta-amyloid (Aβ) from the brain. ApoE is closely associated with various lipoprotein receptors, which contribute to Aβ brain removal via metabolic clearance or transit across the blood–brain barrier (BBB). These receptors are subject to ectodomain shedding at the cell surface, which alters endocytic transport and mitigates Aβ elimination. To further understand the manner in which apoE influences Aβ brain clearance, these studies investigated the effect of apoE on lipoprotein receptor shedding. Consistent with prior reports, we observed an increased shedding of the low-density lipoprotein receptor (LDLR) and the LDLR-related protein 1 (LRP1) following Aβ exposure in human brain endothelial cells. When Aβ was co-treated with each apoE isoform, there was a reduction in Aβ-induced shedding with apoE2 and apoE3, while lipoprotein receptor shedding in the presence of apoE4 remained increased. Likewise, intracranial administration of Aβ to apoE-targeted replacement mice (expressing the human apoE isoforms) resulted in an isoform-dependent effect on lipoprotein receptor shedding in the brain (apoE4 > apoE3 > apoE2). Moreover, these results show a strong inverse correlation with our prior work in apoE transgenic mice in which apoE4 animals showed reduced Aβ clearance across the BBB compared to apoE3 animals. Based on these results, apoE4 appears less efficient than other apoE isoforms in regulating lipoprotein receptor shedding, which may explain the differential effects of these isoforms in removing Aβ from the brain. PMID:25015123

  1. NCI: DCTD: Biometric Research Branch

    Science.gov (United States)

    The Biometric Research Branch (BRB) is the statistical and biomathematical component of the Division of Cancer Treatment, Diagnosis and Centers (DCTDC). Its members provide statistical leadership for the national and international research programs of the division in developmental therapeutics, developmental diagnostics, diagnostic imaging and clinical trials.

  2. Branching of keratin intermediate filaments.

    Science.gov (United States)

    Nafeey, Soufi; Martin, Ines; Felder, Tatiana; Walther, Paul; Felder, Edward

    2016-06-01

    Keratin intermediate filaments (IFs) are crucial to maintain mechanical stability in epithelial cells. Since little is known about the network architecture that provides this stiffness and especially about branching properties of filaments, we addressed this question with different electron microscopic (EM) methods. Using EM tomography of high pressure frozen keratinocytes, we investigated the course of several filaments in a branching of a filament bundle. Moreover we found several putative bifurcations in individual filaments. To verify our observation we also visualized the keratin network in detergent extracted keratinocytes with scanning EM. Here bifurcations of individual filaments could unambiguously be identified additionally to bundle branchings. Interestingly, identical filament bifurcations were also found in purified keratin 8/18 filaments expressed in Escherichia coli which were reassembled in vitro. This excludes that an accessory protein contributes to the branch formation. Measurements of the filament cross sectional areas showed various ratios between the three bifurcation arms. This demonstrates that intermediate filament furcation is very different from actin furcation where an entire new filament is attached to an existing filament. Instead, the architecture of intermediate filament bifurcations is less predetermined and hence consistent with the general concept of IF formation. PMID:27039023

  3. Multi-Level Flow Branching

    Czech Academy of Sciences Publication Activity Database

    Tesař, Václav

    Praha: UTAM AV ČR, 2006 - (Náprstek, J.; Fischer, C.), s. 380-381 ISBN 80-86246-27-2. [Engineering mechanics 2006 : national conference with international participation. Svratka (CZ), 15.05.2006-18.05.2006] Institutional research plan: CEZ:AV0Z20760514 Keywords : branching * respiratory system * aortic system * microfluidics Subject RIV: BK - Fluid Dynamics

  4. Cash efficiency for bank branches.

    Science.gov (United States)

    Cabello, Julia García

    2013-01-01

    Bank liquidity management has become a major issue during the financial crisis as liquidity shortages have intensified and have put pressure on banks to diversity and improve their liquidity sources. While a significant strand of the literature concentrates on wholesale liquidity generation and on the alternative to deposit funding, the management of an inventory of cash holdings within the banks' branches is also a relevant issue as any significant improvement in cash management at the bank distribution channels may have a positive effect in reducing liquidity tensions. In this paper, we propose a simple programme of cash efficiency for the banks' branches, very easy to implement, which conform to a set of instructions to be imposed from the bank to their branches. This model proves to significantly reduce cash holdings at branches thereby providing efficiency improvements in liquidity management. The methodology we propose is based on the definition of some stochastic processes combined with renewal processes, which capture the random elements of the cash flow, before applying suitable optimization programmes to all the costs involved in cash movements. The classical issue of the Transaction Demand for the Cash and some aspects of Inventory Theory are also present. Mathematics Subject Classification (2000) C02, C60, E50. PMID:24010026

  5. Fluid Flow at Branching Junctions

    OpenAIRE

    Sochi, Taha

    2013-01-01

    The flow of fluids at branching junctions plays important kinematic and dynamic roles in most biological and industrial flow systems. The present paper highlights some key issues related to the flow of fluids at these junctions with special emphasis on the biological flow networks particularly blood transportation vasculature.

  6. Risk Factor Assessment Branch (RFAB)

    Science.gov (United States)

    The Risk Factor Assessment Branch (RFAB) focuses on the development, evaluation, and dissemination of high-quality risk factor metrics, methods, tools, technologies, and resources for use across the cancer research continuum, and the assessment of cancer-related risk factors in the population.

  7. NADP-malate Dehydrogenase Isoforms of Wheat Leaves under Drought: Their Localization, and Some physicochemical and Kinetic Properties

    Directory of Open Access Journals (Sweden)

    H.G. Babayev

    2015-09-01

    Full Text Available Changes in sub-cellular localization, isoenzyme spectrum and kinetic characteristics of NADP-malate dehydrogenase (NADP-MDH, EC 1.1.1.82 in Triticum durum Desf. genotypes with contrasting drought tolerance have been studied. In chloroplast and cytosol fractions of mesophyll cells of wheat flag leaves 70-75% and 25-30% of the total NADP-MDH activity were found to be localized, respectively. Three isoforms of NADP-MDH with molecular weights of 66, 74 and 86 kDa were revealed in the chloroplast fraction, whereas in the cytosolic fraction molecular weights of the isoenzymes were found to be 42, 66 and 74 kDa. Drought caused the formation of a new 90 kDa isoform of the enzyme in the chloroplast fraction in anthesis phase of ontogenesis. In the drought-tolerant genotype the appearance of the new isoform in the chloroplast fraction was accompanied by a more rapid increase in Km and Vmax contrary to the chloroplast fraction of the drought-sensitive genotype manifesting a slight decrease in these parameters, suggesting one of the adaptive traits in forming drought tolerance in C3 plants.

  8. Immunohistochemical analysis of aldehyde dehydrogenase isoforms and their association with estrogen-receptor status and disease progression in breast cancer

    Science.gov (United States)

    Opdenaker, Lynn M; Arnold, Kimberly M; Pohlig, Ryan T; Padmanabhan, Jayasree S; Flynn, Daniel C; Sims-Mourtada, Jennifer

    2014-01-01

    In many types of tumors, especially breast tumors, aldehyde dehydrogenase (ALDH) activity has been used to identify cancer stem-like cells within the tumor. The presence and quantity of these cells are believed to predict the response of tumors to chemotherapy. Therefore, identification and eradication of these cells would be necessary to cure the patient. However, there are 19 different ALDH isoforms that could contribute to the enzyme activity. ALDH1A1 and ALDH1A3 are among the isoforms mostly responsible for the increased ALDH activity observed in these stem-like cells, although the main isoforms vary in different tissues and tumor types. In the study reported here, we attempted to determine if ALDH1A1 or ALDH1A3, specifically, correlate with tumor stage, grade, and hormone-receptor status in breast-cancer patients. While there was no significant correlation between ALDH1A1 and any of the parameters tested, we were able to identify a positive correlation between ALDH1A3 and tumor stage in triple-negative cancers. In addition, ALDH1A3 was negatively correlated with estrogen-receptor status. Our data suggest that ALDH1A3 could be utilized as a marker to identify stem-like cells within triple-negative tumors. PMID:25540596

  9. Molecular regulation of skeletal muscle myosin heavy chain isoforms

    OpenAIRE

    Brown, David M.

    2015-01-01

    Research investigating the regulation of muscle fibre type has traditionally been conducted in vivo, analyzing global changes at a whole muscle level. Broadly, this thesis aimed to explore more “molecular” approaches, utilizing molecular and cell biology to understand the expression and regulation of myosin heavy chain (MyHC) isoforms as an indicator of muscle fibre composition. The mRNA expression profile of six MyHC isoform genes during C2C12 myogenesis was elucidated to reveal that the...

  10. A Review of Metallothionein Isoforms and their Role in Pathophysiology

    OpenAIRE

    Senthil kumar M; Manisenthil Kumar KT; Shyam Sunder A; Thirumoorthy N; Ganesh GNK; Chatterjee Malay

    2011-01-01

    Abstract The Metallothionein (MT) is a protein which has several interesting biological effects and has been demonstrated increase focus on the role of MT in various biological systems in the past three decades. The studies on the role of MT were limited with few areas like apoptosis and antioxidants in selected organs even fifty years after its discovery. Now acknowledge the exploration of various isoforms of MT such as MT-I, MT-II, MT-III and MT-IV and other isoforms in various biological s...

  11. The solution structures of two soybean calmodulin isoforms provide a structural basis for their selective target activation properties.

    Science.gov (United States)

    Ishida, Hiroaki; Huang, Hao; Yamniuk, Aaron P; Takaya, Yoshiaki; Vogel, Hans J

    2008-05-23

    The intracellular calcium ion is one of the most important secondary messengers in eukaryotic cells. Ca(2+) signals are translated into physiological responses by EF-hand calcium-binding proteins such as calmodulin (CaM). Multiple CaM isoforms occur in plant cells, whereas only a single CaM protein is found in animals. Soybean CaM isoform 1 (sCaM1) shares 90% amino acid sequence identity with animal CaM (aCaM), whereas sCaM4 is only 78% identical. These two sCaM isoforms have distinct target-enzyme activation properties and physiological functions. sCaM4 is highly expressed during the self-defense reaction of the plant and activates the enzyme nitric-oxide synthase (NOS), whereas sCaM1 is incapable of activating NOS. The mechanism of selective target activation by plant CaM isoforms is poorly understood. We have determined high resolution NMR solution structures of Ca(2+)-sCaM1 and -sCaM4. These were compared with previously determined Ca(2+)-aCaM structures. For the N-lobe of the protein, the solution structures of Ca(2+)-sCaM1, -sCaM4, and -aCaM all closely resemble each other. However, despite the high sequence identity with aCaM, the C-lobe of Ca(2+)-sCaM1 has a more open conformation and consequently a larger hydrophobic target-protein binding pocket than Ca(2+)-aCaM or -sCaM4, the presence of which was further confirmed through biophysical measurements. The single Val-144 --> Met substitution in the C-lobe of Ca(2+)-sCaM1, which restores its ability to activate NOS, alters the structure of the C-lobe to a more closed conformation resembling Ca(2+)-aCaM and -sCaM4. The relationships between the structural differences in the two Ca(2+)-sCaM isoforms and their selective target activation properties are discussed. PMID:18347016

  12. Physiological covalent regulation of rat liver branched-chain alpha-ketoacid dehydrogenase

    International Nuclear Information System (INIS)

    A radiochemical assay was developed for measuring branched-chain alpha-ketoacid dehydrogenase activity of Triton X-100 extracts of freeze-clamped rat liver. The proportion of active (dephosphorylated) enzyme was determined by measuring enzyme activities before and after activation of the complex with a broad-specificity phosphoprotein phosphatase. Hepatic branched-chain alpha-ketoacid dehydrogenase activity in normal male Wistar rats was 97% active but decreased to 33% active after 2 days on low-protein (8%) diet and to 13% active after 4 days on the same diet. Restricting protein intake of lean and obese female Zucker rats also caused inactivation of hepatic branched-chain alpha-ketoacid dehydrogenase complex. Essentially all of the enzyme was in the active state in rats maintained for 14 days on either 30 or 50% protein diets. This was also the case for rats maintained on a commercial chow diet (minimum 23% protein). However, maintaining rats on 20, 8, and 0% protein diets decreased the percentage of the active form of the enzyme to 58, 10, and 7% of the total, respectively. Fasting of chow-fed rats for 48 h had no effect on the activity state of hepatic branched-chain alpha-ketoacid dehydrogenase, i.e., 93% of the enzyme remained in the active state compared to 97% for chow-fed rats. However, hepatic enzyme of rats maintained on 8% protein diet was 10% active before starvation and 83% active after 2 days of starvation. Thus, dietary protein deficiency results in inactivation of hepatic branched-chain alpha-ketoacid dehydrogenase complex, presumably as a consequence of low hepatic levels of branched-chain alpha-ketoacids

  13. Characterization of the First α-(1→3) Branching Sucrases of the GH70 Family.

    Science.gov (United States)

    Vuillemin, Marlène; Claverie, Marion; Brison, Yoann; Séverac, Etienne; Bondy, Pauline; Morel, Sandrine; Monsan, Pierre; Moulis, Claire; Remaud-Siméon, Magali

    2016-04-01

    Leuconostoc citreumNRRL B-742 has been known for years to produce a highly α-(1→3)-branched dextran for which the synthesis had never been elucidated. In this work a gene coding for a putative α-transglucosylase of the GH70 family was identified in the reported genome of this bacteria and functionally characterized. From sucrose alone, the corresponding recombinant protein, named BRS-B, mainly catalyzed sucrose hydrolysis and leucrose synthesis. However, in the presence of sucrose and a dextran acceptor, the enzyme efficiently transferred the glucosyl residue from sucrose to linear α-(1→6) dextrans through the specific formation of α-(1→3) linkages. To date, BRS-B is the first reported α-(1→3) branching sucrase. Using a suitable sucrose/dextran ratio, a comb-like dextran with 50% of α-(1→3) branching was synthesized, suggesting that BRS-B is likely involved in the comb-like dextran produced byL. citreumNRRL B-742. In addition, data mining based on the search for specific sequence motifs allowed the identification of two genes putatively coding for branching sucrases in the genome ofLeuconostoc fallaxKCTC3537 andLactobacillus kunkeeiEFB6. Biochemical characterization of the corresponding recombinant enzymes confirmed their branching specificity, revealing that branching sucrases are not only found inL. citreumspecies. According to phylogenetic analyses, these enzymes are proposed to constitute a new subgroup of the GH70 family. PMID:26763236

  14. Deubiquitylating enzymes and disease

    Directory of Open Access Journals (Sweden)

    Baker Rohan T

    2008-10-01

    Full Text Available Abstract Deubiquitylating enzymes (DUBs can hydrolyze a peptide, amide, ester or thiolester bond at the C-terminus of UBIQ (ubiquitin, including the post-translationally formed branched peptide bonds in mono- or multi-ubiquitylated conjugates. DUBs thus have the potential to regulate any UBIQ-mediated cellular process, the two best characterized being proteolysis and protein trafficking. Mammals contain some 80–90 DUBs in five different subfamilies, only a handful of which have been characterized with respect to the proteins that they interact with and deubiquitylate. Several other DUBs have been implicated in various disease processes in which they are changed by mutation, have altered expression levels, and/or form part of regulatory complexes. Specific examples of DUB involvement in various diseases are presented. While no specific drugs targeting DUBs have yet been described, sufficient functional and structural information has accumulated in some cases to allow their rapid development. Publication history Republished from Current BioData's Targeted Proteins database (TPdb; http://www.targetedproteinsdb.com.

  15. Highly specific antibodies for co-detection of human choline kinase α1 and α2 isoforms.

    Directory of Open Access Journals (Sweden)

    Wei Cun See Too

    Full Text Available BACKGROUND: Choline kinase is the first enzyme in the CDP-choline pathway that synthesizes phosphatidylcholine, the major phospholipid in eukaryotic cell membranes. In humans, choline kinase exists as three isoforms (CKα1, α2, and β. Specific inhibition of CKα has been reported to selectively kill tumoral cells. Monoclonal and polyclonal antibodies against CKα used in previous studies to detect the level of this isozyme in different cellular or biochemical contexts were able to detect either the α1 or the α2 isoform. METHODOLOGY/PRINCIPAL FINDINGS: In this study, an antiserum against CKα was produced by immunizing rabbits with denatured, purified recombinant CKα2 full-length protein. This antiserum was highly specific for CKα when tested with extracts from different cell lines, and there was no cross reactivity with purified CKβ and other related proteins like human ethanolamine kinases (EK and yeast choline or ethanolamine kinases. The antiserum simultaneously detected both CKα1 and α2 isoforms in MCF-7 and HepG2 cell extracts, but not in HeLa, HCT-116, and mouse embryonic stem cell extracts. Subsequent protein dot blot assay of total CKα in a human normal/tumor protein array of 30 tissue samples by using the antiserum showed that CKα was not overexpressed in all tumor tissues when compared to their normal counterparts. Most striking differences between tumor and normal CKα expression levels were observed in kidney (11-fold higher in tumor and liver (15-fold lower in tumor samples. CONCLUSION/SIGNIFICANCE: Apart from its high sensitivity and specificity, the antiserum produced in this work, which does not require further purification, has the advantage of co-detecting both α1 and α2 isoforms in cell extracts for direct comparison of their expression levels.

  16. Branching processes in disease epidemics

    Science.gov (United States)

    Singh, Sarabjeet

    Branching processes have served as a model for chemical reactions, biological growth processes and contagion (of disease, information or fads). Through this connection, these seemingly different physical processes share some common universalities that can be elucidated by analyzing the underlying branching process. In this thesis, we focus on branching processes as a model for infectious diseases spreading between individuals belonging to different populations. The distinction between populations can arise from species separation (as in the case of diseases which jump across species) or spatial separation (as in the case of disease spreading between farms, cities, urban centers, etc). A prominent example of the former is zoonoses -- infectious diseases that spill from animals to humans -- whose specific examples include Nipah virus, monkeypox, HIV and avian influenza. A prominent example of the latter is infectious diseases of animals such as foot and mouth disease and bovine tuberculosis that spread between farms or cattle herds. Another example of the latter is infectious diseases of humans such as H1N1 that spread from one city to another through migration of infectious hosts. This thesis consists of three main chapters, an introduction and an appendix. The introduction gives a brief history of mathematics in modeling the spread of infectious diseases along with a detailed description of the most commonly used disease model -- the Susceptible-Infectious-Recovered (SIR) model. The introduction also describes how the stochastic formulation of the model reduces to a branching process in the limit of large population which is analyzed in detail. The second chapter describes a two species model of zoonoses with coupled SIR processes and proceeds into the calculation of statistics pertinent to cross species infection using multitype branching processes. The third chapter describes an SIR process driven by a Poisson process of infection spillovers. This is posed as a

  17. Role of p53 isoforms and aggregations in cancer.

    Science.gov (United States)

    Kim, SeJin; An, Seong Soo A

    2016-06-01

    p53 is a master regulatory protein that is involved in diverse cellular metabolic processes such as apoptosis, DNA repair, and cell cycle arrest. The protective function of p53 (in its homotetrameric form) as a tumor suppressor is lost in more than 50% of human cancers.Despite considerable experimental evidence suggesting the presence of multiple p53 states, it has been difficult to correlate the status of p53 with cancer response to treatments and clinical outcomes, which suggest the importance of complex but essential p53 regulatory pathways.Recent studies have indicated that the expression pattern of p53 isoforms may play a crucial role in regulating normal and cancer cell fates in response to diverse stresses. The human TP53 gene encodes at least 12 p53 isoforms, which are produced in normal tissue through alternative initiation of translation, usage of alternative promoters, and alternative splicing. Furthermore, some researchers have suggested that the formation of mutant p53 aggregates may be associated with cancer pathogenesis due to loss-of function (LoF), dominant-negative (DN), and gain-of function (GoF) effects.As different isoforms or the aggregation state of p53 may influence tumorigenesis, this review aims to examine the correlation of p53 isoforms and aggregation with cancer. PMID:27368003

  18. APPRIS: annotation of principal and alternative splice isoforms.

    Science.gov (United States)

    Rodriguez, Jose Manuel; Maietta, Paolo; Ezkurdia, Iakes; Pietrelli, Alessandro; Wesselink, Jan-Jaap; Lopez, Gonzalo; Valencia, Alfonso; Tress, Michael L

    2013-01-01

    Here, we present APPRIS (http://appris.bioinfo.cnio.es), a database that houses annotations of human splice isoforms. APPRIS has been designed to provide value to manual annotations of the human genome by adding reliable protein structural and functional data and information from cross-species conservation. The visual representation of the annotations provided by APPRIS for each gene allows annotators and researchers alike to easily identify functional changes brought about by splicing events. In addition to collecting, integrating and analyzing reliable predictions of the effect of splicing events, APPRIS also selects a single reference sequence for each gene, here termed the principal isoform, based on the annotations of structure, function and conservation for each transcript. APPRIS identifies a principal isoform for 85% of the protein-coding genes in the GENCODE 7 release for ENSEMBL. Analysis of the APPRIS data shows that at least 70% of the alternative (non-principal) variants would lose important functional or structural information relative to the principal isoform. PMID:23161672

  19. Antagonistic functions of LMNA isoforms in energy expenditure and lifespan.

    Science.gov (United States)

    Lopez-Mejia, Isabel C; de Toledo, Marion; Chavey, Carine; Lapasset, Laure; Cavelier, Patricia; Lopez-Herrera, Celia; Chebli, Karim; Fort, Philippe; Beranger, Guillaume; Fajas, Lluis; Amri, Ez Z; Casas, Francois; Tazi, Jamal

    2014-05-01

    Alternative RNA processing of LMNA pre-mRNA produces three main protein isoforms, that is, lamin A, progerin, and lamin C. De novo mutations that favor the expression of progerin over lamin A lead to Hutchinson-Gilford progeria syndrome (HGPS), providing support for the involvement of LMNA processing in pathological aging. Lamin C expression is mutually exclusive with the splicing of lamin A and progerin isoforms and occurs by alternative polyadenylation. Here, we investigate the function of lamin C in aging and metabolism using mice that express only this isoform. Intriguingly, these mice live longer, have decreased energy metabolism, increased weight gain, and reduced respiration. In contrast, progerin-expressing mice show increased energy metabolism and are lipodystrophic. Increased mitochondrial biogenesis is found in adipose tissue from HGPS-like mice, whereas lamin C-only mice have fewer mitochondria. Consistently, transcriptome analyses of adipose tissues from HGPS and lamin C-only mice reveal inversely correlated expression of key regulators of energy expenditure, including Pgc1a and Sfrp5. Our results demonstrate that LMNA encodes functionally distinct isoforms that have opposing effects on energy metabolism and lifespan in mammals. PMID:24639560

  20. Comparison of liver oncogenic potential among human RAS isoforms

    Science.gov (United States)

    Chung, Sook In; Moon, Hyuk; Ju, Hye-Lim; Kim, Dae Yeong; Cho, Kyung Joo; Ribback, Silvia; Dombrowski, Frank; Calvisi, Diego F.; Ro, Simon Weonsang

    2016-01-01

    Mutation in one of three RAS genes (i.e., HRAS, KRAS, and NRAS) leading to constitutive activation of RAS signaling pathways is considered a key oncogenic event in human carcinogenesis. Whether activated RAS isoforms possess different oncogenic potentials remains an unresolved question. Here, we compared oncogenic properties among RAS isoforms using liver-specific transgenesis in mice. Hydrodynamic transfection was performed using transposons expressing short hairpin RNA downregulating p53 and an activated RAS isoform, and livers were harvested at 23 days after gene delivery. No differences were found in the hepatocarcinogenic potential among RAS isoforms, as determined by both gross examination of livers and liver weight per body weight ratio (LW/BW) of mice expressing HRASQ61L, KRAS4BG12V and NRASQ61K. However, the tumorigenic potential differed significantly between KRAS splicing variants. The LW/BW ratio in KRAS4AG12V mice was significantly lower than in KRAS4BG12V mice (p mice lived significantly longer than KRRAS4BG12V mice (p mice displayed higher expression of the p16INK4A tumor suppressor when compared with KRAS4BG12V tumors. Forced overexpression of p16INK4A significantly reduced tumor growth in KRAS4BG12V mice, suggesting that upregulation of p16INK4A by KRAS4AG12V presumably delays tumor development driven by the latter oncogene. PMID:26799184

  1. Tropomyosin-binding properties modulate competition between tropomodulin isoforms.

    Science.gov (United States)

    Colpan, Mert; Moroz, Natalia A; Gray, Kevin T; Cooper, Dillon A; Diaz, Christian A; Kostyukova, Alla S

    2016-06-15

    The formation and fine-tuning of cytoskeleton in cells are governed by proteins that influence actin filament dynamics. Tropomodulin (Tmod) regulates the length of actin filaments by capping the pointed ends in a tropomyosin (TM)-dependent manner. Tmod1, Tmod2 and Tmod3 are associated with the cytoskeleton of non-muscle cells and their expression has distinct consequences on cell morphology. To understand the molecular basis of differences in the function and localization of Tmod isoforms in a cell, we compared the actin filament-binding abilities of Tmod1, Tmod2 and Tmod3 in the presence of Tpm3.1, a non-muscle TM isoform. Tmod3 displayed preferential binding to actin filaments when competing with other isoforms. Mutating the second or both TM-binding sites of Tmod3 destroyed its preferential binding. Our findings clarify how Tmod1, Tmod2 and Tmod3 compete for binding actin filaments. Different binding mechanisms and strengths of Tmod isoforms for Tpm3.1 contribute to their divergent functional capabilities. PMID:27091317

  2. Cloning, expression and alternative splicing of the novel isoform of hTCP11 gene

    DEFF Research Database (Denmark)

    Ma, Yong-xin; Zhang, Si-zhong; Wu, Qia-qing;

    2003-01-01

    To identify a novel isoform of hTCP11 gene and investigate its expression and alternative splicing.......To identify a novel isoform of hTCP11 gene and investigate its expression and alternative splicing....

  3. Lipolytic enzymes involving lipolysis in Teleost: Synteny, structure, tissue distribution, and expression in grass carp (Ctenopharyngodon idella).

    Science.gov (United States)

    Sun, Jian; Ji, Hong; Li, Xue-Xian; Shi, Xiao-Chen; Du, Zhen-Yu; Chen, Li-Qiao

    2016-08-01

    Lipolysis is the biochemical pathway responsible for the sequential hydrolysis of triacylglycerols (TAGs) stored in cellular lipid droplets. Three enzymes are known to participate in TAGs hydrolysis, including adipose triglyceride lipase (ATGL), hormone-sensitive lipase (HSL), and monoglyceride lipase (MGL), and each is present in mammals as only one isoform. Here we show that the genome of grass carp (Ctenopharyngodon idella) and other teleosts codes for one ATGL, two HSLs, and one MGL isoforms. Two isoforms of HSL gene, HSLa and HSLb, derived from paralogous genes that could be originated from teleost-specific genome duplication (TSGD) event. The genes encoding for fish ATGL and MGL were conserved and contained nine and seven coding exons, respectively. However, two isoforms of HSL gene had a remarkable variation in gene structure, such as HSLa gene contained ten and HSLb contained thirteen exons. All three enzymes, including two isoforms of HSL, were expressed in a wide range of tissues, but the abundance of each gene mRNA showed the tissue-dependent expression patterns. During fasting, only ATGL and HSLa showed a significant increase in adipose tissue and adipocyte, indicating that ATGL and HSLa may be the main rate-limiting enzymes controlling the hydrolysis of TAGs in fasting-induced lipolysis. Different expression of HSLa and HSLb suggests that they might serve different roles in fasting-induced lipolysis. These results provide evidence about the conservation and divergence of genes of fish lipolytic enzymes. PMID:27131420

  4. Branching problems of unitary representations

    OpenAIRE

    Kobayashi, Toshiyuki

    2003-01-01

    The irreducible decomposition of a unitary representation often contains continuous spectrum when restricted to a non-compact subgroup. The author singles out a nice class of branching problems where each irreducible summand occurs discretely with finite multiplicity (admissible restrictions). Basic theory and new perspectives of admissible restrictions are presented from both analytic and algebraic view points. We also discuss some applications of admissible restrictions to modular varieties...

  5. Cash efficiency for bank branches

    OpenAIRE

    Cabello, Julia García

    2013-01-01

    Bank liquidity management has become a major issue during the financial crisis as liquidity shortages have intensified and have put pressure on banks to diversity and improve their liquidity sources. While a significant strand of the literature concentrates on wholesale liquidity generation and on the alternative to deposit funding, the management of an inventory of cash holdings within the banks’ branches is also a relevant issue as any significant improvement in cash management at the bank ...

  6. Solid State Photovoltaic Research Branch

    Energy Technology Data Exchange (ETDEWEB)

    1990-09-01

    This report summarizes the progress of the Solid State Photovoltaic Research Branch of the Solar Energy Research Institute (SERI) from October 1, 1988, through September 30,l 1989. Six technical sections of the report cover these main areas of SERIs in-house research: Semiconductor Crystal Growth, Amorphous Silicon Research, Polycrystalline Thin Films, III-V High-Efficiency Photovoltaic Cells, Solid-State Theory, and Laser Raman and Luminescence Spectroscopy. Sections have been indexed separately for inclusion on the data base.

  7. Creep of welded branched pipes

    OpenAIRE

    Rayner, Glen

    2004-01-01

    Creep failure of welds in high-temperature power plant steam piping systems is known to be a potential cause of plant failure. Creep behaviour of plain pipes with circumferential welds and cross-weld specimens have received fairly extensive attention. However, research into the creep behaviour of welded thick-walled branched steam pipes has received less attention. Consequently, this thesis addresses improving the understanding of the creep behaviour for this type of geometry. Numerical and a...

  8. Interactions between Axillary Branches of Arabidopsis

    Institute of Scientific and Technical Information of China (English)

    Veronica Ongaro; Katherine Bainbridge; Lisa Williamson; Ottoline Leyser

    2008-01-01

    Studies of apical dominance have benefited greatly from two-branch assays in pea and bean,in which the shoot system is trimmed back to leave only two active cotyledonary axillary branches.In these two-branch shoots,a large body of evidence shows that one actively growing branch is able to inhibit the growth of the other,prompting studies on the nature of the inhibitory signals,which are still poorly understood.Here,we describe the establishment of two-branch assays in Arabidopsis,using consecutive branches on the bolting stem.As with the classical studies in pea and bean,these consecutive branches are able to inhibit one another's growth.Not only can the upper branch inhibit the lower branch,but also the lower branch can inhibit the upper branch,illustrating the bi-directional action of the inhibitory signals.Using mutants,we show that the inhibition is partially dependent on the MAX pathway and that while the inhibition is clearly transmitted across the stem from the active to the inhibited branch,the vascular connectivity of the two branches is weak,and the MAX pathway is capable of acting unilaterally in the stem.

  9. Branching processes and neutral evolution

    CERN Document Server

    Taïb, Ziad

    1992-01-01

    The Galton-Watson branching process has its roots in the problem of extinction of family names which was given a precise formulation by F. Galton as problem 4001 in the Educational Times (17, 1873). In 1875, an attempt to solve this problem was made by H. W. Watson but as it turned out, his conclusion was incorrect. Half a century later, R. A. Fisher made use of the Galton-Watson process to determine the extinction probability of the progeny of a mutant gene. However, it was J. B. S. Haldane who finally gave the first sketch of the correct conclusion. J. B. S. Haldane also predicted that mathematical genetics might some day develop into a "respectable branch of applied mathematics" (quoted in M. Kimura & T. Ohta, Theoretical Aspects of Population Genetics. Princeton, 1971). Since the time of Fisher and Haldane, the two fields of branching processes and mathematical genetics have attained a high degree of sophistication but in different directions. This monograph is a first attempt to apply the current sta...

  10. Multigene families encode the major enzymes of antioxidant metabolism in Eucalyptus grandis L

    OpenAIRE

    Felipe Karam Teixeira; Larissa Menezes-Benavente; Vinícius Costa Galvão; Márcia Margis-Pinheiro

    2005-01-01

    Antioxidant metabolism protects cells from oxidative damage caused by reactive oxygen species (ROS). In plants, several enzymes act jointly to maintain redox homeostasis. Moreover, isoform diversity contributes to the fine tuning necessary for plant responses to both exogenous and endogenous signals influencing antioxidant metabolism. This study aimed to provide a comprehensive view of the major classes of antioxidant enzymes in the woody species Eucalyptus grandis. A careful survey of the FO...

  11. Enzyme detection by microfluidics

    DEFF Research Database (Denmark)

    2013-01-01

    Microfluidic-implemented methods of detecting an enzyme, in particular a DNA-modifying enzyme, are provided, as well as methods for detecting a cell, or a microorganism expressing said enzyme. The enzyme is detected by providing a nucleic acid substrate, which is specifically targeted by that...... enzyme...

  12. Workshop on Branching Processes and Their Applications

    CERN Document Server

    Gonzalez Velasco, Miguel; Martinez, Rodrigo; Molina, Manuel

    2010-01-01

    Contains papers presented at the Workshop on Branching Processes and Their Applications (WBPA09), held in Badajoz, Spain, April 20-23, 2009, which deal with theoretical and practical aspects of branching process theory

  13. Mechanical properties of branched actin filaments

    CERN Document Server

    Razbin, Mohammadhosein; Benetatos, Panayotis; Zippelius, Annette

    2015-01-01

    Cells moving on a two dimensional substrate generate motion by polymerizing actin filament networks inside a flat membrane protrusion. New filaments are generated by branching off existing ones, giving rise to branched network structures. We investigate the force-extension relation of branched filaments, grafted on an elastic structure at one end and pushing with the free ends against the leading edge cell membrane. Single filaments are modeled as worm-like chains, whose thermal bending fluctuations are restricted by the leading edge cell membrane, resulting in an effective force. Branching can increase the stiffness considerably; however the effect depends on branch point position and filament orientation, being most pronounced for intermediate tilt angles and intermediate branch point positions. We describe filament networks without cross-linkers to focus on the effect of branching. We use randomly positioned branch points, as generated in the process of treadmilling, and orientation distributions as measur...

  14. Determination of the class and isoform selectivity of small-molecule histone deacetylase inhibitors

    DEFF Research Database (Denmark)

    Khan, N.; Jeffers, M.; Kumar, S.;

    2008-01-01

    ) against a panel of rhHDAC (recombinant human HDAC) isoforms. Eight rhHDACs were expressed using a baculoviral system, and a Fluor de Lystrade mark (Biomol International) HDAC assay was optimized for each purified isoform. The potency and selectivity of ten HDACs on class I isoforms (rhHDAC1, rhHDAC2, rh...

  15. Carbonic anhydrase activators: gold nanoparticles coated with derivatized histamine, histidine, and carnosine show enhanced activatory effects on several mammalian isoforms.

    Science.gov (United States)

    Saada, Mohamed-Chiheb; Montero, Jean-Louis; Vullo, Daniela; Scozzafava, Andrea; Winum, Jean-Yves; Supuran, Claudiu T

    2011-03-10

    Lipoic acid moieties were attached to amine or amino acids showing activating properties against the zinc enzyme carbonic anhydrase (CA, EC 4.2.1.1). The obtained lipoic acid conjugates of histamine, L-histidine methyl ester, and L-carnosine methyl ester were attached to gold nanoparticles (NPs) by reaction with Au(III) salts in reducing conditions. The CA activators (CAAs)-coated NPs showed low nanomolar activation (K(A)s of 1-9 nM) of relevant cytosolic, membrane-bound, mitochondrial, and transmembrane CA isoforms, such as CA I, II, IV, VA, VII, and XIV. These NPs also effectively activated CAs ex vivo, in whole blood experiments, with an increase of 200-280% of the CA activity. This is the first example of enzyme activation with nanoparticles and may lead to biomedical applications for conditions in which the CA activity is diminished, such as aging, Alzheimer's disease, or CA deficiency syndrome. PMID:21291238

  16. Targeted Expression of Stromelysin-1 in Mammary Gland Provides Evidence for a Role of Proteinases in Branching Morphogenesis and the Requirement for an Intact Basement Membrane for Tissue-specific Gene Expression

    Energy Technology Data Exchange (ETDEWEB)

    Sympson, Carolyn J; Talhouk, Rabih S; Alexander, Caroline M; Chin, Jennie R; Cliff, Shirley M; Bissell, Mina J; Werb, Zena

    1994-05-01

    The extracellular matrix (ECM) is an important regulator of the differentiated phenotype of mammary epithelial cells in culture. Despite the fact that ECM-degrading enzymes have been implicated in morphogenesis and tissue remodeling, there is little evidence for a direct role for such regulation in vivo. We generated transgenic mice that express autoactivated isoforms of the matrix metalloproteinase stromelysin-1, under the control of the whey acidic protein gene promoter, to examine the effect of inappropriate expression of this enzyme. Stromelysin-1 is implicated as the primary player in the loss of basement membrane and loss of function in the mammary gland during involution. The transgene was expressed at low levels in mammary glands of virgin female mice, leading to an unexpected phenotype: The primary ducts had supernumerary branches and showed precocious development of alveoli that expressed beta-casein at levels similar to that of an early- to mid-pregnant gland. Lactating glands showed high levels of transgene expression, with accumulation at the basement membrane, and a decrease in laminin and collagen IV, resulting in a loss of basement membrane integrity; this was accompanied by a dramatic alteration of alveolar morphology, with decreased size and shrunken lumina containing little beta-casein. During pregnancy, expression of endogenous whey acidic protein and beta-casein was reduced in transgenic glands, confirming the observed dependence of milk protein transcription of ECM in mammary epithelial cells in culture. These data provide direct evidence that stromelysin-1 activity can be morphogenic for mammary epithelial cells, inducing hyperproliferation and differentiation in virgin animals, and that its lytic activity can, indeed, disrupt membrane integrity and reduce mammary-specific function. We conclude that the balance of ECM-degrading enzymes with their inhibitors, and the associated regulation of ECM structure, is crucial for tissue-specific gene

  17. Characterization of rabbit CD5 isoforms.

    Science.gov (United States)

    Pospisil, Richard; Kabat, Juraj; Mage, Rose G

    2009-08-01

    Previously described polyclonal or monoclonal antibodies (mAb) to rabbit CD5, raised against expressed recombinant protein or peptides, recognize CD5 on most rabbit B cells. The mAb KEN-5 was originally reported to recognize rabbit CD5. However, KEN-5 binds almost exclusively to T cells and only to a minor population of B cells. We show here that by Enzyme-linked Immunosorbent Assay (ELISA), KEN-5 binds to recombinant rabbit CD5. This interaction is partially inhibited by polyclonal goat anti-CD5 antibody. In addition, immunoprecipitations from lysates of surface biotinylated rabbit lymphocytes with KEN-5 or our anti-CD5 mAb isolate molecules that migrate identically on gels with the same approximate relative molecular mass of 67,000 M(r). By flow cytometric analyses of individual cells from spleen, thymus and appendix, KEN-5 recognizes CD5-like molecules mainly on T cells and on 3-6% of IgM(+) B cells. Immunohistochemical staining of splenic and appendix tissues and confocal immunofluorescent imaging confirm and extend results from flow cytometric analyses. Quantitation of fluorescent colocalization indicates that staining by KEN-5 colocalizes with staining by anti-CD5 on small percentage lymphocytes in splenic tissue sections. As CD5 has both N- and O-linked glycosylation, we hypothesised that differential binding of KEN-5 to T cells and B-cells may be explained by different glycan structures on the CD5 present on T compared to B cells. This hypothesis is supported by ELISA data that show that deglycosylation diminishes the binding of KEN-5 to recombinant rabbit CD5. Screening KEN-5 on an array with 406 glycans was inconclusive. Although we did not identify a strongly binding glycan structure, the data are suggestive that the epitope recognized by KEN-5 may be influenced by glycan structures. The epitope this mAb recognizes may either be the glycan itself, or more likely, is influenced by neighboring glycan structure. Our findings suggest that development

  18. Non-homologous isofunctional enzymes: A systematic analysis of alternative solutions in enzyme evolution

    Directory of Open Access Journals (Sweden)

    Wolf Yuri I

    2010-04-01

    Full Text Available Abstract Background Evolutionarily unrelated proteins that catalyze the same biochemical reactions are often referred to as analogous - as opposed to homologous - enzymes. The existence of numerous alternative, non-homologous enzyme isoforms presents an interesting evolutionary problem; it also complicates genome-based reconstruction of the metabolic pathways in a variety of organisms. In 1998, a systematic search for analogous enzymes resulted in the identification of 105 Enzyme Commission (EC numbers that included two or more proteins without detectable sequence similarity to each other, including 34 EC nodes where proteins were known (or predicted to have distinct structural folds, indicating independent evolutionary origins. In the past 12 years, many putative non-homologous isofunctional enzymes were identified in newly sequenced genomes. In addition, efforts in structural genomics resulted in a vastly improved structural coverage of proteomes, providing for definitive assessment of (nonhomologous relationships between proteins. Results We report the results of a comprehensive search for non-homologous isofunctional enzymes (NISE that yielded 185 EC nodes with two or more experimentally characterized - or predicted - structurally unrelated proteins. Of these NISE sets, only 74 were from the original 1998 list. Structural assignments of the NISE show over-representation of proteins with the TIM barrel fold and the nucleotide-binding Rossmann fold. From the functional perspective, the set of NISE is enriched in hydrolases, particularly carbohydrate hydrolases, and in enzymes involved in defense against oxidative stress. Conclusions These results indicate that at least some of the non-homologous isofunctional enzymes were recruited relatively recently from enzyme families that are active against related substrates and are sufficiently flexible to accommodate changes in substrate specificity. Reviewers This article was reviewed by Andrei

  19. Heterodimeric l-amino acid oxidase enzymes from Egyptian Cerastes cerastes venom: Purification, biochemical characterization and partial amino acid sequencing

    OpenAIRE

    A.E. El Hakim; W.H. Salama; M.B. Hamed; Ali, A. A.; N.M. Ibrahim

    2015-01-01

    Two l-amino acid oxidase enzyme isoforms, Cc-LAAOI and Cc-LAAOII were purified to apparent homogeneity from Cerastes cerastes venom in a sequential two-step chromatographic protocol including; gel filtration and anion exchange chromatography. The native molecular weights of the isoforms were 115 kDa as determined by gel filtration on calibrated Sephacryl S-200 column, while the monomeric molecular weights of the enzymes were, 60, 56 kDa and 60, 53 kDa for LAAOI and LAAOII, respectively. The t...

  20. EFSA Panel on Food Contact Materials, Enzymes, Flavourings and Processing Aids (CEF); Scientific Opinion on Flavouring Group Evaluation 06, Revision 3 (FGE.06Rev3): Straight- and branched-chain aliphatic unsaturated primary alcohols, aldehydes, carboxylic acids, and esters from chemical groups 1

    DEFF Research Database (Denmark)

    Larsen, John Christian; Nørby, Karin Kristiane; Beltoft, Vibe Meister; Lund, Pia; Binderup, Mona-Lise; Frandsen, Henrik Lauritz

    The Panel on Food Contact Materials, Enzymes, Flavourings and Processing Aids of the European Food Safety Authority was requested to evaluate 50 flavouring substances in the Flavouring Group Evaluation 6, Revision 3, using the Procedure in Commission Regulation (EC) No 1565/2000. None of the subs...

