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Sample records for nadph-dependent electron transferase

  1. Constitutive NADPH-dependent electron transferase activity of the Nox4 dehydrogenase domain.

    Science.gov (United States)

    Nisimoto, Yukio; Jackson, Heather M; Ogawa, Hisamitsu; Kawahara, Tsukasa; Lambeth, J David

    2010-03-23

    NADPH oxidase 4 (Nox4) is constitutively active, while Nox2 requires the cytosolic regulatory subunits p47(phox) and p67(phox) and activated Rac with activation by phorbol 12-myristate 13-acetate (PMA). This study was undertaken to identify the domain on Nox4 that confers constitutive activity. Lysates from Nox4-expressing cells exhibited constitutive NADPH- but not NADH-dependent hydrogen peroxide production with a K(m) for NADPH of 55 +/- 10 microM. The concentration of Nox4 in cell lysates was estimated using Western blotting and allowed calculation of a turnover of approximately 200 mol of H(2)O(2) min(-1) (mol of Nox4)(-1). A chimeric protein (Nox2/4) consisting of the Nox2 transmembrane (TM) domain and the Nox4 dehydrogenase (DH) domain showed H(2)O(2) production in the absence of cytosolic regulatory subunits. In contrast, chimera Nox4/2, consisting of the Nox4 TM and Nox2 DH domains, exhibited PMA-dependent activation that required coexpression of regulatory subunits. Nox DH domains from several Nox isoforms were purified and evaluated for their electron transferase activities. Nox1 DH, Nox2 DH, and Nox5 DH domains exhibited barely detectable activities toward artificial electron acceptors, while the Nox4 DH domain exhibited significant rates of reduction of cytochrome c (160 min(-1), largely superoxide dismutase-independent), ferricyanide (470 min(-1)), and other electron acceptors (artificial dyes and cytochrome b(5)). Rates were similar to those observed for H(2)O(2) production by the Nox4 holoenzyme in cell lysates. The activity required added FAD and was seen with NADPH but not NADH. These results indicate that the Nox4 DH domain exists in an intrinsically activated state and that electron transfer from NADPH to FAD is likely to be rate-limiting in the NADPH-dependent reduction of oxygen by holo-Nox4.

  2. Conformational changes of the NADPH-dependent cytochrome P450 reductase in the course of electron transfer to cytochromes P450

    DEFF Research Database (Denmark)

    Laursen, Tomas; Jensen, Kenneth; Møller, Birger Lindberg

    2011-01-01

    The NADPH-dependent cytochrome P450 reductase (CPR) is a key electron donor to eucaryotic cytochromes P450 (CYPs). CPR shuttles electrons from NADPH through the FAD and FMN-coenzymes into the iron of the prosthetic heme-group of the CYP. In the course of these electron transfer reactions, CPR und...... to serve as an effective electron transferring "nano-machine"....

  3. Constitutive NADPH-Dependent Electron Transferase Activity of the Nox4 Dehydrogenase Domain?

    OpenAIRE

    Nisimoto, Yukio; Jackson, Heather M.; Ogawa, Hisamitsu; Kawahara, Tsukasa; Lambeth, J. David

    2010-01-01

    NADPH oxidase 4 (Nox4) is constitutively active, while Nox2 requires the cytosolic regulatory subunits p47 phox and p67 phox and activated Rac with activation by phorbol 12-myristate 13-acetate (PMA). This study was undertaken to identify the domain on Nox4 that confers constitutive activity. Lysates from Nox4-expressing cells exhibited constitutive NADPH- but not NADH-dependent hydrogen peroxide production with a K m for NADPH of 55 ? 10 ?M. The concentration of Nox4 in cell lysates was esti...

  4. Comparative molecular modeling study of Arabidopsis NADPH-dependent thioredoxin reductase and its hybrid protein.

    Directory of Open Access Journals (Sweden)

    Yuno Lee

    Full Text Available 2-Cys peroxiredoxins (Prxs play important roles in the protection of chloroplast proteins from oxidative damage. Arabidopsis NADPH-dependent thioredoxin reductase isotype C (AtNTRC was identified as efficient electron donor for chloroplastic 2-Cys Prx-A. There are three isotypes (A, B, and C of thioredoxin reductase (TrxR in Arabidopsis. AtNTRA contains only TrxR domain, but AtNTRC consists of N-terminal TrxR and C-terminal thioredoxin (Trx domains. AtNTRC has various oligomer structures, and Trx domain is important for chaperone activity. Our previous experimental study has reported that the hybrid protein (AtNTRA-(Trx-D, which was a fusion of AtNTRA and Trx domain from AtNTRC, has formed variety of structures and shown strong chaperone activity. But, electron transfer mechanism was not detected at all. To find out the reason of this problem with structural basis, we performed two different molecular dynamics (MD simulations on AtNTRC and AtNTRA-(Trx-D proteins with same cofactors such as NADPH and flavin adenine dinucleotide (FAD for 50 ns. Structural difference has found from superimposition of two structures that were taken relatively close to average structure. The main reason that AtNTRA-(Trx-D cannot transfer the electron from TrxR domain to Trx domain is due to the difference of key catalytic residues in active site. The long distance between TrxR C153 and disulfide bond of Trx C387-C390 has been observed in AtNTRA-(Trx-D because of following reasons: i unstable and unfavorable interaction of the linker region, ii shifted Trx domain, and iii different or weak interface interaction of Trx domains. This study is one of the good examples for understanding the relationship between structure formation and reaction activity in hybrid protein. In addition, this study would be helpful for further study on the mechanism of electron transfer reaction in NADPH-dependent thioredoxin reductase proteins.