  1. The Effects of a Branch Campus

    Science.gov (United States)

    Lien, Donald; Wang, Yaqin

    2012-01-01

    We examine the effects of a branch campus on the social welfare of the host country and the foreign university. Overall, we find that a branch campus increases both the domestic social welfare (measured by the aggregate student utility) and the tuition revenue of the foreign university. The effect of a branch campus on the brain drain is…

  2. One isoform of Arg/Abl2 tyrosine kinase is nuclear and the other seven cytosolic isoforms differently modulate cell morphology, motility and the cytoskeleton

    Energy Technology Data Exchange (ETDEWEB)

    Bianchi, Cristina; Torsello, Barbara; Di Stefano, Vitalba; Zipeto, Maria A.; Facchetti, Rita; Bombelli, Silvia; Perego, Roberto A., E-mail: roberto.perego@unimib.it

    2013-08-01

    The non-receptor tyrosine kinase Abelson related gene (Arg/Abl2) regulates cell migration and morphogenesis by modulating the cytoskeleton. Arg promotes actin-based cell protrusions and spreading, and inhibits cell migration by attenuating stress fiber formation and contractility via activation of the RhoA inhibitor, p190RhoGAP, and by regulating focal adhesion dynamics also via CrkII phosphorylation. Eight full-length Arg isoforms with different N- and C-termini are endogenously expressed in human cells. In this paper, the eight Arg isoforms, subcloned in the pFLAG-CMV2 vector, were transfected in COS-7 cells in order to study their subcellular distribution and role in cell morphology, migration and cytoskeletal modulation. The transfected 1BSCTS Arg isoform has a nuclear distribution and phosphorylates CrkII in the nucleus, whilst the other isoforms are detected in the cytoplasm. The 1BLCTL, 1BSCTL, 1ASCTS isoforms were able to significantly decrease stress fibers, induce cell shrinkage and filopodia-like protrusions with a significant increase in p190RhoGAP phosphorylation. In contrast, 1ALCTL, 1ALCTS, 1ASCTL and 1BLCTS isoforms do not significantly decrease stress fibers and induce the formation of retraction tail-like protrusions. The 1BLCTL and 1ALCTL isoforms have different effects on cell migration and focal adhesions. All these data may open new perspectives to study the mechanisms of cell invasiveness. -Highlights: • Each of the eight Arg isoforms was transfected in COS-7 cells. • Only the 1BSCTS Arg isoform has a nuclear distribution in transfected cells. • The cytoplasmic isoforms and F-actin colocalize cortically and in cell protrusions. • Arg isoforms differently phosphorylate p190RhoGAP and CrkII. • Arg isoforms differently modulate stress fibers, cell protrusions and motility.

  3. Enzyme immobilization: an update

    OpenAIRE

    Homaei, Ahmad Abolpour; Sariri, Reyhaneh; Vianello, Fabio; Stevanato, Roberto

    2013-01-01

    Compared to free enzymes in solution, immobilized enzymes are more robust and more resistant to environmental changes. More importantly, the heterogeneity of the immo-bilized enzyme systems allows an easy recovery of both enzymes and products, multiple re-use of enzymes, continuous operation of enzymatic processes, rapid termination of reactions, and greater variety of bioreactor designs. This paper is a review of the recent literatures on enzyme immobilization by various techniques, the need...

  4. Differential regulation of macropinocytosis by Abi1/Hssh3bp1 isoforms.

    Directory of Open Access Journals (Sweden)

    Patrycja M Dubielecka

    Full Text Available BACKGROUND: Macropinocytosis, which is a constitutive cellular process of fluid and macromolecule uptake, is regulated by actin cytoskeleton rearrangements near the plasma membrane. Activation of Rac1, which is proposed to act upstream of the actin polymerization regulatory Wave 2 complex, has been found to correlate with enhanced macropinocytosis. One of the components of the Wave 2 complex is Abi1. Multiple, alternatively spliced isoforms of Abi1 are expressed in mammalian cells, but the functional significance of the various isoforms is unknown. PRINCIPAL FINDINGS: Here, using flow cytometric assay analysis for Alexa Fluor 647, we demonstrate that Abi1 isoforms 2 and 3 differentially regulate macropinocytosis. LNCaP cells expressing isoform 3 had increased macropinocytic uptake that correlated with enhanced cell spreading and higher Rac1 activation in comparison to cells expressing isoform 2. Isoform 2 expressing cells had decreased macropinocytic uptake, but demonstrated greater sensitivity to Rac1 activation. Moreover, more isoform 2 was localized within the cytoplasm in comparison to isoform 3, which was more associated with the plasma membrane. Activated Rac1 was found to specifically bind to a site in exon 10 of isoform 2 in vitro. Because of alternative mRNA splicing, exon 10 is absent from isoform 3, precluding similar binding of activated Rac1. Both isoforms, however, bound to inactive Rac1 through the same non-exon 10 site. Thus, Abi1 isoform 3-containing Wave 2 complex exhibited a differential binding to activated vs. inactive Rac1, whereas isoform 2-containing Wave 2 complex bound activated or inactive Rac1 comparably. CONCLUSION: Based on these observations, we postulate that Abi1 isoforms differentially regulate macropinocytosis as a consequence of their different relative affinities for activated Rac1 in Wave 2 complex. These findings also raise the possibility that isoform-specific roles occur in other Abi1 functions.

  5. Photo-control of nitric oxide synthase activity using a caged isoform specific inhibitor.

    Science.gov (United States)

    Montgomery, Heather J; Perdicakis, Basil; Fishlock, Dan; Lajoie, Gilles A; Jervis, Eric; Guy Guillemette, J

    2002-06-01

    Nitric oxide (NO) plays a critical role in a number of physiological processes and is produced in mammalian cells by nitric oxide synthase (NOS) isozymes. Because of the diverse functions of NO, pharmaceutical interventions which seek to abrogate adverse effects of excess NOS activity must not interfere with the normal regulation of NO levels in the body. A method has been developed for the control of NOS enzyme activity using the localized photochemical release of a caged isoform-specific NOS inhibitor. The caged form of an iNOS inhibitor has been synthesized and tested for photosensitivity and potency. UV and multiphoton uncaging were verified using a hemoglobin-based assay. IC(50) values were determined for the inhibitor (70+/-11 nM), the caged inhibitor (1098+/-172 nM), the UV uncaged inhibitor (67+/-26 nM) and the multiphoton uncaged inhibitor (73+/-11 nM). UV irradiation of the caged inhibitor resulted in a 86% reduction in iNOS activity after 5 min. Multiphoton uncaging had an apparent first order time constant of 0.007+/-0.001 min(-1). A therapeutic range exists, with molar excess of inhibitor to enzyme from 3- to 7-fold, over which the full dynamic range of the inhibition can be exploited. PMID:11937350

  6. EGFR soluble isoforms and their transcripts are expressed in meningiomas.

    Science.gov (United States)

    Guillaudeau, Angélique; Durand, Karine; Bessette, Barbara; Chaunavel, Alain; Pommepuy, Isabelle; Projetti, Fabrice; Robert, Sandrine; Caire, François; Rabinovitch-Chable, Hélène; Labrousse, François

    2012-01-01

    The EGFR (epidermal growth factor receptor) is involved in the oncogenesis of many tumors. In addition to the full-length EGFR (isoform a), normal and tumor cells produce soluble EGFR isoforms (sEGFR) that lack the intracellular domain. sEGFR isoforms b, c and d are encoded by EGFR variants 2 (v2), 3 (v3) and 4 (v4) mRNA resulting from gene alternative splicing. Accordingly, the results of EGFR protein expression analysis depend on the domain targeted by the antibodies. In meningiomas, EGFR expression investigations mainly focused on EGFR isoform a. sEGFR and EGFRvIII mutant, that encodes a constitutively active truncated receptor, have not been studied. In a 69 meningiomas series, protein expression was analyzed by immunohistochemistry using extracellular domain targeted antibody (ECD-Ab) and intracellular domain targeted antibody (ICD-Ab). EGFRv1 to v4 and EGFRvIII mRNAs were quantified by RT-PCR and EGFR amplification revealed by MLPA. Results were analyzed with respect to clinical data, tumor resection (Simpson grade), histological type, tumor grade, and patient outcome.Immunochemical staining was stronger with ECD-Ab than with ICD-Ab. Meningiomas expressed EGFRv1 to -v4 mRNAs but not EGFRvIII mutant. Intermediate or high ECD-Ab staining and high EGFRv1 to v4 mRNA levels were associated to a better progression free survival (PFS). PFS was also improved in women, when tumor resection was evaluated as Simpson 1 or 2, in grade I vs. grade II and III meningiomas and when Ki67 labeling index was lower than 10%. Our results suggest that, EGFR protein isoforms without ICD and their corresponding mRNA variants are expressed in meningiomas in addition to the whole isoform a. EGFRvIII was not expressed. High expression levels seem to be related to a better prognosis. These results indicate that the oncogenetic mechanisms involving the EGFR pathway in meningiomas could be different from other tumor types. PMID:22623992

  7. EGFR soluble isoforms and their transcripts are expressed in meningiomas.

    Directory of Open Access Journals (Sweden)

    Angélique Guillaudeau

    Full Text Available The EGFR (epidermal growth factor receptor is involved in the oncogenesis of many tumors. In addition to the full-length EGFR (isoform a, normal and tumor cells produce soluble EGFR isoforms (sEGFR that lack the intracellular domain. sEGFR isoforms b, c and d are encoded by EGFR variants 2 (v2, 3 (v3 and 4 (v4 mRNA resulting from gene alternative splicing. Accordingly, the results of EGFR protein expression analysis depend on the domain targeted by the antibodies. In meningiomas, EGFR expression investigations mainly focused on EGFR isoform a. sEGFR and EGFRvIII mutant, that encodes a constitutively active truncated receptor, have not been studied. In a 69 meningiomas series, protein expression was analyzed by immunohistochemistry using extracellular domain targeted antibody (ECD-Ab and intracellular domain targeted antibody (ICD-Ab. EGFRv1 to v4 and EGFRvIII mRNAs were quantified by RT-PCR and EGFR amplification revealed by MLPA. Results were analyzed with respect to clinical data, tumor resection (Simpson grade, histological type, tumor grade, and patient outcome.Immunochemical staining was stronger with ECD-Ab than with ICD-Ab. Meningiomas expressed EGFRv1 to -v4 mRNAs but not EGFRvIII mutant. Intermediate or high ECD-Ab staining and high EGFRv1 to v4 mRNA levels were associated to a better progression free survival (PFS. PFS was also improved in women, when tumor resection was evaluated as Simpson 1 or 2, in grade I vs. grade II and III meningiomas and when Ki67 labeling index was lower than 10%. Our results suggest that, EGFR protein isoforms without ICD and their corresponding mRNA variants are expressed in meningiomas in addition to the whole isoform a. EGFRvIII was not expressed. High expression levels seem to be related to a better prognosis. These results indicate that the oncogenetic mechanisms involving the EGFR pathway in meningiomas could be different from other tumor types.

  8. Secretory pathway Ca2+/Mn2+-ATPase isoform 2 and lactation: specific localization of plasmalemmal and secretory pathway Ca2+ pump isoforms in the mammary gland

    Energy Technology Data Exchange (ETDEWEB)

    Faddy, Helen M.; Smart, Chanel E.; Xu, Ren; Lee, Genee Y.; Kenny, Paraic A.; Feng, Mingye; Rao, Rajini; Brown, Melissa A.; Bissell, Mina J.; Roberts-Thomson, Sarah J.; Monteith, Gregory R.

    2008-04-09

    The supply of calcium to the developing neonate via milk is an important physiological process. Until recently the mechanism for the enrichment of milk with calcium was thought to be almost entirely mediated via the secretory pathway. However, recent studies suggest that a specific isoform of the plasma membrane calcium ATPase, PMCA2, is the primary mechanism for calcium transport into milk, highlighting a major role for apical calcium transport. We compared the expression of the recently identified secretory calcium ATPase, SPCA2, and SPCA1, in the mouse mammary gland during different stages of development. SPCA2 levels increased over 35 fold during lactation, while SPCA1 increased only a modest two fold. The potential importance of SPCA2 in lactation was also highlighted by its localization to luminal secretory cells of the mammary gland during lactation, while SPCA1 was expressed throughout the cells of the mammary gland. We also observed major differences in the localization of PMCA2 and PMCA1 during lactation. Using the SCp2 mouse mammary epithelial cell 3D culture model, differences in the sub-cellular distribution of PMCA2 and PMCA1 were clear. These studies highlight the likely specific roles of PMCA2 and SPCA2 in lactation, and link the recently characterized SPCA2 calcium pump to the supply of calcium into milk and the regulation of Golgi resident enzymes important in lactation. They also indicate that calcium transport into milk is a complex interplay between apical and secretory pathways.

  9. Stochastic Transition between Turbulent Branch and Thermodynamic Branch of an Inhomogeneous Plasma

    OpenAIRE

    Kawasaki, Mitsuhiro; Itoh, Sanae-I.; Yagi, Masatoshi; Itoh, Kimitaka

    2002-01-01

    Transition phenomena between thermodynamic branch and turbulent branch in submarginal turbulent plasma are analyzed with statistical theory. Time-development of turbulent fluctuation is obtained by numerical simulations of Langevin equation which contains submarginal characteristics. Probability density functions and transition rates between two states are analyzed. Transition from turbulent branch to thermodynamic branch occurs in almost entire region between subcritical bifurcation point an...

  10. Identification and Analysis of the Role of Superoxide Dismutases Isoforms in the Pathogenesis of Paracoccidioides spp.

    Directory of Open Access Journals (Sweden)

    Diana Tamayo

    2016-03-01

    Full Text Available The ability of Paracoccidioides to defend itself against reactive oxygen species (ROS produced by host effector cells is a prerequisite to survive. To counteract these radicals, Paracoccidioides expresses, among different antioxidant enzymes, superoxide dismutases (SODs. In this study, we identified six SODs isoforms encoded by the Paracoccidioides genome. We determined gene expression levels of representative isolates of the phylogenetic lineages of Paracoccidioides spp. (S1, PS2, PS3 and Pb01-like using quantitative RT-PCR. Assays were carried out to analyze SOD gene expression of yeast cells, mycelia cells, the mycelia-to-yeast transition and the yeast-to-mycelia germination, as well as under treatment with oxidative agents and during interaction with phagocytic cells. We observed an increased expression of PbSOD1 and PbSOD3 during the transition process, exposure to oxidative agents and interaction with phagocytic cells, suggesting that these proteins could assist in combating the superoxide radicals generated during the host-pathogen interaction. Using PbSOD1 and PbSOD3 knockdown strains we showed these genes are involved in the response of the fungus against host effector cells, particularly the oxidative stress response, and in a mouse model of infection. Protein sequence analysis together with functional analysis of knockdown strains seem to suggest that PbSOD3 expression is linked with a pronounced extracellular activity while PbSOD1 seems more related to intracellular requirements of the fungus. Altogether, our data suggests that P. brasiliensis actively responds to the radicals generated endogenously during metabolism and counteracts the oxidative burst of immune cells by inducing the expression of SOD isoforms.

  11. Identification and Analysis of the Role of Superoxide Dismutases Isoforms in the Pathogenesis of Paracoccidioides spp.

    Science.gov (United States)

    Tamayo, Diana; Muñoz, José F; Lopez, Ángela; Urán, Martha; Herrera, Juan; Borges, Clayton L; Restrepo, Ángela; Soares, Celia M; Taborda, Carlos P; Almeida, Agostinho J; McEwen, Juan G; Hernández, Orville

    2016-03-01

    The ability of Paracoccidioides to defend itself against reactive oxygen species (ROS) produced by host effector cells is a prerequisite to survive. To counteract these radicals, Paracoccidioides expresses, among different antioxidant enzymes, superoxide dismutases (SODs). In this study, we identified six SODs isoforms encoded by the Paracoccidioides genome. We determined gene expression levels of representative isolates of the phylogenetic lineages of Paracoccidioides spp. (S1, PS2, PS3 and Pb01-like) using quantitative RT-PCR. Assays were carried out to analyze SOD gene expression of yeast cells, mycelia cells, the mycelia-to-yeast transition and the yeast-to-mycelia germination, as well as under treatment with oxidative agents and during interaction with phagocytic cells. We observed an increased expression of PbSOD1 and PbSOD3 during the transition process, exposure to oxidative agents and interaction with phagocytic cells, suggesting that these proteins could assist in combating the superoxide radicals generated during the host-pathogen interaction. Using PbSOD1 and PbSOD3 knockdown strains we showed these genes are involved in the response of the fungus against host effector cells, particularly the oxidative stress response, and in a mouse model of infection. Protein sequence analysis together with functional analysis of knockdown strains seem to suggest that PbSOD3 expression is linked with a pronounced extracellular activity while PbSOD1 seems more related to intracellular requirements of the fungus. Altogether, our data suggests that P. brasiliensis actively responds to the radicals generated endogenously during metabolism and counteracts the oxidative burst of immune cells by inducing the expression of SOD isoforms. PMID:26963091

  12. Identification and Analysis of the Role of Superoxide Dismutases Isoforms in the Pathogenesis of Paracoccidioides spp.

    Science.gov (United States)

    Tamayo, Diana; Muñoz, José F.; Lopez, Ángela; Urán, Martha; Herrera, Juan; Borges, Clayton L.; Restrepo, Ángela; Soares, Celia M.; Taborda, Carlos P.; Almeida, Agostinho J.; McEwen, Juan G.; Hernández, Orville

    2016-01-01

    The ability of Paracoccidioides to defend itself against reactive oxygen species (ROS) produced by host effector cells is a prerequisite to survive. To counteract these radicals, Paracoccidioides expresses, among different antioxidant enzymes, superoxide dismutases (SODs). In this study, we identified six SODs isoforms encoded by the Paracoccidioides genome. We determined gene expression levels of representative isolates of the phylogenetic lineages of Paracoccidioides spp. (S1, PS2, PS3 and Pb01-like) using quantitative RT-PCR. Assays were carried out to analyze SOD gene expression of yeast cells, mycelia cells, the mycelia-to-yeast transition and the yeast-to-mycelia germination, as well as under treatment with oxidative agents and during interaction with phagocytic cells. We observed an increased expression of PbSOD1 and PbSOD3 during the transition process, exposure to oxidative agents and interaction with phagocytic cells, suggesting that these proteins could assist in combating the superoxide radicals generated during the host-pathogen interaction. Using PbSOD1 and PbSOD3 knockdown strains we showed these genes are involved in the response of the fungus against host effector cells, particularly the oxidative stress response, and in a mouse model of infection. Protein sequence analysis together with functional analysis of knockdown strains seem to suggest that PbSOD3 expression is linked with a pronounced extracellular activity while PbSOD1 seems more related to intracellular requirements of the fungus. Altogether, our data suggests that P. brasiliensis actively responds to the radicals generated endogenously during metabolism and counteracts the oxidative burst of immune cells by inducing the expression of SOD isoforms. PMID:26963091

  13. Defects in Peroxisomal 6-Phosphogluconate Dehydrogenase Isoform PGD2 Prevent Gametophytic Interaction in Arabidopsis thaliana.

    Science.gov (United States)

    Hölscher, Christian; Lutterbey, Marie-Christin; Lansing, Hannes; Meyer, Tanja; Fischer, Kerstin; von Schaewen, Antje

    2016-05-01

    We studied the localization of 6-phosphogluconate dehydrogenase (PGD) isoforms of Arabidopsis (Arabidopsis thaliana). Similar polypeptide lengths of PGD1, PGD2, and PGD3 obscured which isoform may represent the cytosolic and/or plastidic enzyme plus whether PGD2 with a peroxisomal targeting motif also might target plastids. Reporter-fusion analyses in protoplasts revealed that, with a free N terminus, PGD1 and PGD3 accumulate in the cytosol and chloroplasts, whereas PGD2 remains in the cytosol. Mutagenesis of a conserved second ATG enhanced the plastidic localization of PGD1 and PGD3 but not PGD2. Amino-terminal deletions of PGD2 fusions with a free C terminus resulted in peroxisomal import after dimerization, and PGD2 could be immunodetected in purified peroxisomes. Repeated selfing of pgd2 transfer (T-)DNA alleles yielded no homozygous mutants, although siliques and seeds of heterozygous plants developed normally. Detailed analyses of the C-terminally truncated PGD2-1 protein showed that peroxisomal import and catalytic activity are abolished. Reciprocal backcrosses of pgd2-1 suggested that missing PGD activity in peroxisomes primarily affects the male gametophyte. Tetrad analyses in the quartet1-2 background revealed that pgd2-1 pollen is vital and in vitro germination normal, but pollen tube growth inside stylar tissues appeared less directed. Mutual gametophytic sterility was overcome by complementation with a genomic construct but not with a version lacking the first ATG. These analyses showed that peroxisomal PGD2 activity is required for guided growth of the male gametophytes and pollen tube-ovule interaction. Our report finally demonstrates an essential role of oxidative pentose-phosphate pathway reactions in peroxisomes, likely needed to sustain critical levels of nitric oxide and/or jasmonic acid, whose biosynthesis both depend on NADPH provision. PMID:26941195

  14. Functional representation of enzymes by specific peptides.

    Directory of Open Access Journals (Sweden)

    Vered Kunik

    2007-08-01

    Full Text Available Predicting the function of a protein from its sequence is a long-standing goal of bioinformatic research. While sequence similarity is the most popular tool used for this purpose, sequence motifs may also subserve this goal. Here we develop a motif-based method consisting of applying an unsupervised motif extraction algorithm (MEX to all enzyme sequences, and filtering the results by the four-level classification hierarchy of the Enzyme Commission (EC. The resulting motifs serve as specific peptides (SPs, appearing on single branches of the EC. In contrast to previous motif-based methods, the new method does not require any preprocessing by multiple sequence alignment, nor does it rely on over-representation of motifs within EC branches. The SPs obtained comprise on average 8.4 +/- 4.5 amino acids, and specify the functions of 93% of all enzymes, which is much higher than the coverage of 63% provided by ProSite motifs. The SP classification thus compares favorably with previous function annotation methods and successfully demonstrates an added value in extreme cases where sequence similarity fails. Interestingly, SPs cover most of the annotated active and binding site amino acids, and occur in active-site neighboring 3-D pockets in a highly statistically significant manner. The latter are assumed to have strong biological relevance to the activity of the enzyme. Further filtering of SPs by biological functional annotations results in reduced small subsets of SPs that possess very large enzyme coverage. Overall, SPs both form a very useful tool for enzyme functional classification and bear responsibility for the catalytic biological function carried out by enzymes.

  15. PERSONALISED DENTURES WITH BRANCHING TECHNIQUE

    Directory of Open Access Journals (Sweden)

    Puneet

    2012-12-01

    Full Text Available ABSTRACT: While the basic process of making dentures has chang ed little over the past several decades, new materials and techniques can help labor atories and clinicians provide functional, esthetic restorations that offer exceptional value t o patients. Unlike the conventional “linear” methods which foll ow specific steps in a cookbook fashion, the Branching Technique is a dynamic conce pt which can be adjusted to meet the clinical needs of each patient. This customized tec hnique allows for a “trial” denture to be worn by each patient that is used to pinpoint and solve th e patient’s denture problems before the final denture is made. This individualized approach helps t o eliminate disappointments or surprises. From the preliminary impressions and the “training” d enture (it’s actually a provisional denture to the functional impressions and porcelain teeth to hold the patient’s vertical dimension, the Branching Technique produces the most esthetically pleasing and functionally stable dentures possible.

  16. Differential regulation of renal phospholipase C isoforms by catecholamines.

    OpenAIRE

    Yu, P Y; Asico, L D; Eisner, G M; Jose, P A

    1995-01-01

    Dopamine and D1 agonists and NE all increase phosphatidyl inositol-specific phospholipase C (PLC) activity, but whereas dopamine produces a natriuresis, NE has an antinatriuretic effect. To determine if catecholamines differentially regulate the expression of PLC isoforms, we infused fenoldopam, a D1 agonist, or pramipexole, a D1/D2 agonist, intravenously or infused fenoldopam or NE into the renal artery of anesthetized rats. After 3-4 h of infusion, when the expected natriuresis (fenoldopam ...

  17. GABAB(1) receptor subunit isoforms differentially regulate stress resilience

    OpenAIRE

    O’Leary, Olivia F.; Felice, Daniela; Galimberti, Stefano; Savignac, Hélène M.; Bravo, Javier A.; Crowley, Tadhg; El Yacoubi, Malika; Vaugeois, Jean-Marie; Gassmann, Martin; Bettler, Bernhard; Dinan, Timothy G.; Cryan, John F.

    2014-01-01

    Stress can increase susceptibility to developing psychiatric disorders, including depression. Understanding the neurobiological mechanisms underlying stress resilience and susceptibility is key to identifying novel targets for the development of more effective treatments for stress-related psychiatric disorders. Here we show that specific isoforms of GABAB receptor subunits differentially regulate stress resilience. Specifically, GABAB(1a)−/− mice are more susceptible whereas GABAB(1b)−/− mic...

  18. Characterisation of CDKL5 Transcript Isoforms in Human and Mouse

    Science.gov (United States)

    Dando, Owen; Landsberger, Nicoletta; Kilstrup-Nielsen, Charlotte; Kind, Peter C.; Bailey, Mark E. S.; Cobb, Stuart R.

    2016-01-01

    Mutations in the X-linked Cyclin-Dependent Kinase-Like 5 gene (CDKL5) cause early onset infantile spasms and subsequent severe developmental delay in affected children. Deleterious mutations have been reported to occur throughout the CDKL5 coding region. Several studies point to a complex CDKL5 gene structure in terms of exon usage and transcript expression. Improvements in molecular diagnosis and more extensive research into the neurobiology of CDKL5 and pathophysiology of CDKL5 disorders necessitate an updated analysis of the gene. In this study, we have analysed human and mouse CDKL5 transcript patterns both bioinformatically and experimentally. We have characterised the predominant brain isoform of CDKL5, a 9.7 kb transcript comprised of 18 exons with a large 6.6 kb 3’-untranslated region (UTR), which we name hCDKL5_1. In addition we describe new exonic regions and a range of novel splice and UTR isoforms. This has enabled the description of an updated gene model in both species and a standardised nomenclature system for CDKL5 transcripts. Profiling revealed tissue- and brain development stage-specific differences in expression between transcript isoforms. These findings provide an essential backdrop for the diagnosis of CDKL5-related disorders, for investigations into the basic biology of this gene and its protein products, and for the rational design of gene-based and molecular therapies for these disorders. PMID:27315173

  19. Comparative genomic analysis reveals a novel mitochondrial isoform of human rTS protein and unusual phylogenetic distribution of the rTS gene

    Directory of Open Access Journals (Sweden)

    McGuire John J

    2005-09-01

    Full Text Available Abstract Background The rTS gene (ENOSF1, first identified in Homo sapiens as a gene complementary to the thymidylate synthase (TYMS mRNA, is known to encode two protein isoforms, rTSα and rTSβ. The rTSβ isoform appears to be an enzyme responsible for the synthesis of signaling molecules involved in the down-regulation of thymidylate synthase, but the exact cellular functions of rTS genes are largely unknown. Results Through comparative genomic sequence analysis, we predicted the existence of a novel protein isoform, rTS, which has a 27 residue longer N-terminus by virtue of utilizing an alternative start codon located upstream of the start codon in rTSβ. We observed that a similar extended N-terminus could be predicted in all rTS genes for which genomic sequences are available and the extended regions are conserved from bacteria to human. Therefore, we reasoned that the protein with the extended N-terminus might represent an ancestral form of the rTS protein. Sequence analysis strongly predicts a mitochondrial signal sequence in the extended N-terminal of human rTSγ, which is absent in rTSβ. We confirmed the existence of rTS in human mitochondria experimentally by demonstrating the presence of both rTSγ and rTSβ proteins in mitochondria isolated by subcellular fractionation. In addition, our comprehensive analysis of rTS orthologous sequences reveals an unusual phylogenetic distribution of this gene, which suggests the occurrence of one or more horizontal gene transfer events. Conclusion The presence of two rTS isoforms in mitochondria suggests that the rTS signaling pathway may be active within mitochondria. Our report also presents an example of identifying novel protein isoforms and for improving gene annotation through comparative genomic analysis.

  20. Purification of a branched-chain keto acid dehydrogenase from Pseudomonas putida.

    OpenAIRE

    Sokatch, J R; McCully, V; Roberts, C M

    1981-01-01

    We purified branched-chain keto acid dehydrogenase to a specific activity of 10 mumol/min per mg of protein from Pseudomonas putida grown on valine. The purified enzyme was active with 2-ketoisovalerate, 2-ketoisocaproate, and 2-keto-3-methylvalerate in a ratio of 1.0:0.8:0.7 but showed no activity with either pyruvate or 2-ketoglutarate. There were four polypeptides in the purified enzyme (molecular weights, 49,000, 46,000, 39,000, and 37,000). The purified enzyme was deficient in the specif...

  1. Structures of multi-branched dextrins produced by saccharifyiing alpha-amylase from starch.

    Science.gov (United States)

    Umeki, K; Yamamoto, T

    1975-11-01

    From the digest of beta-limit dextrin (prepared from glutinous rice starch) with saccharifying alpha-amylase of Bacillus subtilis [EC 3.2.1.1] (BSA), two extensibely branched dextrins consisting of nine (No. 6, Fig. 1) and ten (No 7, Fig.1) glucose units were isolated by paper chromatography. Structural analysis using various enzymes revealed that No. 6 and No. 7 were both mixtures of four triply branched dextrins. They had structures which were built up with 63-alpha-glucosylmaltotriose and/or 62-alpha-glucosylmaltose as a linking unit. However, the branching configuration and the minimum alpha-1, 4-glucosidic linkages existing between two branches followed one of the three structures shown below: (see article). PMID:814118

  2. Molecular mechanical differences between isoforms of contractile actin in the presence of isoforms of smooth muscle tropomyosin.

    OpenAIRE

    Lennart Hilbert; Genevieve Bates; Roman, Horia N.; Jenna L Blumenthal; Zitouni, Nedjma B.; Apolinary Sobieszek; Mackey, Michael C.; Anne-Marie Lauzon

    2013-01-01

    The proteins involved in smooth muscle's molecular contractile mechanism - the anti-parallel motion of actin and myosin filaments driven by myosin heads interacting with actin - are found as different isoforms. While their expression levels are altered in disease states, their relevance to the mechanical interaction of myosin with actin is not sufficiently understood. Here, we analyzed in vitro actin filament propulsion by smooth muscle myosin for [Formula: see text]-actin ([Formula: see text...

  3. Branching diffusions in random environment

    CERN Document Server

    Böinghoff, Christian

    2011-01-01

    We consider the diffusion approximation of branching processes in random environment (BPREs). This diffusion approximation is similar to and mathematically more tractable than BPREs. We obtain the exact asymptotic behavior of the survival probability. As in the case of BPREs, there is a phase transition in the subcritical regime due to different survival opportunities. In addition, we characterize the process conditioned to never go extinct and establish a backbone construction. In the strongly subcritical regime, mean offspring numbers are increased but still subcritical in the process conditioned to never go extinct. Here survival is solely due to an immortal individual, whose offspring are the ancestors of additional families. In the weakly subcritical regime, the mean offspring number is supercritical in the process conditioned to never go extinct. Thus this process survives with positive probability even if there was no immortal individual.

  4. Recursive Branching Simulated Annealing Algorithm

    Science.gov (United States)

    Bolcar, Matthew; Smith, J. Scott; Aronstein, David

    2012-01-01

    This innovation is a variation of a simulated-annealing optimization algorithm that uses a recursive-branching structure to parallelize the search of a parameter space for the globally optimal solution to an objective. The algorithm has been demonstrated to be more effective at searching a parameter space than traditional simulated-annealing methods for a particular problem of interest, and it can readily be applied to a wide variety of optimization problems, including those with a parameter space having both discrete-value parameters (combinatorial) and continuous-variable parameters. It can take the place of a conventional simulated- annealing, Monte-Carlo, or random- walk algorithm. In a conventional simulated-annealing (SA) algorithm, a starting configuration is randomly selected within the parameter space. The algorithm randomly selects another configuration from the parameter space and evaluates the objective function for that configuration. If the objective function value is better than the previous value, the new configuration is adopted as the new point of interest in the parameter space. If the objective function value is worse than the previous value, the new configuration may be adopted, with a probability determined by a temperature parameter, used in analogy to annealing in metals. As the optimization continues, the region of the parameter space from which new configurations can be selected shrinks, and in conjunction with lowering the annealing temperature (and thus lowering the probability for adopting configurations in parameter space with worse objective functions), the algorithm can converge on the globally optimal configuration. The Recursive Branching Simulated Annealing (RBSA) algorithm shares some features with the SA algorithm, notably including the basic principles that a starting configuration is randomly selected from within the parameter space, the algorithm tests other configurations with the goal of finding the globally optimal

  5. Branching process models of cancer

    CERN Document Server

    Durrett, Richard

    2015-01-01

    This volume develops results on continuous time branching processes and applies them to study rate of tumor growth, extending classic work on the Luria-Delbruck distribution. As a consequence, the authors calculate the probability that mutations that confer resistance to treatment are present at detection and quantify the extent of tumor heterogeneity. As applications, the authors evaluate ovarian cancer screening strategies and give rigorous proofs for results of Heano and Michor concerning tumor metastasis. These notes should be accessible to students who are familiar with Poisson processes and continuous time. Richard Durrett is mathematics professor at Duke University, USA. He is the author of 8 books, over 200 journal articles, and has supervised more than 40 Ph.D. students. Most of his current research concerns the applications of probability to biology: ecology, genetics, and most recently cancer.

  6. Selective glucocorticoid receptor translational isoforms reveal glucocorticoid-induced apoptotic transcriptomes.

    Science.gov (United States)

    Wu, I; Shin, S C; Cao, Y; Bender, I K; Jafari, N; Feng, G; Lin, S; Cidlowski, J A; Schleimer, R P; Lu, N Z

    2013-01-01

    Induction of T-cell apoptosis contributes to the anti-inflammatory and antineoplastic benefits of glucocorticoids. The glucocorticoid receptor (GR) translational isoforms have distinct proapoptotic activities in osteosarcoma cells. Here we determined whether GR isoforms selectively induce apoptosis in Jurkat T lymphoblastic leukemia cells. Jurkat cells stably expressing individual GR isoforms were generated and treated with vehicle or dexamethasone (DEX). DEX induced apoptosis in cells expressing the GR-A, -B, or -C, but not the GR-D, isoform. cDNA microarray analyses of cells sensitive (GR-C3) and insensitive (GR-D3) to DEX revealed glucocorticoid-induced proapoptotic transcriptomes. Genes that were regulated by the proapoptotic GR-C3, but not by the GR-D3, isoform likely contributed to glucocorticoid-induced apoptosis. The identified genes include those that are directly involved in apoptosis and those that facilitate cell killing. Chromatin immunoprecipitation assays demonstrated that distinct chromatin modification abilities may underlie the distinct functions of GR isoforms. Interestingly, all GR isoforms, including the GR-D3 isoform, suppressed mitogen-stimulated cytokines. Furthermore, the GR-C isoforms were selectively upregulated in mitogen-activated primary T cells and DEX treatment induced GR-C target genes in activated T cells. Cell-specific expressions and functions of GR isoforms may help to explain the tissue- and individual-selective actions of glucocorticoids and may provide a basis for developing improved glucocorticoids. PMID:23303127

  7. Vegetation survey of PEN Branch wetlands

    Energy Technology Data Exchange (ETDEWEB)

    1991-01-01

    A survey was conducted of vegetation along Pen Branch Creek at Savannah River Site (SRS) in support of K-Reactor restart. Plants were identified to species by overstory, understory, shrub, and groundcover strata. Abundance was also characterized and richness and diversity calculated. Based on woody species basal area, the Pen Branch delta was the most impacted, followed by the sections between the reactor and the delta. Species richness for shrub and groundcover strata were also lowest in the delta. No endangered plant species were found. Three upland pine areas were also sampled. In support of K Reactor restart, this report summarizes a study of the wetland vegetation along Pen Branch. Reactor effluent enters Indian Grove Branch and then flows into Pen Branch and the Pen Branch Delta.

  8. Effect of branch frequency in Aspergillus oryzae on protein secretion and culture viscosity.

    Science.gov (United States)

    Bocking, S P; Wiebe, M G; Robson, G D; Hansen, K; Christiansen, L H; Trinci, A P

    1999-12-20

    Highly branched mutants of two strains of Aspergillus oryzae (IFO4177, which produces alpha-amylase, and a transformant of IFO4177 [AMG#13], which produces heterologous glucoamylase in addition to alpha-amylase) were generated by UV or nitrous acid mutagenesis. Four mutants of the parental strain (IFO4177), which were 10 to 50% more branched than the parental strain, were studied in stirred batch culture and no differences were observed in either the amount or the rate of enzyme production. Five mutants of the transformed parental strain (AMG#13), which were 20 to 58% more branched than the parental strain, were studied in either batch, fed-batch or continuous culture. In batch culture, three of the mutants produced more glucoamylase than the transformed parental strain, although only two mutants produced more glucoamylase and alpha-amylase combined. No increase in enzyme production was observed in either chemostat or fed-batch culture. Cultures of highly branched mutants were less viscous than those of the parental and transformed parental strains. A linear relationship was found between the degree of branching (measured as hyphal growth unit length) and culture viscosity (measured as the torque exerted on the rheometer impeller) for these strains. DOT-controlled fed-batch cultures (in which the medium feed rate was determined by the DOT) were thus inoculated with either the transformed parent or highly branched mutants of the transformed parent to determine whether the reduced viscosity would improve aeration and give higher enzyme yields. The average rate of medium addition was higher for the two highly branched mutants (ca. 8.3 g medium h(-1)) than for the parental strain (5.7 g medium h(-1)). Specific enzyme production in the DOT controlled fed-batch cultures was similar for all three strains (approx. 0.24 g alpha-amylase and glucoamylase [g of biomass](-1)), but one of the highly branched mutants made more total enzyme (24.3 +/- 0.2 g alpha-amylase and

  9. 3D modelling of branching in plants

    OpenAIRE

    Evers, J.B.

    2011-01-01

    Shoot branching is a key determinant of overall aboveground plant form. During plant development, the number of branches formed strongly influences the amount of light absorbed by the plant, and thus the plant’s competitive strength in terms of light capture in relation to neighbouring plants. Branching is regulated by multiple internal factors which are modulated by different environmental signals. A key environmental signal in the context of a plant population is a low red / far-red intensi...

  10. Immunohistochemical analysis of aldehyde dehydrogenase isoforms and their association with estrogen-receptor status and disease progression in breast cancer

    Directory of Open Access Journals (Sweden)

    Opdenaker LM

    2014-12-01

    Full Text Available Lynn M Opdenaker,1,2 Kimberly M Arnold,1,3 Ryan T Pohlig,3,4 Jayasree S Padmanabhan,1 Daniel C Flynn,1,3 Jennifer Sims-Mourtada1–3 1Center for Translational Cancer Research, Helen F Graham Cancer Center, Christiana Care Health Services, Inc., Newark, Delaware, USA; 2Department of Biological Sciences, 3Department of Medical Laboratory Sciences, 4Biostatistics Core Facility, University of Delaware, Newark, Delaware, USA Abstract: In many types of tumors, especially breast tumors, aldehyde dehydrogenase (ALDH activity has been used to identify cancer stem-like cells within the tumor. The presence and quantity of these cells are believed to predict the response of tumors to chemotherapy. Therefore, identification and eradication of these cells would be necessary to cure the patient. However, there are 19 different ALDH isoforms that could contribute to the enzyme activity. ALDH1A1 and ALDH1A3 are among the isoforms mostly responsible for the increased ALDH activity observed in these stem-like cells, although the main isoforms vary in different tissues and tumor types. In the study reported here, we attempted to determine if ALDH1A1 or ALDH1A3, specifically, correlate with tumor stage, grade, and hormone-receptor status in breast-cancer patients. While there was no significant correlation between ALDH1A1 and any of the parameters tested, we were able to identify a positive correlation between ALDH1A3 and tumor stage in triple-negative cancers. In addition, ALDH1A3 was negatively correlated with estrogen-receptor status. Our data suggest that ALDH1A3 could be utilized as a marker to identify stem-like cells within triple-negative tumors. Keywords: breast tumor, ALDH, ALDH1A1, ALDH1A3, stem-like cells, triple-negative cancer

  11. Branched silver nanowires as controllable plasmon routers.

    Science.gov (United States)

    Fang, Yurui; Li, Zhipeng; Huang, Yingzhou; Zhang, Shunping; Nordlander, Peter; Halas, Naomi J; Xu, Hongxing

    2010-05-12

    Using polarization dependent scattering spectroscopy, we investigate plasmon propagation on branched silver nanowires. By controlling the polarization of the incident laser light, the wire plasmons can be routed into different wire branches and result in light emission from the corresponding wire ends. This routing behavior is found to be strongly dependent on the wavelength of light. Thus for certain incident polarizations, light of different wavelength will be routed into different branches. The branched nanowire can thus serve as a controllable router and multiplexer in integrated plasmonic circuits. PMID:20420411

  12. The ENZYME data bank.

    Science.gov (United States)

    Bairoch, A

    1994-01-01

    The ENZYME data bank is a repository of information relative to the nomenclature of enzymes. It is primarily based on the recommendations of the Nomenclature Committee of the International Union of Biochemistry and Molecular Biology (IUBMB) and it contains the following data for each type of characterized enzyme for which an EC (Enzyme Commission) number has been provided: EC number Recommended name Alternative names (if any) Catalytic activity Cofactors (if any) Pointers to the SWISS-PROT protein sequence entrie(s) that correspond to the enzyme (if any) Pointers to human disease(s) associated with a deficiency of the enzyme (if any). PMID:7937072

  13. Altered Alpha-Synuclein, Parkin, and Synphilin Isoform Levels in Multiple System Atrophy Brains

    DEFF Research Database (Denmark)

    Brudek, Tomasz; Winge, Kristian; Bredo Rasmussen, Nadja;

    2016-01-01

    -1 isoforms. In MSA brains, alpha-synuclein140 and alpha-synuclein112 isoform levels were significantly increased,whereas levels of the alpha-synuclein126 isoform were decreased in the substantia nigra, striatum, cerebellar cortex, and nucleus dentatus vs. CONTROLS: Moreover, in MSA cases, we showed...... increased levels of parkin isoforms lacking the N-terminal ubiquitin-like domain and an aggregation-prone synphiln-1A isoform that causes neuronal toxicity in MSA. In PD brains, Parkin transcript variant 3, 7 and 11 were significantly and specifically overexpressed in the striatum and cerebellar cortex......, together with synphilin-1A and 1C. The changes of isoform expression profiles in neurodegenerative diseases suggest alterations in the regulation of transcription and/or splicing events, leading to regional/cellular events that may be important for the highly increased aggregation of alpha-synuclein in the...

  14. PKC isoforms interact with and phosphorylate DNMT1

    Directory of Open Access Journals (Sweden)

    Pradhan Sriharsa

    2011-05-01

    Full Text Available Abstract Background DNA methyltransferase 1 (DNMT1 has been shown to be phosphorylated on multiple serine and threonine residues, based on cell type and physiological conditions. Although recent studies have suggested that protein kinase C (PKC may be involved, the individual contribution of PKC isoforms in their ability to phosphorylate DNMT1 remains unknown. The PKC family consists of at least 12 isoforms that possess distinct differences in structure, substrate requirement, expression and localization. Results Here we show that PKCα, βI, βII, δ, γ, η, ζ and μ preferentially phosphorylate the N-terminal domain of human DNMT1. No such phosphorylation of DNMT1 was observed with PKCε. Using PKCζ as a prototype model, we also found that PKC physically interacts with and phosphorylates DNMT1. In vitro phosphorylation assays conducted with recombinant fragments of DNMT1 showed that PKCζ preferentially phosphorylated the N-terminal region of DNMT1. The interaction of PKCζ with DNMT1 was confirmed by GST pull-down and co-immunoprecipitation experiments. Co-localization experiments by fluorescent microscopy further showed that endogenous PKCζ and DNMT1 were present in the same molecular complex. Endogenous PKCζ activity was also detected when DNMT1 was immunoprecipitated from HEK-293 cells. Overexpression of both PKCζ and DNMT1 in HEK-293 cells, but not of either alone, reduced the methylation status of genes distributed across the genome. Moreover, in vitro phosphorylation of DNMT1 by PKCζ reduced its methytransferase activity. Conclusions Our results indicate that phosphorylation of human DNMT1 by PKC is isoform-specific and provides the first evidence of cooperation between PKCζ and DNMT1 in the control of the DNA methylation patterns of the genome.

  15. Branch mode selection during early lung development.

    Directory of Open Access Journals (Sweden)

    Denis Menshykau

    Full Text Available Many organs of higher organisms, such as the vascular system, lung, kidney, pancreas, liver and glands, are heavily branched structures. The branching process during lung development has been studied in great detail and is remarkably stereotyped. The branched tree is generated by the sequential, non-random use of three geometrically simple modes of branching (domain branching, planar and orthogonal bifurcation. While many regulatory components and local interactions have been defined an integrated understanding of the regulatory network that controls the branching process is lacking. We have developed a deterministic, spatio-temporal differential-equation based model of the core signaling network that governs lung branching morphogenesis. The model focuses on the two key signaling factors that have been identified in experiments, fibroblast growth factor (FGF10 and sonic hedgehog (SHH as well as the SHH receptor patched (Ptc. We show that the reported biochemical interactions give rise to a Schnakenberg-type Turing patterning mechanisms that allows us to reproduce experimental observations in wildtype and mutant mice. The kinetic parameters as well as the domain shape are based on experimental data where available. The developed model is robust to small absolute and large relative changes in the parameter values. At the same time there is a strong regulatory potential in that the switching between branching modes can be achieved by targeted changes in the parameter values. We note that the sequence of different branching events may also be the result of different growth speeds: fast growth triggers lateral branching while slow growth favours bifurcations in our model. We conclude that the FGF10-SHH-Ptc1 module is sufficient to generate pattern that correspond to the observed branching modes.

  16. Branch mode selection during early lung development.

    Science.gov (United States)

    Menshykau, Denis; Kraemer, Conradin; Iber, Dagmar

    2012-01-01

    Many organs of higher organisms, such as the vascular system, lung, kidney, pancreas, liver and glands, are heavily branched structures. The branching process during lung development has been studied in great detail and is remarkably stereotyped. The branched tree is generated by the sequential, non-random use of three geometrically simple modes of branching (domain branching, planar and orthogonal bifurcation). While many regulatory components and local interactions have been defined an integrated understanding of the regulatory network that controls the branching process is lacking. We have developed a deterministic, spatio-temporal differential-equation based model of the core signaling network that governs lung branching morphogenesis. The model focuses on the two key signaling factors that have been identified in experiments, fibroblast growth factor (FGF10) and sonic hedgehog (SHH) as well as the SHH receptor patched (Ptc). We show that the reported biochemical interactions give rise to a Schnakenberg-type Turing patterning mechanisms that allows us to reproduce experimental observations in wildtype and mutant mice. The kinetic parameters as well as the domain shape are based on experimental data where available. The developed model is robust to small absolute and large relative changes in the parameter values. At the same time there is a strong regulatory potential in that the switching between branching modes can be achieved by targeted changes in the parameter values. We note that the sequence of different branching events may also be the result of different growth speeds: fast growth triggers lateral branching while slow growth favours bifurcations in our model. We conclude that the FGF10-SHH-Ptc1 module is sufficient to generate pattern that correspond to the observed branching modes. PMID:22359491

  17. The branch banking boom in Illinois: a byproduct of restrictive branching laws

    OpenAIRE

    Erin Davis; Tara Rice

    2007-01-01

    What’s behind the boom in bank branches across Illinois, particularly in Chicago? The authors explore the history of branch banking within the state and across the nation to help explain this recent trend and discuss its future implications.

  18. Functional differences between L- and T-plastin isoforms

    OpenAIRE

    1994-01-01

    Fimbrins/plastins are a family of highly conserved actin-bundling proteins. They are present in all eukaryotic cells including yeast, but each isoform displays a remarkable tissue specificity. T-plastin is normally found in epithelial and mesenchymal cells while L-plastin is present in hematopoietic cells. However, L-plastin has been also found in tumor cells of non-hematopoietic origin (Lin, C.-S., R. H. Aebersold, S. B. Kent, M. Varma, and J. Leavitt. 1988. Mol. Cell. Biol. 8:4659-4668; Lin...

  19. The FU gene and its possible protein isoforms

    Directory of Open Access Journals (Sweden)

    Nöthen Markus M

    2004-07-01

    Full Text Available Abstract Background FU is the human homologue of the Drosophila gene fused whose product fused is a positive regulator of the transcription factor Cubitus interruptus (Ci. Thus, FU may act as a regulator of the human counterparts of Ci, the GLI transcription factors. Since Ci and GLI are targets of Hedgehog signaling in development and morphogenesis, it is expected that FU plays an important role in Sonic, Desert and/or Indian Hedgehog induced cellular signaling. Results The FU gene was identified on chromosome 2q35 at 217.56 Mb and its exon-intron organization determined. The human developmental disorder Syndactyly type 1 (SD1 maps to this region on chromosome 2 and the FU coding region was sequenced using genomic DNA from an affected individual in a linked family. While no FU mutations were found, three single nucleotide polymorphisms were identified. The expression pattern of FU was thoroughly investigated and all examined tissues express FU. It is also clear that different tissues express transcripts of different sizes and some tissues express more than one transcript. By means of nested PCR of specific regions in RT/PCR generated cDNA, it was possible to verify two alternative splicing events. This also suggests the existence of at least two additional protein isoforms besides the FU protein that has previously been described. This long FU and a much shorter isoform were compared for the ability to regulate GLI1 and GLI2. None of the FU isoforms showed any effects on GLI1 induced transcription but the long form can enhance GLI2 activity. Apparently FU did not have any effect on SUFU induced inhibition of GLI. Conclusions The FU gene and its genomic structure was identified. FU is a candidate gene for SD1, but we have not identified a pathogenic mutation in the FU coding region in a family with SD1. The sequence information and expression analyses show that transcripts of different sizes are expressed and subjected to alternative splicing

  20. GSK-3β phosphorylation of functionally distinct tau isoforms has differential, but mild effects

    Directory of Open Access Journals (Sweden)

    Gamblin T Chris

    2009-05-01

    Full Text Available Abstract Background Tau protein exists as six different isoforms that differ by the inclusion or exclusion of exons 2, 3 and 10. Exon 10 encodes a microtubule binding repeat, thereby resulting in three isoforms with three microtubule binding repeats (3R and three isoforms that have four microtubule binding repeats (4R. In normal adult brain, the relative amounts of 3R tau and 4R tau are approximately equal. These relative protein levels are preserved in Alzheimer's disease, although in other neurodegenerative tauopathies such as progressive supranuclear palsy, corticobasal degeneration and Pick's disease, the ratio of 3R:4R is frequently altered. Because tau isoforms are not equally involved in these diseases, it is possible that they either have inherently unique characteristics owing to their primary structures or that post-translational modification, such as phosphorylation, differentially affects their properties. Results We have determined the effects of phosphorylation by a kinase widely believed to be involved in neurodegenerative processes, glycogen synthase kinase-3β (GSK-3β, on the microtubule binding and inducer-initiated polymerization of these isoforms in vitro. We have found that each isoform has a unique microtubule binding and polymerization profile that is altered by GSK-3β. GSK-3β phosphorylation had differential effects on the isoforms although there were similarities between isoforms and the effects were generally mild. Conclusion These results indicate that tau phosphorylation by a single kinase can have isoform specific outcomes. The mild nature of these changes, however, makes it unlikely that differential effects of GSK-3β phosphorylation on the isoforms are causative in neurodegenerative disease. Instead, the inherent differences in the isoform interactions themselves and local conditions in the diseased cells are likely the major determinant of isoform involvement in various neurodegenerative disorders.

  1. Selective glucocorticoid receptor translational isoforms reveal glucocorticoid-induced apoptotic transcriptomes

    OpenAIRE

    Wu, I; Shin, S. C.; Cao, Y; Bender, I K; N Jafari; Feng, G.; Lin, S.; Cidlowski, J. A.; Schleimer, R. P.; Lu, N Z

    2013-01-01

    Induction of T-cell apoptosis contributes to the anti-inflammatory and antineoplastic benefits of glucocorticoids. The glucocorticoid receptor (GR) translational isoforms have distinct proapoptotic activities in osteosarcoma cells. Here we determined whether GR isoforms selectively induce apoptosis in Jurkat T lymphoblastic leukemia cells. Jurkat cells stably expressing individual GR isoforms were generated and treated with vehicle or dexamethasone (DEX). DEX induced apoptosis in cells expres...

  2. MetaDiff: differential isoform expression analysis using random-effects meta-regression

    OpenAIRE

    Jia, Cheng; Guan, Weihua; Yang, Amy; Xiao, Rui; Tang, W. H. Wilson; Moravec, Christine S.; Margulies, Kenneth B.; Cappola, Thomas P.; Li, Mingyao; Li, Chun

    2015-01-01

    Background RNA sequencing (RNA-Seq) allows an unbiased survey of the entire transcriptome in a high-throughput manner. A major application of RNA-Seq is to detect differential isoform expression across experimental conditions, which is of great biological interest due to its direct relevance to protein function and disease pathogenesis. Detection of differential isoform expression is challenging because of uncertainty in isoform expression estimation owing to ambiguous reads and variability i...

  3. Induction of Chemokine Expression by Adiponectin In Vitro is Isoform-Dependent

    OpenAIRE

    Song, Huijuan; Chan, James; Rovin, Brad H.

    2009-01-01

    Adiponectin is reported to have both pro- and anti-inflammatory effects. Because adiponectin circulates in isoforms of various sizes, and some responses to adiponectin are isoform-dependent, it was postulated that the pro-inflammatory effects of adiponectin may isoform-specific. To test this, peripheral blood mononuclear cells (PBMC), microvascular endothelial cells (MVEC), and human glomerular mesangial cells (HMC) were treated with high or low molecular weight (HMW, LMW) recombinant human a...

  4. Hydrogen peroxide generation and antioxidant enzyme activities in the leaves and roots of wheat cultivars subjected to long-term soil drought stress.

    Science.gov (United States)

    Huseynova, Irada M; Aliyeva, Durna R; Mammadov, Alamdar Ch; Aliyev, Jalal A

    2015-08-01

    The dynamics of the activity of catalase, ascorbate peroxidase, guaiacol peroxidase, and benzidine peroxidase, as well as the level of hydrogen peroxide in the vegetative organs of durum wheat (Triticum durum Desf.) cultivars was studied under long-term soil drought conditions. It was established that hydrogen peroxide generation occurred at early stages of stress in the tolerant variety Barakatli-95, whereas in the susceptible variety Garagylchyg-2 its significant amounts were accumulated only at later stages. Garagylchyg-2 shows a larger reduction of photochemical activity of PS II in both genotypes at all stages of ontogenesis under drought stress than Barakatli-95. The highest activity of catalase which plays a leading role in the neutralization of hydrogen peroxide was observed in the leaves and roots of the drought-tolerant variety Barakatli-95. Despite the fact that the protection system also includes peroxidases, the activity of these enzymes even after synthesis of their new portions is substantially lower compared with catalase. Native PAGE electrophoresis revealed the presence of one isoform of CAT, seven isoforms of APX, three isoforms of GPO, and three isoforms of BPO in the leaves, and also three isoforms of CAT, four isoforms of APX, two isoforms of GPO, and six isoforms of BPO in the roots of wheat. One isoform of CAT was found in the roots when water supply was normal and three isoforms were observed under drought conditions. Stress associated with long-term soil drought in the roots of wheat has led to an increase in the heterogeneity due to the formation of two new sedentary forms of catalase: CAT2 and CAT3. PMID:26008794

  5. Enzyme Therapy: Current Perspectives.

    Science.gov (United States)

    UmaMaheswari, Thiyagamoorthy; Hemalatha, Thiagarajan; Sankaranarayanan, Palavesam; Puvanakrishnan, Rengarajulu

    2016-01-01

    Enzymes control all metabolic processes in human system from simple digestion of food to highly complex immune response. Physiological reactions occuring in healthy individuals are disturbed when enzymes are deficient or absent. Enzymes are administered for normalizing biological function in certain pathologies. Initially, crude proteolytic enzymes were used for the treatment of gastrointestinal disorders. Recent advances have enabled enzyme therapy as a promising tool in the treatment of cardiovascular, oncological and hereditary diseases. Now, a spectrum of other diseases are also covered under enzyme therapy. But, the available information on the use of enzymes as therapeutic agents for different diseases is scanty. This review details the enzymes which have been used to treat various diseases/disorders. PMID:26891548

  6. Branched-Chain Amino Acids

    Directory of Open Access Journals (Sweden)

    Matteo Ghiringhelli

    2015-07-01

    Full Text Available Our study is focused on evaluation and use of the most effective and correct nutrients. In particular, our attention is directed to the role of certain amino acids in cachectic patients.During parenteral nutrition in humans, physician already associates in the PN-bags different formulations including amino acids, lipids and glucose solutions or essential amino acids solution alone or exclusively branched-chain amino acids (BCAA. Studies investigated the effects of dietary BCAA ingestion on different diseases and conditions such as obesity and metabolic disorders, liver disease, muscle atrophy, cancer, impaired immunity or injuries (surgery, trauma, burns, and sepsis. BCAAs have been shown to affect gene expression, protein metabolism, apoptosis and regeneration of hepatocytes, and insulin resistance. They have also been shown to inhibit the proliferation of liver cancer cells in vitro, and are essential for lymphocyte proliferation and dendritic cell maturation. Oral or parenteral administration of these three amino acids will allow us to evaluate the real efficacy of these compounds during a therapy to treat malnutrition in subjects unable to feed themselves.

  7. AVM branch vibration test equipment

    International Nuclear Information System (INIS)

    An inventory of the test equipment of the AVM Branch ''Acoustic and Vibratory Mechanics Analysis Methods'' group has been undertaken. The purpose of this inventory is to enable better acquaintance with the technical characteristics of the equipment, providing an accurate definition of their functionalities, ad to inform potential users of the possibilities and equipment available in this field. The report first summarizes the various experimental surveys conduced. Then, using the AVM equipment database to draw up an exhaustive list of available equipment, it provides a full-scope picture of the vibration measurement systems (sensors, conditioners and exciters) and data processing resources commonly used on industrial sites and in laboratories. A definition is also given of a mobile test unit, called 'shelter', and a test bench used for the testing and performance rating of the experimental analysis methods developed by the group. The report concludes with a description of two fixed installations: - the calibration bench ensuring the requisite quality level for the vibration measurement systems ; - the training bench, whereby know-how acquired in the field in the field of measurement and experimental analysis processes is made available to others. (author). 27 refs., 15 figs., 2 appends

  8. IsoformEx: isoform level gene expression estimation using weighted non-negative least squares from mRNA-Seq data

    Directory of Open Access Journals (Sweden)

    Gupta Ravi

    2011-07-01

    Full Text Available Abstract Background mRNA-Seq technology has revolutionized the field of transcriptomics for identification and quantification of gene transcripts not only at gene level but also at isoform level. Estimating the expression levels of transcript isoforms from mRNA-Seq data is a challenging problem due to the presence of constitutive exons. Results We propose a novel algorithm (IsoformEx that employs weighted non-negative least squares estimation method to estimate the expression levels of transcript isoforms. Validations based on in silico simulation of mRNA-Seq and qRT-PCR experiments with real mRNA-Seq data showed that IsoformEx could accurately estimate transcript expression levels. In comparisons with published methods, the transcript expression levels estimated by IsoformEx showed higher correlation with known transcript expression levels from simulated mRNA-Seq data, and higher agreement with qRT-PCR measurements of specific transcripts for real mRNA-Seq data. Conclusions IsoformEx is a fast and accurate algorithm to estimate transcript expression levels and gene expression levels, which takes into account short exons and alternative exons with a weighting scheme. The software is available at http://bioinformatics.wistar.upenn.edu/isoformex.

  9. Branching out Has So Much to Offer

    Science.gov (United States)

    Murray, Joe

    2012-01-01

    In 1989 there were thirty ATM branches nationally. In January 2012 there were just twelve ATM branches with another three "proposed". How can that happen? How did it happen? Maybe the most pertinent question is: Why did it happen? There is no single answer to the last question, but perhaps it was something to do with the changes that were sweeping…

  10. 12 CFR 741.11 - Foreign branching.

    Science.gov (United States)

    2010-01-01

    ...) Contents of Business Plan. The written business plan must address the following: (1) Analysis of market... regarding the branch (shares, lending, capital, charge-offs, collections); (6) The field of membership or... for branch operations (balance sheet and income and expense projections) for the first and second...

  11. Spectral problem for branching chain quantum graph

    International Nuclear Information System (INIS)

    Branching chain of rings as a quantum graph is considered. We use the transfer matrix method to obtain the spectral equation. The existence of bound states is proved. The discrete spectrum of the Schrödinger operator for the system is described. We find the dependence of the eigenvalues positions on the branching angle.

  12. Proteomic Analysis of Cytokeratin Isoforms Uncovers Association with Survival in Lung Adenocarcinoma

    Directory of Open Access Journals (Sweden)

    Tarek G. Gharib

    2002-01-01

    Full Text Available Cytokeratins. (CK are intermediate filaments whose expression is often altered in epithelial cancer. Systematic identification of lung adenocarcinoma proteins using two-dimensional polyacrylamide gel electrophoresis and mass spectrometry has uncovered numerous CK isoforms. In this study, 93 lung adenocarcinomas. (64 stage I and 29 stage III and 10 uninvolved lung samples were quantitatively examined for protein expression. Fourteen of 21 isoforms of CK 7, 8, 18, 19 occurred at significantly higher levels. (P<.05 in tumors compared to uninvolved adjacent tissue. Specific isoforms of the four types of CK identified correlated with either clinical outcome or individual clinical-pathological parameters. All five of the CK7 isoforms associated with patient survival represented cleavage products. Two of five CK7 isoforms. (nos. 2165 and 2091, one of eight CK8 isoforms. (no. 439, one of three CK19 isoforms. (no. 1955 were associated with survival and significantly correlated to their mRNA levels, suggesting that transcription underlies overexpression of these CK isoforms. Our data indicate substantial heterogeneity among CK in lung adenocarcinomas resulting from posttranslational modifications, some of which correlated with patient survival and other clinical parameters. Therefore, specific isoforms of individual CK may have utility as diagnostic or predictive markers in lung adenocarcinomas.

  13. Enzyme inhibition by iminosugars

    DEFF Research Database (Denmark)

    López, Óscar; Qing, Feng-Ling; Pedersen, Christian Marcus;

    2013-01-01

    Imino- and azasugar glycosidase inhibitors display pH dependant inhibition reflecting that both the inhibitor and the enzyme active site have groups that change protonation state with pH. With the enzyme having two acidic groups and the inhibitor one basic group, enzyme-inhibitor complexes with...

  14. Liquid chromatography/mass spectrometry analysis of branched fructans produced in vitro with C-13-labeled substrates

    DEFF Research Database (Denmark)

    Verspreet, Joran; Hansen, Anders Holmgaard; Dornez, Emmie;

    2014-01-01

    -labeled substrates. Liquid chromatography/mass spectrometry (LC/MS) was used for the structural identification of branched fructans. The MS2 fragmentation of these compounds is described for the first time. Analytes were charged by electrospray ionization in negative mode and a quadrupole mass analyzer was used for...... branch. CONCLUSIONSThe MS2 fragmentation patterns described in the current paper now allow rapid screening of large sample sets for the presence of branched, graminan-type fructans. Furthermore, the data enables the characterization of fructan-metabolizing enzymes by identification of the fructan...

  15. Assignment and expression patterns of porcine muscle-specific isoform of phosphoglycerate mutase gene

    Institute of Scientific and Technical Information of China (English)

    Haifang Qiu; Shuhong Zhao; Xuewen Xu; Martine Yerle; Bang Liu

    2008-01-01

    It has been reported that the muscle-specific isoform (type M, PGAM2) of phosphoglycerate mutase (PGAM) is a housekeeping en-zyme; it catalyzes the conversion of 3-phospboglycerate into 2-phosphoglycerate in the glyeolysis process to release energy. It is encoded by the Pgam2 gene. In this study, the eDNA of the porcine Pgam2 was cloned. This gene contains an open reading frame of 765 bp en-coding a protein of 253 residues, and the predicted protein sequences share high similarity with other mammalians, 96% identity with humans, and 94% identity with mouse and rats. Pgam2 was mapped to SSC18q13-q21 by the RH panel. In this region, there are several QTLs, such as fat ratio, lean percentage, and diameter of muslce fiber, which affect meat production and quality. The reverse transcrip-tase-polymerase chain reaction revealed that the porcine Pgam2 gene was mainly expressed in the muscle tissue (skeletal muscle and cardiac muscle), and was expressed highly at skeletal muscle development stages (embryonic periods: 33, 65, and 90 days post-conception(dpe); postnatal pigs: 4 days and adult). This indicates that the Pgam2 gene plays an important role in muscle growth and development. In addition, it was demonstrated that PGAM2 locates both in cytoplasm and nuclei, and takes part in the glycometabolism process of cyto-plasm and nuclei.

  16. Glyoxylate Reductase Isoform 1 is Localized in the Cytosol and Not Peroxisomes in Plant Cells

    Institute of Scientific and Technical Information of China (English)

    Steven L. K. Ching; Satinder K. Gidda; Amanda Rochon; Owen R. van Cauwenberghe; Barry J. Shelp; Robert T. Mullen

    2012-01-01

    Glyoxylate reductase (GLYR) is a key enzyme in plant metabolism which catalyzes the detoxification of both photorespiratory glyoxylate and succinic semialdehdye,an intermediate of the γ-aminobutyrate (GABA) pathway.Two isoforms of GLYR exist in plants,GLYR1 and GLYR2,and while GLYR2 is known to be localized in plastids,GLYR1 has been reported to be localized in either peroxisomes or the cytosol.Here,we reappraised the intracellular localization of GLYR1 in Arabidopsis thaliana L.Heynh (ecotype Lansberg erecta) using both transiently-transformed suspension cells and stably-transformed plants,in combination with fluorescence microscopy.The results indicate that GLYR1 is localized exclusively to the cytosol regardless of the species,tissue and/or cell type,or exposure of plants to environmental stresses that would increase flux through the GABA pathway.Moreover,the C-terminal tripeptide sequence of GLYR1,-SRE,despite its resemblance to a type 1 peroxisomal targeting signal,is not sufficient for targeting to peroxisomes.Collectively,these results define the cytosol as the intracellular location of GLYR1 and provide not only important insight to the metabolic roles of GLYR1 and the compartmentation of the GABA and photorespiratory pathways in plant cells,but also serve as a useful reference for future studies of proteins proposed to be localized to peroxisomes and/or the cytosol.

  17. Data mining of enzymes using specific peptides

    Directory of Open Access Journals (Sweden)

    Lavi Yair

    2009-12-01

    Full Text Available Abstract Background Predicting the function of a protein from its sequence is a long-standing challenge of bioinformatic research, typically addressed using either sequence-similarity or sequence-motifs. We employ the novel motif method that consists of Specific Peptides (SPs that are unique to specific branches of the Enzyme Commission (EC functional classification. We devise the Data Mining of Enzymes (DME methodology that allows for searching SPs on arbitrary proteins, determining from its sequence whether a protein is an enzyme and what the enzyme's EC classification is. Results We extract novel SP sets from Swiss-Prot enzyme data. Using a training set of July 2006, and test sets of July 2008, we find that the predictive power of SPs, both for true-positives (enzymes and true-negatives (non-enzymes, depends on the coverage length of all SP matches (the number of amino-acids matched on the protein sequence. DME is quite different from BLAST. Comparing the two on an enzyme test set of July 2008, we find that DME has lower recall. On the other hand, DME can provide predictions for proteins regarded by BLAST as having low homologies with known enzymes, thus supplying complementary information. We test our method on a set of proteins belonging to 10 bacteria, dated July 2008, establishing the usefulness of the coverage-length cutoff to determine true-negatives. Moreover, sifting through our predictions we find that some of them have been substantiated by Swiss-Prot annotations by July 2009. Finally we extract, for production purposes, a novel SP set trained on all Swiss-Prot enzymes as of July 2009. This new set increases considerably the recall of DME. The new SP set is being applied to three metagenomes: Sargasso Sea with over 1,000,000 proteins, producing predictions of over 220,000 enzymes, and two human gut metagenomes. The outcome of these analyses can be characterized by the enzymatic profile of the metagenomes, describing the relative

  18. Acetylation Targets the M2 Isoform of Pyruvate Kinase for Degradation through Chaperone-Mediated Autophagy and Promotes Tumor Growth

    Science.gov (United States)

    Lv, Lei; Li, Dong; Zhao, Di; Lin, Ruiting; Chu, Yajing; Zhang, Heng; Zha, Zhengyu; Liu, Ying; Li, Zi; Xu, Yanping; Wang, Gang; Huang, Yiran; Xiong, Yue; Guan, Kun-Liang; Lei, Qun-Ying

    2016-01-01

    SUMMARY Most tumor cells take up more glucose than normal cells but metabolize glucose via glycolysis even in the presence of normal levels of oxygen, a phenomenon known as the Warburg effect. Tumor cells commonly express the embryonic M2 isoform of pyruvate kinase (PKM2) that may contribute to the metabolism shift from oxidative phosphorylation to aerobic glycolysis and tumorigenesis. Here we show that PKM2 is acetylated on lysine 305 and that this acetylation is stimulated by high glucose concentration. PKM2 K305 acetylation decreases PKM2 enzyme activity and promotes its lysosomal-dependent degradation via chaperone-mediated autophagy (CMA). Acetylation increases PKM2 interaction with HSC70, a chaperone for CMA, and association with lysosomes. Ectopic expression of an acetylation mimetic K305Q mutant accumulates glycolytic intermediates and promotes cell proliferation and tumor growth. These results reveal an acetylation regulation of pyruvate kinase and the link between lysine acetylation and CMA. PMID:21700219

  19. A Review of Metallothionein Isoforms and their Role in Pathophysiology

    Directory of Open Access Journals (Sweden)

    Senthil kumar M

    2011-05-01

    Full Text Available Abstract The Metallothionein (MT is a protein which has several interesting biological effects and has been demonstrated increase focus on the role of MT in various biological systems in the past three decades. The studies on the role of MT were limited with few areas like apoptosis and antioxidants in selected organs even fifty years after its discovery. Now acknowledge the exploration of various isoforms of MT such as MT-I, MT-II, MT-III and MT-IV and other isoforms in various biological systems. Strong evidence exists that MT modulates complex diseases and the immune system in the body but the primary function of MT still remains unknown. This review's main objective is to explore the capability to specifically manipulate MT levels in cells and in animals to provide answers regarding how MT could impact those complex disease scenarios. The experimental result mentioned in this review related among MT, zinc, cadmium, diabetic, heart disease, bone retardation, neuro toxicity, kidney dysfunction, cancer, and brain suggest novel method for exploration and contribute significantly to the growing scientist to research further in this field.

  20. Entropy-based model for miRNA isoform analysis.

    Directory of Open Access Journals (Sweden)

    Shengqin Wang

    Full Text Available MiRNAs have been widely studied due to their important post-transcriptional regulatory roles in gene expression. Many reports have demonstrated the evidence of miRNA isoform products (isomiRs in high-throughput small RNA sequencing data. However, the biological function involved in these molecules is still not well investigated. Here, we developed a Shannon entropy-based model to estimate isomiR expression profiles of high-throughput small RNA sequencing data extracted from miRBase webserver. By using the Kolmogorov-Smirnov statistical test (KS test, we demonstrated that the 5p and 3p miRNAs present more variants than the single arm miRNAs. We also found that the isomiR variant, except the 3' isomiR variant, is strongly correlated with Minimum Free Energy (MFE of pre-miRNA, suggesting the intrinsic feature of pre-miRNA should be one of the important factors for the miRNA regulation. The functional enrichment analysis showed that the miRNAs with high variation, particularly the 5' end variation, are enriched in a set of critical functions, supporting these molecules should not be randomly produced. Our results provide a probabilistic framework for miRNA isoforms analysis, and give functional insights into pre-miRNA processing.

  1. A New View of Ras Isoforms in Cancers.

    Science.gov (United States)

    Nussinov, Ruth; Tsai, Chung-Jung; Chakrabarti, Mayukh; Jang, Hyunbum

    2016-01-01

    Does small GTPase K-Ras4A have a single state or two states, one resembling K-Ras4B and the other N-Ras? A recent study of K-Ras4A made the remarkable observation that even in the absence of the palmitoyl, K-Ras4A can be active at the plasma membrane. Importantly, this suggests that K-Ras4A may exist in two distinct signaling states. In state 1, K-Ras4A is only farnesylated, like K-Ras4B; in state 2, farnesylated and palmitoylated, like N-Ras. The K-Ras4A hypervariable region sequence is positively charged, in between K-Ras4B and N-Ras. Taken together, this raises the possibility that the farnesylated but nonpalmitoylated state 1, like K-Ras4B, binds calmodulin and is associated with colorectal and other adenocarcinomas like lung cancer and pancreatic ductal adenocarcinoma. On the other hand, state 2 may be associated with melanoma and other cancers where N-Ras is a major contributor, such as acute myeloid leukemia. Importantly, H-Ras has two, singly and doubly, palmitoylated states that may also serve distinct functional roles. The multiple signaling states of palmitoylated Ras isoforms question the completeness of small GTPase Ras isoform statistics in different cancer types and call for reevaluation of concepts and protocols. They may also call for reconsideration of oncogenic Ras therapeutics. PMID:26659836

  2. Role of cysteines in mammalian VDAC isoforms' function.

    Science.gov (United States)

    De Pinto, Vito; Reina, Simona; Gupta, Ankit; Messina, Angela; Mahalakshmi, Radhakrishnan

    2016-08-01

    In this mini-review, we analyze the influence of cysteines in the structure and activity of mitochondrial outer membrane mammalian VDAC isoforms. The three VDAC isoforms show conserved sequences, similar structures and the same gene organization. The meaning of three proteins encoded in different chromosomes must thus be searched for subtle differences at the amino acid level. Among others, cysteine content is noticeable. In humans, VDAC1 has 2, VDAC2 has 9 and VDAC3 has 6 cysteines. Recent works have shown that, at variance from VDAC1, VDAC2 and VDAC3 exhibit cysteines predicted to protrude towards the intermembrane space, making them a preferred target for oxidation by ROS. Mass spectrometry in VDAC3 revealed that a disulfide bridge can be formed and other cysteine oxidations are also detectable. Both VDAC2 and VDAC3 cysteines were mutagenized to highlight their role in vitro and in complementation assays in Δporin1 yeast. Chemico-physical techniques revealed an important function of cysteines in the structural stabilization of the pore. In conclusion, the works available on VDAC cysteines support the notion that the three proteins are paralogs with a similar pore-function and slightly different, but important, ancillary biological functions. 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:26947058

  3. Phylogenomic relationships between amylolytic enzymes from 85 strains of fungi.

    Science.gov (United States)

    Chen, Wanping; Xie, Ting; Shao, Yanchun; Chen, Fusheng

    2012-01-01

    Fungal amylolytic enzymes, including α-amylase, gluocoamylase and α-glucosidase, have been extensively exploited in diverse industrial applications such as high fructose syrup production, paper making, food processing and ethanol production. In this paper, amylolytic genes of 85 strains of fungi from the phyla Ascomycota, Basidiomycota, Chytridiomycota and Zygomycota were annotated on the genomic scale according to the classification of glycoside hydrolase (GH) from the Carbohydrate-Active enZymes (CAZy) Database. Comparisons of gene abundance in the fungi suggested that the repertoire of amylolytic genes adapted to their respective lifestyles. Amylolytic enzymes in family GH13 were divided into four distinct clades identified as heterologous α-amylases, eukaryotic α-amylases, bacterial and fungal α-amylases and GH13 α-glucosidases. Family GH15 had two branches, one for gluocoamylases, and the other with currently unknown function. GH31 α-glucosidases showed diverse branches consisting of neutral α-glucosidases, lysosomal acid α-glucosidases and a new clade phylogenetically related to the bacterial counterparts. Distribution of starch-binding domains in above fungal amylolytic enzymes was related to the enzyme source and phylogeny. Finally, likely scenarios for the evolution of amylolytic enzymes in fungi based on phylogenetic analyses were proposed. Our results provide new insights into evolutionary relationships among subgroups of fungal amylolytic enzymes and fungal evolutionary adaptation to ecological conditions. PMID:23166747

  4. Plasma biomarker for detection of early stage pancreatic cancer and risk factors for pancreatic malignancy using antibodies for apolipoprotein-AII isoforms

    OpenAIRE

    Kazufumi Honda; Michimoto Kobayashi; Takuji Okusaka; Jo Ann Rinaudo; Ying Huang; Tracey Marsh; Mitsuaki Sanada; Yoshiyuki Sasajima; Shoji Nakamori; Masashi Shimahara; Takaaki Ueno; Akihiko Tsuchida; Naohiro Sata; Tatsuya Ioka; Yohichi Yasunami

    2015-01-01

    We recently reported that circulating apolipoprotein AII (apoAII) isoforms apoAII-ATQ/AT (C-terminal truncations of the apoAII homo-dimer) decline significantly in pancreatic cancer and thus might serve as plasma biomarkers for the early detection of this disease. We report here the development of novel enzyme-linked immunosorbent assays (ELISAs) for measurement of apoAII-ATQ/AT and their clinical applicability for early detection of pancreatic cancer. Plasma and serum concentrations of apoAI...

  5. Isoforms of thyroxine-binding globulin as a model for molecular epidemiology of human cancer risk

    International Nuclear Information System (INIS)

    The novel field of molecular epidemiology of human cancer risk has added a new branch to classical epidemiology by providing a direct link between human cancer and carcinogen exposure. It was estimated that about 80% of cancers are due to environmental factors. The blood proteins are almost certainly targets for modification in human cancer, and their identification and characterization will be of primary importance in the development of the new and rapidly evolving field of molecular epidemiology. Among blood proteins that are altered in human cancer, TBG occupies a special place because the level of human blood TBG is the most sensitive to intensification of biosynthesis and proliferation processes in organisms in different types of cancer. The increase of TBG concentration in cancer can be result from both activation of TBG biosynthesis in liver or altering of post translation glycosylation that prolongs protein survival time. The molecular basis for the change in the properties of TBG in cancer is unknown. These distinctive changes could have important consequences for the function of TBG in cancer and may help to develop more precise markers for monitoring pathological progression in this disease. Considerable variability and subtlety can occur in the carbohydrate composition and structure of serum glycoproteins in disease. This can be either as a major change, such as an increase in the number of oligosaccharide branches at a particular glycosylation site or as a minor change such as the addition of an extra fucose or sialic acid residue. Increased fucosylation has also been reported for transferrin and alpha-fetoprotein in liver cancer; thyroglobulin in thyroid cancer, IgG in myeloma, haptoglobin in ovarian cancer. The last own studies have shown that in clinically healthy teenagers born in Khojniki (137 Cs 185-555 kBq/m), we have found an unusual thyroid profile exhibiting increased levels of total triiodothyronine (T3), total thyroxine (T4), and thyroxine

  6. [Croatian Medical Association--Branch Zagreb].

    Science.gov (United States)

    Kaić, Zvonimir; Sain, Snjezana; Gulić, Mirjana; Mahovlić, Vjekoslav; Krznarić, Zeljko

    2014-01-01

    The available literature shows us that "Druztvo ljeciteljah u Zagrebus (the Society of Healers in Zagreb) was founded as far back as the year 1845 by a total of thirteen members. This data allows us to follow the role of doctors and health workers in Zagreb through their everyday profession, research, organizational and social work as well as management through a period of over one hundred to seventy years. The Branch Zagreb was active before the official establishment of subsidiaries of CMA which is evident from the minutes of the regular annual assembly of the Croatian Medical Association on 21 March 1948. Until the end of 1956, there was no clear division of labor, functions and competencies between the Branch and the Main Board. Their actions were instead consolidated and the Branch operated within and under the name of Croatian Medical Association. In that year the Branch became independent. The Branch Zagreb is the largest and one of the most active branches of the Croatian Medical Association. At the moment, the Branch brings together 3621 members, regular members--doctors of medicine (2497), doctors of dental medicine (384), retired physicians (710), and associate members (30 specialists with higher education who are not doctors). The Branch is especially accomplished in its activities in the area of professional development of its members and therefore organizes a series of scientific conferences in the framework of continuous education of physicians, allowing its members to acquire necessary points for the extension of their operating license. The choir "Zagrebacki lijecnici pjevaci" (Zagreb Physicians' Choir) of the Croatian Medical Music Society of the CMA and its activities are inseparable from the Branch Zagreb. The Branch is firmly linked to the parent body, the CMA, and thus has a visible impact on the strategy and the activities of the Association as a whole. Most professional societies of the CMA have their headquarters in Zagreb and this is

  7. Altered levels of LIL3 isoforms in Arabidopsis lead to disturbed pigment-protein assembly and chlorophyll synthesis, chlorotic phenotype and impaired photosynthetic performance.

    Science.gov (United States)

    Lohscheider, Jens N; Rojas-Stütz, Marc C; Rothbart, Maxi; Andersson, Ulrica; Funck, Dietmar; Mendgen, Kurt; Grimm, Bernhard; Adamska, Iwona

    2015-10-01

    Light-harvesting complex (LHC)-like (LIL) proteins contain two transmembrane helices of which the first bears a chlorophyll (Chl)-binding motif. They are widespread in photosynthetic organisms, but almost nothing is known about their expression and physiological functions. We show that two LIL3 paralogues (LIL3:1 and LIL3:2) in Arabidopsis thaliana are expressed in photosynthetically active tissues and their expression is differentially influenced by light stress. Localization studies demonstrate that both isoforms are associated with subcomplexes of LHC antenna of photosystem II. Transgenic plants with reduced amounts of LIL3:1 exhibited a slightly impaired growth and have reduced Chl and carotenoid contents as compared to wild-type plants. Ectopic overexpression of either paralogue led to a developmentally regulated switch to co-suppression of both LIL3 isoforms, resulting in a circular chlorosis of the leaf rosettes. Chlorotic sectors show severely diminished levels of LIL3 isoforms and other proteins, and thylakoid morphology was changed. Additionally, the levels of enzymes involved in Chl biosynthesis are altered in lil3 mutant plants. Our data support a role of LIL3 paralogues in the regulation of Chl biosynthesis under light stress and under standard growth conditions as well as in a coordinated ligation of newly synthesized and/or rescued Chl molecules to their target apoproteins. PMID:25808681

  8. The branching gene RAMOSUS1 mediates interactions among two novel signals and auxin in pea.

    Science.gov (United States)

    Foo, Eloise; Bullier, Erika; Goussot, Magali; Foucher, Fabrice; Rameau, Catherine; Beveridge, Christine Anne

    2005-02-01

    In Pisum sativum, the RAMOSUS genes RMS1, RMS2, and RMS5 regulate shoot branching via physiologically defined mobile signals. RMS1 is most likely a carotenoid cleavage enzyme and acts with RMS5 to control levels of an as yet unidentified mobile branching inhibitor required for auxin inhibition of branching. Our work provides molecular, genetic, and physiological evidence that RMS1 plays a central role in a shoot-to-root-to-shoot feedback system that regulates shoot branching in pea. Indole-3-acetic acid (IAA) positively regulates RMS1 transcript level, a potentially important mechanism for regulation of shoot branching by IAA. In addition, RMS1 transcript levels are dramatically elevated in rms3, rms4, and rms5 plants, which do not contain elevated IAA levels. This degree of upregulation of RMS1 expression cannot be achieved in wild-type plants by exogenous IAA application. Grafting studies indicate that an IAA-independent mobile feedback signal contributes to the elevated RMS1 transcript levels in rms4 plants. Therefore, the long-distance signaling network controlling branching in pea involves IAA, the RMS1 inhibitor, and an IAA-independent feedback signal. Consistent with physiological studies that predict an interaction between RMS2 and RMS1, rms2 mutations appear to disrupt this IAA-independent regulation of RMS1 expression. PMID:15659639

  9. Mechanisms of side branching and tip splitting in a model of branching morphogenesis.

    Directory of Open Access Journals (Sweden)

    Yina Guo

    Full Text Available Recent experimental work in lung morphogenesis has described an elegant pattern of branching phenomena. Two primary forms of branching have been identified: side branching and tip splitting. In our previous study of lung branching morphogenesis, we used a 4 variable partial differential equation (PDE, due to Meinhardt, as our mathematical model to describe the reaction and diffusion of morphogens creating those branched patterns. By altering key parameters in the model, we were able to reproduce all the branching styles and the switch between branching modes. Here, we attempt to explain the branching phenomena described above, as growing out of two fundamental instabilities, one in the longitudinal (growth direction and the other in the transverse direction. We begin by decoupling the original branching process into two semi-independent sub-processes, 1 a classic activator/inhibitor system along the growing stalk, and 2 the spatial growth of the stalk. We then reduced the full branching model into an activator/inhibitor model that embeds growth of the stalk as a controllable parameter, to explore the mechanisms that determine different branching patterns. We found that, in this model, 1 side branching results from a pattern-formation instability of the activator/inhibitor subsystem in the longitudinal direction. This instability is far from equilibrium, requiring a large inhomogeneity in the initial conditions. It successively creates periodic activator peaks along the growing stalk, each of which later on migrates out and forms a side branch; 2 tip splitting is due to a Turing-style instability along the transversal direction, that creates the spatial splitting of the activator peak into 2 simultaneously-formed peaks at the growing tip, the occurrence of which requires the widening of the growing stalk. Tip splitting is abolished when transversal stalk widening is prevented; 3 when both instabilities are satisfied, tip bifurcation occurs

  10. Mechanisms of Side Branching and Tip Splitting in a Model of Branching Morphogenesis

    Science.gov (United States)

    Guo, Yina; Sun, Mingzhu; Garfinkel, Alan; Zhao, Xin

    2014-01-01

    Recent experimental work in lung morphogenesis has described an elegant pattern of branching phenomena. Two primary forms of branching have been identified: side branching and tip splitting. In our previous study of lung branching morphogenesis, we used a 4 variable partial differential equation (PDE), due to Meinhardt, as our mathematical model to describe the reaction and diffusion of morphogens creating those branched patterns. By altering key parameters in the model, we were able to reproduce all the branching styles and the switch between branching modes. Here, we attempt to explain the branching phenomena described above, as growing out of two fundamental instabilities, one in the longitudinal (growth) direction and the other in the transverse direction. We begin by decoupling the original branching process into two semi-independent sub-processes, 1) a classic activator/inhibitor system along the growing stalk, and 2) the spatial growth of the stalk. We then reduced the full branching model into an activator/inhibitor model that embeds growth of the stalk as a controllable parameter, to explore the mechanisms that determine different branching patterns. We found that, in this model, 1) side branching results from a pattern-formation instability of the activator/inhibitor subsystem in the longitudinal direction. This instability is far from equilibrium, requiring a large inhomogeneity in the initial conditions. It successively creates periodic activator peaks along the growing stalk, each of which later on migrates out and forms a side branch; 2) tip splitting is due to a Turing-style instability along the transversal direction, that creates the spatial splitting of the activator peak into 2 simultaneously-formed peaks at the growing tip, the occurrence of which requires the widening of the growing stalk. Tip splitting is abolished when transversal stalk widening is prevented; 3) when both instabilities are satisfied, tip bifurcation occurs together with side

  11. Branch architecture, light interception and crown development in saplings of a plagiotropically branching tropical tree, Polyalthia jenkinsii (Annonaceae).