  5. Sex-related differences in NADPH-dependent lipid peroxidation induced by cadmium

    Energy Technology Data Exchange (ETDEWEB)

    Sato, Masao; Nagai, Yasushi

    1986-10-01

    Male and female rats were dosed once a day for 2 days with injections of 1.5 mg Cd/kg. Formation of thiobarbituric acid reactive substances (TBA-RS) was significantly increased in male rat liver but not in the females. NADPH-dependent lipid peroxidation in vitro in microsomes derived from untreated rat liver was greater in males than in females. Furthermore, addition of cadmium (Cd) to microsomes isolated from male rat liver produced a dose-dependent potentiation of NADPH-dependent lipid peroxidation from low concentrations of CD. In microsomes derived from females a significant increase in lipid peroxidation was observed only at high Cd concentrations. NADPH-dependent lipid peroxidation enhanced by Cd was greater in the males than in the females. These data suggest that a sex-related difference in the ability of Cd to induce lipid peroxidation in vivo in rat liver appears to be mediated partly through differences in hepatic microsomal NADPH-dependent lipid peroxidation.

  6. Chloroplast NADPH-Dependent Thioredoxin Reductase from Chlorella vulgaris Alleviates Environmental Stresses in Yeast Together with 2-Cys Peroxiredoxin

    Science.gov (United States)

    Machida, Takeshi; Ishibashi, Akiko; Kirino, Ai; Sato, Jun-ichi; Kawasaki, Shinji; Niimura, Youichi; Honjoh, Ken-ichi; Miyamoto, Takahisa

    2012-01-01

    Chloroplast NADPH-dependent thioredoxin reductase (NTRC) catalyzes the reduction of 2-Cys peroxiredoxin (2-Cys Prx) and, thus, probably functions as an antioxidant system. The functions of the enzyme in oxidative and salt stresses have been reported previously. We have previously identified and characterized NTRC in Chlorella vulgaris. In the present study, we isolated a full-length cDNA clone encoding 2-Cys Prx from C. vulgaris and investigated the involvement of Chlorella NTRC/2-Cys Prx system in several environmental stress tolerances by using yeast as a eukaryotic model. Deduced Chlorella 2-Cys Prx was homologous to those of chloroplast 2-Cys Prxs from plants, and two conserved cysteine residues were found in the deduced sequence. Enzyme assay showed that recombinant mature C. vulgaris NTRC (mCvNTRC) transferred electrons from NADPH to recombinant mature C. vulgaris 2-Cys Prx (mCvPrx), and mCvPrx decomposed hydrogen peroxide, tert-butyl hydroperoxide, and peroxynitrite by cooperating with mCvNTRC. Based on the results, the mCvNTRC/mCvPrx antioxidant system was identified in Chlorella. The antioxidant system genes were expressed in yeast separately or coordinately. Stress tolerances of yeast against freezing, heat, and menadione-induced oxidative stresses were significantly improved by expression of mCvNTRC, and the elevated tolerances were more significant when both mCvNTRC and mCvPrx were co-expressed. Our results reveal a novel feature of NTRC: it functions as an antioxidant system with 2-Cys Prx in freezing and heat stress tolerances. PMID:23029353

  7. Structure of Hordeum vulgare NADPH-dependent thioredoxin reductase 2. Unwinding the reaction mechanism

    Energy Technology Data Exchange (ETDEWEB)

    Kirkensgaard, Kristine G. [Carlsberg Laboratory (Denmark); Enzyme and Protein Chemistry, Department of Systems BioIogy, Technical University of Denmark (Denmark); Hägglund, Per; Finnie, Christine; Svensson, Birte [Enzyme and Protein Chemistry, Department of Systems BioIogy, Technical University of Denmark (Denmark); Henriksen, Anette, E-mail: anette@crc.dk [Carlsberg Laboratory (Denmark)

    2009-09-01

    The first crystal structure of a cereal NTR, a protein involved in seed development and germination, has been determined. The structure is in a conformation that excludes NADPH binding and indicates that a domain reorientation facilitated by Trx binding precedes NADPH binding in the reaction mechanism. Thioredoxins (Trxs) are protein disulfide reductases that regulate the intracellular redox environment and are important for seed germination in plants. Trxs are in turn regulated by NADPH-dependent thioredoxin reductases (NTRs), which provide reducing equivalents to Trx using NADPH to recycle Trxs to the active form. Here, the first crystal structure of a cereal NTR, HvNTR2 from Hordeum vulgare (barley), is presented, which is also the first structure of a monocot plant NTR. The structure was determined at 2.6 Å resolution and refined to an R{sub cryst} of 19.0% and an R{sub free} of 23.8%. The dimeric protein is structurally similar to the structures of AtNTR-B from Arabidopsis thaliana and other known low-molecular-weight NTRs. However, the relative position of the two NTR cofactor-binding domains, the FAD and the NADPH domains, is not the same. The NADPH domain is rotated by 25° and bent by a 38% closure relative to the FAD domain in comparison with AtNTR-B. The structure may represent an intermediate between the two conformations described previously: the flavin-oxidizing (FO) and the flavin-reducing (FR) conformations. Here, analysis of interdomain contacts as well as phylogenetic studies lead to the proposal of a new reaction scheme in which NTR–Trx interactions mediate the FO to FR transformation.

  8. Thioredoxin and NADPH-Dependent Thioredoxin Reductase C Regulation of Tetrapyrrole Biosynthesis.

    Science.gov (United States)

    Da, Qingen; Wang, Peng; Wang, Menglong; Sun, Ting; Jin, Honglei; Liu, Bing; Wang, Jinfa; Grimm, Bernhard; Wang, Hong-Bin

    2017-10-01

    In chloroplasts, thioredoxin (TRX) isoforms and NADPH-dependent thioredoxin reductase C (NTRC) act as redox regulatory factors involved in multiple plastid biogenesis and metabolic processes. To date, less is known about the functional coordination between TRXs and NTRC in chlorophyll biosynthesis. In this study, we aimed to explore the potential functions of TRX m and NTRC in the regulation of the tetrapyrrole biosynthesis (TBS) pathway. Silencing of three genes, TRX m1 , TRX m2 , and TRX m4 ( TRX ms ), led to pale-green leaves, a significantly reduced 5-aminolevulinic acid (ALA)-synthesizing capacity, and reduced accumulation of chlorophyll and its metabolic intermediates in Arabidopsis ( Arabidopsis thaliana ). The contents of ALA dehydratase, protoporphyrinogen IX oxidase, the I subunit of Mg-chelatase, Mg-protoporphyrin IX methyltransferase (CHLM), and NADPH-protochlorophyllide oxidoreductase were decreased in triple TRX m- silenced seedlings compared with the wild type, although the transcript levels of the corresponding genes were not altered significantly. Protein-protein interaction analyses revealed a physical interaction between the TRX m isoforms and CHLM. 4-Acetoamido-4-maleimidylstilbene-2,2-disulfonate labeling showed the regulatory impact of TRX ms on the CHLM redox status. Since CHLM also is regulated by NTRC (Richter et al., 2013), we assessed the concurrent functions of TRX m and NTRC in the control of CHLM. Combined deficiencies of three TRX m isoforms and NTRC led to a cumulative decrease in leaf pigmentation, TBS intermediate contents, ALA synthesis rate, and CHLM activity. We discuss the coordinated roles of TRX m and NTRC in the redox control of CHLM stability with its corollary activity in the TBS pathway. © 2017 American Society of Plant Biologists. All Rights Reserved.

  9. Structure of Hordeum vulgare NADPH-dependent thioredoxin reductase 2. Unwinding the reaction mechanism

    International Nuclear Information System (INIS)

    Kirkensgaard, Kristine G.; Hägglund, Per; Finnie, Christine; Svensson, Birte; Henriksen, Anette

    2009-01-01

    The first crystal structure of a cereal NTR, a protein involved in seed development and germination, has been determined. The structure is in a conformation that excludes NADPH binding and indicates that a domain reorientation facilitated by Trx binding precedes NADPH binding in the reaction mechanism. Thioredoxins (Trxs) are protein disulfide reductases that regulate the intracellular redox environment and are important for seed germination in plants. Trxs are in turn regulated by NADPH-dependent thioredoxin reductases (NTRs), which provide reducing equivalents to Trx using NADPH to recycle Trxs to the active form. Here, the first crystal structure of a cereal NTR, HvNTR2 from Hordeum vulgare (barley), is presented, which is also the first structure of a monocot plant NTR. The structure was determined at 2.6 Å resolution and refined to an R cryst of 19.0% and an R free of 23.8%. The dimeric protein is structurally similar to the structures of AtNTR-B from Arabidopsis thaliana and other known low-molecular-weight NTRs. However, the relative position of the two NTR cofactor-binding domains, the FAD and the NADPH domains, is not the same. The NADPH domain is rotated by 25° and bent by a 38% closure relative to the FAD domain in comparison with AtNTR-B. The structure may represent an intermediate between the two conformations described previously: the flavin-oxidizing (FO) and the flavin-reducing (FR) conformations. Here, analysis of interdomain contacts as well as phylogenetic studies lead to the proposal of a new reaction scheme in which NTR–Trx interactions mediate the FO to FR transformation

  10. Crystallization and preliminary X-ray analysis of the NADPH-dependent 3-quinuclidinone reductase from Rhodotorula rubra

    International Nuclear Information System (INIS)

    Takeshita, Daijiro; Kataoka, Michihiko; Miyakawa, Takuya; Miyazono, Ken-ichi; Uzura, Atsuko; Nagata, Koji; Shimizu, Sakayu; Tanokura, Masaru

    2009-01-01

    The NADPH-dependent 3-quinuclidinone reductase from Rhodotorula rubra was expressed, purified, and crystallized and X-ray diffraction data of this crystal were collected to 2.2 Å resolution. (R)-3-Quinuclidinol is a useful compound that is applicable to the synthesis of various pharmaceuticals. The NADPH-dependent carbonyl reductase 3-quinuclidinone reductase from Rhodotorula rubra catalyzes the stereospecific reduction of 3-quinuclidinone to (R)-3-quinuclidinol and is expected to be utilized in industrial production of this alcohol. 3-Quinuclidinone reductase from R. rubra was expressed in Escherichia coli and purified using Ni-affinity and ion-exchange column chromatography. Crystals of the protein were obtained by the sitting-drop vapour-diffusion method using PEG 8000 as the precipitant. The crystals belonged to space group P4 1 2 1 2, with unit-cell parameters a = b = 91.3, c = 265.4 Å, and diffracted X-rays to 2.2 Å resolution. The asymmetric unit contained four molecules of the protein and the solvent content was 48.4%

  11. Crystallization and preliminary X-ray analysis of the NADPH-dependent 3-quinuclidinone reductase from Rhodotorula rubra

    Science.gov (United States)

    Takeshita, Daijiro; Kataoka, Michihiko; Miyakawa, Takuya; Miyazono, Ken-ichi; Uzura, Atsuko; Nagata, Koji; Shimizu, Sakayu; Tanokura, Masaru