    Science.gov (United States)

    Osada, Noriyuki; Takeda, Hiroshi

    2003-01-01

    To investigate crown development patterns, branch architecture, branch-level light interception, and leaf and branch dynamics were studied in saplings of a plagiotropically branching tree species, Polyalthia jenkinsii Hk. f. & Thoms. (Annonaceae) in a Malaysian rain forest. Lengths of branches and parts of the branches lacking leaves ('bare' branches) were smaller in upper branches than in lower branches within crowns, whereas lengths of 'leafy' parts and the number of leaves per branch were larger in intermediate than in upper and lower branches. Maximum diffuse light absorption (DLA) of individual leaves was not related to sapling height or branch position within crowns, whereas minimum DLA was lower in tall saplings. Accordingly, branch-level light interception was higher in intermediate than in upper and lower branches. The leaf production rate was higher and leaf loss rate was smaller in upper than in intermediate and lower branches. Moreover, the branch production rate of new first-order branches was larger in the upper crowns. Thus, leaf and branch dynamics do not correspond to branch-level light interception in the different canopy zones. As a result of architectural constraints, branches at different vertical positions experience predictable light microenvironments in plagiotropic species. Accordingly, this pattern of carbon allocation among branches might be particularly important for growth and crown development in plagiotropic species. PMID:12495920

  12. Non-redundant functions of two proline dehydrogenase isoforms in Arabidopsis

    Directory of Open Access Journals (Sweden)

    Müller Gudrun

    2010-04-01

    Full Text Available Abstract Background Proline (Pro accumulation is a widespread response of prokaryotic and eukaryotic cells subjected to osmotic stress or dehydration. When the cells are released from stress, Pro is degraded to glutamate by Pro-dehydrogenase (ProDH and Pyrroline-5-carboxylate dehydrogenase (P5CDH, which are both mitochondrial enzymes in eukaryotes. While P5CDH is a single copy gene in Arabidopsis, two ProDH genes have been identified in the genome. Until now, only ProDH1 (At3g30775 had been functionally characterised. Results We demonstrate vasculature specific expression of the Arabidopsis ProDH2 gene (At5g38710 as well as enzymatic activity and mitochondrial localisation of the encoded protein. Expression levels of ProDH2 are generally low, but increased in senescent leaves and in the abscission zone of floral organs. While sucrose represses ProDH2 expression, Pro and NaCl were identified as inducers. Endogenous ProDH2 expression was not able to overcome Pro sensitivity of ProDH1 mutants, but overexpression of a GFP-tagged form of ProDH2 enabled the utilisation of Pro as single nitrogen source for growth. Amongst two intronic insertion mutants, one was identified as a null allele, whereas the other still produced native ProDH2 transcripts. Conclusions Arabidopsis possesses two functional ProDHs, which have non-redundant, although partially overlapping physiological functions. The two ProDH isoforms differ with respect to spatial, developmental and environmental regulation of expression. While ProDH1 appears to be the dominant isoform under most conditions and in most tissues, ProDH2 was specifically upregulated during salt stress, when ProDH1 was repressed. The characterisation of ProDH2 as a functional gene requires a careful re-analysis of mutants with a deletion of ProDH1, which were so far considered to be devoid of ProDH activity. We hypothesise that ProDH2 plays an important role in Pro homeostasis in the vasculature, especially under

  13. Agents in branching space-times

    Institute of Scientific and Technical Information of China (English)

    Nuel; Belnap

    2003-01-01

    The aim of this essay Is to make some brief suggestions on the beginnings of a theory of agentsand agency In branching spacetlmes.foe thought Is to combine the ideas of agency as developed agalnst the relatively simple background of branching time with the richer notions of Indetermlnlsm asstructured In the theory of branching space-times.My plan Is to say a little about agency In branchingtime and a little about branchlg spacetlmes,and then ask how the two can be brought together.Atthe end there Is an ...

  14. Measurement of Prominent eta Decay Branching Fractions

    CERN Document Server

    López, A; Méndez, H; Ramírez, J; Ge, J Y; Miller, D H; Sanghi, B; Shipsey, I P J; Xin, B; Adams, G S; Anderson, M; Cummings, J P; Danko, I; Hu, D; Moziak, B; Napolitano, J; He, Q; Insler, J; Muramatsu, H; Park, C S; Thorndike, E H; Yang, F; Artuso, M; Blusk, S; Khalil, S; Li, J; Menaa, N; Mountain, R; Nisar, S; Randrianarivony, K; Sia, R; Skwarnicki, T; Stone, S; Wang, J C; Bonvicini, G; Cinabro, D; Dubrovin, M; Lincoln, A; Asner, D M; Edwards, K W; Naik, P; Briere, R A; Ferguson, T; Tatishvili, G; Vogel, H; Watkins, M E; Rosner, J L; Adam, N E; Alexander, J P; Cassel, D G; Duboscq, J E; Ehrlich, R; Fields, L; Galik, R S; Gibbons, L; Gray, R; Gray, S W; Hartill, D L; Heltsley, B K; Hertz, D; Jones, C D; Kandaswamy, J; Kreinick, D L; Kuznetsov, V E; Mahlke-Krüger, H; Mohapatra, D; Onyisi, P U E; Patterson, J R; Peterson, D; Riley, D; Ryd, A; Sadoff, A J; Shi, X; Stroiney, S; Sun, W M; Wilksen, T; Athar, S B; Patel, R; Yelton, J; Rubin, P; Eisenstein, B I; Karliner, I; Lowrey, N; Selen, M; White, E J; Wiss, J; Mitchell, R E; Shepherd, M R; Besson, D; Pedlar, T K; Cronin-Hennessy, D; Gao, K Y; Hietala, J; Kubota, Y; Klein, T; Lang, B W; Poling, R; Scott, A W; Zweber, P; Dobbs, S; Metreveli, Z; Seth, K K; Tomaradze, A G; Ernst, J; Ecklund, K M; Severini, H; Love, W; Savinov, V

    2007-01-01

    The decay psi(2S) --> eta J/psi is used to measure, for the first time, all prominent eta-meson branching fractions with the same experiment in the same dataset, thereby providing a consistent treatment of systematics across branching fractions. We present results for eta decays to gamma gamma, pi+pi-pi0, 3 pi0, pi+ pi- gamma, and e+ e- gamma, accounting for 99.9% of all eta decays. The precisions for several of the branching fractions and their ratios are improved. Two channels, pi+ pi- gamma and e+ e- gamma, show results that differ at the level of three standard deviations from those previously determined.

  15. Different phosphoinositide 3-kinase isoforms mediate carrageenan nociception and inflammation.

    Science.gov (United States)

    Pritchard, Rory A; Falk, Lovissa; Larsson, Mathilda; Leinders, Mathias; Sorkin, Linda S

    2016-01-01

    Phosphoinositide 3-kinases (PI3Ks) participate in signal transduction cascades that can directly activate and sensitize nociceptors and enhance pain transmission. They also play essential roles in chemotaxis and immune cell infiltration leading to inflammation. We wished to determine which PI3K isoforms were involved in each of these processes. Lightly anesthetized rats (isoflurane) were injected subcutaneously with carrageenan in their hind paws. This was preceded by a local injection of 1% DMSO vehicle or an isoform-specific antagonist to PI3K-α (compound 15-e), -β (TGX221), -δ (Cal-101), or -γ (AS252424). We measured changes in the mechanical pain threshold and spinal c-Fos expression (4 hours after injection) as indices of nociception. Paw volume, plasma extravasation (Evans blue, 0.3 hours after injection), and neutrophil (myeloperoxidase; 1 hour after injection) and macrophage (CD11b+; 4 hour after injection) infiltration into paw tissue were the measured inflammation endpoints. Only PI3K-γ antagonist before treatment reduced the carrageenan-induced pain behavior and spinal expression of c-Fos (P ≤ 0.01). In contrast, pretreatment with PI3K-α, -δ, and-γ antagonists reduced early indices of inflammation. Plasma extravasation PI3K-α (P ≤ 0.05), -δ (P ≤ 0.05), and -γ (P ≤ 0.01), early (0-2 hour) edema -α (P ≤ 0.05), -δ (P ≤ 0.001), and -γ (P ≤ 0.05), and neutrophil infiltration (all P ≤ 0.001) were all reduced compared to vehicle pretreatment. Later (2-4 hour), edema and macrophage infiltration (P ≤ 0.05) were reduced by only the PI3K-δ and -γ isoform antagonists, with the PI3K-δ antagonist having a greater effect on edema. PI3K-β antagonism was ineffective in all paradigms. These data indicate that pain and clinical inflammation are pharmacologically separable and may help to explain clinical conditions in which inflammation naturally wanes or goes into remission, but pain continues unabated. PMID:26313408

  16. Activation of antithrombin III isoforms by heparan sulphate glycosaminoglycans and other sulphated polysaccharides.

    Science.gov (United States)

    Carlson, T H; Kolman, M R; Piepkorn, M

    1995-07-01

    Antithrombin III occurs naturally as two functionally distinct molecular species that differ in glycosylation at Asn135. Whereas the predominant, glycosylated isoform has high affinity for heparin, a quantitatively minor isoform lacking glycosylation at that site displays relatively higher affinity for both heparins and heparinoids. We characterized the ability of various sulphated polysaccharides to potentiate the rates of thrombin inhibition by the isoforms. High-molecular-weight dextran sulphate was the most effective of those studied, increasing thrombin inhibition by the higher-affinity antithrombin III isoform up to five-fold more efficiently than did heparin fractions with low-affinity for antithrombin III. In addition, dextran sulphate activated the higher-affinity isoform as much as twelve times more effectively than it did the lower-affinity isoform. Pentosan polysulphate was up to three-fold, and some heparan sulphate fractions up to two-fold, more effective with the higher, compared with the lower affinity, isoform. Heparan sulphate preparations less effectively increased the rate of thrombin inhibition than did the other low-affinity polysaccharides. Structure-function studies indicated positive correlations between activity and both polymer length and anionic group density of low-affinity sulphated polysaccharides. The observed effects of the heparan sulphates on this anticoagulant pathway, although of low potency, are consistent with the hypotheses that these substances naturally regulate blood homeostasis in vascular tissues and that much of this function may be mediated by the higher-affinity antithrombin III isoform. PMID:8589216

  17. Activation of AMPK alpha and gamma-isoform complexes in the intact ischemic rat heart

    Science.gov (United States)

    AMP-activated protein kinase (AMPK) plays a key role in modulating cellular metabolic processes. AMPK, a serine-threonine kinase, is a heterotrimeric complex of catalytic alpha-subunits and regulatory beta- and gamma-subunits with multiple isoforms. Mutations in the cardiac gamma(2)-isoform have bee...

  18. Comprehensive analysis of tropomyosin isoforms in skeletal muscles by top-down proteomics.

    Science.gov (United States)

    Jin, Yutong; Peng, Ying; Lin, Ziqing; Chen, Yi-Chen; Wei, Liming; Hacker, Timothy A; Larsson, Lars; Ge, Ying

    2016-04-01

    Mammalian skeletal muscles are heterogeneous in nature and are capable of performing various functions. Tropomyosin (Tpm) is a major component of the thin filament in skeletal muscles and plays an important role in controlling muscle contraction and relaxation. Tpm is known to consist of multiple isoforms resulting from different encoding genes and alternative splicing, along with post-translational modifications. However, a systematic characterization of Tpm isoforms in skeletal muscles is still lacking. Therefore, we employed top-down mass spectrometry (MS) to identify and characterize Tpm isoforms present in different skeletal muscles from multiple species, including swine, rat, and human. Our study revealed that Tpm1.1 and Tpm2.2 are the two major Tpm isoforms in swine and rat skeletal muscles, whereas Tpm1.1, Tpm2.2, and Tpm3.12 are present in human skeletal muscles. Tandem MS was utilized to identify the sequences of the major Tpm isoforms. Furthermore, quantitative analysis revealed muscle-type specific differences in the abundance of un-modified and modified Tpm isoforms in rat and human skeletal muscles. This study represents the first systematic investigation of Tpm isoforms in skeletal muscles, which not only demonstrates the capabilities of top-down MS for the comprehensive characterization of skeletal myofilament proteins but also provides the basis for further studies on these Tpm isoforms in muscle-related diseases. PMID:27090236

  19. Molecular cloning and pharmacology of functionally distinct isoforms of the human histamine H(3) receptor

    DEFF Research Database (Denmark)

    Wellendorph, Petrine; Goodman, M W; Burstein, E S;

    2002-01-01

    The pharmacology of histamine H(3) receptors suggests the presence of distinct receptor isoforms or subtypes. We herein describe multiple, functionally distinct, alternatively spliced isoforms of the human H(3) receptor. Combinatorial splicing at three different sites creates at least six distinc...

  20. Roles of the troponin isoforms during indirect flight muscle development in Drosophila

    Indian Academy of Sciences (India)

    Salam Herojeet Singh; Prabodh Kumar; Nallur B. Ramachandra; Upendra Nongthomba

    2014-08-01

    Troponin proteins in cooperative interaction with tropomyosin are responsible for controlling the contraction of the striated muscles in response to changes in the intracellular calcium concentration. Contractility of the muscle is determined by the constituent protein isoforms, and the isoforms can switch over from one form to another depending on physiological demands and pathological conditions. In Drosophila, amajority of themyofibrillar proteins in the indirect flight muscles (IFMs) undergo post-transcriptional and post-translational isoform changes during pupal to adult metamorphosis to meet the high energy and mechanical demands of flight. Using a newly generated Gal4 strain (UH3-Gal4) which is expressed exclusively in the IFMs, during later stages of development, we have looked at the developmental and functional importance of each of the troponin subunits (troponin-I, troponin-T and troponin-C) and their isoforms. We show that all the troponin subunits are required for normal myofibril assembly and flight, except for the troponin-C isoform 1 (TnC1). Moreover, rescue experiments conducted with troponin-I embryonic isoform in the IFMs, where flies were rendered flightless, show developmental and functional differences of TnI isoforms and importance of maintaining the right isoform.

  1. Translational control of C/EBPalpha and C/EBPbeta isoform expression

    NARCIS (Netherlands)

    Calkhoven, C F; Müller, C; Leutz, A

    2000-01-01

    Transcription factors derived from CCAAT/enhancer binding protein (C/EBP)alpha and C/EBPbeta genes control differentiation and proliferation in a number of cell types. Various C/EBP isoforms arise from unique C/EBPbeta and C/EBPalpha mRNAs by differential initiation of translation. These isoforms re

  2. Regulation of carnitine palmitoyltransferase I (CPT-Iα) gene expression by the peroxisome proliferator activated receptor gamma coactivator (PGC-1) isoforms

    Science.gov (United States)

    Sadana, Prabodh; Zhang, Yi; Song, Shulan; Cook, George A.; Elam, Marshall B.; Park, Edwards A.

    2007-01-01

    Summary The peroxisome proliferator-activated receptor gamma coactivators (PGC-1) have important roles in mitochondrial biogenesis and metabolic control in a variety of tissues. There are multiple isoforms of PGC-1 including PGC-1α and PGC-1β. Both the PGC-1α and β isoforms promote mitochondrial biogenesis and fatty acid oxidation, but only PGC-1α stimulates gluconeogenesis in the liver. Carnitine palmitoyltransferase I (CPT-I) is a key enzyme regulating mitochondrial fatty acid oxidation. In these studies, we determined that PGC-1β stimulated expression of the “liver” isoform of CPT-I (CPT-Iα) but that PGC-1β did not induce pyruvate dehydrogenase kinase 4 (PDK4) which is a regulator of pyruvate metabolism. The CPT-Iα gene is induced by thyroid hormone. We found that T3 increased the expression of PGC-1β and that PGC-1β enhanced the T3 induction of CPT-Iα. The thyroid hormone receptor interacts with PGC-1β in a ligand dependent manner. Unlike PGC-1α, the interaction of PGC-1β and the T3 receptor does not occur exclusively through the leucine-X-X-leucine-leucine motif in PGC-1β. We have found that PGC-1β is associated with the CPT-Iα gene in vivo. Overall, our results demonstrate that PGC-1β is a coactivator in the T3 induction of CPT-Iα and that PGC-1β has similarities and differences with the PGC-1α isoform. PMID:17239528

  3. Enzymes for improved biomass conversion

    Energy Technology Data Exchange (ETDEWEB)

    Brunecky, Roman; Himmel, Michael E.

    2016-02-02

    Disclosed herein are enzymes and combinations of the enzymes useful for the hydrolysis of cellulose and the conversion of biomass. Methods of degrading cellulose and biomass using enzymes and cocktails of enzymes are also disclosed.

  4. Enzymes and muscle diseases

    Directory of Open Access Journals (Sweden)

    M. Plebani

    2011-09-01

    Full Text Available Skeletal muscle disorders may result in release of muscle enzymes into the circulation and give increased serum enzyme activity. A variety of enzymes routinely determined in the clinical laboratory may be elevated, but creatine kinase is the enzyme present in the highest concentration in muscle, and in every variety of muscle disease is the serum enzyme which shows the greatest incidence and degree of elevation. Aspartate aminotransferase is the enzyme associated most significantly with inflammation. A diagnostic algorithm based on the combined measurement of creatine kinase, aspartate aminotransferase and aldolase has been found to discriminate muscular distrophies from polymyositis and other myopathies. This combination of laboratory tests has diagnostic application and thus allows the clinician to better select patients who need to have a skeletal muscle biopsy as a diagnostic procedure.

  5. Exo70 Isoform Switching upon Epithelial-Mesenchymal Transition Mediates Cancer Cell Invasion

    Science.gov (United States)

    Lu, Hezhe; Liu, Jianglan; Liu, Shujing; Zeng, Jingwen; Ding, Deqiang; Carstens, Russ P.; Cong, Yusheng; Xu, Xiaowei; Guo, Wei

    2014-01-01

    Summary Epithelial-mesenchymal transition (EMT) is an important developmental process hijacked by cancer cells for their dissemination. Here we show that Exo70, a component of the exocyst complex, undergoes isoform switching mediated by ESRP1, a pre-mRNA splicing factor that regulates EMT. Expression of the epithelial isoform of Exo70 affects the levels of key EMT transcriptional regulators such as Snail and ZEB2, and is sufficient to drive the transition to epithelial phenotypes. Differential Exo70 isoforms expression in human tumors correlates with cancer progression, and increased expression of the epithelial isoform of Exo70 inhibits tumor metastasis in mice. At the molecular level, the mesenchymal but not the epithelial isoform of Exo70 interacts with the Arp2/3 complex and stimulates actin polymerization for tumor invasion. Our findings provide a mechanism by which the exocyst function and actin dynamics are modulated for EMT and tumor invasion. PMID:24331928

  6. Pharmacological targeting of PI3K isoforms as a therapeutic strategy in chronic lymphocytic leukaemia

    Science.gov (United States)

    Blunt, Matthew D.; Steele, Andrew J.

    2015-01-01

    PI3Kδ inhibitors such as idelalisib are providing improved therapeutic options for the treatment of chronic lymphocytic leukaemia (CLL). However under certain conditions, inhibition of a single PI3K isoform can be compensated by the other PI3K isoforms, therefore PI3K inhibitors which target multiple PI3K isoforms may provide greater efficacy. The development of compounds targeting multiple PI3K isoforms (α, β, δ, and γ) in CLL cells, in vitro, resulted in sustained inhibition of BCR signalling but with enhanced cytotoxicity and the potential for improve clinical responses. This review summarises the progress of PI3K inhibitor development and describes the rationale and potential for targeting multiple PI3K isoforms. PMID:26500849

  7. Pharmacological targeting of PI3K isoforms as a therapeutic strategy in chronic lymphocytic leukaemia

    Directory of Open Access Journals (Sweden)

    Matthew D. Blunt

    2015-01-01

    Full Text Available PI3Kδ inhibitors such as idelalisib are providing improved therapeutic options for the treatment of chronic lymphocytic leukaemia (CLL. However under certain conditions, inhibition of a single PI3K isoform can be compensated by the other PI3K isoforms, therefore PI3K inhibitors which target multiple PI3K isoforms may provide greater efficacy. The development of compounds targeting multiple PI3K isoforms (α, β, δ, and γ in CLL cells, in vitro, resulted in sustained inhibition of BCR signalling but with enhanced cytotoxicity and the potential for improve clinical responses. This review summarises the progress of PI3K inhibitor development and describes the rationale and potential for targeting multiple PI3K isoforms.

  8. Isoform-specific phosphorylation-dependent regulation of connexin hemichannels

    DEFF Research Database (Denmark)

    Alstrøm, Jette Skov; Hansen, Daniel Bloch; Nielsen, Morten Schak;

    2015-01-01

    Connexins form gap junction channels made up of two connexons (hemichannels) from adjacent cells. Unopposed hemichannels may open toward the extracellular space upon stimulation by, e.g., removal of divalent cations from the extracellular solution and allow isoform-specific transmembrane flux of...... fluorescent dyes and physiologically relevant molecules, such as ATP and ions. Connexin (Cx)43 and Cx30 are the major astrocytic connexins. Protein kinase C (PKC) regulates Cx43 in its cell-cell gap junction configuration and may also act to keep Cx43 hemichannels closed. In contrast, the regulation of Cx30...... hemichannels by PKC is unexplored. To determine phosphorylation-dependent regulation of Cx30 and Cx43 hemichannels, these were heterologously expressed in Xenopus laevis oocytes and opened with divalent cation-free solution. Inhibition of PKC activity did not affect hemichannel opening of either connexin. PKC...

  9. Branching processes with immigration and related topics

    OpenAIRE

    Li, Zenghu

    2006-01-01

    This is a survey on recent progresses in the study of branching processes with immigration, generalized Ornstein-Uhlenbeck processes and affine Markov processes. We mainly focus on the applications of skew convolution semigroups and the connections in those processes.

  10. Applied Aeroscience and CFD Branch Overview

    Science.gov (United States)

    LeBeau, Gerald J.; Kirk, Benjamin S.

    2014-01-01

    The principal mission of NASA Johnson Space Center is Human Spaceflight. In support of the mission the Applied Aeroscience and CFD Branch has several technical competencies that include aerodynamic characterization, aerothermodynamic heating, rarefied gas dynamics, and decelerator (parachute) systems.

  11. FY 1990 Applied Sciences Branch annual report

    Energy Technology Data Exchange (ETDEWEB)

    Keyes, B.M.; Dippo, P.C. [eds.

    1991-11-01

    The Applied Sciences Branch actively supports the advancement of DOE/SERI goals for the development and implementation of the solar photovoltaic technology. The primary focus of the laboratories is to provide state-of-the-art analytical capabilities for materials and device characterization and fabrication. The branch houses a comprehensive facility which is capable of providing information on the full range of photovoltaic components. A major objective of the branch is to aggressively pursue collaborative research with other government laboratories, universities, and industrial firms for the advancement of photovoltaic technologies. Members of the branch disseminate research findings to the technical community in publications and presentations. This report contains information on surface and interface analysis, materials characterization, development, electro-optical characterization module testing and performance, surface interactions and FTIR spectroscopy.

  12. Residence times of branching diffusion processes

    Science.gov (United States)

    Dumonteil, E.; Mazzolo, A.

    2016-07-01

    The residence time of a branching Brownian process is the amount of time that the mother particle and all its descendants spend inside a domain. Using the Feynman-Kac formalism, we derive the residence-time equation as well as the equations for its moments for a branching diffusion process with an arbitrary number of descendants. This general approach is illustrated with simple examples in free space and in confined geometries where explicit formulas for the moments are obtained within the long time limit. In particular, we study in detail the influence of the branching mechanism on those moments. The present approach can also be applied to investigate other additive functionals of branching Brownian process.

  13. Code 672 observational science branch computer networks

    Science.gov (United States)

    Hancock, D. W.; Shirk, H. G.

    1988-01-01

    In general, networking increases productivity due to the speed of transmission, easy access to remote computers, ability to share files, and increased availability of peripherals. Two different networks within the Observational Science Branch are described in detail.

  14. Molecular Basis of Inhibitory Activities of Berberine against Pathogenic Enzymes in Alzheimer's Disease

    OpenAIRE

    Hong-Fang Ji; Liang Shen

    2012-01-01

    The natural isoquinoline alkaloid berberine possesses potential to treat Alzheimer's disease (AD) by targeting multiple pathogenic factors. In the present study, docking simulations were performed to gain deeper insights into the molecular basis of berberine's inhibitory effects against the important pathogenic enzymes of AD, that is, acetylcholinesterase, butyrylcholinesterase, and two isoforms of monoamine oxidase. It was found that the theoretical binding affinities of berberine to the fou...

  15. Arachidonic Acid-metabolizing Cytochrome P450 Enzymes Are Targets of ω-3 Fatty Acids*

    OpenAIRE

    Arnold, Cosima; Markovic, Marija; Blossey, Katrin; Wallukat, Gerd; Fischer, Robert; Dechend, Ralf; Konkel, Anne; von Schacky, Clemens; Luft, Friedrich C.; Muller, Dominik N.; Rothe, Michael; Schunck, Wolf-Hagen

    2010-01-01

    Eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) protect against cardiovascular disease by largely unknown mechanisms. We tested the hypothesis that EPA and DHA may compete with arachidonic acid (AA) for the conversion by cytochrome P450 (CYP) enzymes, resulting in the formation of alternative, physiologically active, metabolites. Renal and hepatic microsomes, as well as various CYP isoforms, displayed equal or elevated activities when metabolizing EPA or DHA instead of AA. CYP2C/2J...

  16. Glutamine Synthetase in Legumes: Recent Advances in Enzyme Structure and Functional Genomics

    OpenAIRE

    Marco Betti; Margarita García-Calderón; Pérez-Delgado, Carmen M.; Alfredo Credali; Guillermo Estivill; Francisco Galván; Vega, José M.; Márquez, Antonio J.

    2012-01-01

    Glutamine synthetase (GS) is the key enzyme involved in the assimilation of ammonia derived either from nitrate reduction, N2 fixation, photorespiration or asparagine breakdown. A small gene family is encoding for different cytosolic (GS1) or plastidic (GS2) isoforms in legumes. We summarize here the recent advances carried out concerning the quaternary structure of GS, as well as the functional relationship existing between GS2 and processes such as nodulation, photore...

  17. Troponin T isoform expression is modulated during Atlantic Halibut metamorphosis

    Directory of Open Access Journals (Sweden)

    Llewellyn Lynda

    2007-06-01

    Full Text Available Abstract Background Flatfish metamorphosis is a thyroid hormone (TH driven process which leads to a dramatic change from a symmetrical larva to an asymmetrical juvenile. The effect of THs on muscle and in particular muscle sarcomer protein genes is largely unexplored in fish. The change in Troponin T (TnT, a pivotal protein in the assembly of skeletal muscles sarcomeres and a modulator of calcium driven muscle contraction, during flatfish metamophosis is studied. Results In the present study five cDNAs for halibut TnT genes were cloned; three were splice variants arising from a single fast TnT (fTnT gene; a fourth encoded a novel teleost specific fTnT-like cDNA (AfTnT expressed exclusively in slow muscle and the fifth encoded the teleost specific sTnT2. THs modified the expression of halibut fTnT isoforms which changed from predominantly basic to acidic isoforms during natural and T4 induced metamorphosis. In contrast, expression of red muscle specific genes, AfTnT and sTnT2, did not change during natural metamorphosis or after T4 treatment. Prior to and after metamorphosis no change in the dorso-ventral symmetry or temporal-spatial expression pattern of TnT genes and muscle fibre organization occurred in halibut musculature. Conclusion Muscle organisation in halibut remains symmetrical even after metamorphosis suggesting TH driven changes are associated with molecular adaptations. We hypothesize that species specific differences in TnT gene expression in teleosts underlies different larval muscle developmental programs which better adapts them to the specific ecological constraints.

  18. p53 isoforms regulate astrocyte-mediated neuroprotection and neurodegeneration.

    Science.gov (United States)

    Turnquist, C; Horikawa, I; Foran, E; Major, E O; Vojtesek, B; Lane, D P; Lu, X; Harris, B T; Harris, C C

    2016-09-01

    Bidirectional interactions between astrocytes and neurons have physiological roles in the central nervous system and an altered state or dysfunction of such interactions may be associated with neurodegenerative diseases, such as Alzheimer's disease (AD) and amyotrophic lateral sclerosis (ALS). Astrocytes exert structural, metabolic and functional effects on neurons, which can be either neurotoxic or neuroprotective. Their neurotoxic effect is mediated via the senescence-associated secretory phenotype (SASP) involving pro-inflammatory cytokines (e.g., IL-6), while their neuroprotective effect is attributed to neurotrophic growth factors (e.g., NGF). We here demonstrate that the p53 isoforms Δ133p53 and p53β are expressed in astrocytes and regulate their toxic and protective effects on neurons. Primary human astrocytes undergoing cellular senescence upon serial passaging in vitro showed diminished expression of Δ133p53 and increased p53β, which were attributed to the autophagic degradation and the SRSF3-mediated alternative RNA splicing, respectively. Early-passage astrocytes with Δ133p53 knockdown or p53β overexpression were induced to show SASP and to exert neurotoxicity in co-culture with neurons. Restored expression of Δ133p53 in near-senescent, otherwise neurotoxic astrocytes conferred them with neuroprotective activity through repression of SASP and induction of neurotrophic growth factors. Brain tissues from AD and ALS patients possessed increased numbers of senescent astrocytes and, like senescent astrocytes in vitro, showed decreased Δ133p53 and increased p53β expression, supporting that our in vitro findings recapitulate in vivo pathology of these neurodegenerative diseases. Our finding that Δ133p53 enhances the neuroprotective function of aged and senescent astrocytes suggests that the p53 isoforms and their regulatory mechanisms are potential targets for therapeutic intervention in neurodegenerative diseases. PMID:27104929

  19. Locomotion in Lymphocytes is Altered by Differential PKC Isoform Expression

    Science.gov (United States)

    Sundaresan, A.; Risin, D.; Pellis, N. R.

    1999-01-01

    Lymphocyte locomotion is critical for proper elicitation of the immune response. Locomotion of immune cells via the interstitium is essential for optimal immune function during wound healing, inflammation and infection. There are conditions which alter lymphocyte locomotion and one of them is spaceflight. Lymphocyte locomotion is severely inhibited in true spaceflight (true microgravity) and in rotating wall vessel culture (modeled microgravity). When lymphocytes are activated prior to culture in modeled microgravity, locomotion is not inhibited and the levels are comparable to those of static cultured lymphocytes. When a phorbol ester (PMA) is used in modeled microgravity, lymphocyte locomotion is restored by 87%. This occurs regardless if PMA is added after culture in the rotating wall vessel or during culture. Inhibition of DNA synthesis also does not alter restoration of lymphocyte locomotion by PMA. PMA is a direct activator of (protein kinase C) PKC . When a calcium ionophore, ionomycin is used it does not possess any restorative properties towards locomotion either alone or collectively with PMA. Since PMA brings about restoration without help from calcium ionophores (ionomycin), it is infer-red that calcium independent PKC isoforms are involved. Changes were perceived in the protein levels of PKC 6 where levels of the protein were downregulated at 24,72 and 96 hours in untreated rotated cultures (modeled microgravity) compared to untreated static (1g) cultures. At 48 hours there is an increase in the levels of PKC & in the same experimental set up. Studies on transcriptional and translational patterns of calcium independent isoforms of PKC such as 8 and E are presented in this study.

  20. Soils of Walker Branch Watershed

    Energy Technology Data Exchange (ETDEWEB)

    Lietzke, D.A.

    1994-01-01

    The soil survey of Walker Branch Watershed (WBW) utilized the most up-to-date knowledge of soils, geology, and geohydrology in building the soils data base needed to reinterpret past research and to begin new research in the watershed. The soils of WBW were also compared with soils mapped elsewhere along Chestnut Ridge on the Oak Ridge Reservation to (1) establish whether knowledge obtained elsewhere could be used within the watershed, (2) determine whether there were any soils restricted to the watershed, and (3) evaluate geologic formation lateral variability. Soils, surficial geology, and geomorphology were mapped at a scale of 1:1200 using a paper base map having 2-ft contour intervals. Most of the contours seemed to reasonably represent actual landform configurations, except for dense wooded areas. For example, the very large dolines or sinkholes were shown on the contour base map, but numerous smaller ones were not. In addition, small drainageways and gullies were often not shown. These often small but important features were located approximately as soil mapping progressed. WBW is underlain by dolostones of the Knox Group, but only a very small part of the surface area contains outcroppings of rock and most outcrops were located in the lower part. Soil mapping revealed the presence of both ancient alluvium and ancient colluvium deposits, not recognized in previous soil surveys, that have been preserved in high-elevation stable portions of present-day landforms. An erosional geomorphic process of topographic inversion requiring several millions of years within the Pleistocene is necessary to bring about the degree of inversion that is expressed in the watershed. Indeed, some of these ancient alluvial and colluvial remnants may date back into the Tertiary. Also evident in the watershed, and preserved in the broad, nearly level bottoms of dolines, are multiple deposits of silty material either devoid or nearly devoid of coarse fragments. Recent research

  1. From configurations to branched configurations and beyond

    OpenAIRE

    Magnot, Jean-Pierre

    2013-01-01

    We propose here a geometric and topological setting for the study of branching effects arising in various fields of research, e.g. in statistical mechanics and turbulence theory. We describe various aspects that appear key points to us, and finish with a limit of such a construction which stand in the dynamics on probability spaces where it seems that branching effects can be fully studied without any adaptation of the framework.

  2. CAST Xi'an Branch Established

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    @@ China Academy of Space Technology (CAST) Xi'an Branch was established on April 8,2008.The new branch consists of the headquarters of the former Xi'an Institute of Space Radio Technology (XISRT) of CAST and five former subsidiaries of the XISRT,i.e.the Institute of Space Electronic Technology,the Institute of Microwave Technology,the Institute of Space Antenna Technology,the Manufacturing Center for Space Electronics and Civilian High-tech Company.

  3. AGB (asymptotic giant branch): Star evolution

    Energy Technology Data Exchange (ETDEWEB)

    Becker, S.A.

    1987-01-01

    Asymptotic giant branch stars are red supergiant stars of low-to-intermediate mass. This class of stars is of particular interest because many of these stars can have nuclear processed material brought up repeatedly from the deep interior to the surface where it can be observed. A review of recent theoretical and observational work on stars undergoing the asymptotic giant branch phase is presented. 41 refs.

  4. Branch retinal artery occlusion in Susac's syndrome

    Directory of Open Access Journals (Sweden)

    Ricardo Evangelista Marrocos de Aragão

    2015-02-01

    Full Text Available Susac's syndrome is a rare disease attribuited to a microangiopathy involving the arterioles of the cochlea, retina and brain. Encefalopathy, hearing loss, and visual deficits are the hallmarks of the disease. Visual loss is due to multiple, recurrent branch arterial retinal occlusions. We report a case of a 20-year-old women with Susac syndrome presented with peripheral vestibular syndrome, hearing loss, ataxia, vertigo, and vision loss due occlusion of the retinal branch artery.

  5. Branch and Bound Algorithm for Multiprocessor Scheduling

    OpenAIRE

    Rahman, Mostafizur

    2009-01-01

    The multiprocessor task graph scheduling problem has been extensively studied asacademic optimization problem which occurs in optimizing the execution time of parallelalgorithm with parallel computer. The problem is already being known as one of the NPhardproblems. There are many good approaches made with many optimizing algorithmto find out the optimum solution for this problem with less computational time. One ofthem is branch and bound algorithm.In this paper, we propose a branch and bound...

  6. AGB [asymptotic giant branch]: Star evolution

    International Nuclear Information System (INIS)

    Asymptotic giant branch stars are red supergiant stars of low-to-intermediate mass. This class of stars is of particular interest because many of these stars can have nuclear processed material brought up repeatedly from the deep interior to the surface where it can be observed. A review of recent theoretical and observational work on stars undergoing the asymptotic giant branch phase is presented. 41 refs

  7. Comparative efficiency analysis of Portuguese bank branches

    OpenAIRE

    Portela, Maria; Thanassoulis, Emmanuel

    2006-01-01

    The advent of Internet banking and phone banking is changing the role of bank branches from a predominantly transaction- based one to a sales-oriented role. This paper reports on an assessment of the branches of a Portuguese bank in terms of their performance in their new roles in three different areas: Their efficiency in fostering the use of new transaction channels, their efficiency in increasing sales and their customer base, and their efficiency in generating profits. Service qu...

  8. Comparative performance analysis of portuguese bank branches

    OpenAIRE

    Portela, Maria; Thanassoulis, Emmanuel

    2003-01-01

    The advent of Internet banking and phone banking is changing the role of bank branches from a predominantly transaction-based one to a sales-oriented role. This paper reports on an assessment of the branches of a Portuguese bank in terms of their performance in their new roles in three different areas: Their effectiveness in fostering the use of new transaction channels such as the internet and the telephone, their effectiveness in increasing sales and their customer base, and the...

  9. Fiber-Optic Communication Technology Branching Devices

    Science.gov (United States)

    Williams, J. C.

    1985-02-01

    This tutorial review of fiber-optic branching devices covers example uses of branching devices, device types, device-performance characteristics, examples of current technology, and system-design methodology. The discussion is limited to passive single- and multimode devices fabricated from optical fibers or graded-index components. Integrated-optic, wavelength-division-multiplexing, and polarization-selective devices are not specifically addressed.

  10. Characterization of ductal and lobular breast carcinomas using novel prolactin receptor isoform specific antibodies

    International Nuclear Information System (INIS)

    Prolactin is a polypeptide hormone responsible for proliferation and differentiation of the mammary gland. More recently, prolactin's role in mammary carcinogenesis has been studied with greater interest. Studies from our laboratory and from others have demonstrated that three specific isoforms of the prolactin receptor (PRLR) are expressed in both normal and cancerous breast cells and tissues. Until now, reliable isoform specific antibodies have been lacking. We have prepared and characterized polyclonal antibodies against each of the human PRLR isoforms that can effectively be used to characterize human breast cancers. Rabbits were immunized with synthetic peptides of isoform unique regions and immune sera affinity purified prior to validation by Western blot and immunohistochemical analyses. Sections of ductal and lobular carcinomas were stained with each affinity purified isoform specific antibody to determine expression patterns in breast cancer subclasses. We show that the rabbit antibodies have high titer and could specifically recognize each isoform of PRLR. Differences in PRLR isoform expression levels were observed and quantified using histosections from xenografts of established human breast cancer cells lines, and ductal and lobular carcinoma human biopsy specimens. In addition, these results were verified by real-time PCR with isoform specific primers. While nearly all tumors contained LF and SF1b, the majority (76%) of ductal carcinoma biopsies expressed SF1a while the majority of lobular carcinomas lacked SF1a staining (72%) and 27% had only low levels of expression. Differences in the receptor isoform expression profiles may be critical to understanding the role of PRL in mammary tumorigenesis. Since these antibodies are specifically directed against each PRLR isoform, they are valuable tools for the evaluation of breast cancer PRLR content and have potential clinical importance in treatment of this disease by providing new reagents to study

  11. Plasma branched chain amino acid abnormalities in sake-treated rats.

    Directory of Open Access Journals (Sweden)

    Watanabe,Akiharu

    1985-02-01

    Full Text Available Plasma amino acid abnormalities in rats treated with large doses of sake and whisky for 3 days were investigated under adequate nutritional conditions. A significant decrease in plasma branched-chain amino acid (BCAA levels was observed in sake- but not whisky-treated rats. However, known factors affecting BCAA levels, such as serum insulin and plasma glucagon levels ahd BCAA-metabolizing enzyme (BCAA transaminase and branched chain alpha-ketoacid dehydrogenase activities in the liver and skeletal muscle, were not significantly altered in the sake group. Furthermore, ethanol-metabolizing enzyme (alcohol and aldehyde dehydrogenases and the microsomal ethanol-oxidizing system activities in the liver were not altered in the sake group. Other mechanisms need to be considered for explaining the diminished levels of plasma BCAA in sake-treated rats.

  12. All change at the CERN UBS branch

    CERN Multimedia

    Antonella Del Rosso

    2012-01-01

    UBS branches across the country are being modernised, and the CERN branch is no exception. The Bulletin brings you a preview of the project, which will get under way in January 2013.   Mock-up of the renovated UBS branch. The changes at the UBS branch in CERN's Main Building will be no simple facelift. The entire bank will be renovated, transforming the present relatively confined premises into an open and attractive area. "The renovation of the UBS branches is part of a wider campaign designed to further enhance our customer relations," explains Ezio Mangia, the head of the CERN branch.  The UBS bank currently occupies three sets of premises in CERN's Main Building (two on the ground floor and one in the basement). "By the end of the work, which is scheduled to be completed by the middle of next year, CERN customers will benefit from a new area with open-plan counters and "hole-in-the-wall" machines accessible to...

  13. Branched-Chain Aminotransferases Control TORC1 Signaling in Saccharomyces cerevisiae.

    Directory of Open Access Journals (Sweden)

    Joanne M Kingsbury

    2015-12-01

    Full Text Available The conserved target of rapamycin complex 1 (TORC1 integrates nutrient signals to orchestrate cell growth and proliferation. Leucine availability is conveyed to control TORC1 activity via the leu-tRNA synthetase/EGOC-GTPase module in yeast and mammals, but the mechanisms sensing leucine remain only partially understood. We show here that both leucine and its α-ketoacid metabolite, α-ketoisocaproate, effectively activate the yeast TORC1 kinase via both EGOC GTPase-dependent and -independent mechanisms. Leucine and α-ketoisocaproate are interconverted by ubiquitous branched-chain aminotransferases (BCAT, which in yeast are represented by the mitochondrial and cytosolic enzymes Bat1 and Bat2, respectively. BCAT yeast mutants exhibit severely compromised TORC1 activity, which is partially restored by expression of Bat1 active site mutants, implicating both catalytic and structural roles of BCATs in TORC1 control. We find that Bat1 interacts with branched-chain amino acid metabolic enzymes and, in a leucine-dependent fashion, with the tricarboxylic acid (TCA-cycle enzyme aconitase. BCAT mutation perturbed TCA-cycle intermediate levels, consistent with a TCA-cycle block, and resulted in low ATP levels, activation of AMPK, and TORC1 inhibition. We propose the biosynthetic capacity of BCAT and its role in forming multicomplex metabolons connecting branched-chain amino acids and TCA-cycle metabolism governs TCA-cycle flux to activate TORC1 signaling. Because mammalian mitochondrial BCAT is known to form a supramolecular branched-chain α-keto acid dehydrogenase enzyme complex that links leucine metabolism to the TCA-cycle, these findings establish a precedent for understanding TORC1 signaling in mammals.