    2009-01-01

    (R)-3-Quinuclidinol is a useful compound that is applicable to the synthesis of various pharmaceuticals. The NADPH-dependent carbonyl reductase 3-­quinuclidinone reductase from Rhodotorula rubra catalyzes the stereospecific reduction of 3-quinuclidinone to (R)-3-quinuclidinol and is expected to be utilized in industrial production of this alcohol. 3-Quinuclidinone reductase from R. rubra was expressed in Escherichia coli and purified using Ni-affinity and ion-exchange column chromatography. Crystals of the protein were obtained by the sitting-drop vapour-diffusion method using PEG 8000 as the precipitant. The crystals belonged to space group P41212, with unit-cell parameters a = b = 91.3, c = 265.4 Å, and diffracted X-rays to 2.2 Å resolution. The asymmetric unit contained four molecules of the protein and the solvent content was 48.4%. PMID:19478454

  12. NADPH-dependent glutamate dehydrogenase in Penicillium chrysogenum is involved in regulation of beta-lactam production

    DEFF Research Database (Denmark)

    Thykær, Jette; Kildegaard, Kanchana Rueksomtawin; Noorman, H.

    2008-01-01

    was detected in either of the Delta gdhA strains. Supplementation with glutamate restored growth but no beta-lactam production was detected for the constructed strains. Cultures with high ammonium concentrations (repressing conditions) and with proline as nitrogen source (de-repressed conditions) showed......The interactions between the ammonium assimilatory pathways and beta-lactam production were investigated by disruption of the NADPH-dependent glutamate dehydrogenase gene (gdhA) in two industrial beta-lactam-producing strains of Penicillium chrysogenum. The strains used were an adipoyl-7-ADCA...... continued beta-lactam production for the reference strains whereas the Delta gdhA strains remained non-productive under all conditions. By overexpressing the NAD-dependent glutamate dehydrogenase, the specific growth rate could be restored, but still no beta-lactam production was detected. The results...

  13. NADPH-dependent D-aldose reductases and xylose fermentation in Fusarium oxysporum

    DEFF Research Database (Denmark)

    Panagiotou, Gianni; Christakopoulos, P.

    2004-01-01

    for NADPH over NADH. In this study, the influence of aeration and the response to the addition of electron acceptors on xylose fermentation by F. oxysporum were also studied. The batch cultivation of F. oxysporum on xylose was performed under aerobic, anaerobic and oxygen-limited conditions in stirred tank...... conditions (0.3 vvm). When the artificial electron acceptor acetoin was added to an anaerobic batch fermentation of xylose by F. oxysporum, the ethanol yield increased while xylitol excretion was also decreased....

  14. Biochemical and molecular characterization of an azoreductase from Staphylococcus aureus, a tetrameric NADPH-dependent flavoprotein

    Science.gov (United States)

    Chen, Huizhong; Hopper, Sherryll L.; Cerniglia, Carl E.

    2018-01-01

    Azo dyes are a predominant class of colourants used in tattooing, cosmetics, foods and consumer products. A gene encoding NADPH-flavin azoreductase (Azo1) from the skin bacterium Staphylococcus aureus ATCC 25923 was identified and overexpressed in Escherichia coli. RT-PCR results demonstrated that the azo1 gene was constitutively expressed at the mRNA level in S. aureus. Azo1 was found to be a tetramer with a native molecular mass of 85 kDa containing four non-covalently bound FMN. Azo1 requires NADPH, but not NADH, as an electron donor for its activity. The enzyme was resolved to dimeric apoprotein by removing the flavin prosthetic groups using hydrophobic-interaction chromatography. The dimeric apoprotein was reconstituted on-column and in free stage with FMN, resulting in the formation of a fully functional native-like tetrameric enzyme. The enzyme cleaved the model azo dye 2-[4-(dimethylamino)phenylazo]benzoic acid (Methyl Red) into N,N-dimethyl-p-phenylenediamine and 2-aminobenzoic acid. The apparent Km values for NADPH and Methyl Red substrates were 0·;074 and 0·057 mM, respectively. The apparent Vmax was 0·4 µM min−1 (mg protein)−1. Azo1 was also able to metabolize Orange II, Amaranth, Ponceau BS and Ponceau S azo dyes. Azo1 represents the first azoreductase to be identified and characterized from human skin microflora. PMID:15870453

  15. The protein inhibitor of nNOS (PIN/DLC1/LC8) binding does not inhibit the NADPH-dependent heme reduction in nNOS, a key step in NO synthesis.

    Science.gov (United States)

    Parhad, Swapnil S; Jaiswal, Deepa; Ray, Krishanu; Mazumdar, Shyamalava

    2016-03-25

    The neuronal nitric oxide synthase (nNOS) is an essential enzyme involved in the synthesis of nitric oxide (NO), a potent neurotransmitter. Although previous studies have indicated that the dynein light chain 1 (DLC1) binding to nNOS could inhibit the NO synthesis, the claim is challenged by contradicting reports. Thus, the mechanism of nNOS regulation remained unclear. nNOS has a heme-bearing, Cytochrome P450 core, and the functional enzyme is a dimer. The electron flow from NADPH to Flavin, and finally to the heme of the paired nNOS subunit within a dimer, is facilitated upon calmodulin (CaM) binding. Here, we show that DLC1 binding to nNOS-CaM complex does not affect the electron transport from the reductase to the oxygenase domain. Therefore, it cannot inhibit the rate of NADPH-dependent heme reduction in nNOS, which results in l-Arginine oxidation. Also, the NO release activity does not decrease with increasing DLC1 concentration in the reaction mix, which further confirmed that DLC1 does not inhibit nNOS activity. These findings suggest that the DLC1 binding may have other implications for the nNOS function in the cell. Copyright © 2016 Elsevier Inc. All rights reserved.