  14. Northwestern Branch of Mangala Vallis

    Science.gov (United States)

    2002-01-01

    (Released 12 June 2002) The Science One of the many branches of the Mangala Vallis channel system is seen in this image. The water that likely carved the channels emerged from a huge graben or fracture almost 1000 km to the south. The THEMIS image shows where one of the channels exits the cratered highlands terrain onto the lowland plains. A bright scarp marks the transition between the two terrain types and demonstrates that in this location the highlands terrain is being eroded back. Note how the floor of the main channel appears to be at the same level as the lowland terrain, suggestive of a base level where erosion is no longer effective. Most of the steep slope faces in the image display darker slope streaks that are thought to be dust avalanche scars and indicate that a relatively thick mantle of dust is present in this region. Wind-sculpted ridges known as yardangs cover many of the surfaces throughout the area as shown by images from the Mars Global Surveyor mission. Most of them are at the limit of resolution in the THEMIS image but some are evident on the floor of the main channel at the point at which a smaller side channel enters. In this location they appear to extend right up to the base of the channel wall, giving the appearance that they are emerging from underneath the thick pile of material into which the channel is eroded. This suggests a geologic history in which a preexisting landscape of eroded yardangs was covered over by a thick pile of younger material that is now eroding back down to the original level. Alternatively, it is possible that the yardangs formed more recently at the abrupt transition between the channel floor and wall. More analysis is necessary to sort out the story. The Story This channel system is named 'Mangala,' the word for Mars in Sanskrit, a language of the Hindus of India that goes back more than 4,000 years, with written literature almost as long. Great epic tales have been written in this language, and Odyssey is

  15. Enzyme-catalysed assembly of DNA hydrogel

    Science.gov (United States)

    Um, Soong Ho; Lee, Jong Bum; Park, Nokyoung; Kwon, Sang Yeon; Umbach, Christopher C.; Luo, Dan

    2006-10-01

    DNA is a remarkable polymer that can be manipulated by a large number of molecular tools including enzymes. A variety of geometric objects, periodic arrays and nanoscale devices have been constructed. Previously we synthesized dendrimer-like DNA and DNA nanobarcodes from branched DNA via ligases. Here we report the construction of a hydrogel entirely from branched DNA that are three-dimensional and can be crosslinked in nature. These DNA hydrogels were biocompatible, biodegradable, inexpensive to fabricate and easily moulded into desired shapes and sizes. The distinct difference of the DNA hydrogel to other bio-inspired hydrogels (including peptide-based, alginate-based and DNA (linear)-polyacrylamide hydrogels) is that the crosslinking is realized via efficient, ligase-mediated reactions. The advantage is that the gelling processes are achieved under physiological conditions and the encapsulations are accomplished in situ-drugs including proteins and even live mammalian cells can be encapsulated in the liquid phase eliminating the drug-loading step and also avoiding denaturing conditions. Fine tuning of these hydrogels is easily accomplished by adjusting the initial concentrations and types of branched DNA monomers, thus allowing the hydrogels to be tailored for specific applications such as controlled drug delivery, tissue engineering, 3D cell culture, cell transplant therapy and other biomedical applications.

  16. Magnetically responsive enzyme powders

    International Nuclear Information System (INIS)

    Powdered enzymes were transformed into their insoluble magnetic derivatives retaining their catalytic activity. Enzyme powders (e.g., trypsin and lipase) were suspended in various liquid media not allowing their solubilization (e.g., saturated ammonium sulfate and highly concentrated polyethylene glycol solutions, ethanol, methanol, 2-propanol) and subsequently cross-linked with glutaraldehyde. Magnetic modification was successfully performed at low temperature in a freezer (−20 °C) using magnetic iron oxides nano- and microparticles prepared by microwave-assisted synthesis from ferrous sulfate. Magnetized cross-linked enzyme powders were stable at least for two months in water suspension without leakage of fixed magnetic particles. Operational stability of magnetically responsive enzymes during eight repeated reaction cycles was generally without loss of enzyme activity. Separation of magnetically modified cross-linked powdered enzymes from reaction mixtures was significantly simplified due to their magnetic properties. - Highlights: • Cross-linked enzyme powders were prepared in various liquid media. • Insoluble enzymes were magnetized using iron oxides particles. • Magnetic iron oxides particles were prepared by microwave-assisted synthesis. • Magnetic modification was performed under low (freezing) temperature. • Cross-linked powdered trypsin and lipase can be used repeatedly for reaction

  17. HYDRATION AND ENZYME ACTIVITY

    OpenAIRE

    Poole, P.

    1984-01-01

    Hydration induced conformation and dynamic changes are followed using a variety of experimental techniques applied to hen egg white lysozyme. These changes are completed just before the onset of enzyme activity, which occurs before all polar groups are hydrated, and before monolayer coverage is attained. We suggest that these hydration induced changes are necessary for the return of enzyme activity.

  18. Directed Evolution of Enzymes

    OpenAIRE

    Doucet, Nicolas; Pelletier, Joelle,

    2004-01-01

    This brief technological report presents an overview of techniques and applications in the field of directed evolution of enzyme catalysts. These techniques allow for the creation of modified enzymes that are better adapted to many industrial contexts. Recent applications in organic synthesis as well as commercial, biomedical, and environmental usage of these modified catalysts will be presented.

  19. Magnetically responsive enzyme powders

    Energy Technology Data Exchange (ETDEWEB)

    Pospiskova, Kristyna, E-mail: kristyna.pospiskova@upol.cz [Regional Centre of Advanced Technologies and Materials, Palacky University, Slechtitelu 11, 783 71 Olomouc (Czech Republic); Safarik, Ivo, E-mail: ivosaf@yahoo.com [Regional Centre of Advanced Technologies and Materials, Palacky University, Slechtitelu 11, 783 71 Olomouc (Czech Republic); Department of Nanobiotechnology, Institute of Nanobiology and Structural Biology of GCRC, Na Sadkach 7, 370 05 Ceske Budejovice (Czech Republic)

    2015-04-15

    Powdered enzymes were transformed into their insoluble magnetic derivatives retaining their catalytic activity. Enzyme powders (e.g., trypsin and lipase) were suspended in various liquid media not allowing their solubilization (e.g., saturated ammonium sulfate and highly concentrated polyethylene glycol solutions, ethanol, methanol, 2-propanol) and subsequently cross-linked with glutaraldehyde. Magnetic modification was successfully performed at low temperature in a freezer (−20 °C) using magnetic iron oxides nano- and microparticles prepared by microwave-assisted synthesis from ferrous sulfate. Magnetized cross-linked enzyme powders were stable at least for two months in water suspension without leakage of fixed magnetic particles. Operational stability of magnetically responsive enzymes during eight repeated reaction cycles was generally without loss of enzyme activity. Separation of magnetically modified cross-linked powdered enzymes from reaction mixtures was significantly simplified due to their magnetic properties. - Highlights: • Cross-linked enzyme powders were prepared in various liquid media. • Insoluble enzymes were magnetized using iron oxides particles. • Magnetic iron oxides particles were prepared by microwave-assisted synthesis. • Magnetic modification was performed under low (freezing) temperature. • Cross-linked powdered trypsin and lipase can be used repeatedly for reaction.

  20. Development of 3-(4-aminosulphonyl)-phenyl-2-mercapto-3H-quinazolin-4-ones as inhibitors of carbonic anhydrase isoforms involved in tumorigenesis and glaucoma.

    Science.gov (United States)

    Alafeefy, Ahmed M; Carta, Fabrizio; Ceruso, Mariangela; Al-Tamimi, Abdul-Malek S; Al-Kahtani, Abdulla A; Supuran, Claudiu T

    2016-03-15

    A series of heterocyclic benzenesulfonamides incorporating 2-mercapto-3H-quinazolin-4-one tails were prepared by condensation of substituted anthranilic acids with 4-isothiocyanato-benzenesulfonamide. These sulfonamides were investigated as inhibitors of the human carbonic anhydrase (hCA, EC 4.2.1.1) isoforms hCA I and II (cytosolic isozymes), as well as hCA IX and XII (trans-membrane, tumor-associated enzymes). They acted as medium potency inhibitors of hCA I (KIs of 81.0-3084 nM), being highly effective as hCA II (KIs in the range of 0.25-10.8 nM), IX (KIs of 3.7-50.4 nM) and XII (KIs of 0.60-52.9 nM) inhibitors. These compounds should thus be of interest as preclinical candidates in pathologies in which the activity of these enzymes should be inhibited, such as glaucoma (CA II and XII as targets) or some tumors in which the activity of three isoforms (CA II, IX and XII) is dysregulated. PMID:26875933

  1. Implication of cytochrome P-450 1A isoforms and the AH receptor in the genotoxicity of coal-tar fume condensate and bitumen fume condensates.

    Science.gov (United States)

    Genevois, C; Pfohl-Leszkowicz, A; Boillot, K; Brandt, H; Castegnaro, M

    1998-06-01

    During the hot application of bitumen- or coal-tar-containing materials, fumes are emitted that contain polycyclic aromatic compounds. Although workers' exposure to these fumes is low, it might lead to health problems. No study has reported the metabolic pathways involved in the genotoxicity of coal tar or bitumen fume condensates (CTFC, BFCs). We have therefore studied the DNA adducts formed by incubation of CTFC or BFCs with liver microsomes from several type of mice and with yeast microsomes expressing individual human CYP enzymes. Our results demonstrates that: (1) the aryl hydrocarbon receptor (AHR) plays an important role in the biotransformation of BFCs and to a lesser extent of CTFC; (2) for CTFC, both cytochrome P450 (CYP) 1A isoforms are involved in the formation of genotoxic compounds, and the reactive metabolites formed via CYP 1A1, are substrates for epoxide hydrolase (mEH); (3) for BFCs, the genotoxicity is partially dependent upon CYP 1A1 and the reactive metabolites are not substrates for mEH; (4) CYP 1A isoforms are not exclusively responsible for the genotoxicity of the CTFC and BFCs as other CYPs and also enzymes of the [AH] gene battery, may play an important role. PMID:21781875

  2. Artificial Enzymes, "Chemzymes"

    DEFF Research Database (Denmark)

    Bjerre, Jeannette; Rousseau, Cyril Andre Raphaël; Pedersen, Lavinia Georgeta M;

    2008-01-01

    Enzymes have fascinated scientists since their discovery and, over some decades, one aim in organic chemistry has been the creation of molecules that mimic the active sites of enzymes and promote catalysis. Nevertheless, even today, there are relatively few examples of enzyme models that...... successfully perform Michaelis-Menten catalysis under enzymatic conditions (i.e., aqueous medium, neutral pH, ambient temperature) and for those that do, very high rate accelerations are seldomly seen. This review will provide a brief summary of the recent developments in artificial enzymes, so called...... "Chemzymes", based on cyclodextrins and other molecules. Only the chemzymes that have shown enzyme-like activity that has been quantified by different methods will be mentioned. This review will summarize the work done in the field of artificial glycosidases, oxidases, epoxidases, and esterases, as well as...

  3. Studies of the cytosolic thymidine kinase in human cells and comparison to the recombinantly expressed enzyme

    DEFF Research Database (Denmark)

    Kock Jensen, Helle

    Thymidine kinase (TK) is a key enzyme in the salvage pathway of the nucleoside metabolism catalyzing the first phosphorylation step in TTP synthesis. Human cytosolic TK (TKl) is highly cell cycle regulated. TKl is regulated on many different levels of expression and isoforms with altered enzymatic...... identical but further investigations showed some interesting differences. Recombinant TKl is about 10 fold more sensitive towards TTP as inhibitor. Furthermore the effect of removal of ATP from the native TKl on the enzyme kinetics and native molecular weight was not found for recombinant TKl. Native TKl...

  4. VEGF-A isoforms program differential VEGFR2 signal transduction, trafficking and proteolysis

    Directory of Open Access Journals (Sweden)

    Gareth W. Fearnley

    2016-05-01

    Full Text Available Vascular endothelial growth factor A (VEGF-A binding to the receptor tyrosine kinase VEGFR2 triggers multiple signal transduction pathways, which regulate endothelial cell responses that control vascular development. Multiple isoforms of VEGF-A can elicit differential signal transduction and endothelial responses. However, it is unclear how such cellular responses are controlled by isoform-specific VEGF-A–VEGFR2 complexes. Increasingly, there is the realization that the membrane trafficking of receptor–ligand complexes influences signal transduction and protein turnover. By building on these concepts, our study shows for the first time that three different VEGF-A isoforms (VEGF-A165, VEGF-A121 and VEGF-A145 promote distinct patterns of VEGFR2 endocytosis for delivery into early endosomes. This differential VEGFR2 endocytosis and trafficking is linked to VEGF-A isoform-specific signal transduction events. Disruption of clathrin-dependent endocytosis blocked VEGF-A isoform-specific VEGFR2 activation, signal transduction and caused substantial depletion in membrane-bound VEGFR1 and VEGFR2 levels. Furthermore, such VEGF-A isoforms promoted differential patterns of VEGFR2 ubiquitylation, proteolysis and terminal degradation. Our study now provides novel insights into how different VEGF-A isoforms can bind the same receptor tyrosine kinase and elicit diverse cellular outcomes.

  5. Androgen receptor isoforms in human prostatic cancer tissue and LNCaP cell line

    Institute of Scientific and Technical Information of China (English)

    Shu-Jie XIA; Xiao-Da TANG; Qing-Zheng MA

    2001-01-01

    Aim: To investigate the androgen receptor (AR) isoform expressions in human prostatic cancer tissue and LNCaP cell line. Methods: With high resolution isoelectric focusing (IEF) method we demonstrated the different expressions of AR isoforms in human prostatic cancer tissues and LNCaP cell line. Results: Data were obtained from three prostatic cancer specimens and the LNCaP cell line. Three types of AR isoforms were detected with pI values at 6.5,6.0, and 5.3. For the 3 prostatic cancer specimens, 1 sample showed all the three types of AR isoforms, the second specimen expressed at 6.5 and 6.0, and the third failed to show any type of isoforms. The LNCaP cell line expressed all the three AR isoforms. Binding of 3H-dihydrotestosterone (3H-DHT) to these three isoforms was inhibited by the addition ofl00-fold excess of DHT or testosterone, while not by progesterone, oestradiol and diethylstilboestrol. Conclusion: The expression of AR isofonns is different in different prostate cancer tissues, which may be related to the difference in the effect of anti-androgen therapy in different patients.

  6. Dysregulation of miRNA isoform level at 5' end in Alzheimer's disease.

    Science.gov (United States)

    Wang, Shengqin; Xu, Yuming; Li, Musheng; Tu, Jing; Lu, Zuhong

    2016-06-15

    Alzheimer's disease (AD) is the most common form of dementia, whose mechanism is still not yet fully understood. A miRNA-based signature method, commonly according to the changes of expression levels, is widely used for AD analysis in previous studies. Recently, miRNA isoforms called as isomiR variants, which is considered to play important biological roles, have been demonstrated as the applications of high throughput sequencing platforms. Here, we presented an entropy-based model to detect the miRNA isoform level at the 5' end, and found many miRNAs with significant changes of isoform levels between the early stage and the late stage of AD by the application of this model to the public data. The statistical significance of the overlap between isoform-level changed miRNAs and AD related miRNAs extracted from HMDD2 supports that these miRNA isoforms are not degradation products. Based on the most common isomiR seed analysis of isoform-level changed AD related miRNAs, the predicted targets are also found to be enriched for genes involved in transcriptional regulation and the nervous system. After comparing with the expression level based method, we detected that changes of 5' isoform levels are more stable than those of expression levels for AD related miRNA detecting. PMID:26899870

  7. The isolation of parvalbumin isoforms from the tail muscle of the American alligator (Alligator mississipiensis).

    Science.gov (United States)

    Laney, E L; Shabanowitz, J; King, G; Hunt, D F; Nelson, D J

    1997-04-01

    Multiple parvalbumin isoforms have been detected in the tail (skeletal) muscle of the American alligator (Alligator mississipiensis). One of these isoforms (APV-1) has been highly purified and partially characterized. Protein purification involved mainly gel filtration and anion exchange chromatography, and characterization included gel electrophoresis, amino acid composition analysis, metal ion analysis, MALDI-TOF and ESI mass spectrometry, ultraviolet and fluorescence spectroscopy, and one- and two-dimensional 500 MHz proton NMR spectroscopy. The alligator isoforms are rich in phenylalanine and deficient in the other aromatic residues as is typical for parvalbumins. In fact, the one highly purified isoform that forms the basis of this study has only phenyl-alanine as an aromatic residue. Ion exchange chromatography further indicates that this isoform has a relatively high isoelectric point (pl approximately 5.0), indicating that it is an alpha-lineage parvalbumin. This alligator parvalbumin isoform is unusual in that it has an atypically high Ca2+ content (almost 3.0 mole of Ca2+ per mole of protein) following purification, a fact supported by terbium fluorescence titration experiments. Preliminary comparative analysis of the highly purified alligator parvalbumin isoform (in the Ca2-loaded state) by two-dimensional 1H-NMR (2D 1H TOCSY and 2D 1H NOESY) indicates that there is considerable similarity in structure between the alligator protein and a homologous protein obtained from the silver hake (a saltwater fish species). PMID:9076974

  8. Linear and Branched PEIs (Polyethylenimines) and Their Property Space

    Science.gov (United States)

    Lungu, Claudiu N.; Diudea, Mircea V.; Putz, Mihai V.; Grudziński, Ireneusz P.

    2016-01-01

    A chemical property space defines the adaptability of a molecule to changing conditions and its interaction with other molecular systems determining a pharmacological response. Within a congeneric molecular series (compounds with the same derivatization algorithm and thus the same brute formula) the chemical properties vary in a monotonic manner, i.e., congeneric compounds share the same chemical property space. The chemical property space is a key component in molecular design, where some building blocks are functionalized, i.e., derivatized, and eventually self-assembled in more complex systems, such as enzyme-ligand systems, of which (physico-chemical) properties/bioactivity may be predicted by QSPR/QSAR (quantitative structure-property/activity relationship) studies. The system structure is determined by the binding type (temporal/permanent; electrostatic/covalent) and is reflected in its local electronic (and/or magnetic) properties. Such nano-systems play the role of molecular devices, important in nano-medicine. In the present article, the behavior of polyethylenimine (PEI) macromolecules (linear LPEI and branched BPEI, respectively) with respect to the glucose oxidase enzyme GOx is described in terms of their (interacting) energy, geometry and topology, in an attempt to find the best shape and size of PEIs to be useful for a chosen (nanochemistry) purpose. PMID:27089324

  9. Isoform-specific upregulation of palladin in human and murine pancreas tumors.

    Directory of Open Access Journals (Sweden)

    Silvia M Goicoechea

    Full Text Available Pancreatic ductal adenocarcinoma (PDA is a lethal disease with a characteristic pattern of early metastasis, which is driving a search for biomarkers that can be used to detect the cancer at an early stage. Recently, the actin-associated protein palladin was identified as a candidate biomarker when it was shown that palladin is mutated in a rare inherited form of PDA, and overexpressed in many sporadic pancreas tumors and premalignant precursors. In this study, we analyzed the expression of palladin isoforms in murine and human PDA and explored palladin's potential use in diagnosing PDA. We performed immunohistochemistry and immunoblot analyses on patient samples and tumor-derived cells using an isoform-selective monoclonal antibody and a pan-palladin polyclonal antibody. Immunoblot and real-time quantitative reverse transcription-PCR were used to quantify palladin mRNA levels in human samples. We show that there are two major palladin isoforms expressed in pancreas: 65 and 85-90 kDa. The 65 kDa isoform is expressed in both normal and neoplastic ductal epithelial cells. The 85-90 kDa palladin isoform is highly overexpressed in tumor-associated fibroblasts (TAFs in both primary and metastatic tumors compared to normal pancreas, in samples obtained from either human patients or genetically engineered mice. In tumor-derived cultured cells, expression of palladin isoforms follows cell-type specific patterns, with the 85-90 kDa isoform in TAFs, and the 65 kDa isoform predominating in normal and neoplastic epithelial cells. These results suggest that upregulation of 85-90 kDa palladin isoform may play a role in the establishment of the TAF phenotype, and thus in the formation of a desmoplastic tumor microenvironment. Thus, palladin may have a potential use in the early diagnosis of PDA and may have much broader significance in understanding metastatic behavior.

  10. Learning-dependent gene expression of CREB1 isoforms in the molluscan brain

    Directory of Open Access Journals (Sweden)

    Hisayo Sadamoto

    2010-05-01

    Full Text Available Cyclic AMP-responsive element binding protein1 (CREB1 has multiple functions in gene regulation. Various studies have reported that CREB1-dependent gene induction is necessary for memory formation and long-lasting behavioral changes in both vertebrates and invertebrates. In the present study, we characterized Lymnaea CREB1 (LymCREB1 mRNA isoforms of spliced variants in the central nervous system (CNS of the pond snail Lymnaea stagnalis. Among these spliced variants, the three isoforms that code a whole LymCREB1 protein are considered to be the activators for gene regulation. The other four isoforms, which code truncated LymCREB1 proteins with no kinase inducible domain, are the repressors. For a better understanding of the possible roles of different LymCREB1 isoforms, the expression level of these isoform mRNAs was investigated by a real-time quantitative RT-PCR method. Further, we examined the changes in gene expression for all the isoforms in the CNS after conditioned taste aversion (CTA learning or backward conditioning as a control. The results showed that CTA learning increased LymCREB1 gene expression, but it did not change the activator/repressor ratio. Our findings showed that the repressor isoforms, as well as the activator ones, are expressed in large amounts in the CNS, and the gene expression of CREB1 isoforms appeared to be specific for the given stimulus. This was the first quantitative analysis of the expression patterns of CREB1 isoforms at the mRNA level and their association with learning behavior.

  11. Myosin isoform fiber type and fiber size in the tail of the Virginia opossum (Didelphis virginiana).

    Science.gov (United States)

    Hazimihalis, P J; Gorvet, M A; Butcher, M T

    2013-01-01

    Muscle fiber type is a well studied property in limb muscles, however, much less is understood about myosin heavy chain (MHC) isoform expression in caudal muscles of mammalian tails. Didelphid marsupials are an interesting lineage in this context as all species have prehensile tails, but show a range of tail-function depending on either their arboreal or terrestrial locomotor habits. Differences in prehensility suggest that MHC isoform fiber types may also be different, in that terrestrial opossums may have a large distribution of oxidative fibers for object carrying tasks instead of faster, glycolytic fiber types expected in mammals with long tails. To test this hypothesis, MHC isoform fiber type and their regional distribution (proximal/transitional/distal) were determined in the tail of the Virginia opossum (Didelphis virginiana). Fiber types were determined by a combination of myosin-ATPase histochemistry, immunohistochemistry, and SDS-PAGE. Results indicate a predominance of the fast MHC-2A and -2X isoforms in each region of the tail. The presence of two fast isoforms, in addition to the slow MHC-1 isoform, was confirmed by SDS-PAGE analysis. The overall MHC isoform fiber type distribution for the tail was: 25% MHC-1, 71% MHC-2A/X hybrid, and 4% MHC-1/2A hybrid. Oxidative MHC-2A/X isoform fibers were found to be relatively large in cross-section compared to slow, oxidative MHC-1 and MHC-1/2A hybrid fibers. A large percentage of fast MHC-2A/X hybrids fibers may be suggestive of an evolutionary transition in MHC isoform distribution (fast-to-slow fiber type) in the tail musculature of an opossum with primarily a terrestrial locomotor habit and adaptive tail-function. PMID:23152195

  12. PML isoforms in response to arsenic: high-resolution analysis of PML body structure and degradation.

    Science.gov (United States)

    Hands, Katherine J; Cuchet-Lourenco, Delphine; Everett, Roger D; Hay, Ronald T

    2014-01-15

    Arsenic is a clinically effective treatment for acute promyelocytic leukaemia (APL) in which the promyelocytic leukaemia (PML) protein is fused to retinoic receptor alpha (RARα). PML-RARα is degraded by the proteasome by a SUMO-dependent, ubiquitin-mediated pathway in response to arsenic treatment, curing the disease. Six major PML isoforms are expressed as a result of alternative splicing, each of which encodes a unique C-terminal region. Using a system in which only a single EYFP-linked PML isoform is expressed, we demonstrate that PMLI, PMLII and PMLVI accumulate in the cytoplasm following arsenic treatment, whereas PMLIII, PMLIV and PMLV do not. 3D structured illumination was used to obtain super-resolution images of PML bodies, revealing spherical shells of PML along with associated SUMO. Arsenic treatment results in dramatic isoform-specific changes to PML body ultrastructure. After extended arsenic treatment most PML isoforms are degraded, leaving SUMO at the core of the nuclear bodies. A high-content imaging assay identifies PMLV as the isoform most readily degraded following arsenic treatment, and PMLIV as relatively resistant to degradation. Immunoprecipitation analysis demonstrates that all PML isoforms are modified by SUMO and ubiquitin after arsenic treatment, and by using siRNA, we demonstrate that arsenic-induced degradation of all PML isoforms is dependent on the ubiquitin E3 ligase RNF4. Intriguingly, depletion of RNF4 results in marked accumulation of PMLV, suggesting that this isoform is an optimal substrate for RNF4. Thus the variable C-terminal domain influences the rate and location of degradation of PML isoforms following arsenic treatment. PMID:24190887

  13. Airway branching morphogenesis in three dimensional culture

    Directory of Open Access Journals (Sweden)

    Gudjonsson Thorarinn

    2010-11-01

    Full Text Available Abstract Background Lungs develop from the fetal digestive tract where epithelium invades the vascular rich stroma in a process called branching morphogenesis. In organogenesis, endothelial cells have been shown to be important for morphogenesis and the maintenance of organ structure. The aim of this study was to recapitulate human lung morphogenesis in vitro by establishing a three dimensional (3D co-culture model where lung epithelial cells were cultured in endothelial-rich stroma. Methods We used a human bronchial epithelial cell line (VA10 recently developed in our laboratory. This cell line cell line maintains a predominant basal cell phenotype, expressing p63 and other basal markers such as cytokeratin-5 and -14. Here, we cultured VA10 with human umbilical vein endothelial cells (HUVECs, to mimic the close interaction between these cell types during lung development. Morphogenesis and differentiation was monitored by phase contrast microscopy, immunostainings and confocal imaging. Results We found that in co-culture with endothelial cells, the VA10 cells generated bronchioalveolar like structures, suggesting that lung epithelial branching is facilitated by the presence of endothelial cells. The VA10 derived epithelial structures display various complex patterns of branching and show partial alveolar type-II differentiation with pro-Surfactant-C expression. The epithelial origin of the branching VA10 colonies was confirmed by immunostaining. These bronchioalveolar-like structures were polarized with respect to integrin expression at the cell-matrix interface. The endothelial-induced branching was mediated by soluble factors. Furthermore, fibroblast growth factor receptor-2 (FGFR-2 and sprouty-2 were expressed at the growing tips of the branching structures and the branching was inhibited by the FGFR-small molecule inhibitor SU5402. Discussion In this study we show that a human lung epithelial cell line can be induced by endothelial cells to

  14. Photocontrol of nitric oxide production in cell culture using a caged isoform selective inhibitor.

    Science.gov (United States)

    Perdicakis, Basil; Montgomery, Heather J; Abbott, Glenn L; Fishlock, Dan; Lajoie, Gilles A; Guillemette, J Guy; Jervis, Eric

    2005-01-01

    Over the past decade, multiphoton microscopy has progressed from a photonic novelty to a technique whose application is currently experiencing exponential growth in the biological sciences. A novel application of this technology with significant therapeutic potential is the control of drug activity by multiphoton photolysis of caged therapeutics. As an initial case study, the potent isoform selective inhibitor N-(3-(aminomethyl)benzyl) acetamidine (1400W) of inducible nitric oxide synthase (iNOS) has been conjugated to a caging molecule 6-bromo-7-hydroxy-4-hydroxyquinoline-2-ylmethyl acetyl ester (Bhc). Here we present the first report of a bulk therapeutic effect, inhibition of nitric oxide production, in mammalian cell culture by multiphoton photolysis of a caged drug, Bhc-1400W. Mouse macrophage RAW 264.7 cells induced with bacterial lipopolysaccharides to express iNOS were used to assess the therapeutic value of the conjugated inhibitor. Both 1400W and Bhc-1400W are stable in metabolically active cells and an optimal time interval for the photorelease of the inhibitor was determined. The ratios of the IC(50) values of Bhc-1400W over 1400W calculated in the presence of iNOS enzyme and in RAW 264.7 cell culture are 19 and 100, respectively, indicating that a broad therapeutic range exists in cell culture. Multiphoton uncaging protocols and therapeutic doses of inhibitors were not cytotoxic. Photocontrol of LPS induced nitric oxide production was achieved in mammalian cell culture using a single laser focal volume. This technology has the potential to control active drug concentrations in vivo, a lack of which is one of the main problems currently associated with systemic drug administration. PMID:15582451

  15. Detergents profoundly affect inhibitor potencies against both cyclo-oxygenase isoforms.

    Science.gov (United States)

    Ouellet, Marc; Falgueyret, Jean-Pierre; Percival, M David

    2004-02-01

    The sensitivity of Coxs (cyclo-oxygenases) to inhibition is known to be highly dependent on assay conditions. In the present study, the inhibitor sensitivities of purified Cox-1 and -2 were determined in a colorimetric assay using the reducing agent N, N, N ', N '-tetramethyl- p -phenylenediamine. With the detergent genapol X-100 (2 mM) present, the potencies of nimesulide, ibuprofen, flufenamic acid, niflumic acid and naproxen were increased over 100-fold against Cox-2 and titration curve shapes changed, so that maximal inhibition now approached 100%. Indomethacin, diclofenac and flosulide were not changed in potency. Similar effects of genapol were observed with inhibitors of Cox-1. DuP-697 and two analogues became more than 10-fold less potent against Cox-2 with genapol present. Tween-20, Triton X-100 and phosphatidylcholine, but not octylglucoside, gave qualitatively similar effects as genapol. Similar detergent-dependent changes in inhibitor potency were also observed using a [(14)C]arachidonic acid HPLC assay. The increases in potency of ibuprofen, flufenamic acid, isoxicam and niflumic acid towards Cox-2 and ibuprofen towards Cox-1 were accompanied by a change from time-independent to time-dependent inhibition. The interactions of Cox inhibitors has been described in terms of multiple binding step mechanisms. The genapol-dependent increase in inhibitor potency for ketoprofen was associated with an increase in the rate constant for the conversion of the initial enzyme-inhibitor complex to a second, more tightly bound form. The loss of potency for some inhibitors is probably due to inhibitor partitioning into detergent micelles. The present study identifies detergents as another factor that must be considered when determining inhibitor potencies against both Cox isoforms. PMID:14510637

  16. Metabolic Reprogramming by Hexosamine Biosynthetic and Golgi N-Glycan Branching Pathways.

    Science.gov (United States)

    Ryczko, Michael C; Pawling, Judy; Chen, Rui; Abdel Rahman, Anas M; Yau, Kevin; Copeland, Julia K; Zhang, Cunjie; Surendra, Anu; Guttman, David S; Figeys, Daniel; Dennis, James W

    2016-01-01

    De novo uridine-diphosphate-N-acetylglucosamine (UDP-GlcNAc) biosynthesis requires glucose, glutamine, acetyl-CoA and uridine, however GlcNAc salvaged from glycoconjugate turnover and dietary sources also makes a significant contribution to the intracellular pool. Herein we ask whether dietary GlcNAc regulates nutrient transport and intermediate metabolism in C57BL/6 mice by increasing UDP-GlcNAc and in turn Golgi N-glycan branching. GlcNAc added to the drinking water showed a dose-dependent increase in growth of young mice, while in mature adult mice fat and body-weight increased without affecting calorie-intake, activity, energy expenditure, or the microbiome. Oral GlcNAc increased hepatic UDP-GlcNAc and N-glycan branching on hepatic glycoproteins. Glucose homeostasis, hepatic glycogen, lipid metabolism and response to fasting were altered with GlcNAc treatment. In cultured cells GlcNAc enhanced uptake of glucose, glutamine and fatty-acids, and enhanced lipid synthesis, while inhibition of Golgi N-glycan branching blocked GlcNAc-dependent lipid accumulation. The N-acetylglucosaminyltransferase enzymes of the N-glycan branching pathway (Mgat1,2,4,5) display multistep ultrasensitivity to UDP-GlcNAc, as well as branching-dependent compensation. Indeed, oral GlcNAc rescued fat accumulation in lean Mgat5(-/-) mice and in cultured Mgat5(-/-) hepatocytes, consistent with N-glycan branching compensation. Our results suggest GlcNAc reprograms cellular metabolism by enhancing nutrient uptake and lipid storage through the UDP-GlcNAc supply to N-glycan branching pathway. PMID:26972830

  17. The Future of Washington's Branch Campuses. HECB Report on Branch Campus Development Plans-HB 2707

    Science.gov (United States)

    Washington Higher Education Coordinating Board, 2005

    2005-01-01

    Washington's research university branch campuses were created in 1989 to increase opportunities for students in several regions of the state to complete their baccalaureate and graduate-level studies at public universities close to their homes. Currently, the University of Washington operates branch campuses in Bothell and Tacoma. The Bothell…

  18. Measurement of tau lepton branching fractions

    International Nuclear Information System (INIS)

    We present τ- lepton branching fraction measurements based on data from the TPC/Two-Gamma detector at PEP. Using a sample ofτ- → ντK-π+π- events, we examine the resonance structure of the K-π+π- system and obtain the first measurements of branching fractions for τ- → ντK1-(1270) and τ- → ντK1-(1400). We also describe a complete set of branching fraction measurements in which all the decays of the τ- lepton are separated into classes defined by the identities of the charged particles and an estimate of the number of neutrals. This is the first such global measurement with decay classes defined by the four possible charged particle species, e, μ, π, and K

  19. Geometry optimization of branchings in vascular networks

    Science.gov (United States)

    Khamassi, Jamel; Bierwisch, Claas; Pelz, Peter

    2016-06-01

    Progress has been made in developing manufacturing technologies which enable the fabrication of artificial vascular networks for tissue cultivation. However, those networks are rudimentary designed with respect to their geometry. This restricts long-term biological functionality of vascular cells which depends on geometry-related fluid mechanical stimuli and the avoidance of vessel occlusion. In the present work, a bioinspired geometry optimization for branchings in artificial vascular networks has been conducted. The analysis could be simplified by exploiting self-similarity properties of the system. Design rules in the form of two geometrical parameters, i.e., the branching angle and the radius ratio of the daughter branches, are derived using the wall shear stress as command variable. The numerical values of these parameters are within the range of experimental observations. Those design rules are not only beneficial for tissue engineering applications. Moreover, they can be used as indicators for diagnoses of vascular diseases or for the layout of vascular grafts.

  20. Branched chain amino acid transaminase and branched chain alpha-ketoacid dehydrogenase activity in the brain, liver and skele­tal muscle of acute hepatic failure rats

    Directory of Open Access Journals (Sweden)

    Takei,Nobuyuki

    1985-02-01

    Full Text Available Branched chain amino acid (BCAA transaminase activity increased in both the mitochondrial and supernatant fractions of brain from hepatic failure rats, in which a partial hepatectomy was performed 24h following carbon tetrachloride (CCl4 administration, although the activity of liver and skeletal muscle was the same as in control rats. The elevation of mitochondrial BCAA transaminase activity in liver-injured rats was partly due to increased activity of brain specific Type III isozyme. Branched chain alpha-ketoacid (BCKA dehydrogenase in the brain homogenates was not significantly altered in acute hepatic failure rats, while the liver enzyme activity was markedly diminished. BCKA dehydrogenase activity in the brain homogenates was inhibited by adding ATP to the assay system, and was activated in vitro by preincubating the brain homogenate at 37 degrees C for 15 min. These findings suggest that brain BCAA catabolism is accelerated in acute hepatic failure rats.

  1. Starch modification with microbial alpha-glucanotransferase enzymes

    OpenAIRE

    van der Maarel, Marc J. E. C.; Leemhuis, Hans

    2013-01-01

    Starch is an agricultural raw material used in many food and industrial products. It is present in granules that vary in shape in the form of amylose and amylopectin. Starch-degrading enzymes are used on a large scale in the production of sweeteners (high fructose corn syrup) and concentrated glucose syrups as substrate for the fermentative production of bioethanol and basic chemicals. Over the last two decades alpha-glucanotransferases (EC 2.4.1.xx), such as branching enzyme (EC 2.4.1.18) an...

  2. Enzymes in Analytical Chemistry.

    Science.gov (United States)

    Fishman, Myer M.

    1980-01-01

    Presents tabular information concerning recent research in the field of enzymes in analytic chemistry, with methods, substrate or reaction catalyzed, assay, comments and references listed. The table refers to 128 references. Also listed are 13 general citations. (CS)

  3. Membrane Assisted Enzyme Fractionation

    DEFF Research Database (Denmark)

    Yuan, Linfeng

    difference. In this thesis, separations using crossflow elecro-membrane filtration (EMF) of amino acids, bovine serum albumin (BSA) and industrial enzymes from Novozymes were performed. The main objective of this study was to investigate the technological feasibility of EMF in the application of industrial...... enzyme fractionation, such as removal of a side activity from the main enzyme activity. As a proof-of-concept, amino acids were used as model solution to test the feasibility of EMF in the application of amphoteric molecule separation. A single amino acid was used to illustrate the effect of an electric...... TMP on the separation performance were very small in the investigated range. The mass transport of each enzyme can be well explained by the Extended-Nernst-Planck equation. Better separation was observed at lower feed concentration, higher solution pH in the investigated range and with a polysulfone...

  4. Effects of prolonged recombinant human erythropoietin administration on muscle membrane transport systems and metabolic marker enzymes

    DEFF Research Database (Denmark)

    Juel, C; Thomsen, J J; Rentsch, R L;

    2007-01-01

    the expression of muscle membrane transport proteins. Likewise, improvements in performance may involve upregulation of metabolic enzymes. Since Epo is known to augment performance we tested the effect of rHuEpo on some marker enzymes that are related to aerobic capacity. For these purposes eight...... performance by approximately 54%. Membrane transport systems and carbonic anhydrases involved in pH regulation remained unchanged. Of the Na(+), K(+)-pump isoforms only the density of the alpha2 subunit was decreased (by 22%) after treatment. The marker enzymes cytochrom c and hexokinase remained unchanged...... with the treatment. In conclusion, changes in muscle membrane transport proteins and selected muscle enzymes do not contribute to the Epo-induced improvement in performance....

  5. Enzyme with rhamnogalacturonase activity.

    OpenAIRE

    Kofod, L.V.; Andersen, L N; Dalboge, H; Kauppinen, M.S.; Christgau, S; Heldt-Hansen, H.P.; Christophersen, C.; Nielsen, P.M.; Voragen, A. G. J.; Schols, H.A.

    1998-01-01

    An enzyme exhibiting rhamnogalacturonase activity, capable of cleaving a rhamnogalacturonan backbone in such a manner that galacturonic acids are left as the non-reducing ends, and which exhibits activity on hairy regions from a soy bean material and/or on saponified hairy regions from a sugar beet material. The enzyme has the amino acid sequence of SEQ ID NO:2 and is encoded by the DNA sequence of SEQ ID NO:1

  6. Overproduction of ligninolytic enzymes

    Energy Technology Data Exchange (ETDEWEB)

    Elisashvili, Vladimir; Kachlishvili, Eva; Torok, Tamas

    2014-06-17

    Methods, compositions, and systems for overproducing ligninolytic enzymes from the basidiomycetous fungus are described herein. As described, the method can include incubating a fungal strain of Cerrena unicolor IBB 303 in a fermentation system having growth medium which includes lignocellulosic material and then cultivating the fungal strain in the fermentation system under conditions wherein the fungus expresses the ligninolytic enzymes. In some cases, the lignocellulosic material is mandarin peel, ethanol production residue, walnut pericarp, wheat bran, wheat straw, or banana peel.

  7. A Synaptotagmin Isoform Switch during the Development of an Identified CNS Synapse.

    Science.gov (United States)

    Kochubey, Olexiy; Babai, Norbert; Schneggenburger, Ralf

    2016-06-01

    Various Synaptotagmin (Syt) isoform genes are found in mammals, but it is unknown whether Syts can function redundantly in a given nerve terminal, or whether isoforms can be switched during the development of a nerve terminal. Here, we investigated the possibility of a developmental Syt isoform switch using the calyx of Held as a model synapse. At mature calyx synapses, fast Ca(2+)-driven transmitter release depended entirely on Syt2, but the release phenotype of Syt2 knockout (KO) mice was weaker at immature calyces, and absent at pre-calyceal synapses early postnatally. Instead, conditional genetic inactivation shows that Syt1 mediates fast release at pre-calyceal synapses, as well as a fast release component resistant to Syt2 deletion in immature calyces. This demonstrates a developmental Syt1-Syt2 isoform switch at an identified synapse, a mechanism that could fine-tune the speed, reliability, and plasticity of transmitter release at fast releasing CNS synapses. PMID:27210552

  8. Isoform-selective regulation of glycogen phosphorylase by energy deprivation and phosphorylation in astrocytes

    DEFF Research Database (Denmark)

    Müller, Margit S; Pedersen, Sofie E; Walls, Anne B;

    2015-01-01

    Glycogen phosphorylase (GP) is activated to degrade glycogen in response to different stimuli, to support both the astrocyte's own metabolic demand and the metabolic needs of neurons. The regulatory mechanism allowing such a glycogenolytic response to distinct triggers remains incompletely...... understood. In the present study, we used siRNA-mediated differential knockdown of the two isoforms of GP expressed in astrocytes, muscle isoform (GPMM), and brain isoform (GPBB), to analyze isoform-specific regulatory characteristics in a cellular setting. Subsequently, we tested the response of each...... determination of glycogen content showing an increase in glycogen levels following knockdown of either GPMM or GPBB. NE triggered glycogenolysis within 15 min in control cells and after GPBB knockdown. However, astrocytes in which expression of GPMM had been silenced showed a delay in response to NE, with...

  9. Branch and Bound Experiments in Convex Nonlinear Integer Programming

    OpenAIRE

    Omprakash K. Gupta; Ravindran, A

    1985-01-01

    The branch and bound principle has long been established as an effective computational tool for solving mixed integer linear programming problems. This paper investigates the computational feasibility of branch and bound methods in solving convex nonlinear integer programming problems. The efficiency of a branch and bound method often depends on the rules used for selecting the branching variables and branching nodes. Among others, the concepts of pseudo-costs and estimations are implemented ...

  10. Effect of left bundle branch block on TIMI frame count

    OpenAIRE

    Hatice Tolunay; Ahmet Kasapkara; İsa Öner Yüksel; Nurcan Başar; Ayşe Saatcı Yaşar; Mehmet Bilge

    2010-01-01

    Aim: Left bundle branch block is an independent risk factorfor cardiac mortality. In this study we aimed to evaluatecoronary blood flow with TIMI frame count in patients with left bundle branch block and angiographically proven normal coronary arteries.Materials and methods: We retrospectively studied 17 patients with left bundle branch block and as a control group 16 patients without left bundle branch block. All patientshad angiographically proven normal coronary arteries.Left bundle branch...

  11. Network-Based Isoform Quantification with RNA-Seq Data for Cancer Transcriptome Analysis.

    Science.gov (United States)

    Zhang, Wei; Chang, Jae-Woong; Lin, Lilong; Minn, Kay; Wu, Baolin; Chien, Jeremy; Yong, Jeongsik; Zheng, Hui; Kuang, Rui

    2015-12-01

    High-throughput mRNA sequencing (RNA-Seq) is widely used for transcript quantification of gene isoforms. Since RNA-Seq data alone is often not sufficient to accurately identify the read origins from the isoforms for quantification, we propose to explore protein domain-domain interactions as prior knowledge for integrative analysis with RNA-Seq data. We introduce a Network-based method for RNA-Seq-based Transcript Quantification (Net-RSTQ) to integrate protein domain-domain interaction network with short read alignments for transcript abundance estimation. Based on our observation that the abundances of the neighboring isoforms by domain-domain interactions in the network are positively correlated, Net-RSTQ models the expression of the neighboring transcripts as Dirichlet priors on the likelihood of the observed read alignments against the transcripts in one gene. The transcript abundances of all the genes are then jointly estimated with alternating optimization of multiple EM problems. In simulation Net-RSTQ effectively improved isoform transcript quantifications when isoform co-expressions correlate with their interactions. qRT-PCR results on 25 multi-isoform genes in a stem cell line, an ovarian cancer cell line, and a breast cancer cell line also showed that Net-RSTQ estimated more consistent isoform proportions with RNA-Seq data. In the experiments on the RNA-Seq data in The Cancer Genome Atlas (TCGA), the transcript abundances estimated by Net-RSTQ are more informative for patient sample classification of ovarian cancer, breast cancer and lung cancer. All experimental results collectively support that Net-RSTQ is a promising approach for isoform quantification. Net-RSTQ toolbox is available at http://compbio.cs.umn.edu/Net-RSTQ/. PMID:26699225

  12. Expression of Two Novel Alternatively Spliced COL2A1 Isoforms During Chondrocyte Differentiation

    OpenAIRE

    McAlinden, Audrey; Johnstone, Brian; Kollar, John; Kazmi, Najam; Hering, Thomas M.