  16. Resistance of Saccharomyces cerevisiae to High Concentrations of Furfural Is Based on NADPH-Dependent Reduction by at Least Two Oxireductases ▿ †

    Science.gov (United States)

    Heer, Dominik; Heine, Daniel; Sauer, Uwe

    2009-01-01

    Biofuels derived from lignocellulosic biomass hold promises for a sustainable fuel economy, but several problems hamper their economical feasibility. One important problem is the presence of toxic compounds in processed lignocellulosic hydrolysates, with furfural as a key toxin. While Saccharomyces cerevisiae has some intrinsic ability to reduce furfural to the less-toxic furfuryl alcohol, higher resistance is necessary for process conditions. By comparing an evolved, furfural-resistant strain and its parent in microaerobic, glucose-limited chemostats at increasing furfural challenge, we elucidate key mechanism and the molecular basis of both natural and high-level furfural resistance. At lower concentrations of furfural, NADH-dependent oxireductases are the main defense mechanism. At furfural concentrations above 15 mM, however, 13C-flux and global array-based transcript analysis demonstrated that the NADPH-generating flux through the pentose phosphate pathway increases and that NADPH-dependent oxireductases become the major resistance mechanism. The transcript analysis further revealed that iron transmembrane transport is upregulated in response to furfural. While these responses occur in both strains, high-level resistance in the evolved strain was based on strong induction of ADH7, the uncharacterized open reading frame (ORF) YKL071W, and four further, likely NADPH-dependent, oxireductases. By overexpressing the ADH7 gene and the ORF YKL071W, we inversely engineered significantly increased furfural resistance in the parent strain, thereby demonstrating that these two enzymes are key elements of the resistance phenotype. PMID:19854918

  17. Mechanism of mercuric chloride resistance in microorganisms. II. NADPH-dependent reduction of mercuric chloride and vaporization of mercury from mercuric chloride by a multiple drug resistant strain of Escherichia coli

    Energy Technology Data Exchange (ETDEWEB)

    Komura, I; Funaba, T; Izaki, K

    1971-01-01

    The activity to vaporize a /sup 203/Hg compound from /sup 203/HgCl/sub 2/ was demonstrated in crude cell-free extracts of a strain of Escherichia coli W2252, which had acquired the multiple drug resistance. NADPH was essential for the vaporization, while NADH had only a slight stimulating effect and NADP/sup +/ had no effect. The oxidation of NADPH dependent on HgCl/sub 2/ was also demonstrated in the crude extracts, but the HgCl/sub 2/-dependent NADH oxidation could be demonstrated only when a partially purified enzyme preparation was used. The rate of NADH oxidation was much slower than that of NADPH oxidation. It was concluded that NADPH, and to a lesser extent NADH, act as electron donors for the enzymatic reduction of HgCl/sub 2/ and the vaporization occurs after this reduction. This reduction and subsequent vaporization seem to provide a mechanism of resistance to HgCl/sub 2/ in E. coli strains having the multiple drug resistance. 15 references, 4 figures, 4 tables.

  18. The protein inhibitor of nNOS (PIN/DLC1/LC8) binding does not inhibit the NADPH-dependent heme reduction in nNOS, a key step in NO synthesis

    International Nuclear Information System (INIS)

    Parhad, Swapnil S.; Jaiswal, Deepa; Ray, Krishanu; Mazumdar, Shyamalava

    2016-01-01

    The neuronal nitric oxide synthase (nNOS) is an essential enzyme involved in the synthesis of nitric oxide (NO), a potent neurotransmitter. Although previous studies have indicated that the dynein light chain 1 (DLC1) binding to nNOS could inhibit the NO synthesis, the claim is challenged by contradicting reports. Thus, the mechanism of nNOS regulation remained unclear. nNOS has a heme-bearing, Cytochrome P450 core, and the functional enzyme is a dimer. The electron flow from NADPH to Flavin, and finally to the heme of the paired nNOS subunit within a dimer, is facilitated upon calmodulin (CaM) binding. Here, we show that DLC1 binding to nNOS-CaM complex does not affect the electron transport from the reductase to the oxygenase domain. Therefore, it cannot inhibit the rate of NADPH-dependent heme reduction in nNOS, which results in L-Arginine oxidation. Also, the NO release activity does not decrease with increasing DLC1 concentration in the reaction mix, which further confirmed that DLC1 does not inhibit nNOS activity. These findings suggest that the DLC1 binding may have other implications for the nNOS function in the cell. - Highlights: • The effect of interaction of nNOS with DLC1 has been debatable with contradicting reports in literature. • Purified DLC1 has no effect on electron transport between reductase and oxygenase domain of purified nNOS-CaM. • The NO release activity of nNOS was not altered by DLC1, supporting that DLC1 does not inhibit the enzyme. • These findings suggest that the DLC1 binding may have other implications for the nNOS function in the cell.

  19. The protein inhibitor of nNOS (PIN/DLC1/LC8) binding does not inhibit the NADPH-dependent heme reduction in nNOS, a key step in NO synthesis

    Energy Technology Data Exchange (ETDEWEB)

    Parhad, Swapnil S. [Tata Institute of Fundamental Research (TIFR), Homi Bhabha Road, Mumbai 400 005 (India); Jaiswal, Deepa [Tata Institute of Fundamental Research (TIFR), Homi Bhabha Road, Mumbai 400 005 (India); TIFR Centre for Interdisciplinary Sciences, 21 Brundavan Colony, Narsingi, Hyderabad 500075 (India); Ray, Krishanu, E-mail: krishanu@tifr.res.in [Tata Institute of Fundamental Research (TIFR), Homi Bhabha Road, Mumbai 400 005 (India); Mazumdar, Shyamalava, E-mail: shyamal@tifr.res.in [Tata Institute of Fundamental Research (TIFR), Homi Bhabha Road, Mumbai 400 005 (India)

    2016-03-25

    The neuronal nitric oxide synthase (nNOS) is an essential enzyme involved in the synthesis of nitric oxide (NO), a potent neurotransmitter. Although previous studies have indicated that the dynein light chain 1 (DLC1) binding to nNOS could inhibit the NO synthesis, the claim is challenged by contradicting reports. Thus, the mechanism of nNOS regulation remained unclear. nNOS has a heme-bearing, Cytochrome P450 core, and the functional enzyme is a dimer. The electron flow from NADPH to Flavin, and finally to the heme of the paired nNOS subunit within a dimer, is facilitated upon calmodulin (CaM) binding. Here, we show that DLC1 binding to nNOS-CaM complex does not affect the electron transport from the reductase to the oxygenase domain. Therefore, it cannot inhibit the rate of NADPH-dependent heme reduction in nNOS, which results in L-Arginine oxidation. Also, the NO release activity does not decrease with increasing DLC1 concentration in the reaction mix, which further confirmed that DLC1 does not inhibit nNOS activity. These findings suggest that the DLC1 binding may have other implications for the nNOS function in the cell. - Highlights: • The effect of interaction of nNOS with DLC1 has been debatable with contradicting reports in literature. • Purified DLC1 has no effect on electron transport between reductase and oxygenase domain of purified nNOS-CaM. • The NO release activity of nNOS was not altered by DLC1, supporting that DLC1 does not inhibit the enzyme. • These findings suggest that the DLC1 binding may have other implications for the nNOS function in the cell.

  20. Transferases in Polymer Chemistry

    NARCIS (Netherlands)

    van der Vlist, Jeroen; Loos, Katja; Palmans, ARA; Heise, A

    2010-01-01

    Transferases are enzymes that catalyze reactions in which a group is transferred from one compound to another. This makes these enzymes ideal catalysts for polymerization reactions. In nature, transferases are responsible for the synthesis of many important natural macromolecules. In synthetic

  1. Overexpression of Arabidopsis NADPH-dependent thioredoxin reductase C (AtNTRC) confers freezing and cold shock tolerance to plants

    Energy Technology Data Exchange (ETDEWEB)

    Moon, Jeong Chan [National Institute of Ecology, 1210 Geumgang-ro, Maseo-myeon, Seocheon-gun 325-813 (Korea, Republic of); Lee, Sangmin [Department of Biochemistry, College of Natural Sciences, Kangwon National University, Chuncheon (Korea, Republic of); Shin, Su Young [National Institute of Ecology, 1210 Geumgang-ro, Maseo-myeon, Seocheon-gun 325-813 (Korea, Republic of); Chae, Ho Byoung; Jung, Young Jun [Division of Applied Life Science - BK21+ program, PMBBRC, Gyeongsang National University, Jinju (Korea, Republic of); Jung, Hyun Suk [Department of Biochemistry, College of Natural Sciences, Kangwon National University, Chuncheon (Korea, Republic of); Lee, Kyun Oh [Division of Applied Life Science - BK21+ program, PMBBRC, Gyeongsang National University, Jinju (Korea, Republic of); Lee, Jung Ro, E-mail: leejr73@nie.re.kr [National Institute of Ecology, 1210 Geumgang-ro, Maseo-myeon, Seocheon-gun 325-813 (Korea, Republic of); Department of Biochemistry and Biophysics, Texas A& M University, College Station, TX (United States); Lee, Sang Yeol, E-mail: sylee@gnu.ac.kr [Division of Applied Life Science - BK21+ program, PMBBRC, Gyeongsang National University, Jinju (Korea, Republic of)

    2015-08-07

    Overexpression of AtNTRC (AtNTRC{sup OE}) in Arabidopsis thaliana led to a freezing and cold stress tolerance, whereas a knockout mutant (atntrc) showed a stress-sensitive phenotype. Biochemical analyses showed that the recombinant AtNTRC proteins exhibited a cryoprotective activity for malate dehydrogenase and lactic dehydrogenase. Furthermore, conclusive evidence of its interaction with nucleic acids in vitro is provided here on the basis of gel shift and electron microscopy analysis. Recombinant AtNTRC efficiently protected RNA and DNA from RNase A and metal catalyzed oxidation damage, respectively. The C-terminal thioredoxin domain is required for the nucleic acid–protein complex formation. From these results, it can be hypothesized that AtNTRC, which is known to be an electron donor of peroxiredoxin, contributes the stability of macromolecules under cold stress. - Highlights: • AtNTRC has a cryoprotective activity in vitro. • Overexpression of AtNTRC increases tolerance to freezing and cold shock stresses. • Thioredoxin domain of AtNTRC protects nucleic acids in vitro. • AtNTRC inhibits protein aggregation under freezing stress in vitro.

  2. Overexpression of Arabidopsis NADPH-dependent thioredoxin reductase C (AtNTRC) confers freezing and cold shock tolerance to plants

    International Nuclear Information System (INIS)

    Moon, Jeong Chan; Lee, Sangmin; Shin, Su Young; Chae, Ho Byoung; Jung, Young Jun; Jung, Hyun Suk; Lee, Kyun Oh; Lee, Jung Ro; Lee, Sang Yeol