    2007-01-01

    Alternative splicing of the type II procollagen gene (COL2A1) is developmentally-regulated during chondrogenesis. Type IIA procollagen (+ exon 2) is synthesized by chondroprogenitor cells while type IIB procollagen (- exon 2) is synthesized by differentiated chondrocytes. Here, we report expression of two additional alternatively spliced COL2A1 isoforms during chondrocyte differentiation of bone marrow derived mesenchymal stem cells (MSCs). One isoform, named IIC, contains only the first 34 n...

  13. Myosin heavy-chain isoforms in the flight and leg muscles of hummingbirds and zebra finches

    OpenAIRE

    Velten, Brandy P.; Welch, Kenneth C.

    2014-01-01

    Myosin heavy chain (MHC) isoform complement is intimately related to a muscle's contractile properties, yet relatively little is known about avian MHC isoforms or how they may vary with fiber type and/or the contractile properties of a muscle. The rapid shortening of muscles necessary to power flight at the high wingbeat frequencies of ruby-throated hummingbirds and zebra finches (25–60 Hz), along with the varied morphology and use of the hummingbird hindlimb, provides a unique opportunity to...

  14. Sodium pump isoform specificity for the digitalis-like factor isolated from human peritoneal dialysate.

    Science.gov (United States)

    Tao, Q F; Hollenberg, N K; Price, D A; Graves, S W

    1997-03-01

    We have isolated a labile, specific sodium pump inhibitor or digitalis-like factor from the peritoneal dialysate of volume-expanded renal failure patients whose levels correlated closely with volume status and blood pressure. This study characterizes the inhibitory profile of this agent compared with that of ouabain against the three alpha-isoforms of the sodium pump. We prepared microsomal Na,K-ATPase from rat tissues representing the highest proportion of one of the alpha-isoforms. Both Northern and Western blot analyses confirmed that kidney had predominantly the alpha1-isoform, skeletal muscle the alpha2-isoform, and fetal brain the alpha3-isoform. Ouabain (5 x 10(-6) mol/L) produced little inhibition of kidney Na,K-ATPase (3.4+/-2.0%) but significant inhibition of skeletal muscle (37.2+/-3.7%, P<.001) and fetal brain (38.8+/-3.5%, P<.001) activity. In contrast, the labile digitalis-like factor, causing comparable inhibition of fetal brain Na,K-ATPase activity (33.3+/-4.7%), produced markedly greater inhibition of kidney (42.5+/-5.6%, P<.001) and moderately greater inhibition of skeletal muscle pump activity (57.7+/-6.3%, P<.05). In addition, the labile digitalis-like factor produced a marked concentration-dependent inhibition of the alpha2- and alpha3-isoforms (r=.79, P=.00005). Experiments combining the labile digitalis-like factor and ouabain confirmed that digitalis-like factor, unlike ouabain, was an effective inhibitor of all three isoforms in rat, in particular alpha2. The different pattern of isoform sensitivity displayed by the labile digitalis-like factor and ouabain further differentiates the two agents and raises some interesting possibilities about the functional implications of the endogenous factor. PMID:9052901

  15. Genomic organization and the tissue distribution of alternatively spliced isoforms of the mouse Spatial gene

    Directory of Open Access Journals (Sweden)

    Mattei Marie-Geneviève

    2004-07-01

    Full Text Available Abstract Background The stromal component of the thymic microenvironment is critical for T lymphocyte generation. Thymocyte differentiation involves a cascade of coordinated stromal genes controlling thymocyte survival, lineage commitment and selection. The "Stromal Protein Associated with Thymii And Lymph-node" (Spatial gene encodes a putative transcription factor which may be involved in T-cell development. In the testis, the Spatial gene is also expressed by round spermatids during spermatogenesis. Results The Spatial gene maps to the B3-B4 region of murine chromosome 10 corresponding to the human syntenic region 10q22.1. The mouse Spatial genomic DNA is organised into 10 exons and is alternatively spliced to generate two short isoforms (Spatial-α and -γ and two other long isoforms (Spatial-δ and -ε comprising 5 additional exons on the 3' site. Here, we report the cloning of a new short isoform, Spatial-β, which differs from other isoforms by an additional alternative exon of 69 bases. This new exon encodes an interesting proline-rich signature that could confer to the 34 kDa Spatial-β protein a particular function. By quantitative TaqMan RT-PCR, we have shown that the short isoforms are highly expressed in the thymus while the long isoforms are highly expressed in the testis. We further examined the inter-species conservation of Spatial between several mammals and identified that the protein which is rich in proline and positive amino acids, is highly conserved. Conclusions The Spatial gene generates at least five alternative spliced variants: three short isoforms (Spatial-α, -β and -γ highly expressed in the thymus and two long isoforms (Spatial-δ and -ε highly expressed in the testis. These alternative spliced variants could have a tissue specific function.

  16. Recombinant erythropoietin in humans has a prolonged effect on circulating erythropoietin isoform distribution.

    Directory of Open Access Journals (Sweden)

    Niels Jacob Aachmann-Andersen

    Full Text Available The membrane-assisted isoform immunoassay (MAIIA quantitates erythropoietin (EPO isoforms as percentages of migrated isoforms (PMI. We evaluated the effect of recombinant human EPO (rhEPO on the distribution of EPO isoforms in plasma in a randomized, placebo-controlled, double-blinded, cross-over study. 16 healthy subjects received either low-dose Epoetin beta (5000 IU on days 1, 3, 5, 7, 9, 11 and 13; high-dose Epoetin beta (30.000 IU on days 1, 2 and 3 and placebo on days 5, 7, 9, 11 and 13; or placebo on all days. PMI on days 4, 11 and 25 was determined by interaction of N-acetyl glucosamine with the glycosylation dependent desorption of EPO isoforms. At day 25, plasma-EPO in both rhEPO groups had returned to values not different from the placebo group. PMI with placebo, reflecting the endogenous EPO isoforms, averaged 82.5 (10.3 % (mean (SD. High-dose Epoetin beta decreased PMI on days 4 and 11 to 31.0 (4.2% (p<0.00001 and 45.2 (7.3% (p<0.00001. Low-dose Epoetin beta decreased PMI on days 4 and 11 to 46.0 (12.8% (p<0.00001 and 46.1 (10.4% (p<0.00001. In both rhEPO groups, PMI on day 25 was still decreased (high-dose Epoetin beta: 72.9 (19.4% (p=0.029; low-dose Epoetin beta: 73.1 (17.8% (p=0.039. In conclusion, Epoetin beta leaves a footprint in the plasma-EPO isoform pattern. MAIIA can detect changes in EPO isoform distribution up til at least three weeks after administration of Epoetin beta even though the total EPO concentration has returned to normal.

  17. Expression of mdr isoforms in mice during estrous cycle and under hormone stimulation

    OpenAIRE

    Marion Schiengold; Lavínia Schwantes; Ribeiro, Maria F; Nívia Lothhammer; Gonzalez, Tatiana P.; Jose Artur Bogo Chies; Nardi, Nance B

    2006-01-01

    The multidrug resistance (MDR) phenotype is associated with the expression of P-glycoprotein (Pgp), coded by the multigenic mdr family. Mice present the isoforms mdr1 and mdr3, which are responsible for multidrug resistance, and mdr2, that is involved in the transport of phospholipids. mdr1 expression has more recently been associated also with the secretion of steroid hormones. This work presents an RT-PCR analysis of the expression of mdr isoforms, in several organs of mice during different...

  18. HIF1α isoforms in benign and malignant prostate tissue and their correlation to neuroendocrine differentiation

    International Nuclear Information System (INIS)

    Neuroendocrine (NE) differentiation in prostate cancer has been correlated with a poor prognosis and hormone refractory disease. In a previous report, we demonstrated the presence of immunoreactive cytoplasmic hypoxia inducible factor 1α (HIF1α), in both benign and malignant NE prostate cells. HIF1α and HIF1β are two subunits of HIF1, a transcription factor important for angiogenesis. The aim of this study was to elucidate whether the cytoplasmic stabilization of HIF1α in androgen independent NE differentiated prostate cancer is due to the presence of certain HIF1α isoforms. We studied the HIF1α isoforms present in 8 cases of benign prostate hyperplasia (BPH) and 43 cases of prostate cancer with and without NE differentiation using RT-PCR, sequencing analysis, immunohistochemistry and in situ hybridization. We identified multiple isoforms in both benign and malignant prostate tissues. One of these isoforms, HIF1α1.2, which was previously reported to be testis specific, was found in 86% of NE-differentiated prostate tumors, 92% of HIF1α immunoreactive prostate tumors and 100% of cases of benign prostate hyperplasia. Immunohistochemistry and in situ hybridization results showed that this isoform corresponds to the cytoplasmic HIF1α present in androgen-independent NE cells of benign and malignant prostate tissue and co-localizes with immunoreactive cytoplasmic HIF1β. Our results indicate that the cytoplasmic stabilization of HIF1α in NE-differentiated cells in benign and malignant prostate tissue is due to presence of an HIF1α isoform, HIF1α1.2. Co-localization of this isoform with HIF1β indicates that the HIF1α1.2 isoform might sequester HIF1β in the cytoplasm

  19. Differential and Conditional Activation of PKC-Isoforms Dictates Cardiac Adaptation during Physiological to Pathological Hypertrophy

    OpenAIRE

    Shaon Naskar; Kaberi Datta; Arkadeep Mitra; Kanchan Pathak; Ritwik Datta; Trisha Bansal; Sagartirtha Sarkar

    2014-01-01

    A cardiac hypertrophy is defined as an increase in heart mass which may either be beneficial (physiological hypertrophy) or detrimental (pathological hypertrophy). This study was undertaken to establish the role of different protein kinase-C (PKC) isoforms in the regulation of cardiac adaptation during two types of cardiac hypertrophy. Phosphorylation of specific PKC-isoforms and expression of their downstream proteins were studied during physiological and pathological hypertrophy in 24 week ...

  20. Progesterone receptor isoform A may regulate the effects of neoadjuvant aglepristone in canine mammary carcinoma

    OpenAIRE

    Guil-Luna, Silvia; Stenvang, Jan; Brünner, Nils; De Andrés, Francisco Javier; Rollón, Eva; Domingo, Víctor; Sánchez-Céspedes, Raquel; Millán, Yolanda; Mulas, Juana Martín de las

    2014-01-01

    Background Progesterone receptors play a key role in the development of canine mammary tumours, and recent research has focussed on their possible value as therapeutic targets using antiprogestins. Cloning and sequencing of the progesterone receptor gene has shown that the receptor has two isoforms, A and B, transcribed from a single gene. Experimental studies in human breast cancer suggest that the differential expression of progesterone receptor isoforms has implications for hormone therapy...

  1. Thermal Denaturation and Aggregation of Myosin Subfragment 1 Isoforms with Different Essential Light Chains

    OpenAIRE

    Zubov, Eugene O.; Nikolaeva, Olga P.; Kurganov, Boris I.; Dmitrii I. Levitsky; Markov, Denis I.

    2010-01-01

    We compared thermally induced denaturation and aggregation of two isoforms of the isolated myosin head (myosin subfragment 1, S1) containing different “essential” (or “alkali”) light chains, A1 or A2. We applied differential scanning calorimetry (DSC) to investigate the domain structure of these two S1 isoforms. For this purpose, a special calorimetric approach was developed to analyze the DSC profiles of irreversibly denaturing multidomain proteins. Using this approach, we revealed two calor...

  2. Differential Contributions of Nonmuscle Myosin II Isoforms and Functional Domains to Stress Fiber Mechanics

    OpenAIRE

    Ching-Wei Chang; Sanjay Kumar

    2015-01-01

    While is widely acknowledged that nonmuscle myosin II (NMMII) enables stress fibers (SFs) to generate traction forces against the extracellular matrix, little is known about how specific NMMII isoforms and functional domains contribute to SF mechanics. Here we combine biophotonic and genetic approaches to address these open questions. First, we suppress the NMMII isoforms MIIA and MIIB and apply femtosecond laser nanosurgery to ablate and investigate the viscoelastic retraction of individual ...

  3. Electrophoretic Mobility of Cardiac Myosin Heavy Chain Isoforms Revisited: Application of MALDI TOF/TOF Analysis

    Czech Academy of Sciences Publication Activity Database

    Arnoštová, P.; Jedelsky, P. L.; Soukup, Tomáš; Žurmanová, J.

    2011-01-01

    Roč. 2011, - (2011), e634253. ISSN 1110-7243 R&D Projects: GA AV ČR IAAX01110901; GA ČR(CZ) GA304/08/0256 Institutional research plan: CEZ:AV0Z50110509 Keywords : cardiac MyHC isoforms * MyHC isoform mobility * effect of thyroid hormones * mass spectrometry * SDS-PAGE and western blot Subject RIV: ED - Physiology Impact factor: 2.436, year: 2011

  4. Kinetics of local and systemic isoforms of serum amyloid A in bovine mastitic milk

    DEFF Research Database (Denmark)

    Jacobsen, Stine; Niewold, T.A.; Kornalijnslijper, E.;

    2005-01-01

    The aim of the present study was to characterise the serum amyloid A (SAA) response to intramammary inoculation of Escherichia coli and to examine the distribution of hepatically and extrahepatically pruduced SAA isoforms in plasma and milk fra cows with mastitis.......The aim of the present study was to characterise the serum amyloid A (SAA) response to intramammary inoculation of Escherichia coli and to examine the distribution of hepatically and extrahepatically pruduced SAA isoforms in plasma and milk fra cows with mastitis....

  5. On the red giant branch mass loss in 47 Tucanae: Constraints from the horizontal branch morphology

    CERN Document Server

    Salaris, Maurizio; Pietrinferni, Adriano

    2016-01-01

    We obtain stringent constraints on the actual efficiency of mass loss for red giant branch stars in the Galactic globular cluster 47 Tuc, by comparing synthetic modeling based on stellar evolution tracks with the observed distribution of stars along the horizontal branch in the colour-magnitude-diagram. We confirm that the observed, wedge-shaped distribution of the horizontal branch can be reproduced only by accounting for a range of initial He abundances --in agreement with inferences from the analysis of the main sequence-- and a red giant branch mass loss with a small dispersion. We have carefully investigated several possible sources of uncertainty that could affect the results of the horizontal branch modeling, stemming from uncertainties in both stellar model computations and the cluster properties such as heavy element abundances, reddening and age. We determine a firm lower limit of ~0.17$Mo for the mass lost by red giant branch stars, corresponding to horizontal branch stellar masses between ~0.65Mo ...

  6. Inhibitors of Testosterone Biosynthetic and Metabolic Activation Enzymes

    Directory of Open Access Journals (Sweden)

    Leping Ye

    2011-12-01

    Full Text Available The Leydig cells of the testis have the capacity to biosynthesize testosterone from cholesterol. Testosterone and its metabolically activated product dihydrotestosterone are critical for the development of male reproductive system and spermatogenesis. At least four steroidogenic enzymes are involved in testosterone biosynthesis: Cholesterol side chain cleavage enzyme (CYP11A1 for the conversion of cholesterol into pregnenolone within the mitochondria, 3β-hydroxysteroid dehydrogenase (HSD3B, for the conversion of pregnenolone into progesterone, 17α-hydroxylase/17,20-lyase (CYP17A1 for the conversion of progesterone into androstenedione and 17β-hydroxysteroid dehydrogenase (HSD17B3 for the formation of testosterone from androstenedione. Testosterone is also metabolically activated into more potent androgen dihydrotestosterone by two isoforms 5α-reductase 1 (SRD5A1 and 2 (SRD5A2 in Leydig cells and peripheral tissues. Many endocrine disruptors act as antiandrogens via directly inhibiting one or more enzymes for testosterone biosynthesis and metabolic activation. These chemicals include industrial materials (perfluoroalkyl compounds, phthalates, bisphenol A and benzophenone and pesticides/biocides (methoxychlor, organotins, 1,2-dibromo-3-chloropropane and prochloraz and plant constituents (genistein and gossypol. This paper reviews these endocrine disruptors targeting steroidogenic enzymes.

  7. Medial branch neurotomy in low back pain

    International Nuclear Information System (INIS)

    This study aimed to assess the effectiveness of pulsed radiofrequency medial branch dorsal ramus neurotomy in patients with facet joint syndrome. From January 2008 to April 2010, 92 patients with facet joint syndrome diagnosed by strict inclusion criteria and controlled diagnostic blocks undergone medial branch neurotomy. We did not exclude patients with failed back surgery syndrome (FBSS). Electrodes (20G) with 5-mm active tip were placed under fluoroscopy guide parallel to medial branch. Patients were followed up by physical examination and by Visual Analog Scale and Oswestry Disability Index at 1, 6, and 12 months. In all cases, pain improvement was statistically significant and so quality of life. Three non-FBSS patients had to undergo a second neurotomy because of non-satisfactory pain decrease. Complications were reported in no case. Medial branch radiofrequency neurotomy has confirmed its well-established effectiveness in pain and quality of life improvement as long as strict inclusion criteria be fulfilled and nerve ablation be accomplished by parallel electrode positioning. This statement can be extended also to FBSS patients. (orig.)

  8. Medial branch neurotomy in low back pain

    Energy Technology Data Exchange (ETDEWEB)

    Masala, Salvatore; Mammucari, Matteo; Simonetti, Giovanni [Interventional Radiology and Radiotherapy University ' ' Tor Vergata' ' , Department of Diagnostic and Molecular Imaging, Rome (Italy); Nano, Giovanni [Interventional Radiology and Radiotherapy University ' ' Tor Vergata' ' , Department of Diagnostic and Molecular Imaging, Rome (Italy); University ' ' Tor Vergata' ' , Department of Radiology, Rome (Italy); Marcia, Stefano [S. Giovanni di Dio Hospital, Department of Diagnostic and Molecular Imaging, Cagliari (Italy)

    2012-07-15

    This study aimed to assess the effectiveness of pulsed radiofrequency medial branch dorsal ramus neurotomy in patients with facet joint syndrome. From January 2008 to April 2010, 92 patients with facet joint syndrome diagnosed by strict inclusion criteria and controlled diagnostic blocks undergone medial branch neurotomy. We did not exclude patients with failed back surgery syndrome (FBSS). Electrodes (20G) with 5-mm active tip were placed under fluoroscopy guide parallel to medial branch. Patients were followed up by physical examination and by Visual Analog Scale and Oswestry Disability Index at 1, 6, and 12 months. In all cases, pain improvement was statistically significant and so quality of life. Three non-FBSS patients had to undergo a second neurotomy because of non-satisfactory pain decrease. Complications were reported in no case. Medial branch radiofrequency neurotomy has confirmed its well-established effectiveness in pain and quality of life improvement as long as strict inclusion criteria be fulfilled and nerve ablation be accomplished by parallel electrode positioning. This statement can be extended also to FBSS patients. (orig.)

  9. Measurement of the eta decay branching ratio

    CERN Document Server

    Bargholtz, C; Bogoslawsky, D; Calén, H; Capellaro, F; Clement, H; Demirörs, L; Ekström, C; Fransson, K; Geren, L; Gustafsson, L; Höistad, Bo; Ivanov, G; Jacewicz, M; Jiganov, E; Johansson, T; Keleta, S; Koch, I; Kullander, Sven; Kupsc, A; Kuznetsov, A; Laukhin, I V; Lindberg, K; Marciniewski, P; Meier, R; Morosov, B; Oelert, W; Pauly, C; Pettersson, H; Petukhov, Yu P; Povtorejko, A; Ruber, Roger J M Y; Schönning, K; Scobel, W; Shafigullin, R; Shwartz, B; Skorodko, T Yu; Sopov, V; Stepaniak, J; Chernyshov, V; Tegnér, P E; Engblom, P T; Tikhomirov, V; Turowiecki, A; Wagner, G J; Wolke, M; Yamamoto, A; Zabierowski, J; Zartova, I; Zlomanczuk, Yu; Bargholtz, Chr.

    2007-01-01

    The reaction pd->3He eta at threshold was used to provide a clean source of eta mesons for decay studies with the WASA detector at CELSIUS. The branching ratio of the decay eta->pi+pi-e+e- is measured to be (4.6+/-1.4+/-0.5)x10^-4.

  10. 76 FR 60757 - Executive Branch Qualified Trusts

    Science.gov (United States)

    2011-09-30

    ... qualified trusts provisions for the executive branch in subparts D and E of 5 CFR part 2634 (see 57 FR 11800.... 12674, 54 FR 15159, 3 CFR, 1989 Comp., p. 215, as modified by E.O. 12731, 55 FR 42547, 3 CFR, 1990 Comp... been in business, its policies and philosophy in managing assets, the types of clients it serves,...

  11. Infrared studies of asymptotic giant branch stars

    International Nuclear Information System (INIS)

    In this thesis studies are presented of asymptotic giant branch stars, which are thought to be an important link in the evolution of the galaxy. The studies were performed on the basis of data collected by the IRAS, the infrared astronomical satelite. 233 refs.; 33 figs.; 16 tabs

  12. Recombinant erythropoietin in humans has a prolonged effect on circulating erythropoietin isoform distribution

    DEFF Research Database (Denmark)

    Aachmann-Andersen, Niels Jacob; Just Christensen, Søren; Lisbjerg, Kristian;

    2014-01-01

    The membrane-assisted isoform immunoassay (MAIIA) quantitates erythropoietin (EPO) isoforms as percentages of migrated isoforms (PMI). We evaluated the effect of recombinant human EPO (rhEPO) on the distribution of EPO isoforms in plasma in a randomized, placebo-controlled, double-blinded, cross......-over study. 16 healthy subjects received either low-dose Epoetin beta (5000 IU on days 1, 3, 5, 7, 9, 11 and 13); high-dose Epoetin beta (30.000 IU on days 1, 2 and 3 and placebo on days 5, 7, 9, 11 and 13); or placebo on all days. PMI on days 4, 11 and 25 was determined by interaction of N......-acetyl glucosamine with the glycosylation dependent desorption of EPO isoforms. At day 25, plasma-EPO in both rhEPO groups had returned to values not different from the placebo group. PMI with placebo, reflecting the endogenous EPO isoforms, averaged 82.5 (10.3) % (mean (SD)). High-dose Epoetin beta decreased PMI on...

  13. Distribution of tropomyosin isoforms in different types of single fibers isolated from bovine skeletal muscles.

    Science.gov (United States)

    Oe, M; Ojima, K; Nakajima, I; Chikuni, K; Shibata, M; Muroya, S

    2016-08-01

    To clarify the relationship between myosin heavy chain (MyHC) isoforms and tropomyosin (TPM) isoforms in single fibers, 64 single fibers were isolated from each of bovine three muscles (masseter, semispinalis and semitendinosus). mRNA expressions of MyHC and TPM isoforms were analyzed by real-time PCR. All single fibers from the masseter expressed MyHC-slow. The fibers from the semispinalis expressed both MyHC-slow and 2a. The fibers from the semitendinosus expressed MyHC-slow, 2a and 2x. TPM-1 and TPM-2 were co-expressed in 2a and 2x type fibers, and TPM-2 and TPM-3 were co-expressed in slow type fibers. The expression pattern of TPM isoforms in each fiber type was similar between fibers isolated from different muscles. These results suggest that TPM-1 and TPM-3 isoforms correspond to the function of 2a or 2x type fibers and slow type fibers, respectively, with TPM-2 in common. Furthermore, the patterns of MyHC and TPM isoform combinations did not vary among single fibers isolated from the individual muscles examined. PMID:27105153

  14. Expression of mdr isoforms in mice during estrous cycle and under hormone stimulation

    Directory of Open Access Journals (Sweden)

    Marion Schiengold

    2006-01-01

    Full Text Available The multidrug resistance (MDR phenotype is associated with the expression of P-glycoprotein (Pgp, coded by the multigenic mdr family. Mice present the isoforms mdr1 and mdr3, which are responsible for multidrug resistance, and mdr2, that is involved in the transport of phospholipids. mdr1 expression has more recently been associated also with the secretion of steroid hormones. This work presents an RT-PCR analysis of the expression of mdr isoforms, in several organs of mice during different phases of the estrous cycle. Additionally, females were ovariectomized, submitted to different hormone treatments, and their uterus was analyzed for the expression of mdr isoforms. The results show that in the adrenal gland and ovaries mdr1 is the main isoform during proestrus, and that progesterone or a combination of progesterone and estrogen induce the expression of all mdr isoforms in the uterus of ovariectomized females. We suggest that the functions of mdr1 and mdr3 are overlapping, that mdr3 may be the more efficient isoform in the detoxification function, and that mdr1 may be more closely related to the secretion of steroid hormones.

  15. The AFCRL Lunar amd Planetary Research Branch

    Science.gov (United States)

    Price, Stephan D.

    2011-07-01

    The Lunar and Planetary research program led by Dr John (Jack) Salisbury in the 1960s at the United States Air Force Cambridge Research Laboratories (AFCRL) investigated the surface characteristics of Solar System bodies. The Branch was one of the first groups to measure the infrared spectra of likely surface materials in the laboratory under appropriate vacuum and temperature conditions. The spectral atlases created from the results were then compared to photometric and spectral measurements obtained from ground- and balloon-based telescopes to infer the mineral compositions and physical conditions of the regoliths of the Moon, Mars and asteroids. Starting from scratch, the Branch initially sponsored observations of other groups while its in-house facilities were being constructed. The earliest contracted efforts include the spatially-resolved mapping of the Moon in the first half of the 1960s by Richard W. Shorthill and John W. Saari of the Boeing Scientific Research Laboratories in Seattle. This effort ultimately produced isophotal and isothermal contour maps of the Moon during a lunation and time-resolved thermal images of the eclipsed Moon. The Branch also sponsored probe rocket-based experiments flown by Riccardo Giacconi and his group at American Science and Engineering Inc. that produced the first observations of X-ray stars in 1962 and later the first interferometric measurement of the ozone and C02 emission in the upper atmosphere. The Branch also made early use of balloon-based measurements. This was a singular set of experiments, as these observations are among the very few mid-infrared astronomical measurements obtained from a balloon platform. Notable results of the AFCRL balloon flights were the mid-infrared spectra of the spatially-resolved Moon obtained with the University of Denver mid-infrared spectrometer on the Branch's balloon-borne 61-cm telescope during a 1968 flight. These observations remain among the best available. Salisbury also funded

  16. Concerted suppression of all starch branching enzyme genes in barley produces amylose-only starch granules

    DEFF Research Database (Denmark)

    Carciofi, Massimiliano; Blennow, Andreas; Jensen, Susanne L;

    2012-01-01

    endosperm. This trait was segregating 3:1. Amylose-only starch granules were irregularly shaped and showed peculiar thermal properties and crystallinity. Transgenic lines retained high-yield possibly due to a pleiotropic upregualtion of other starch biosynthetic genes compensating the SBEs loss. For...

  17. Enzyme inhibitor studies reveal complex control of methyl-D-erythritol 4-phosphate (MEP pathway enzyme expression in Catharanthus roseus.

    Directory of Open Access Journals (Sweden)

    Mei Han

    Full Text Available In Catharanthus roseus, the monoterpene moiety exerts a strong flux control for monoterpene indole alkaloid (MIA formation. Monoterpene synthesis depends on the methyl-D-erythritol 4-phosphate (MEP pathway. Here, we have explored the regulation of this pathway in response to developmental and environmental cues and in response to specific enzyme inhibitors. For the MEP pathway entry enzyme 1-deoxy-D-xylulose 5-phosphate synthase (DXS, a new (type I DXS isoform, CrDXS1, has been cloned, which, in contrast to previous reports on type II CrDXS, was not transcriptionally activated by the transcription factor ORCA3. Regulation of the MEP pathway in response to metabolic perturbations has been explored using the enzyme inhibitors clomazone (precursor of 5-ketochlomazone, inhibitor of DXS and fosmidomycin (inhibitor of deoxyxylulose 5-phosphate reductoisomerase (DXR, respectively. Young leaves of non-flowering plants were exposed to both inhibitors, adopting a non-invasive in vivo technique. Transcripts and proteins of DXS (3 isoforms, DXR, and hydroxymethylbutenyl diphosphate synthase (HDS were monitored, and protein stability was followed in isolated chloroplasts. Transcripts for DXS1 were repressed by both inhibitors, whereas transcripts for DXS2A&B, DXR and HDS increased after clomazone treatment but were barely affected by fosmidomycin treatment. DXS protein accumulated in response to both inhibitors, whereas DXR and HDS proteins were less affected. Fosmidomycin-induced accumulation of DXS protein indicated substantial posttranscriptional regulation. Furthermore, fosmidomycin effectively protected DXR against degradation in planta and in isolated chloroplasts. Thus our results suggest that DXR protein stability may be affected by substrate binding. In summary, the present results provide novel insight into the regulation of DXS expression in C. roseus in response to MEP-pathway perturbation.

  18. Enzyme Inhibitor Studies Reveal Complex Control of Methyl-D-Erythritol 4-Phosphate (MEP) Pathway Enzyme Expression in Catharanthus roseus

    Science.gov (United States)

    Han, Mei; Heppel, Simon C.; Su, Tao; Bogs, Jochen; Zu, Yuangang; An, Zhigang; Rausch, Thomas

    2013-01-01

    In Catharanthus roseus, the monoterpene moiety exerts a strong flux control for monoterpene indole alkaloid (MIA) formation. Monoterpene synthesis depends on the methyl-D-erythritol 4-phosphate (MEP) pathway. Here, we have explored the regulation of this pathway in response to developmental and environmental cues and in response to specific enzyme inhibitors. For the MEP pathway entry enzyme 1-deoxy-D-xylulose 5-phosphate synthase (DXS), a new (type I) DXS isoform, CrDXS1, has been cloned, which, in contrast to previous reports on type II CrDXS, was not transcriptionally activated by the transcription factor ORCA3. Regulation of the MEP pathway in response to metabolic perturbations has been explored using the enzyme inhibitors clomazone (precursor of 5-ketochlomazone, inhibitor of DXS) and fosmidomycin (inhibitor of deoxyxylulose 5-phosphate reductoisomerase (DXR)), respectively. Young leaves of non-flowering plants were exposed to both inhibitors, adopting a non-invasive in vivo technique. Transcripts and proteins of DXS (3 isoforms), DXR, and hydroxymethylbutenyl diphosphate synthase (HDS) were monitored, and protein stability was followed in isolated chloroplasts. Transcripts for DXS1 were repressed by both inhibitors, whereas transcripts for DXS2A&B, DXR and HDS increased after clomazone treatment but were barely affected by fosmidomycin treatment. DXS protein accumulated in response to both inhibitors, whereas DXR and HDS proteins were less affected. Fosmidomycin-induced accumulation of DXS protein indicated substantial posttranscriptional regulation. Furthermore, fosmidomycin effectively protected DXR against degradation in planta and in isolated chloroplasts. Thus our results suggest that DXR protein stability may be affected by substrate binding. In summary, the present results provide novel insight into the regulation of DXS expression in C. roseus in response to MEP-pathway perturbation. PMID:23650515

  19. Molecular characterization of human thyroid hormone receptor β isoform 4.

    Science.gov (United States)

    Moriyama, Kenji; Yamamoto, Hiroyuki; Futawaka, Kumi; Atake, Asami; Kasahara, Masato; Tagami, Tetsuya

    2016-01-01

    Thyroid hormone exerts a pleiotropic effect on development, differentiation, and metabolism through thyroid hormone receptor (TR). A novel thyroid hormone receptor β isoform (TRβ4) was cloned using PCR from a human pituitary cDNA library as a template. We report here the characterization of TRβ4 from a molecular basis. Temporal expression of TRβ4 during the fetal period is abundant in the brain and kidney, comparable with the adult pattern. Western blot analysis revealed that TRs are ubiquitination labile proteins, while TRβ1 is potentially stable. TRβ1, peroxisome proliferator-activated receptors (PPAR), and vitamin D receptor (VDR), which belong to class II transcription factors that function via the formation of heterodimeric complexes with retinoid X receptor (RXR), were suppressed by TRβ4 in a dose-dependent manner. Thus, TRβ4 exhibits ligand-independent transcriptional silencing, possibly as a substitute for dimerized RXR. In this study, TRβ1 and TRβ4 transcripts were detected in several cell lines. Quantitative RT-PCR assay showed that the expression of TRβ4 in human embryonic carcinoma cells of the testis was suppressed by sex hormone in a reciprocal manner to TRβ1. In contrast, TRβ4 was expressed under a high dose of triiodothyronine (T3) in a reciprocal manner to TRβ1. Finally, in transiently transfected NIH-3T3 cells, green fluorescence protein (GFP)-tagged TRβ4 was mostly nuclear in both the absence and the presence of T3. By mutating defined regions of both TRβs, we found that both TRβ1 and TRβ4 had altered nuclear/cytoplasmic distribution as compared with wild-type, and different to T3 and the nuclear receptor corepressor (NCoR). Thus, site-specific DNA binding is not essential for maintaining TRβs within the nucleus. PMID:26513165

  20. Hyphenated techniques for the characterization and quantification of metallothionein isoforms

    Energy Technology Data Exchange (ETDEWEB)

    Prange, Andreas; Schaumloeffel, Dirk [GKSS Research Center, Institute for Coastal Research/Physical and Chemical Analysis, Max-Planck-Strasse, 21502 Geesthacht (Germany)

    2002-07-01

    Recent developments in the coupling of highly selective separation techniques such as capillary electrophoresis (CE) and high-performance liquid chromatography (HPLC) to element-specific and molecule-specific detectors, such as inductively-coupled plasma mass spectrometry (ICP-MS) and electrospray ionization-tandem mass spectrometry (ESI-MS/MS) for the characterization and quantification of metallothioneins (MTs) are critically reviewed and discussed. This review gives an update based on the literature over the last five years. The coupling of CE to ICP-MS is especially highlighted. As a result of progress in new interface technologies for CE-ICP-MS, research topics presented in the literature are changing from ''the characterization of interfaces by metallothioneins'' to the ''characterization of metallothioneins by CE-ICP-MS''. New applications of CE-ICP-MS to the analysis of MTs in real samples are summarized. The potential of the on-line isotope dilution technique for the quantification of MTs and for the determination of the stoichiometric composition of metalloprotein complexes is discussed. Furthermore, a selection of relevant papers dealing with HPLC-ICP-MS for MT analysis are summarized and compared to those dealing with CE-ICP-MS. In particular, the use of size-exclusion (SE)-HPLC as a preliminary separation step for metallothioneins in real samples prior to further chromatographic or electrophoretic separations is considered. Additionally, the application of electrospray ionisation-tandem mass spectrometry (ESI-MS/MS) for the identification of metallothionein isoforms following electrophoretic or chromatographic separation is discussed. (orig.)

  1. The H{sub 1}–H{sub 2} domain of the α{sub 1} isoform of Na{sup +}–K{sup +}–ATPase is involved in ouabain toxicity in rat ventricular myocytes

    Energy Technology Data Exchange (ETDEWEB)

    Xiong, Chen; Li, Jun-xia; Guo, Hui-cai; Zhang, Li-nan; Guo, Wei; Meng, Jing; Wang, Yong-li, E-mail: wangyongli@gmail.com

    2012-07-01

    The composition of different isoforms of Na{sup +}-K{sup +}-ATPase (NKA, Na/K pump) in ventricular myocytes is an important factor in determining the therapeutic effect and toxicity of cardiac glycosides (CGs) on heart failure. The mechanism whereby CGs cause these effects is still not completely clear. In the present study, we prepared two site-specific antibodies (SSA78 and WJS) against the H{sub 1}–H{sub 2} domain of α{sub 1} and α{sub 2} isoforms of NKA in rat heart, respectively, and compared their influences on the effect of ouabain (OUA) in isolated rat ventricular myocytes. SSA78 or WJS, which can specifically bind with the α{sub 1} or α{sub 2} isoform, were assessed with enzyme linked immunosorbent assay (ELISA), Western blot and immunofluorescent staining methods. Preincubation of myocytes with SSA78 inhibited low OUA affinity pump current but not high OUA affinity pump current, reduced the rise in cytosolic calcium concentration ([Ca{sup 2+}]{sub i}), attenuated mitochondrial Ca{sup 2+} overload, restored mitochondrial membrane potential reduction, and delayed the decrease of the myocardial contractile force as well as the occurrence of arrhythmic contraction induced by high concentrations (1 mM) but not low concentrations (1 μM) of OUA. Similarly, preincubation of myocytes with WJS inhibited high OUA affinity pump current, reduced the increase of [Ca{sup 2+}]{sub i} and the contractility induced by 1 μM but not that induced by 1 mM OUA. These results indicate that the H{sub 1}–H{sub 2} domain of the NKA α{sub 1} isoform mediates OUA-induced cardiac toxicity in rat ventricular myocytes, and inhibitors for this binding site may be used as an adjunct to CGs treatment for cardiovascular disease. -- Highlights: ► We prepared two antibodies against the H{sub 1}-H{sub 2} domain of α{sub 1} and α{sub 2} isoforms of NKA. ► The H{sub 1}-H{sub 2} domain of the NKA α{sub 1} isoform mediates OUA-induced cardiac toxicity. ► The H{sub 1}-H{sub 2

  2. Expression of 14-3-3 protein isoforms in mouse oocytes, eggs and ovarian follicular development

    Directory of Open Access Journals (Sweden)

    De Santanu

    2012-01-01

    Full Text Available Abstract Background The 14-3-3 (YWHA proteins are a highly conserved, ubiquitously expressed family of proteins. Seven mammalian isoforms of 14-3-3 are known (β, γ, ε, ζ, η, τ and, σ. These proteins associate with many intracellular proteins involved in a variety of cellular processes including regulation of the cell cycle, metabolism and protein trafficking. We are particularly interested in the role of 14-3-3 in meiosis in mammalian eggs and the role 14-3-3 proteins may play in ovarian function. Therefore, we examined the expression of 14-3-3 proteins in mouse oocyte and egg extracts by Western blotting after polyacrylamide gel electrophoresis, viewed fixed cells by indirect immunofluorescence, and examined mouse ovarian cells by immunohistochemical staining to study the expression of the different 14-3-3 isoforms. Results We have determined that all of the mammalian 14-3-3 isoforms are expressed in mouse eggs and ovarian follicular cells including oocytes. Immunofluorescence confocal microscopy of isolated oocytes and eggs confirmed the presence of all of the isoforms with characteristic differences in some of their intracellular localizations. For example, some isoforms (β, ε, γ, and ζ are expressed more prominently in peripheral cytoplasm compared to the germinal vesicles in oocytes, but are uniformly dispersed within eggs. On the other hand, 14-3-3η is diffusely dispersed in the oocyte, but attains a uniform punctate distribution in the egg with marked accumulation in the region of the meiotic spindle apparatus. Immunohistochemical staining detected all isoforms within ovarian follicles, with some similarities as well as notable differences in relative amounts, localizations and patterns of expression in multiple cell types at various stages of follicular development. Conclusions We found that mouse oocytes, eggs and follicular cells within the ovary express all seven isoforms of the 14-3-3 protein. Examination of the

  3. Analyzing the Interaction of Andrographolide and Neoandrographolide, Diterpenoid Compounds From Andrographis Paniculata (Burm.F) Nees, to Cyclooxygenase-2 Enzyme by Docking Simulation

    OpenAIRE

    Jutti Levita; Enade P. Istyastono; As’ari Nawawi; Abdul Mutholib; de Esch, Iwan J. P.; Slamet Ibrahim Surantaatmadja

    2009-01-01

    Cyclooxygenase (COX), an enzyme involved in the conversion of arachidonic acid to prostaglandins, exists in two isoforms, which are COX-1 and COX-2. Despite the similarities of COX-1 and COX-2, the two isoforms show subtle differences in amino acid composition at the active sites. Since COX-1 has isoleucine, a bulkier amino acid at position 523 than COX-2’s valine, it allows COX-2 to have a larger space in its active site. Andrographolide reduces COX-2 expression induced by PAF and fMLP in...

  4. Production of ACAT1 56-kDa isoform in human cells via trans-splicing involving the ampicillin resistance gene

    Institute of Scientific and Technical Information of China (English)

    Guang-Jing Hu; Jia Chen; Xiao-Nan Zhao; Jia-Jia Xu; Dong-Qing Guo; Ming Lu; Ming Zhu

    2013-01-01

    Trans-splicing,a process involving the cleavage and joining of two separate transcripts,can expand the transcriptome and proteome in eukaryotes.Chimeric RNAs generated by trans-splicing are increasingly described in literatures.The widespread presence of antibiotic resistance genes in natural environments and human intestines is becoming an important challenge for public health.Certain antibiotic resistance genes,such as ampicillin resistance gene (Amp),are frequently used in recombinant plasmids.Until now,trans-splicing involving recombinant plasmid-derived exogenous transcripts and endogenous cellular RNAs has not been reported.Acyl-CoA:cholesterol acyltransferase 1 (ACAT1) is a key enzyme involved in cellular cholesterol homeostasis.The 4.3-kb human ACAT1 chimeric mRNA can produce 50-kDa and 56-kDa isoforms with different enzymatic activities.Here,we show that human ACAT1 56-kDa isoform is produced from an mRNA species generated through the trans-splicing of an exogenous transcript encoded by the antisense strand of Ampr (asAmp) present in common Ampr-plasmids and the 4.3-kb endogenous ACAT1 chimeric mRNA,which is presumably processed through a prior event of interchromosomal trans-splicing.Strikingly,DNA fragments containing the asAmp with an upstream recombined cryptic promoter and the corresponding exogenous asAmp transcripts have been detected in human cells.Our findings shed lights on the mechanism of human ACAT1 56-kDa isoform production,reveal an exogenous-endogenous trans-splicing system,in which recombinant plasmid-derived exogenous transcripts are linked with endogenous cellular RNAs in human cells,and suggest that exogenous DNA might affect human gene expression at both DNA and RNA levels.

  5. Angiotensin-converting enzyme

    DEFF Research Database (Denmark)

    Sørensen, P G; Rømer, F K; Cortes, D

    1984-01-01

    In order to evaluate bleomycin-associated lung damage in humans, lung function parameters and serum levels of the endothelial-bound angiotensin-converting enzyme (ACE) were determined by serial measurements in 11 patients who were treated for testicular cancer. None developed clinical or radiolog......In order to evaluate bleomycin-associated lung damage in humans, lung function parameters and serum levels of the endothelial-bound angiotensin-converting enzyme (ACE) were determined by serial measurements in 11 patients who were treated for testicular cancer. None developed clinical or...

  6. The small chemical enzyme inhibitor 5-phenylnicotinic acid/CD13 inhibits cell migration and invasion of tartrate-resistant acid phosphatase/ACP5-overexpressing MDA-MB-231 breast cancer cells.