    2015-01-01

    Overexpression of AtNTRC (AtNTRC OE ) in Arabidopsis thaliana led to a freezing and cold stress tolerance, whereas a knockout mutant (atntrc) showed a stress-sensitive phenotype. Biochemical analyses showed that the recombinant AtNTRC proteins exhibited a cryoprotective activity for malate dehydrogenase and lactic dehydrogenase. Furthermore, conclusive evidence of its interaction with nucleic acids in vitro is provided here on the basis of gel shift and electron microscopy analysis. Recombinant AtNTRC efficiently protected RNA and DNA from RNase A and metal catalyzed oxidation damage, respectively. The C-terminal thioredoxin domain is required for the nucleic acid–protein complex formation. From these results, it can be hypothesized that AtNTRC, which is known to be an electron donor of peroxiredoxin, contributes the stability of macromolecules under cold stress. - Highlights: • AtNTRC has a cryoprotective activity in vitro. • Overexpression of AtNTRC increases tolerance to freezing and cold shock stresses. • Thioredoxin domain of AtNTRC protects nucleic acids in vitro. • AtNTRC inhibits protein aggregation under freezing stress in vitro

  3. NADPH-dependent thioredoxin reductase C plays a role in nonhost disease resistance against Pseudomonas syringae pathogens by regulating chloroplast-generated reactive oxygen species

    Directory of Open Access Journals (Sweden)

    Yasuhiro Ishiga

    2016-04-01

    Full Text Available Chloroplasts are cytoplasmic organelles for photosynthesis in eukaryotic cells. In addition, recent studies have shown that chloroplasts have a critical role in plant innate immunity against invading pathogens. Hydrogen peroxide is a toxic by-product from photosynthesis, which also functions as a signaling compound in plant innate immunity. Therefore, it is important to regulate the level of hydrogen peroxide in response to pathogens. Chloroplasts maintain components of the redox detoxification system including enzymes such as 2-Cys peroxiredoxins (2-Cys Prxs, and NADPH-dependent thioredoxin reductase C (NTRC. However, the significance of 2-Cys Prxs and NTRC in the molecular basis of nonhost disease resistance is largely unknown. We evaluated the roles of Prxs and NTRC using knock-out mutants of Arabidopsis in response to nonhost Pseudomonas syringae pathogens. Plants lacking functional NTRC showed localized cell death (LCD accompanied by the elevated accumulation of hydrogen peroxide in response to nonhost pathogens. Interestingly, the Arabidopsis ntrc mutant showed enhanced bacterial growth and disease susceptibility of nonhost pathogens. Furthermore, the expression profiles of the salicylic acid (SA and jasmonic acid (JA-mediated signaling pathways and phytohormone analyses including SA and JA revealed that the Arabidopsis ntrc mutant shows elevated JA-mediated signaling pathways in response to nonhost pathogen. These results suggest the critical role of NTRC in plant innate immunity against nonhost P. syringae pathogens.

  4. Silencing of NADPH-Dependent Oxidoreductase Genes (yqhD and dkgA) in Furfural-Resistant Ethanologenic Escherichia coli▿

    Science.gov (United States)

    Miller, E. N.; Jarboe, L. R.; Yomano, L. P.; York, S. W.; Shanmugam, K. T.; Ingram, L. O.

    2009-01-01

    Low concentrations of furfural are formed as a side product during the dilute acid hydrolysis of hemicellulose. Growth is inhibited by exposure to furfural but resumes after the complete reduction of furfural to the less toxic furfuryl alcohol. Growth-based selection was used to isolate a furfural-resistant mutant of ethanologenic Escherichia coli LY180, designated strain EMFR9. Based on mRNA expression levels in the parent and mutant in response to furfural challenge, genes encoding 12 oxidoreductases were found to vary by more than twofold (eight were higher in EMFR9; four were higher in the parent). All 12 genes were cloned. When expressed from plasmids, none of the eight genes in the first group increased furfural tolerance in the parent (LY180). Expression of three of the silenced genes (yqhD, dkgA, and yqfA) in EMFR9 was found to decrease furfural tolerance compared to that in the parent. Purified enzymes encoded by yqhD and dkgA were shown to have NADPH-dependent furfural reductase activity. Both exhibited low Km values for NADPH (8 μM and 23 μM, respectively), similar to those of biosynthetic reactions. Furfural reductase activity was not associated with yqfA. Deleting yqhD and dkgA in the parent (LY180) increased furfural tolerance, but not to the same extent observed in the mutant EMFR9. Together, these results suggest that the process of reducing furfural by using an enzyme with a low Km for NADPH rather than a direct inhibitory action is the primary cause for growth inhibition by low concentrations of furfural. PMID:19429550

  5. Overexpression of chloroplast NADPH-dependent thioredoxin reductase in Arabidopsis enhances leaf growth and elucidates in-vivo function of reductase and thioredoxin domains

    Directory of Open Access Journals (Sweden)

    Jouni eToivola

    2013-10-01

    Full Text Available Plant chloroplasts have versatile thioredoxin systems including two thioredoxin reductases and multiple types of thioredoxins. Plastid-localized NADPH-dependent thioredoxin reductase (NTRC contains both reductase (NTRd and thioredoxin (TRXd domains in a single polypeptide and forms homodimers. To study the action of NTRC and NTRC domains in vivo, we have complemented the ntrc knockout line of Arabidopsis with the wild type and full-length NTRC genes, in which 2-Cys motifs either in NTRd, or in TRXd were inactivated. The ntrc line was also transformed either with the truncated NTRd or TRXd alone. Overexpression of wild-type NTRC promoted plant growth by increasing leaf size and biomass yield of the rosettes. Complementation of the ntrc line with the full-length NTRC gene containing an active reductase but an inactive thioredoxin domain, or vice versa, recovered wild-type chloroplast phenotype and, partly, rosette biomass production, indicating that the NTRC domains are capable of interacting with other chloroplast thioredoxin systems. Overexpression of truncated NTRd or TRXd in ntrc background did not restore wild-type phenotype. Modelling of the 3-dimensional structure of the NTRC dimer indicates extensive interactions between the NTR domains and the TRX domains further stabilize the dimeric structure. The long linker region between the NTRd and TRXd, however, allows flexibility for the position of the TRXd in the dimer. Supplementation of the TRXd in the NTRC homodimer model by free chloroplast thioredoxins indicated that TRXf is the most likely partner to interact with NTRC. We propose that overexpression of NTRC promotes plant biomass yield both directly by stimulation of chloroplast biosynthetic and protected pathways controlled by NTRC and indirectly via free chloroplast thioredoxins. Our data indicate that overexpression of chloroplast thiol redox-regulator has a potential to increase biofuel yield in plant and algal species suitable for

  6. Identification and cloning of an NADPH-dependent hydroxycinnamoyl-CoA double bond reductase involved in dihydrochalcone formation in Malus×domestica Borkh.