    Science.gov (United States)

    Krumpel, Michael; Reithmeier, Anja; Senge, Teresa; Baeumler, Toni Andreas; Frank, Martin; Nyholm, Per-Georg; Ek-Rylander, Barbro; Andersson, Göran

    2015-11-15

    Tartrate-resistant acid phosphatase (TRAP/ACP5/uteroferrin/purple acid phosphatase/PP5) has received considerable attention as a newly discovered proinvasion metastasis driver associated with different malignancies. This renders TRAP an interesting target for novel anti-cancer therapy approaches. TRAP exists as two isoforms, 5a and 5b, where the 5a isoform represents an enzymatically less active monomeric precursor to the more enzymatically active 5b isoform generated by proteolytic excision of a repressive loop domain. Recently, three novel lead compounds were identified by fragment-based screening and demonstrated to be efficient TRAP enzyme inhibitors in vitro. We conclude that one of the three compounds i.e. 5-phenylnicotinic acid (CD13) was efficient as a TRAP inhibitor with Kic values in the low micromolar range towards the TRAP 5b isoform, but was not able to inhibit the TRAP 5a isoform. Structure-based docking revealed similar interactions of CD13 with the active site in both TRAP isoforms. In stably TRAP-overexpressing MDA-MB-231 breast cancer cells, CD13 inhibited intracellular TRAP activity and showed no cytotoxicity at 200 µM. Furthermore, CD13 selectively blocked the TRAP 5b isoform compared to the TRAP 5a in cultured cells, indicating the usefulness of CD13 for assessing the different biological functions of the two TRAP isoforms 5a and 5b in cell systems. Moreover, inhibition of cell migration and invasion of stably TRAP-overexpressing MDA-MB-231 by CD13 was observed. These data establish a proof of principle that a small chemical inhibitor of the TRAP enzyme can block TRAP-dependent functions in cancer cells. PMID:26428664

  7. Hyperthermophilic Enzymes with Industrial Applications

    OpenAIRE

    Mojsov, Kiro; Janevski, Aco; Andronikov, Darko; Zezova, Silvana

    2014-01-01

    Hyperthermophilic enzymes are typically thermostable and are optimally active at high temperatures. Hyperthermophilic enzymes are very similar to their mesophilic homologues. No single mechanism that is responsible for the remarkable stability of hyperthermophilic enzymes. Increased thermo stability must be found in a small number of highly specific alterations. In this review are described current uses and potential applications of thermophilic and hyperthermophilic enzymes as ...

  8. The ENZYME database in 2000.

    Science.gov (United States)

    Bairoch, A

    2000-01-01

    The ENZYME database is a repository of information related to the nomenclature of enzymes. In recent years it has became an indispensable resource for the development of metabolic databases. The current version contains information on 3705 enzymes. It is available through the ExPASy WWW server (http://www.expasy.ch/enzyme/ ). PMID:10592255

  9. Transferrin and Its Isoforms from Normal Human Serum Revealed by Several Analytical Techniques

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    Transferrin(TF) and its isoforms have been widely reported via various analytical techniques, including a noticeable increased number of isoforms with low content of sialic acid(asialo-, monosialo-, and disialo-transferrin) and asialo-TF as well as disialo-TF, with one or several oligosaccharides released in human serum transferrin(hTf). Here,hTf has been purified by native gradient polyacrylamide gel electrophoresis(PAGENG) before use. The hTf extracted with the electron-transfer approach showed a single subunit band(77.1 Da) in the SDS-PAGE gel, but it exhibited two bands in the native and denatured isoelectric focusing(IEF) gels, namely, hTf-2Fe3+ and apo-hTf, without finding any other transferrin isoforms. A reversed phase HPLC(RP-HPLC) equipped with a C18 column effectively separated hTf and its polymers and combined off-line techniques, including peptide mass fingerprinting(PMF), matrix-assisted laser desorption/ionization time-of-flight mass spectrometry(MALDI-TOF-MS) and database search, and identified the high homology among hTf, apo-hTf, and their isoforms. Moreover, the elution solution consisting of acetonitrile and formic acid could easily denature both hTf and apo-hTf to form various isoforms during separation with HPLC, indicating that chemical factors lead to the formation of various isoforms in transferrin, artificially, during extraction and separation.The authors claimed that only two transferrin isoforms existed in the NHS, namely, hTf-2Fe3+ and apo-hTf, which could be employed in biomarkers, to distinguish the healthy population from many disease sufferers, such as,carbohydrate-deficient transferrin(CDT).

  10. Identification of alternatively translated Tetherin isoforms with differing antiviral and signaling activities.

    Directory of Open Access Journals (Sweden)

    Luis J Cocka

    2012-09-01

    Full Text Available Tetherin (BST-2/CD317/HM1.24 is an IFN induced transmembrane protein that restricts release of a broad range of enveloped viruses. Important features required for Tetherin activity and regulation reside within the cytoplasmic domain. Here we demonstrate that two isoforms, derived by alternative translation initiation from highly conserved methionine residues in the cytoplasmic domain, are produced in both cultured human cell lines and primary cells. These two isoforms have distinct biological properties. The short isoform (s-Tetherin, which lacks 12 residues present in the long isoform (l-Tetherin, is significantly more resistant to HIV-1 Vpu-mediated downregulation and consequently more effectively restricts HIV-1 viral budding in the presence of Vpu. s-Tetherin Vpu resistance can be accounted for by the loss of serine-threonine and tyrosine motifs present in the long isoform. By contrast, the l-Tetherin isoform was found to be an activator of nuclear factor-kappa B (NF-κB signaling whereas s-Tetherin does not activate NF-κB. Activation of NF-κB requires a tyrosine-based motif found within the cytoplasmic tail of the longer species and may entail formation of l-Tetherin homodimers since co-expression of s-Tetherin impairs the ability of the longer isoform to activate NF-κB. These results demonstrate a novel mechanism for control of Tetherin antiviral and signaling function and provide insight into Tetherin function both in the presence and absence of infection.

  11. PI3K isoform-selective inhibitors: next-generation targeted cancer therapies.

    Science.gov (United States)

    Wang, Xiang; Ding, Jian; Meng, Ling-hua

    2015-10-01

    The pivotal roles of phosphatidylinositol 3-kinases (PI3Ks) in human cancers have inspired active development of small molecules to inhibit these lipid kinases. However, the first-generation pan-PI3K and dual-PI3K/mTOR inhibitors have encountered problems in clinical trials, with limited efficacies as a monotherapeutic agent as well as a relatively high rate of side effects. It is increasingly recognized that different PI3K isoforms play non-redundant roles in particular tumor types, which has prompted the development of isoform-selective inhibitors for pre-selected patients with the aim for improving efficacy while decreasing undesirable side effects. The success of PI3K isoform-selective inhibitors is represented by CAL101 (Idelalisib), a first-in-class PI3Kδ-selective small-molecule inhibitor that has been approved by the FDA for the treatment of chronic lymphocytic leukemia, indolent B-cell non-Hodgkin's lymphoma and relapsed small lymphocytic lymphoma. Inhibitors targeting other PI3K isoforms are also being extensively developed. This review focuses on the recent progress in development of PI3K isoform-selective inhibitors for cancer therapy. A deeper understanding of the action modes of novel PI3K isoform-selective inhibitors will provide valuable information to further validate the concept of targeting specific PI3K isoforms, while the identification of biomarkers to stratify patients who are likely to benefit from the therapy will be essential for the success of these agents. PMID:26364801

  12. Photovoltaic Program Branch annual report, FY 1989

    Energy Technology Data Exchange (ETDEWEB)

    Summers, K A [ed.

    1990-03-01

    This report summarizes the progress of the Photovoltaic (PV) Program Branch of the Solar Energy Research Institute (SERI) from October 1, 1988, through September 30, 1989. The branch is responsible for managing the subcontracted portion of SERI's PV Advanced Research and Development Project. In fiscal year (FY) 1989, this included nearly 50 subcontracts, with a total annualized funding of approximately $13.1 million. Approximately two-thirds of the subcontracts were with universities, at a total funding of nearly $4 million. The six technical sections of the report cover the main areas of the subcontracted program: Amorphous Silicon Research, Polycrystalline Thin Films, Crystalline Silicon Materials Research, High-Efficiency Concepts, New Ideas, and University Participation. Technical summaries of each of the subcontracted programs provide a discussion of approaches, major accomplishments in FY 1989, and future research directions. Each report will be cataloged individually.

  13. The Higgs Branch of Impurity Theories

    CERN Document Server

    Kapustin, A A; Kapustin, Anton; Sethi, Savdeep

    1998-01-01

    We consider supersymmetric gauge theories with impurities in various dimensions. These systems arise in the study of intersecting branes. Unlike conventional gauge theories, the Higgs branch of an impurity theory can have compact directions. For models with eight supercharges, the Higgs branch is a hyperKahler manifold given by the moduli space of solutions of certain differential equations. These equations are the dimensional reductions of self-duality equations with boundary conditions determined by the impurities. They can also be interpreted as Nahm transforms of self-duality equations on toroidally compactified spaces. We discuss the application of our results to the light-cone formulation of Yang-Mills theories and to the solution of certain N=2 d=4 gauge theories.

  14. Branch management into micropipeline joint dot

    Directory of Open Access Journals (Sweden)

    Dimitar Tyanev

    2011-11-01

    Full Text Available This paper considers problems related to hardware implementation of computational process with conditional jumps. Hardware refers to asynchronous pipeline organization at microoperational level. Exploration is dedicated to one of the tasks presented in (Tyanev, D., 2009 concerning to micropipeline controller design to control micropipeline stage into joint dot of branch algorithm. Joint dot is the point at which few preceding branches are combined. It appears inevitably into conditional jump structures and this is the reason for the actuality of its problem. Analysis of this new task is presented and request arbitration functioning principles are formulated for the incoming to joint dot requests. The arbiter is responsible for the fair choice on which depends steady peformance of separate pipeline brances. Paper also describes pipeline controller synthesis and analysis of its operation in two variants: about 2-phase and 4-phase data transfer protocol. The synthesized asynchronous arbiter scheme is invariant to the type of pipeline protocol.

  15. Branching Dynamics of Viral Information Spreading

    CERN Document Server

    Iribarren, José Luis

    2011-01-01

    Despite its importance for rumors or innovations propagation, peer-to-peer collaboration, social networking or Marketing, the dynamics of information spreading is not well understood. Since the diffusion depends on the heterogeneous patterns of human behavior and is driven by the participants' decisions, its propagation dynamics shows surprising properties not explained by traditional epidemic or contagion models. Here we present a detailed analysis of our study of real Viral Marketing campaigns where tracking the propagation of a controlled message allowed us to analyze the structure and dynamics of a diffusion graph involving over 31,000 individuals. We found that information spreading displays a non-Markovian branching dynamics that can be modeled by a two-step Bellman-Harris Branching Process that generalizes the static models known in the literature and incorporates the high variability of human behavior. It explains accurately all the features of information propagation under the "tipping-point" and can...

  16. The surface science of enzymes

    DEFF Research Database (Denmark)

    Rod, Thomas Holm; Nørskov, Jens Kehlet

    2002-01-01

    One of the largest challenges to science in the coming years is to find the relation between enzyme structure and function. Can we predict which reactions an enzyme catalyzes from knowledge of its structure-or from its amino acid sequence? Can we use that knowledge to modify enzyme function? To...... solve these problems we must understand in some detail how enzymes interact with reactants from its surroundings. These interactions take place at the surface of the enzyme and the question of enzyme function can be viewed as the surface science of enzymes. In this article we discuss how to describe...... catalysis by enzymes, and in particular the analogies between enzyme catalyzed reactions and surface catalyzed reactions. We do this by discussing two concrete examples of reactions catalyzed both in nature (by enzymes) and in industrial reactors (by inorganic materials), and show that although analogies...

  17. Characterisation of branched gluco-oligosaccharides to study the mode-of-action of a glucoamylase from Hypocrea jecorina.

    Science.gov (United States)

    Jonathan, M C; van Brussel, M; Scheffers, M S; Kabel, M A

    2015-11-01

    In the conversion of starch to fermentable glucose for bioethanol production, hydrolysis of amylopectin by α-amylases and glucoamylases is the slowest step. In this process, α-1,6-branched gluco-oligosaccharides accumulate and are slowly degraded. Glucoamylases that are able to degrade such branched oligosaccharides faster are economically beneficial. This research aimed at the isolation and characterisation of branched gluco-oligosaccharides produced from amylopectin digestion by α-amylase, to be used as substrates for comparing their degradation by glucoamylases. Branched gluco-oligosaccharides with a DP between five and twelve were purified using size exclusion chromatography. These structures were characterised after labelling with 2-aminobenzamide using UHPLC-MS(n) analysis. Further, the purified oligosaccharides were used to evaluate the mode-of-action of a glucoamylase from Hypocrea jecorina. The enzyme cleaves the α-1,4-linkage adjacent to the α-1,6-linkage at a lower rate than that of α-1,4-linkages in linear oligosaccharides. Hence, the branched gluco-oligosaccharides are a suitable substrate to evaluate glucoamylase activity on branched structures. PMID:26256324

  18. Minimum disparity estimation in controlled branching processes

    OpenAIRE

    Gonzalez, Miguel; Minuesa, Carmen; del Puerto, Ines

    2015-01-01

    Minimum disparity estimation in controlled branching processes is dealt with by assuming that the offspring law belongs to a general parametric family. Under some regularity conditions it is proved that the minimum disparity estimators proposed -based on the nonparametric maximum likelihood estimator of the offspring law when the entire family tree is observed- are consistent and asymptotic normally distributed. Moreover, it is discussed the robustness of the estimators proposed. Through a si...

  19. Modelling an observer's branch of extremal consciousness

    OpenAIRE

    Polley, L.

    2012-01-01

    Extreme-order statistics is applied to the branches of an observer in a many-worlds framework. A unitary evolution operator for a step of time is constructed, generating pseudostochastic behaviour with a power-law distribution when applied repeatedly to a particular initial state. The operator models the generation of records, their dating, the splitting of the wavefunction at quantum events, and the recalling of records by the observer. Due to the huge ensemble near an observer's end, the br...

  20. New branching rules induced by plethysm

    International Nuclear Information System (INIS)

    We derive group branching laws for formal characters of subgroups Hπ of GL(n) leaving invariant an arbitrary tensor Tπ of Young symmetry type π where π is an integer partition. The branchings GL(n) ↓ GL(n-1), GL(n) ↓ O(n) and GL(2n) ↓ Sp(2n) fixing a vector vi, a symmetric tensor gij = gji and an antisymmetric tensor fij = -fji, respectively, are obtained as special cases. All new branchings are governed by Schur function series obtained from plethysms of the Schur function sπ ≡ {π} by the basic M series of complete symmetric functions and the L = M-1 series of elementary symmetric functions. Our main technical tool is that of Hopf algebras and our main result is the derivation of a coproduct for any Schur function series obtained by plethysm from another such series. Therefrom one easily obtains π-generalized Newell-Littlewood formulae and the algebra of the formal group characters of these subgroups is established. Concrete examples and extensive tabulations are displayed for H13, H21 and H3, showing their involved and nontrivial representation theory. The nature of the subgroups is shown to be in general affine and in some instances non-reductive. We discuss the complexity of the coproduct formula and give a graphical notation to cope with it. We also discuss the way in which the group branching laws can be reinterpreted as twisted structures deformed by highly nontrivial 2-cocycles. The algebra of subgroup characters is identified as a cliffordization of the algebra of symmetric functions for GL(n) formal characters. Modification rules are beyond the scope of the present paper, but are briefly discussed

  1. Recursive algorithms, branching coefficients and applications

    CERN Document Server

    Lyakhovsky, Vladimir

    2010-01-01

    Recurrent relations for branching coefficients in affine Lie algebras integrable highest weight modules are studied. The decomposition algorithm based on the injection fan technique is adopted to the situation where the Weyl denominator becomes singular with respect to a reductive subalgebra. We study some modifications of the injection fan technique and demonstrate that it is possible to define the "subtracted fans" that play the role similar to the original ones. Possible applications of subtracted fans in CFT models are considered.

  2. Branch Retinal Vein Occlusion and Its Management

    Institute of Scientific and Technical Information of China (English)

    Desmond; Archer

    1992-01-01

    The natural course of Branch Retinal Vein Occlusion is determined by the site and completeness of the occlusion, the integrity of arterial perfusion to the affected sector and the efficiency of the developing collateral circulation. Most patients with tributary vein occlusion have some capillary fall out and microvascular incompetence in the distribution of the affected retina and vision is significantly compromised in over 50% of patients who have either chronic macular oedema or ischemia involving the...

  3. Computers in Some Branches of Applied Physiology .

    Directory of Open Access Journals (Sweden)

    S.S. Verma

    1994-04-01

    Full Text Available This paper reviews the applications of computers in the evaluation of different types of problems occuring in some branches of applied physiology. The recent applications of computers to perform advanced multivariate regression analysis for developing regression models in applied physiology are also highlighted. The regression models are practical significance for screening personnel in defence services, mines, industrial works, sports and the like.

  4. The making of a branching annelid

    OpenAIRE

    Aguado, M. Teresa; Glasby, Christopher J.; Schroeder, Paul C.; Weigert, Anne; Bleidorn, Christoph

    2015-01-01

    Ramisyllis multicaudata is a member of Syllidae (Annelida, Errantia, Phyllodocida) with a remarkable branching body plan. Using a next-generation sequencing approach, the complete mitochondrial genomes of R. multicaudata and Trypanobia sp. are sequenced and analysed, representing the first ones from Syllidae. The gene order in these two syllids does not follow the order proposed as the putative ground pattern in Errantia. The phylogenetic relationships of R. multicaudata are discerned usin...

  5. [Geometry and algebra of branches of the middle cerebral artery].

    Science.gov (United States)

    Blinkov, S M

    1986-01-01

    A classification of the cortical branches of the middle cerebral artery (MCA) is suggested by means of which each branch in any hemisphere can be qualified and identified in any variant of MCA branching. The principle of the classification consists in grouping the branches into arteries and trunks of the second, third, etc. order. Branches supplying blood to a certain sector of the lateral surface of the hemisphere are designated arteries. Their number and zone of branching are constant. Branches giving rise to 2 and more arteries are named trunks. Branching of the trunks, the number of trunks of the second, third, etc. order, and the site and type of origin of the arteries are extremely variable. Each trunk can be designated by a formula stating its order and the name of the artery supplied by this trunk. The arrangement of the MCA branches on the surface of the gyri and deep in the sulci, represented on the map of the lateral surface of the hemisphere, is designated conditionally as geometry of MCA branches. The order of branching of the trunks and the type of origin of the arteries, represented on abstract maps of the lateral surface of the hemisphere, are designated conditionally as algebra of the MCA branches. The variability of the geometry and algebra of the MCA branches must be taken into consideration in operations for extra-intracranial microanastomosis and in endovasal intervention on the MCA. PMID:3811741

  6. The Catalytic Machinery of a Key Enzyme in Amino Acid Biosynthesis

    OpenAIRE

    Ronald E. Viola; Faehnle, Christopher R.; Julio Blanco; Moore, Roger A.; Xuying Liu; Arachea, Buenafe T.; Pavlovsky, Alexander G.

    2010-01-01

    The aspartate pathway of amino acid biosynthesis is essential for all microbial life but is absent in mammals. Characterizing the enzyme-catalyzed reactions in this pathway can identify new protein targets for the development of antibiotics with unique modes of action. The enzyme aspartate β -semialdehyde dehydrogenase (ASADH) catalyzes an early branch point reaction in the aspartate pathway. Kinetic, mutagenic, and structural studies of ASADH from various microbial species have been used to ...

  7. Metabolic evolution of a deep-branching hyperthermophilic chemoautotrophic bacterium.

    Science.gov (United States)

    Braakman, Rogier; Smith, Eric

    2014-01-01

    Aquifex aeolicus is a deep-branching hyperthermophilic chemoautotrophic bacterium restricted to hydrothermal vents and hot springs. These characteristics make it an excellent model system for studying the early evolution of metabolism. Here we present the whole-genome metabolic network of this organism and examine in detail the driving forces that have shaped it. We make extensive use of phylometabolic analysis, a method we recently introduced that generates trees of metabolic phenotypes by integrating phylogenetic and metabolic constraints. We reconstruct the evolution of a range of metabolic sub-systems, including the reductive citric acid (rTCA) cycle, as well as the biosynthesis and functional roles of several amino acids and cofactors. We show that A. aeolicus uses the reconstructed ancestral pathways within many of these sub-systems, and highlight how the evolutionary interconnections between sub-systems facilitated several key innovations. Our analyses further highlight three general classes of driving forces in metabolic evolution. One is the duplication and divergence of genes for enzymes as these progress from lower to higher substrate specificity, improving the kinetics of certain sub-systems. A second is the kinetic optimization of established pathways through fusion of enzymes, or their organization into larger complexes. The third is the minimization of the ATP unit cost to synthesize biomass, improving thermodynamic efficiency. Quantifying the distribution of these classes of innovations across metabolic sub-systems and across the tree of life will allow us to assess how a tradeoff between maximizing growth rate and growth efficiency has shaped the long-term metabolic evolution of the biosphere. PMID:24516572

  8. Metabolic evolution of a deep-branching hyperthermophilic chemoautotrophic bacterium.

    Directory of Open Access Journals (Sweden)

    Rogier Braakman

    Full Text Available Aquifex aeolicus is a deep-branching hyperthermophilic chemoautotrophic bacterium restricted to hydrothermal vents and hot springs. These characteristics make it an excellent model system for studying the early evolution of metabolism. Here we present the whole-genome metabolic network of this organism and examine in detail the driving forces that have shaped it. We make extensive use of phylometabolic analysis, a method we recently introduced that generates trees of metabolic phenotypes by integrating phylogenetic and metabolic constraints. We reconstruct the evolution of a range of metabolic sub-systems, including the reductive citric acid (rTCA cycle, as well as the biosynthesis and functional roles of several amino acids and cofactors. We show that A. aeolicus uses the reconstructed ancestral pathways within many of these sub-systems, and highlight how the evolutionary interconnections between sub-systems facilitated several key innovations. Our analyses further highlight three general classes of driving forces in metabolic evolution. One is the duplication and divergence of genes for enzymes as these progress from lower to higher substrate specificity, improving the kinetics of certain sub-systems. A second is the kinetic optimization of established pathways through fusion of enzymes, or their organization into larger complexes. The third is the minimization of the ATP unit cost to synthesize biomass, improving thermodynamic efficiency. Quantifying the distribution of these classes of innovations across metabolic sub-systems and across the tree of life will allow us to assess how a tradeoff between maximizing growth rate and growth efficiency has shaped the long-term metabolic evolution of the biosphere.

  9. Measurements of $\\Xi_c^{+}$ Branching Ratios

    CERN Document Server

    Link, J M; Anjos, J C; Bediaga, I; Göbel, C; Magnin, J; Massafferri, A; De Miranda, J M; Pepe, I M; Polycarpo, E; Dos Reis, A C; Carrillo, S; Casimiro, E; Cuautle, E; Sánchez-Hernández, A; Uribe, C; Vázquez, F; Agostino, L; Cinquini, L; Cumalat, J P; O'Reilly, B; Segoni, I; Wahl, M; Butler, J N; Cheung, H W K; Chiodini, G; Gaines, I; Garbincius, P H; Garren, L A; Gottschalk, E E; Kasper, P H; Kreymer, A E; Kutschke, R; Wang, M; Benussi, L; Bertani, M; Bianco, S; Fabbri, Franco Luigi; Zallo, A; Reyes, M; Cawlfield, C; Kim, D Y; Rahimi, A; Wiss, J; Gardner, R; Kryemadhi, A; Chung, Y S; Kang, J S; Ko, B R; Kwak, J W; Lee, K B; Cho, K; Park, H; Alimonti, G; Barberis, S; Boschini, M; Cerutti, A; D'Angelo, P; Di Corato, M; Dini, P; Edera, L; Erba, S; Giammarchi, M; Inzani, P; Leveraro, F; Malvezzi, S; Menasce, D; Mezzadri, M; Moroni, L; Pedrini, D; Pontoglio, C; Prelz, F; Rovere, M; Sala, S; Davenport, T F; Arena, V; Boca, G; Bonomi, G; Gianini, G; Liguori, G; Lopes-Pegna, D; Merlo, M M; Pantea, D; Ratti, S P; Riccardi, C; Vitulo, P; Hernández, H; López, A M; Luiggi, E; Méndez, H; Paris, A; Quinones, J; Ramírez, J E; Zhang, Y; Wilson, J R; Handler, T; Mitchell, R; Engh, D; Hosack, M; Johns, W E; Nehring, M S; Sheldon, P D; Stenson, K; Vaandering, E W; Webster, M; Sheaff, M

    2003-01-01

    Using data collected by the fixed target Fermilab experiment FOCUS, we measure the branching ratios of the Cabibbo favored decays $\\Xi_c^+ \\to \\Sigma^+K^-\\pi^+$, $\\Xi_c^+ \\to \\Sigma^+ \\bar{K}^{*}(892)^0$, and $\\Xi_c^+ \\to \\Lambda^0K^-\\pi^+\\pi^+$ relative to $\\Xi_c^+ \\to \\Xi^-\\pi^+\\pi^+$ to be $0.91\\pm0.11\\pm0.04$, $0.78\\pm0.16\\pm0.06$, and $0.28\\pm0.06\\pm0.06$, respectively. We report the first observation of the Cabibbo suppressed decay $\\Xi_c^+ \\to \\Sigma^+K^+K^-$ and we measure the branching ratio relative to $\\Xi_c^+ \\to \\Sigma^+K^-\\pi^+$ to be $0.16\\pm0.06\\pm0.01$. We also set 90% confidence level upper limits for $\\Xi_c^+ \\to \\Sigma^+ \\phi$ and $\\Xi_c^+ \\to \\Xi^*(1690)^0(\\Sigma^+ K^-) K^+$ relative to $\\Xi_c^+ \\to \\Sigma^+K^-\\pi^+$ to be 0.12 and 0.05, respectively. We find an indication of the decays $\\Xi_c^+ \\to \\Omega^-K^{+}\\pi^+$ and $\\Xi_c^+ \\to \\Sigma^{*}(1385)^+ \\bar{K}^0$ and set 90% confidence level upper limits for the branching ratios with respect to $\\Xi_c^+ \\to \\Xi^-\\pi^+\\pi^+$ to be 0.12 a...

  10. Pebbles and Branching Programs for Tree Evaluation

    CERN Document Server

    Cook, Stephen; Wehr, Dustin; Braverman, Mark; Santhanam, Rahul

    2010-01-01

    We introduce the Tree Evaluation Problem, show that it is in logDCFL (and hence in P), and study its branching program complexity in the hope of eventually proving a superlogarithmic space lower bound. The input to the problem is a rooted, balanced d-ary tree of height h, whose internal nodes are labeled with d-ary functions on [k] = {1,...,k}, and whose leaves are labeled with elements of [k]. Each node obtains a value in [k] equal to its d-ary function applied to the values of its d children. The output is the value of the root. We show that the standard black pebbling algorithm applied to the binary tree of height h yields a deterministic k-way branching program with Theta(k^h) states solving this problem, and we prove that this upper bound is tight for h=2 and h=3. We introduce a simple semantic restriction called "thrifty" on k-way branching programs solving tree evaluation problems and show that the same state bound of Theta(k^h) is tight (up to a constant factor) for all h >= 2 for deterministic thrift...

  11. The root of branching river networks.

    Science.gov (United States)

    Perron, J Taylor; Richardson, Paul W; Ferrier, Ken L; Lapôtre, Mathieu

    2012-12-01

    Branching river networks are one of the most widespread and recognizable features of Earth's landscapes and have also been discovered elsewhere in the Solar System. But the mechanisms that create these patterns and control their spatial scales are poorly understood. Theories based on probability or optimality have proven useful, but do not explain how river networks develop over time through erosion and sediment transport. Here we show that branching at the uppermost reaches of river networks is rooted in two coupled instabilities: first, valleys widen at the expense of their smaller neighbours, and second, side slopes of the widening valleys become susceptible to channel incision. Each instability occurs at a critical ratio of the characteristic timescales for soil transport and channel incision. Measurements from two field sites demonstrate that our theory correctly predicts the size of the smallest valleys with tributaries. We also show that the dominant control on the scale of landscape dissection in these sites is the strength of channel incision, which correlates with aridity and rock weakness, rather than the strength of soil transport. These results imply that the fine-scale structure of branching river networks is an organized signature of erosional mechanics, not a consequence of random topology. PMID:23222614

  12. Magnetically responsive enzyme powders

    Czech Academy of Sciences Publication Activity Database

    Pospišková, K.; Šafařík, Ivo

    2015-01-01

    Roč. 380, APR 2015 (2015), s. 197-200. ISSN 0304-8853 R&D Projects: GA MŠk(CZ) LD13021 Institutional support: RVO:67179843 Keywords : enzyme powders * cross-linking * magnetic modification * magnetic separation * magnetic iron oxides particles * microwave-assisted synthesis Subject RIV: CE - Biochemistry Impact factor: 1.970, year: 2014

  13. Enzymes in Forest Soils

    Czech Academy of Sciences Publication Activity Database

    Baldrian, Petr; Štursová, Martina

    Heidelberg, Dordrecht, NY: Springer, 2011 - (Shukla, G.; Varma, A.), s. 61-73 ISBN 978-3-642-14225-3 R&D Projects: GA ČR GA526/08/0751; GA MŠk OC08050 Institutional research plan: CEZ:AV0Z50200510 Keywords : forest soils * soil ecology * enzymes Subject RIV: EE - Microbiology, Virology

  14. Enzymes of Saprotrophic Basidiomycetes

    Czech Academy of Sciences Publication Activity Database

    Baldrian, Petr

    Amsterdam: Academic Press, 2007, s. 19-41. ISBN 978-0-12-374185-1 R&D Projects: GA AV ČR KJB600200516; GA ČR GA526/05/0168; GA MŠk LC06066 Institutional research plan: CEZ:AV0Z50200510 Keywords : saprotrophic basidiomycetes * extracellular enzymes * polymers Subject RIV: EE - Microbiology, Virology

  15. Computational enzyme design

    Science.gov (United States)

    Bolon, Daniel N.

    2002-08-01

    The long-term objective of computational enzyme design is the ability to generate efficient protein catalysts for any chemical reaction. This thesis develops and experimentally validates a general computational approach for the design of enzymes with novel function. In order to include catalytic mechanism in protein design, a high-energy state (HES) rotamer (side chain representation) was constructed. In this rotamer, substrate atoms are in a HES. In addition, at least one amino acid side chain is positioned to interact favorably with substrate atoms in their HES and facilitate the reaction. Including an amino acid side chain in the HES rotamer automatically positions substrate relative to a protein scaffold and allows protein design algorithms to search for sequences capable of interacting favorably with the substrate. Because chemical similarity exists between the transition state and the high-energy state, optimizing the protein sequence to interact favorably with the HES rotamer should lead to transition state stabilization. In addition, the HES rotamer model focuses the subsequent computational active site design on a relevant phase space where an amino acid is capable of interacting in a catalytically active geometry with substrate. Using a HES rotamer model of the histidine mediated nucleophilic hydrolysis of p-nitrophenyl acetate, the catalytically inert 108 residue E. coli thioredoxin as a scaffold, and the ORBIT protein design software to compute sequences, an active site scan identified two promising active site designs. Experimentally, both candidate ?protozymes? demonstrated catalytic activity significantly above background. In addition, the rate enhancement of one of these ?protozymes? was the same order of magnitude as the first catalytic antibodies. Because polar groups are frequently buried at enzyme-substrate interfaces, improved modeling of buried polar interactions may benefit enzyme design. By studying native protein structures, rules have been

  16. A specific isoform of poly(ADP-ribose) glycohydrolase is targeted to the mitochondrial matrix by a N-terminal mitochondrial targeting sequence

    International Nuclear Information System (INIS)

    Poly(ADP-ribose) polymerases (PARPs) convert NAD to polymers of ADP-ribose that are converted to free ADP-ribose by poly(ADP-ribose) glycohydrolase (PARG). The activation of the nuclear enzyme PARP-1 following genotoxic stress has been linked to release of apoptosis inducing factor from the mitochondria, but the mechanisms by which signals are transmitted between nuclear and mitochondrial compartments are not well understood. The study reported here has examined the relationship between PARG and mitochondria in HeLa cells. Endogenous PARG associated with the mitochondrial fraction migrated in the range of 60 kDa. Transient transfection of cells with PARG expression constructs with amino acids encoded by exon 4 at the N-terminus was targeted to the mitochondria as demonstrated by subcellular fractionation and immunofluorescence microscopy of whole cells. Deletion and missense mutants allowed identification of a canonical N-terminal mitochondrial targeting sequence consisting of the first 16 amino acids encoded by PARG exon 4. Sub-mitochondrial localization experiments indicate that this mitochondrial PARG isoform is targeted to the mitochondrial matrix. The identification of a PARG isoform as a component of the mitochondrial matrix raises several interesting possibilities concerning mechanisms of nuclear-mitochondrial cross talk involved in regulation of cell death pathways.

  17. High-throughput screening of inhibitory effects of Bo-yang-hwan-o-tang on human cytochrome P450 isoforms in vitro using UPLC/MS/MS.

    Science.gov (United States)

    Lee, Miran; Park, Jeonghyeon; Lim, Mi-sun; Seong, Sook Jin; Lee, Joomi; Seo, Jeong Ju; Park, Yong-Ki; Lee, Hae Won; Yoon, Young-Ran

    2012-01-01

    Bo-yang-hwan-o-tang (BHT) is an oriental herbal medicine for treating brain disorders such as cerebral ischemia. The objective of this study was to develop an economically feasible and time-saving high-throughput screening method to monitor the potential inhibitory effects of BHT on human cytochrome P450 (CYP) enzymes in vitro. Two cocktail sets were used for incubation of human liver microsomes: Cocktail A: 6 probe substrates for CYP1A2, CYP2A6, CYP2C8, CYP2C19, CYP2D6, CYP3A4; Cocktail B: 3 for CYP2B6, CYP2C9, CYP2E1. The concentrations of the substrate metabolites were simultaneously analyzed using UPLC/MS/MS. The BHT extract had almost negligible inhibitory effects on the nine human CYP isoforms tested, with the half-maximal inhibitory concentration value ranged from 3624.99 to 45412.44 μg/ml. The results suggest that BHT extract has no inhibitory effects on CYP isoforms within the clinically recommended dosage range. We conclude that BHT might be free of drug-herb interactions when co-administered with other medicines. However, more in vivo human studies are needed to confirm these results. The high-throughput screening method can be a useful tool for drug discovery and for understanding drug interactions. PMID:23232241

  18. Male-specific Fruitless isoforms have different regulatory roles conferred by distinct zinc finger DNA binding domains

    OpenAIRE

    Dalton, Justin E.; Fear, Justin M.; Knott, Simon; Baker, Bruce S.; McIntyre, Lauren M.; Arbeitman, Michelle N.

    2013-01-01

    Background Drosophila melanogaster adult males perform an elaborate courtship ritual to entice females to mate. fruitless (fru), a gene that is one of the key regulators of male courtship behavior, encodes multiple male-specific isoforms (FruM). These isoforms vary in their carboxy-terminal zinc finger domains, which are predicted to facilitate DNA binding. Results By over-expressing individual FruM isoforms in fru-expressing neurons in either males or females and assaying the global transcri...

  19. Functional Analysis of the Short Isoform of Orf Virus Protein OV20.0

    Science.gov (United States)

    Tseng, Yeu-Yang; Lin, Fong-Yuan; Cheng, Sun-Fang; Chulakasian, Songkhla; Chou, Chia-Chi; Liu, Ya-Fen; Chang, Wei-Shan; Wong, Min-Liang

    2015-01-01

    ABSTRACT Orf virus (ORFV) OV20.0L is an ortholog of vaccinia virus (VACV) gene E3L. The function of VACV E3 protein as a virulence factor is well studied, but OV20.0 has received less attention. Here we show that like VACV E3L, OV20.0L encodes two proteins, a full-length protein and a shorter form (sh20). The shorter sh20 is an N-terminally truncated OV20.0 isoform generated when a downstream AUG codon is used for initiating translation. These isoforms differed in cellular localization, with full-length OV20.0 and sh20 found throughout the cell and predominantly in the cytoplasm, respectively. Nonetheless, both OV20.0 isoforms were able to bind double-stranded RNA (dsRNA)-activated protein kinase (PKR) and dsRNA. Moreover, both isoforms strongly inhibited PKR activation as shown by decreased phosphorylation of the translation initiation factor eIF2α subunit and protection of Sindbis virus infection against the activity of interferon (IFN). In spite of this apparent conservation of function in vitro, a recombinant ORFV that was able to express only the sh20 isoform was attenuated in a mouse model. IMPORTANCE The OV20.0 protein of orf virus (ORFV) has two isoforms and contributes to virulence, but the roles of the two forms are not known. This study shows that the shorter isoform (sh20) arises due to use of a downstream initiation codon and is amino-terminally truncated. The sh20 form also differs in expression kinetics and cellular localization from full-length OV20.0. Similar to the full-length isoform, sh20 is able to bind dsRNA and PKR, inactivate PKR, and thus act as an antagonist of the interferon response in vitro. In vivo, however, wild-type OV20.0 could not be replaced with sh20 alone without a loss of virulence, suggesting that the functions of the isoforms are not simply redundant. PMID:25694596

  20. Evaluation of allowable external pressure for branch pipe connections

    International Nuclear Information System (INIS)

    Although the allowable external pressure for branch pipe connections is estimated by evaluation procedure for straight pipe, the applicability of this procedure has not been clear. That was discussed in this study. The buckling pressure of branch pipe connections was estimated by linear eigenvalue analysis with FEA (Finite Element Analysis). The types of branch pipe connections were T-joints and T-pipes. The ranges of configurations in branch pipe connections for FEA were determined by survey of branch pipe connections in the Japanese nuclear power plants. The results of the FEA showed that the buckling strength of branch pipe was found sufficiently to be superior to that of straight piping from comparisons of estimations by FEA for branch pipe connections and those by formula for straight pipe in the design code. It was concluded that the formula for straight pipe is applicable for the evaluation of allowable external pressure for branch pipe connections. (author)

  1. Differentiation of crack branching types in fractured glass

    Science.gov (United States)

    Rabinovitch, A.; Bahat, D.

    2011-07-01

    The influence of different physical factors on the fracture branching phenomenon was investigated on a fractured brittle (glass) bottle. A fully branched fracture tree consisting of 144 initiation sites was induced. Branching sites were concentrated in two zones: close to the fracture origin and further away from it along the bottle axis, which was shown to be related to reflection of transverse acoustic waves from the bottle's shoulder. Two types of branching formed, complete full branches (FB), and attempted branching (AB) when one branch was shorter than 5 mm. Branching in the first zone was shown to be regulated by both the distribution of flaw lengths and the distances from each FB to its nearest fracture neighbor. Fracture spacing was found to be proportional to the stress at the propagating fracture tips.

  2. Insulin receptor isoforms: an integrated view focused on gestational diabetes mellitus.

    Science.gov (United States)

    Westermeier, F; Sáez, T; Arroyo, P; Toledo, F; Gutiérrez, J; Sanhueza, C; Pardo, F; Leiva, A; Sobrevia, L

    2016-05-01

    The human insulin receptor (IR) exists in two isoforms that differ by the absence (IR-A) or the presence (IR-B) of a 12-amino acid segment encoded by exon 11. Both isoforms are functionally distinct regarding their binding affinities and intracellular signalling. However, the underlying mechanisms related to their cellular functions in several tissues are only partially understood. In this review, we summarize the current knowledge in this field regarding the alternative splicing of IR isoform, tissue-specific distribution and signalling both in physiology and disease, with an emphasis on the human placenta in gestational diabetes mellitus (GDM). Furthermore, we discuss the clinical relevance of IR isoforms highlighted by findings that show altered insulin signalling due to differential IR-A and IR-B expression in human placental endothelium in GDM pregnancies. Future research and clinical studies focused on the role of IR isoform signalling might provide novel therapeutic targets for treating GDM to improve the adverse maternal and neonatal outcomes. Copyright © 2015 John Wiley & Sons, Ltd. PMID:26431063

  3. Nesprin-2 epsilon: A novel nesprin isoform expressed in human ovary and Ntera-2 cells

    Energy Technology Data Exchange (ETDEWEB)

    Lam, Le Thanh [Wolfson Centre for Inherited Neuromuscular Disease, RJAH Orthopaedic Hospital, Oswestry SY10 7AG (United Kingdom); Boehm, Sabrina V.; Roberts, Roland G. [Department of Medical and Molecular Genetics, King' s College London, London SE1 9RT (United Kingdom); Morris, Glenn E., E-mail: glennmanc@hotmail.com [Wolfson Centre for Inherited Neuromuscular Disease, RJAH Orthopaedic Hospital, Oswestry SY10 7AG (United Kingdom); Institute of Science and Technology in Medicine, Keele University, Keele ST5 5BG (United Kingdom)

    2011-08-26

    Highlights: {yields} A novel epsilon isoform of nesprin-2 has been discovered. {yields} This 120 kDa protein was predicted by bioinformatic analysis, but has not previously been observed. {yields} It is the main isoform expressed in a teratocarcinoma cell line and is also found in ovary. {yields} Like other nesprins, it is located at the nuclear envelope. {yields} We suggest it may have a role in very early development or in some ovary-specific function. -- Abstract: The nuclear envelope-associated cytoskeletal protein, nesprin-2, is encoded by a large gene containing several internal promoters that produce shorter isoforms. In a study of Ntera-2 teratocarcinoma cells, a novel isoform, nesprin-2-epsilon, was found to be the major mRNA and protein product of the nesprin-2 gene. Its existence was predicted by bioinformatic analysis, but this is the first direct demonstration of both the mRNA and the 120 kDa protein which is located at the nuclear envelope. In a panel of 21 adult and foetal human tissues, the nesprin-2-epsilon mRNA was strongly expressed in ovary but was a minor isoform elsewhere. The expression pattern suggests a possible link with very early development and a likely physiological role in ovary.

  4. Most highly expressed protein-coding genes have a single dominant isoform.

    Science.gov (United States)

    Ezkurdia, Iakes; Rodriguez, Jose Manuel; Carrillo-de Santa Pau, Enrique; Vázquez, Jesús; Valencia, Alfonso; Tress, Michael L

    2015-04-01

    Although eukaryotic cells express a wide range of alternatively spliced transcripts, it is not clear whether genes tend to express a range of transcripts simultaneously across cells, or produce dominant isoforms in a manner that is either tissue-specific or regardless of tissue. To date, large-scale investigations into the pattern of transcript expression across distinct tissues have produced contradictory results. Here, we attempt to determine whether genes express a dominant splice variant at the protein level. We interrogate peptides from eight large-scale human proteomics experiments and databases and find that there is a single dominant protein isoform, irrespective of tissue or cell type, for the vast majority of the protein-coding genes in these experiments, in partial agreement with the conclusions from the most recent large-scale RNAseq study. Remarkably, the dominant isoforms from the experimental proteomics analyses coincided overwhelmingly with the reference isoforms selected by two completely orthogonal sources, the consensus coding sequence variants, which are agreed upon by separate manual genome curation teams, and the principal isoforms from the APPRIS database, predicted automatically from the conservation of protein sequence, structure, and function. PMID:25732134

  5. Computational Estimates of Binding Affinities for Estrogen Receptor Isoforms in Rainbow Trout

    CERN Document Server

    Shyu, Conrad

    2009-01-01

    Molecular dynamics simulations are performed to determine the binding affinities between the hormone 17 beta-estradiol (E2) and different estrogen receptor (ER) isoforms in the rainbow trout (Oncorhynchus mykiss). Previous studies have demonstrated that a recent, unique gene duplication of the ER alpha subtype created two isoforms ER alpha 1 and ER alpha 2, and an early secondary split of ER beta produced two distinct isoforms of ER beta 1 and ER beta 2 based on the phylogenetic analysis. The objective of our computational studies is to provide insight into the underlying evolutionary selection pressure on the ER isoforms. Our results show that E2 binds preferentially to ER alpha 1. This finding corresponds to the experimental results as the ERs evolved from gene duplication events are frequently free from selective pressure and should exhibit no deleterious effects. The E2, however, only binds slightly better to ER beta 2. Both isoforms remain competitive. This finding reflects the fact that since ER beta 2 ...

  6. Electrophoretic Mobility of Cardiac Myosin Heavy Chain Isoforms Revisited: Application of MALDI TOF/TOF Analysis

    Directory of Open Access Journals (Sweden)

    Petra Arnostova

    2011-01-01

    Full Text Available The expression of two cardiac myosin heavy chain (MyHC isoforms in response to the thyroid status was studied in left ventricles (LVs of Lewis rats. Major MyHC isoform in euthyroid and hyperthyroid LVs had a higher mobility on SDS-PAGE, whereas hypothyroid LVs predominantly contained a MyHC isoform with a lower mobility corresponding to that of the control soleus muscle. By comparing the MyHC profiles obtained under altered thyroid states together with the control soleus, we concluded that MyHCα was represented by the lower band with higher mobility and MyHCβ by the upper band. The identity of these two bands in SDS-PAGE gels was confirmed by western blot and mass spectrometry. Thus, in contrast to the literature data, we found that the MyHCα possessed a higher mobility rate than the MyHCβ isoform. Our data highlighted the importance of the careful identification of the MyHCα and MyHCβ isoforms analyzed by the SDS-PAGE.

  7. Nesprin-2 epsilon: A novel nesprin isoform expressed in human ovary and Ntera-2 cells

    International Nuclear Information System (INIS)

    Highlights: → A novel epsilon isoform of nesprin-2 has been discovered. → This 120 kDa protein was predicted by bioinformatic analysis, but has not previously been observed. → It is the main isoform expressed in a teratocarcinoma cell line and is also found in ovary. → Like other nesprins, it is located at the nuclear envelope. → We suggest it may have a role in very early development or in some ovary-specific function. -- Abstract: The nuclear envelope-associated cytoskeletal protein, nesprin-2, is encoded by a large gene containing several internal promoters that produce shorter isoforms. In a study of Ntera-2 teratocarcinoma cells, a novel isoform, nesprin-2-epsilon, was found to be the major mRNA and protein product of the nesprin-2 gene. Its existence was predicted by bioinformatic analysis, but this is the first direct demonstration of both the mRNA and the 120 kDa protein which is located at the nuclear envelope. In a panel of 21 adult and foetal human tissues, the nesprin-2-epsilon mRNA was strongly expressed in ovary but was a minor isoform elsewhere. The expression pattern suggests a possible link with very early development and a likely physiological role in ovary.

  8. Quantitative profiling of Drosophila melanogaster Dscam1 isoforms reveals no changes in splicing after bacterial exposure.

    Directory of Open Access Journals (Sweden)

    Sophie A O Armitage

    Full Text Available The hypervariable Dscam1 (Down syndrome cell adhesion molecule 1 gene can produce thousands of different ectodomain isoforms via mutually exclusive alternative splicing. Dscam1 appears to be involved in the immune response of some insects and crustaceans. It has been proposed that the diverse isoforms may be involved in the recognition of, or the defence against, diverse parasite epitopes, although evidence to support this is sparse. A prediction that can be generated from this hypothesis is that the gene expression of specific exons and/or isoforms is influenced by exposure to an immune elicitor. To test this hypothesis, we for the first time, use a long read RNA sequencing method to directly investigate the Dscam1 splicing pattern after exposing adult Drosophila melanogaster and a S2 cell line to live Escherichia coli. After bacterial exposure both models showed increased expression of immune-related genes, indicating that the immune system had been activated. However there were no changes in total Dscam1 mRNA expression. RNA sequencing further showed that there were no significant changes in individual exon expression and no changes in isoform splicing patterns in response to bacterial exposure. Therefore our studies do not support a change of D. melanogaster Dscam1 isoform diversity in response to live E. coli. Nevertheless, in future this approach could be used to identify potentially immune-related Dscam1 splicing regulation in other host species or in response to other pathogens.

  9. Two isoforms of aquaporin 2 responsive to hypertonic stress in the bottlenose dolphin.

    Science.gov (United States)

    Suzuki, Miwa; Wakui, Hitomi; Itou, Takuya; Segawa, Takao; Inoshima, Yasuo; Maeda, Ken; Kikuchi, Kiyoshi

    2016-04-15

    This study investigated the expression of aquaporin 2 (AQP2) and its newly found alternatively spliced isoform (alternative AQP2) and the functions of these AQP2 isoforms in the cellular hyperosmotic tolerance in the bottlenose dolphin, ITALIC! Tursiops truncatus mRNA sequencing revealed that alternative AQP2 lacks the fourth exon and instead has a longer third exon that includes a part of the original third intron. The portion of the third intron, now part of the coding region of alternative AQP2, is highly conserved among many species of the order Cetacea but not among terrestrial mammals. Semi-quantitative PCR revealed that AQP2 was expressed only in the kidney, similar to terrestrial mammals. In contrast, alternative AQP2 was expressed in all organs examined, with strong expression in the kidney. In cultured renal cells, expression of both AQP2 isoforms was upregulated by the addition to the medium of NaCl but not by the addition of mannitol, indicating that the expression of both isoforms is induced by hypersalinity. Treatment with small interfering RNA for both isoforms resulted in a decrease in cell viability in hypertonic medium (500 mOsm kg(-1)) when compared with controls. These findings indicate that the expression of alternatively spliced AQP2 is ubiquitous in cetacean species, and it may be one of the molecules important for cellular osmotic tolerance throughout the body. PMID:26944501

  10. The relationship between gene isoform multiplicity, number of exons and protein divergence.

    Directory of Open Access Journals (Sweden)

    Jordi Morata

    Full Text Available At present we know that phenotypic differences between organisms arise from a variety of sources, like protein sequence divergence, regulatory sequence divergence, alternative splicing, etc. However, we do not have yet a complete view of how these sources are related. Here we address this problem, studying the relationship between protein divergence and the ability of genes to express multiple isoforms. We used three genome-wide datasets of human-mouse orthologs to study the relationship between isoform multiplicity co-occurrence between orthologs (the fact that two orthologs have more than one isoform and protein divergence. In all cases our results showed that there was a monotonic dependence between these two properties. We could explain this relationship in terms of a more fundamental one, between exon number of the largest isoform and protein divergence. We found that this last relationship was present, although with variations, in other species (chimpanzee, cow, rat, chicken, zebrafish and fruit fly. In summary, we have identified a relationship between protein divergence and isoform multiplicity co-occurrence and explained its origin in terms of a simple gene-level property. Finally, we discuss the biological implications of these findings for our understanding of inter-species phenotypic differences.

  11. iReckon: simultaneous isoform discovery and abundance estimation from RNA-seq data.

    Science.gov (United States)

    Mezlini, Aziz M; Smith, Eric J M; Fiume, Marc; Buske, Orion; Savich, Gleb L; Shah, Sohrab; Aparicio, Sam; Chiang, Derek Y; Goldenberg, Anna; Brudno, Michael

    2013-03-01

    High-throughput RNA sequencing (RNA-seq) promises to revolutionize our understanding of genes and their role in human disease by characterizing the RNA content of tissues and cells. The realization of this promise, however, is conditional on the development of effective computational methods for the identification and quantification of transcripts from incomplete and noisy data. In this article, we introduce iReckon, a method for simultaneous determination of the isoforms and estimation of their abundances. Our probabilistic approach incorporates multiple biological and technical phenomena, including novel isoforms, intron retention, unspliced pre-mRNA, PCR amplification biases, and multimapped reads. iReckon utilizes regularized expectation-maximization to accurately estimate the abundances of known and novel isoforms. Our results on simulated and real data demonstrate a superior ability to discover novel isoforms with a significantly reduced number of false-positive predictions, and our abundance accuracy prediction outmatches that of other state-of-the-art tools. Furthermore, we have applied iReckon to two cancer transcriptome data sets, a triple-negative breast cancer patient sample and the MCF7 breast cancer cell line, and show that iReckon is able to reconstruct the complex splicing changes that were not previously identified. QT-PCR validations of the isoforms detected in the MCF7 cell line confirmed all of iReckon's predictions and also showed strong agreement (r(2) = 0.94) with the predicted abundances. PMID:23204306

  12. Secretion of PDGF isoforms during osteoclastogenesis and its modulation by anti-osteoclast drugs.

    Science.gov (United States)

    Rahman, M Motiur; Matsuoka, Kazuhiko; Takeshita, Sunao; Ikeda, Kyoji

    2015-06-26

    In an attempt to identify secretory products of osteoclasts that mediate the coupling of bone formation to resorption, we found that along with osteoclast differentiation, PDGF-A gene expression increase occurred first, by 12 h after stimulation of bone marrow macrophages with M-CSF and RANKL, and peaked at 36 h. This was next followed by a progressive increase in PDGF-B gene expression until a peak at 60 h, when mature osteoclasts formed. Isoform-specific ELISA of the conditioned medium collected every 24 h revealed that all three of the isoforms of PDGF-AA, AB and BB were secreted, in this temporal order as differentiation proceeded. Their secretion was enhanced when osteoclasts were activated by placing them on dentin slices. The secretion of all three isoforms was decreased in cathepsin K-deficient osteoclasts compared with wild-type osteoclasts. Pharmacological inhibition of cathepsin K with odanacatib also inhibited the secretion of all three isoforms, as was also the case with alendronate treatment. The secretion of sphingosine-1-phosphate, which increased during osteoclastogenesis, was reduced from cathepsin K-deficient osteoclasts, and was inhibited by treatment with odanacatib more profoundly than with alendronate. Thus, all three isoforms of PDGF, which are secreted at distinct differentiation stages of osteoclasts, appear to have distinct roles in the cell-cell communication that takes place in the microenvironment of bone remodeling, especially from the osteoclast lineage to mesenchymal cells and vascular cells, thereby stimulating osteogenesis and angiogenesis. PMID:25951977

  13. Thermal Denaturation and Aggregation of Myosin Subfragment 1 Isoforms with Different Essential Light Chains

    Directory of Open Access Journals (Sweden)

    Eugene O. Zubov

    2010-10-01

    Full Text Available We compared thermally induced denaturation and aggregation of two isoforms of the isolated myosin head (myosin subfragment 1, S1 containing different “essential” (or “alkali” light chains, A1 or A2. We applied differential scanning calorimetry (DSC to investigate the domain structure of these two S1 isoforms. For this purpose, a special calorimetric approach was developed to analyze the DSC profiles of irreversibly denaturing multidomain proteins. Using this approach, we revealed two calorimetric domains in  the S1 molecule, the more thermostable domain denaturing in two steps. Comparing the DSC data with temperature dependences of intrinsic fluorescence parameters and S1 ATPase inactivation, we have identified these two calorimetric domains as motor domain and regulatory domain of the myosin head, the motor domain being more thermostable. Some difference between the two S1 isoforms was only revealed by DSC in thermal denaturation of the regulatory domain. We also applied dynamic light scattering (DLS to analyze the aggregation of S1 isoforms induced by their thermal denaturation. We have found no appreciable difference between these S1 isoforms in their aggregation properties under ionic strength conditions close to those in the muscle fiber (in the presence of 100 mM KCl. Under these conditions kinetics of this process was independent of protein concentration, and the aggregation rate was limited by irreversible denaturation of the S1 motor domain.

  14. Alterations of Lymphoid Enhancer Factor-1 Isoform Expression in Solid Tumors and Acute Leukemias

    Institute of Scientific and Technical Information of China (English)

    Wenbing WANG; Carsten M(U)LLER-TIDOW; Ping JI; Bj(o)rn STEFFEN; Ralf METZGER; Paul M. SCHNEIDER; Hartmut HALFTER; Mark SCHRADER; Wolfgang E. BERDEL; Hubert SERVE

    2005-01-01

    Two major transcripts of lymphoid enhancer factor-1 (LEF-1) have been described. The long isoform with β-catenin binding domain functions as a transcriptional enhancer factor. The short isoform derives from an intronic promoter and exhibits dominant negative activity. Recently, alterations of LEF- 1isoforms distribution have been described in colon cancer. In the current study we employed a quantitative real-time reverse transcription PCR method (TaqMan) to analyze expression of LEF-1 isoforms in a large cohort of human tumor (n=304) and tumor-free control samples (n=56). The highest expression level of LEF-1 was found in carcinoma samples whereas brain cancer samples expressed little. Expression of LEF1 was different in distinct cancer types. For example, the mRNA level of LEF-1 was lower in testicular tumor samples compared with tumor-free control samples. Besides epithelial cancers, significant LEF-1expression was also found in hematopoietic cells. In hematological malignancies, overall LEF-1 level was higher in lymphocytic leukemias compared with myeloid leukemias and normal hematopoiesis. However,acute myeloid leukemia and acute lymphocytic leukemia showed a significantly increased fraction of the oncogenic LEF-1 compared with chronic lymphocytic leukemia and chronic myeloid leukemia. Taken together,these data suggest that LEF-1 is abundantly expressed in human tumors and the ratio of the oncogenic and the dominant negative short isoform altered not only in carcinomas but also in leukemia.

  15. Serum amyloid A isoforms in serum and synovial fluid from spontaneously diseased dogs with joint diseases or other conditions

    DEFF Research Database (Denmark)

    Kjelgaard-Hansen, Mads Jens; Christensen, Michelle B.; Lee, Marcel Huisung;

    2007-01-01

    in samples obtained from dogs (n = 16) suffering from different inflammatory or non-inflammatory conditions, which were either related or unrelated to joints. Expression of SAA isoforms was visualized by denaturing isoelectric focusing and Western blotting. Serum amyloid A was present in serum from all dogs...... with systemic inflammatory activity, and up to four major isoforms with apparent isoelectric points between 6.1 and 7.9 were identified. In synovial fluid from inflamed joints one or more highly alkaline SAA isoforms (with apparent isoelectric points above 9.3) were identified, with data suggesting local...... production of these isoforms in the canine inflamed joint....

  16. Glucocorticoid receptor translational isoforms underlie maturational stage-specific glucocorticoid sensitivities of dendritic cells in mice and humans

    OpenAIRE

    Cao, Yun; Bender, Ingrid K.; Konstantinidis, Athanasios K.; Shin, Soon Cheon; Jewell, Christine M.; Cidlowski, John A; Schleimer, Robert P.; Lu, Nick Z.

    2013-01-01

    Mature, but not immature, dendritic cells are sensitive to glucocorticoid-induced apoptosis.Mature, but not immature, dendritic cells express proapoptotic glucocorticoid receptor translational isoforms.

  17. Heterodimeric l-amino acid oxidase enzymes from Egyptian Cerastes cerastes venom: Purification, biochemical characterization and partial amino acid sequencing

    Directory of Open Access Journals (Sweden)

    A.E. El Hakim

    2015-12-01

    Full Text Available Two l-amino acid oxidase enzyme isoforms, Cc-LAAOI and Cc-LAAOII were purified to apparent homogeneity from Cerastes cerastes venom in a sequential two-step chromatographic protocol including; gel filtration and anion exchange chromatography. The native molecular weights of the isoforms were 115 kDa as determined by gel filtration on calibrated Sephacryl S-200 column, while the monomeric molecular weights of the enzymes were, 60, 56 kDa and 60, 53 kDa for LAAOI and LAAOII, respectively. The tryptic peptides of the two isoforms share high sequence homology with other snake venom l-amino acid oxidases. The optimal pH and temperature values of Cc-LAAOI and Cc-LAAOII were 7.8, 50 °C and 7, 60 °C, respectively. The two isoenzymes were thermally stable up to 70 °C. The Km and Vmax values were 0.67 mM, 0.135 μmol/min for LAAOI and 0.82 mM, 0.087 μmol/min for LAAOII. Both isoenzymes displayed high catalytic preference to long-chain, hydrophobic and aromatic amino acids. The Mn2+ ion markedly increased the LAAO activity for both purified isoforms, while Na+, K+, Ca2+, Mg2+ and Ba2+ ions showed a non-significant increase in the enzymatic activity of both isoforms. Furthermore, Zn2+, Ni2+, Co2+, Cu2+ and AL3+ ions markedly inhibited the LAAOI and LAAOII activities. l-Cysteine and reduced glutathione completely inhibited the LAAO activity of both isoenzymes, whereas, β-mercaptoethanol, O-phenanthroline and PMSF completely inhibited the enzymatic activity of LAAOII. Furthermore, iodoacitic acid inhibited the enzymatic activity of LAAOII by 46% and had no effect on the LAAOI activity.

  18. Calcium as a branching signal in Neurospora crassa.

    OpenAIRE

    Reissig, J L; Kinney, S G

    1983-01-01

    The divalent cation ionophore A23187 was found to induce apical branching in Neurospora crassa. Optimal effects were obtained by treatment with 0.1 mM ionophore for 30 min. Branching first became manifest during or shortly after treatment; successive rounds of branching could be observed at later times. Calcium starvation of the mycelium markedly reduced its subsequent response to the ionophore, whereas starvation for other divalent cations had no detectable effect. The branching response was...

  19. Statistics of branched flow in a weak correlated random potential

    OpenAIRE

    Kaplan, Lev

    2002-01-01

    Recent images of electron flow through a two-dimensional electron gas (2DEG) device show branching behavior that is reproduced in numerical simulations of motion in a correlated random potential [cond-mat/0010348]. We show how such branching naturally arises from caustics in the classical flow and find a simple scaling behavior of the branching under variation of the random potential strength. Analytic results describing the statistical properties of the branching are confirmed by classical a...

  20. Characterizing Weighted MSO for Trees by Branching Transitive Closure Logics

    OpenAIRE

    Fülöp, Zoltán; Vogler, Heiko

    2012-01-01

    We introduce the branching transitive closure operator on weighted monadic second-order logic formulas where the branching corresponds in a natural way to the branching inherent in trees. For arbitrary commutative semirings, we prove that weighted monadic second order logics on trees is equivalent to the definability by formulas which start with one of the following operators: (i) a branching transitive closure or (ii) an existential second-order quantifier followed by one universal first-ord...

  1. Exploration of Novel Inhibitors for Class I Histone Deacetylase Isoforms by QSAR Modeling and Molecular Dynamics Simulation Assays.

    Directory of Open Access Journals (Sweden)

    Zainab Noor

    Full Text Available Histone deacetylases (HDAC are metal-dependent enzymes and considered as important targets for cell functioning. Particularly, higher expression of class I HDACs is common in the onset of multiple malignancies which results in deregulation of many target genes involved in cell growth, differentiation and survival. Although substantial attempts have been made to control the irregular functioning of HDACs by employing various inhibitors with high sensitivity towards transformed cells, limited success has been achieved in epigenetic cancer therapy. Here in this study, we used ligand-based pharmacophore and 2-dimensional quantitative structure activity relationship (QSAR modeling approaches for targeting class I HDAC isoforms. Pharmacophore models were generated by taking into account the known IC50 values and experimental energy scores with extensive validations. The QSAR model having an external R2 value of 0.93 was employed for virtual screening of compound libraries. 10 potential lead compounds (C1-C10 were short-listed having strong binding affinities for HDACs, out of which 2 compounds (C8 and C9 were able to interact with all members of class I HDACs. The potential binding modes of HDAC2 and HDAC8 to C8 were explored through molecular dynamics simulations. Overall, bioactivity and ligand efficiency (binding energy/non-hydrogen atoms profiles suggested that proposed hits may be more effective inhibitors for cancer therapy.

  2. 40 CFR 721.10094 - Decene, branched and linear.

    Science.gov (United States)

    2010-07-01

    ... 40 Protection of Environment 30 2010-07-01 2010-07-01 false Decene, branched and linear. 721.10094... Substances § 721.10094 Decene, branched and linear. (a) Chemical substance and significant new uses subject to reporting. (1) The chemical substance identified as decene, branched and linear (PMN P-03-272;...

  3. Targeted mutations in the Na,K-ATPase α 2 isoform confer ouabain resistance and result in abnormal behavior in mice.

    Science.gov (United States)

    Schaefer, Tori L; Lingrel, Jerry B; Moseley, Amy E; Vorhees, Charles V; Williams, Michael T

    2011-06-01

    Sodium and potassium-activated adenosine triphosphatases (Na,K-ATPase) are ubiquitous, participate in osmotic balance and membrane potential, and are composed of α, β, and γ subunits. The α subunit is required for the catalytic and transport properties of the enzyme and contains binding sites for cations, ATP, and digitalis-like compounds including ouabain. There are four known α isoforms; three that are expressed in the CNS in a regional and cell-specific manner. The α2 isoform is most commonly found in astrocytes, pyramidal cells of the hippocampus in adults, and developmentally in several other neuronal types. Ouabain-like compounds are thought to be produced endogenously in mammals, bind the Na,K-ATPase, and function as a stress-related hormone, however, the impact of the Na,K-ATPase ouabain binding site on neurobehavioral function is largely unknown. To determine if the ouabain binding site of the α2 isoform plays a physiological role in CNS function, we examined knock-in mice in which the normally ouabain-sensitive α2 isoform was made resistant (α2(R/R) ) while still retaining basal Na,K-ATPase enzymatic function. Egocentric learning (Cincinnati water maze) was impaired in adult α2(R/R) mice compared to wild type (WT) mice. They also exhibited decreased locomotor activity in a novel environment and increased responsiveness to a challenge with an indirect sympathomimetic agonist (methamphetamine) relative to WT mice. The α2(R/R) mice also demonstrated a blunted acoustic startle reflex and a failure to habituate to repeated acoustic stimuli. The α2(R/R) mice showed no evidence of altered anxiety (elevated zero maze) nor were they impaired in spatial learning or memory in the Morris water maze and neither group could learn in a large Morris maze. These results suggest that the ouabain binding site is involved in specific types of learning and the modulation of dopamine-mediated locomotor behavior. PMID:20936682

  4. Antibody-Mediated Inhibition of the FGFR1c Isoform Induces a Catabolic Lean State in Siberian Hamsters.

    Science.gov (United States)

    Samms, Ricardo J; Lewis, Jo E; Lory, Alex; Fowler, Maxine J; Cooper, Scott; Warner, Amy; Emmerson, Paul; Adams, Andrew C; Luckett, Jeni C; Perkins, Alan C; Wilson, Dana; Barrett, Perry; Tsintzas, Kostas; Ebling, Francis J P

    2015-11-16

    Hypothalamic tanycytes are considered to function as sensors of peripheral metabolism. To facilitate this role, they express a wide range of receptors, including fibroblast growth factor receptor 1 (FGFR1). Using a monoclonal antibody (IMC-H7) that selectively antagonizes the FGFR1c isoform, we investigated possible actions of FGFR1c in a natural animal model of adiposity, the Siberian hamster. Infusion of IMC-H7 into the third ventricle suppressed appetite and increased energy expenditure. Likewise, peripheral treatment with IMC-H7 decreased appetite and body weight and increased energy expenditure and fat oxidation. A greater reduction in body weight and caloric intake was observed in response to IMC-H7 during the long-day fat state as compared to the short-day lean state. This enhanced response to IMC-H7 was also observed in calorically restricted hamsters maintained in long days, suggesting that it is the central photoperiodic state rather than the peripheral adiposity that determines the response to FGFR1c antagonism. Hypothalamic thyroid hormone availability is controlled by deiodinase enzymes (DIO2 and DIO3) expressed in tanycytes and is the key regulator of seasonal cycles of energy balance. Therefore, we determined the effect of IMC-H7 on hypothalamic expression of these deiodinase enzymes. The reductions in food intake and body weight were always associated with decreased expression of DIO2 in the hypothalamic ependymal cell layer containing tanycytes. These data provide further support for the notion the tanycytes are an important component of the mechanism by which the hypothalamus integrates central and peripheral signals to regulate energy intake and expenditure. PMID:26549257

  5. Digestive Enzymes in the Crayfish Cherax albidus: Polymorphism and Partial Characterization

    Directory of Open Access Journals (Sweden)

    Elena Coccia

    2011-01-01

    Full Text Available We will deal with the partial characterization and the activity of the following digestive enzymes: amylase, pectinase, alginase, lipase, and protease present in the digestive tract of juvenile freshwater crayfish Cherax albidus. Gastric juices, the hepatopancreas, and the intestine were sampled for enzyme analysis. Among carbohydratases, amylase activity was the highest. It was significantly higher in the intestine than in the gastric juice and hepatopancreas. Lipase activity was significantly higher in the hepatopancreas and the intestine compared to the gastric juice. Both alkaline and acid proteases were found. Alkaline proteases were characterized by employing specific protease inhibitors. Both trypsin and chymotrypsin activity was detected. The majority of alkaline protease activity was ascribable to trypsin. Several isoforms of digestive enzymes were identified by gel electrophoresis. This work provides basic information to study the digestive abilities of crayfish.

  6. Inhibitors of the Hydrolytic Enzyme Dimethylarginine Dimethylaminohydrolase (DDAH): Discovery, Synthesis and Development.

    Science.gov (United States)

    Murphy, Rhys B; Tommasi, Sara; Lewis, Benjamin C; Mangoni, Arduino A

    2016-01-01

    Dimethylarginine dimethylaminohydrolase (DDAH) is a highly conserved hydrolytic enzyme found in numerous species, including bacteria, rodents, and humans. In humans, the DDAH-1 isoform is known to metabolize endogenous asymmetric dimethylarginine (ADMA) and monomethyl arginine (l-NMMA), with ADMA proposed to be a putative marker of cardiovascular disease. Current literature reports identify the DDAH family of enzymes as a potential therapeutic target in the regulation of nitric oxide (NO) production, mediated via its biochemical interaction with the nitric oxide synthase (NOS) family of enzymes. Increased DDAH expression and NO production have been linked to multiple pathological conditions, specifically, cancer, neurodegenerative disorders, and septic shock. As such, the discovery, chemical synthesis, and development of DDAH inhibitors as potential drug candidates represent a growing field of interest. This review article summarizes the current knowledge on DDAH inhibition and the derived pharmacokinetic parameters of the main DDAH inhibitors reported in the literature. Furthermore, current methods of development and chemical synthetic pathways are discussed. PMID:27187323

  7. Effect of Potato Virus Y on the NADP-Malic Enzyme from Nicotiana tabacum L.: mRNA, Expressed Protein and Activity

    OpenAIRE

    Helena Ryšlavá; Petra Spoustová; Helena Synková; Noemi Čeřovská; Karel Müller; Veronika Doubnerová

    2009-01-01

    The effect of biotic stress induced by viral infection (Potato virus Y, strain NTN and O) on NADP-malic enzyme (EC 1.1.1.40) in tobacco plants (Nicotiana tabacum L., cv. Petit Havana, SR1) was tested at the transcriptional, translational and activity level. The increase of enzyme activity in infected leaves was correlated with the increased amount of expressed protein and with mRNA of cytosolic NADP-ME isoform. Transcription of the chloroplastic enzyme was not influenced by viral infection. T...

  8. Murine branched chain alpha-ketoacid dehydrogenase kinase; cDNA cloning, tissue distribution, and temporal expression during embryonic development.

    Science.gov (United States)

    Doering, C B; Coursey, C; Spangler, W; Danner, D J

    1998-06-01

    These studies were designed to demonstrate the structural and functional similarity of murine branched chain alpha-ketoacid dehydrogenase and its regulation by the complex-specific kinase. Nucleotide sequence and deduced amino acid sequence for the kinase cDNA demonstrate a highly conserved coding sequence between mouse and human. Tissue-specific expression in adult mice parallels that reported in other mammals. Kinase expression in female liver is influenced by circadian rhythm. Of special interest is the fluctuating expression of this kinase during embryonic development against the continuing increase in the catalytic subunits of this mitochondrial complex during development. The need for regulation of the branched chain alpha-ketoacid dehydrogenase complex by kinase expression during embryogenesis is not understood. However, the similarity of murine branched chain alpha-ketoacid dehydrogenase and its kinase to the human enzyme supports the use of this animal as a model for the human system. PMID:9611264

  9. Altered CD45 isoform expression affects lymphocyte function in CD45 Tg mice.

    Science.gov (United States)

    Tchilian, Elma Z; Dawes, Ritu; Hyland, Lisa; Montoya, Maria; Le Bon, Agnes; Borrow, Persephone; Hou, Sam; Tough, David; Beverley, Peter C L

    2004-09-01

    Transgenic mice have been constructed expressing high (CD45RABC) and low (CD45R0) molecular weight CD45 isoforms on a CD45-/- background. Phenotypic analysis and in vivo challenge of these mice with influenza and lymphocytic choriomeningitis viruses shows that T cell differentiation and peripheral T cell function are related to the level of CD45 expression but not to which CD45 isoform is expressed. In contrast, B cell differentiation is not restored, irrespective of the level of expression of a single isoform. All CD45 trangenic mice have T cells with an activated phenotype and increased T cell turnover. These effects are more prominent in CD8 than CD4 cells. The transgenic mice share several properties with humans expressing variant CD45 alleles and provide a model to understand immune function in variant individuals. PMID:15302847

  10. Structural and Functional Characterization of Recombinant Isoforms of the Lentil Lipid Transfer Protein.

    Science.gov (United States)

    Bogdanov, I V; Finkina, E I; Balandin, S V; Melnikova, D N; Stukacheva, E A; Ovchinnikova, T V

    2015-01-01

    The recombinant isoforms Lc-LTP1 and Lc-LTP3 of the lentil lipid transfer protein were overexpressed in E. coli cells. It was confirmed that both proteins are stabilized by four disulfide bonds and characterized by a high proportion of the α-helical structure. It was found that Lc-LTP1 and Lc-LTP3 possess antimicrobial activity and can bind fatty acids. Both isoforms have the ability to bind specific IgE from sera of patients with food allergies, which recognize similar epitopes of the major peach allergen Pru p 3. Both isoforms were shown to have immunological properties similar to those of other plant allergenic LTPs, but Lc-LTP3 displayed a less pronounced immunoreactivity. PMID:26483961

  11. Exposing the specific roles of the invariant chain isoforms in shaping the MHC class II peptidome

    Directory of Open Access Journals (Sweden)

    Jean-Simon eFortin

    2013-12-01

    Full Text Available The peptide repertoire (peptidome associated with MHC class II molecules (MHCIIs is influenced by the polymorphic nature of the peptide binding groove but also by cell-intrinsic factors. The invariant chain (Ii chaperones MHCIIs, affecting their folding and trafficking. Recent discoveries relating to Ii functions have provided insights as to how it edits the MHCII peptidome. In humans, the Ii gene encodes four different isoforms for which structure-function analyses have highlighted common properties but also some non-redundant roles. Another layer of complexity arises from the fact that Ii heterotrimerizes, a characteristic that has the potential to affect the maturation of associated MHCIIs in many different ways, depending on the isoform combinations. Here, we emphasize the peptide editing properties of Ii and discuss the impact of the various isoforms on the MHCII peptidome.

  12. Quorum quenching enzymes.

    Science.gov (United States)

    Fetzner, Susanne

    2015-05-10

    Bacteria use cell-to-cell communication systems based on chemical signal molecules to coordinate their behavior within the population. These quorum sensing systems are potential targets for antivirulence therapies, because many bacterial pathogens control the expression of virulence factors via quorum sensing networks. Since biofilm maturation is also usually influenced by quorum sensing, quenching these systems may contribute to combat biofouling. One possibility to interfere with quorum sensing is signal inactivation by enzymatic degradation or modification. Such quorum quenching enzymes are wide-spread in the bacterial world and have also been found in eukaryotes. Lactonases and acylases that hydrolyze N-acyl homoserine lactone (AHL) signaling molecules have been investigated most intensively, however, different oxidoreductases active toward AHLs or 2-alkyl-4(1H)-quinolone signals as well as other signal-converting enzymes have been described. Several approaches have been assessed which aim at alleviating virulence, or biofilm formation, by reducing the signal concentration in the bacterial environment. These involve the application or stimulation of signal-degrading bacteria as biocontrol agents in the protection of crop plants against soft-rot disease, the use of signal-degrading bacteria as probiotics in aquaculture, and the immobilization or entrapment of quorum quenching enzymes or bacteria to control biofouling in membrane bioreactors. While most approaches to use quorum quenching as antivirulence strategy are still in the research phase, the growing number of organisms and enzymes known to interfere with quorum sensing opens up new perspectives for the development of innovative antibacterial strategies. PMID:25220028

  13. Effects of Weak Light on Starch Accumulation and Starch Synthesis Enzyme Activities in Rice at the Grain Filling Stage

    Institute of Scientific and Technical Information of China (English)

    LI Tian; Ryu OHSUGI; Tohru YAMAGISHI; Haruto SASAKI

    2006-01-01

    Dynamic changes of starch, amylose, sucrose contents and the activities of starch synthesis enzymes under shading treatments after flowering were studied using two dce varieties IR72 (indica) and Nipponbare (japonica) as materials. Under shading treatments, the starch,amylose and sucrose contents decreased, while ADP-glucose pyrophosphorylase (ADPGPPase) activity only changed a little, soluble starch synthase activity and granule bound starch synthase activity decreased, soluble starch branching enzyme (SSBE, Q-enzyme) activity and granule bound starch branching enzyme (GBSBE, Q-enzyme) activity increased, and starch debranching enzyme (DBE, R-enzyme) activity vaded with varieties. Correlation analyses showed that the changes of starch content were positively and significantly correlated with the changes of sucrose content in the weak light. Both ADPGPPase activity and SSBE activity were positively and significantly correlated with starch accumulation rate. It was implied that the decline of starch synthase activities was related to the decrease of starch content and the increase of the activity of starch branching enzyme played an important role in the decrease of the ratio of amylose to the total starch under the weak light.

  14. Unraveling complex interplay between heat shock factor 1 and 2 splicing isoforms.

    Directory of Open Access Journals (Sweden)

    Sylvain Lecomte

    Full Text Available Chaperone synthesis in response to proteotoxic stress is dependent on a family of transcription factors named heat shock factors (HSFs. The two main factors in this family, HSF1 and HSF2, are co-expressed in numerous tissues where they can interact and form heterotrimers in response to proteasome inhibition. HSF1 and HSF2 exhibit two alternative splicing isoforms, called α and β, which contribute to additional complexity in HSF transcriptional regulation, but remain poorly examined in the literature. In this work, we studied the transcriptional activity of HSF1 and HSF2 splicing isoforms transfected into immortalized Mouse Embryonic Fibroblasts (iMEFs deleted for both Hsf1 and Hsf2, under normal conditions and after proteasome inhibition. We found that HSF1α is significantly more active than the β isoform after exposure to the proteasome inhibitor MG132. Furthermore, we clearly established that, while HSF2 had no transcriptional activity by itself, short β isoform of HSF2 exerts a negative role on HSF1β-dependent transactivation. To further assess the impact of HSF2β inhibition on HSF1 activity, we developed a mathematical modelling approach which revealed that the balance between each HSF isoform in the cell regulated the strength of the transcriptional response. Moreover, we found that cellular stress such as proteasome inhibition could regulate the splicing of Hsf2 mRNA. All together, our results suggest that relative amounts of each HSF1 and HSF2 isoforms quantitatively determine the cellular level of the proteotoxic stress response.

  15. The α and Δ isoforms of CREB1 are required to maintain normal pulmonary vascular resistance.

    Directory of Open Access Journals (Sweden)

    Lili Li

    Full Text Available Chronic hypoxia causes pulmonary hypertension associated with structural alterations in pulmonary vessels and sustained vasoconstriction. The transcriptional mechanisms responsible for these distinctive changes are unclear. We have previously reported that CREB1 is activated in the lung in response to alveolar hypoxia but not in other organs. To directly investigate the role of α and Δ isoforms of CREB1 in the regulation of pulmonary vascular resistance we examined the responses of mice in which these isoforms of CREB1 had been inactivated by gene mutation, leaving only the β isoform intact (CREB(αΔ mice. Here we report that expression of CREB regulated genes was altered in the lungs of CREB(αΔ mice. CREB(αΔ mice had greater pulmonary vascular resistance than wild types, both basally in normoxia and following exposure to hypoxic conditions for three weeks. There was no difference in rho kinase mediated vasoconstriction between CREB(αΔ and wild type mice. Stereological analysis of pulmonary vascular structure showed characteristic wall thickening and lumen reduction in hypoxic wild-type mice, with similar changes observed in CREB(αΔ. CREB(αΔ mice had larger lungs with reduced epithelial surface density suggesting increased pulmonary compliance. These findings show that α and Δ isoforms of CREB1 regulate homeostatic gene expression in the lung and that normal activity of these isoforms is essential to maintain low pulmonary vascular resistance in both normoxic and hypoxic conditions and to maintain the normal alveolar structure. Interventions that enhance the actions of α and Δ isoforms of CREB1 warrant further investigation in hypoxic lung diseases.

  16. Comparative proteomics reveals a significant bias toward alternative protein isoforms with conserved structure and function.

    Science.gov (United States)

    Ezkurdia, Iakes; del Pozo, Angela; Frankish, Adam; Rodriguez, Jose Manuel; Harrow, Jennifer; Ashman, Keith; Valencia, Alfonso; Tress, Michael L

    2012-09-01

    Advances in high-throughput mass spectrometry are making proteomics an increasingly important tool in genome annotation projects. Peptides detected in mass spectrometry experiments can be used to validate gene models and verify the translation of putative coding sequences (CDSs). Here, we have identified peptides that cover 35% of the genes annotated by the GENCODE consortium for the human genome as part of a comprehensive analysis of experimental spectra from two large publicly available mass spectrometry databases. We detected the translation to protein of "novel" and "putative" protein-coding transcripts as well as transcripts annotated as pseudogenes and nonsense-mediated decay targets. We provide a detailed overview of the population of alternatively spliced protein isoforms that are detectable by peptide identification methods. We found that 150 genes expressed multiple alternative protein isoforms. This constitutes the largest set of reliably confirmed alternatively spliced proteins yet discovered. Three groups of genes were highly overrepresented. We detected alternative isoforms for 10 of the 25 possible heterogeneous nuclear ribonucleoproteins, proteins with a key role in the splicing process. Alternative isoforms generated from interchangeable homologous exons and from short indels were also significantly enriched, both in human experiments and in parallel analyses of mouse and Drosophila proteomics experiments. Our results show that a surprisingly high proportion (almost 25%) of the detected alternative isoforms are only subtly different from their constitutive counterparts. Many of the alternative splicing events that give rise to these alternative isoforms are conserved in mouse. It was striking that very few of these conserved splicing events broke Pfam functional domains or would damage globular protein structures. This evidence of a strong bias toward subtle differences in CDS and likely conserved cellular function and structure is remarkable and

  17. Branched-chain fatty acid biosynthesis in a branched-chain amino acid aminotransferase mutant of Staphylococcus carnosus

    DEFF Research Database (Denmark)

    Beck, Hans Christian

    2005-01-01

    Fatty acid biosynthesis by a mutant strain of Staphylococcus carnosus deficient in branched-chain amino acid aminotransferase (IlvE) activity was analysed. This mutant was unable to produce the appropriate branched-chain alpha-ketoacid precursors for branched-chain fatty acid biosynthesis from th...

  18. Geology of the Cane Branch and Helton Branch watershed areas, McCreary County, Kentucky

    Science.gov (United States)

    Lyons, Erwin J.

    1957-01-01

    Cane Branch and Helton Branch in McCreary County, Kentucky, are about 1.4 miles apart (fig. 1). Can Branch, which is about 2.1 miles long, emptied into Hughes Fork of Beaver Creek. Its watershed area of about 1.5 square miles lies largely in the Wiborf 7 1/2-minute quadrangle (SW/4 Cumberland Falls 15-minute quadrangle), but the downstream part of the area extends northward into the Hail 7 1/2-minute quadrangle (NW/4 Cumberland Falls 15-minute quadrangle). Helton Branch, which is about 1.1 miles long, has two tributaries and empties into Little Hurricane Fork of Beaver Creek. It drains an area of about 0.8 square mile of while about 0.5 square mile is in the Hail quadrangle and the remainder in the Wilborg quadrangle. The total relief in the Can Branch area is about 500 feet and in the Helton Branch area about 400 feet. Narrow, steep-sided to canyon-like valley and winding ridges, typical of the Pottsville escarpment region, are characteristic of both areas. Thick woods and dense undergrowth cover much of the two areas. Field mapping was done on U.S. Geological Survey 7 1/2-minute maps having a scale of 1:24,000 and a contour interval of 20 feet. Elevations of lithologic contacts were determined with a barometer and a hand level. Aerial photographs were used principally to trace the cliffs formed by sandstone and conglomerate ledges. Exposures, except for those of the cliff- and ledge-forming sandstone and conglomerates, are not abundant. The most complete stratigraphic sections (secs. 3 and 4, fig. 2) in the two areas are exposed in cuts of newly completed Forest Service roads, but the rick in the upper parts of the exposures is weathered. To supplement these sections, additional sections were measured in cuts along the railroad and main highways in nor near the watersheds.

  19. A novel family of katanin-like 2 protein isoforms (KATNAL2), interacting with nucleotide-binding proteins Nubp1 and Nubp2, are key regulators of different MT-based processes in mammalian cells.

    Science.gov (United States)

    Ververis, Antonis; Christodoulou, Andri; Christoforou, Maria; Kamilari, Christina; Lederer, Carsten W; Santama, Niovi

    2016-01-01

    Katanins are microtubule (MT)-severing AAA proteins with high phylogenetic conservation throughout the eukaryotes. They have been functionally implicated in processes requiring MT remodeling, such as spindle assembly in mitosis and meiosis, assembly/disassembly of flagella and cilia and neuronal morphogenesis. Here, we uncover a novel family of katanin-like 2 proteins (KATNAL2) in mouse, consisting of five alternatively spliced isoforms encoded by the Katnal2 genomic locus. We further demonstrate that in vivo these isoforms are able to interact with themselves, with each other and moreover directly and independently with MRP/MinD-type P-loop NTPases Nubp1 and Nubp2, which are integral components of centrioles, negative regulators of ciliogenesis and implicated in centriole duplication in mammalian cells. We find KATNAL2 localized on interphase MTs, centrioles, mitotic spindle, midbody and the axoneme and basal body of sensory cilia in cultured murine cells. shRNAi of Katnal2 results in inefficient cytokinesis and severe phenotypes of enlarged cells and nuclei, increased numbers of centrioles and the manifestation of aberrant multipolar mitotic spindles, mitotic defects, chromosome bridges, multinuclearity, increased MT acetylation and an altered cell cycle pattern. Silencing or stable overexpression of KATNAL2 isoforms drastically reduces ciliogenesis. In conclusion, KATNAL2s are multitasking enzymes involved in the same cell type in critically important processes affecting cytokinesis, MT dynamics, and ciliogenesis and are also implicated in cell cycle progression. PMID:26153462

  20. Investigating the role of the physiological isoform switch of cytochrome c oxidase subunits in reversible mitochondrial disease.

    Science.gov (United States)

    Boczonadi, Veronika; Giunta, Michele; Lane, Maria; Tulinius, Mar; Schara, Ulrike; Horvath, Rita

    2015-06-01

    Reversible infantile respiratory chain deficiency is characterised by spontaneous recovery of mitochondrial myopathy in infants. We studied whether a physiological isoform switch of nuclear cytochrome c oxidase subunits contributes to the age-dependent manifestation and spontaneous recovery in reversible mitochondrial disease. Some nuclear-encoded subunits of cytochrome c oxidase are present as tissue-specific isoforms. Isoforms of subunits COX6A and COX7A expressed in heart and skeletal muscle are different from isoforms expressed in the liver, kidney and brain. Furthermore, in skeletal muscle both the heart and liver isoforms of subunit COX7A have been demonstrated with variable levels, indicating that the tissue-specific expression of nuclear-encoded subunits could provide a basis for the fine-tuning of cytochrome c oxidase activity to the specific metabolic needs of the different tissues. We demonstrate a developmental isoform switch of COX6A and COX7A subunits in human and mouse skeletal muscle. While the liver type isoforms are more present soon after birth, the heart/muscle isoforms gradually increase around 3 months of age in infants, 4 weeks of age in mice, and these isoforms persist in muscle throughout life. Our data in follow-up biopsies of patients with reversible infantile respiratory chain deficiency indicate that the physiological isoform switch does not contribute to the clinical manifestation and to the spontaneous recovery of this disease. However, understanding developmental changes of the different cytochrome c oxidase isoforms may have implications for other mitochondrial diseases. This article is part of a Directed Issue entitled: Energy Metabolism Disorders and Therapies. PMID:25666558