    Science.gov (United States)

    Ibdah, Mwafaq; Berim, Anna; Martens, Stefan; Valderrama, Andrea Lorena Herrera; Palmieri, Luisa; Lewinsohn, Efraim; Gang, David R

    2014-11-01

    The apple tree (Malus sp.) is an agriculturally and economically important source of food and beverages. Many of the health beneficial properties of apples are due to (poly)phenolic metabolites that they contain, including various dihydrochalcones. Although many of the genes and enzymes involved in polyphenol biosynthesis are known in many plant species, the specific reactions that lead to the biosynthesis of the dihydrochalcone precursor, p-dihydrocoumaroyl-CoA (3), are unknown. To identify genes involved in the synthesis of these metabolites, existing genome databases of the Rosaceae were screened for apple genes with significant sequence similarity to Arabidopsis alkenal double bond reductases. Herein described are the isolation and characterization of a Malus hydroxycinnamoyl-CoA double bond reductase, which catalyzed the NADPH-dependent reduction of p-coumaroyl-CoA and feruloyl-CoA to p-dihydrocoumaroyl-CoA and dihydroferuloyl-CoA, respectively. Its apparent Km values for p-coumaroyl-CoA, feruloyl-CoA and NADPH were 96.6, 92.9 and 101.3μM, respectively. The Malus double bond reductase preferred feruloyl-CoA to p-coumaroyl-CoA as a substrate by a factor of 2.1 when comparing catalytic efficiencies in vitro. Expression analysis of the hydroxycinnamoyl-CoA double bond reductase gene revealed that its transcript levels showed significant variation in tissues of different developmental stages, but was expressed when expected for involvement in dihydrochalcone formation. Thus, the hydroxycinnamoyl-CoA double bond reductase appears to be responsible for the reduction of the α,β-unsaturated double bond of p-coumaroyl-CoA, the first step of dihydrochalcone biosynthesis in apple tissues, and may be involved in the production of these compounds. Copyright © 2014 Elsevier Ltd. All rights reserved.

  7. Change of the NADPH depending superoxide producing and ferri hemoglobin reducing activities of cytochrome b558 from spleen cells and erythrocytes membranes induced by the radiation of different character

    International Nuclear Information System (INIS)

    Melkonyan, L.G.; Simonyan, R.M.; Simonyan, M.A.; Sekoyan, E.S.

    2009-01-01

    After the X radiation, UVA radiation and ultrasound radiation of new isoforms of cytochrome cyt b 5 58 from rats erythrocyte membranes - EM (cyt b 5 58III) and from spleen cell membranes (SCM) in vitro, as well as after the radiation of EM ex vivo, the suppression of both NADPH depending O 2 - producing and ferrihemoglobin (ferriHb)-reducing activities of cyt b 5 58 from EM and SCM in homogeneous (in solution) and heterogeneous phases (in EM and SCM) at various scopes takes place. These changes are associated with the destabilization of EM and SCM, conditioned by the change of the aggregation degree of these hemoproteins in EM and SCM, hemoproteins as a result of the influence of the hydrogen peroxide formed during radiolysis and photolysis of the water medium. After He-Ne laser radiation of the cyt b 5 58 from EM and SCM in vitro an increase of the NADPH depending O 2 - producing and ferriHb-reducing activities of the cyt b 5 58 from EM and SCM in homogenous and heterogeneous phases (in membranes) takes place. It is supposed that the suppression (by X-, UVA- and US-radiation) and the stimulation (by He-Ne laser radiation) of the immune system activity and the oxygen homeostasis are associated with the corresponding decrease and increase of the NADPH depending O 2 - producing and ferriHb-reducings activity of the new isoforms of cyt b 5 58 from EM and SCM in homogeneous and heterogeneous phases

  8. Molecular cloning and functional characterization of NADPH-dependent cytochrome P450 reductase from the green microalga Botryococcus braunii, B race.

    Science.gov (United States)

    Tsou, Chung-Yau; Matsunaga, Shigeki; Okada, Shigeru

    2018-01-01

    The green microalga Botryococcus braunii of the B race accumulates various lipophilic compounds containing a 10,11-oxidosqualene epoxide moiety in addition to large amounts of triterpene hydrocarbons. While 2,3-squalene epoxidases have already been isolated and characterized from the alga, the enzyme that catalyzes the 10,11-epoxidation of squalene has remained elusive. In order to obtain a molecular tool to explore a 10,11-squalene epoxidase, cDNA cloning of an NADPH-dependent cytochrome P450 reductase (CPR) that is required by both squalene epoxidases and cytochrome P450 enzymes was carried out. The isolated cDNA contained an open reading frame (1998 bp) that encoded for a protein with 665 amino acid residues with a predicted molecular weight of 71.46 kDa and a theoretical pI of 5.49. Analysis of the deduced amino acid sequence revealed the presence of conserved motifs, including FMN, FAD, and NADPH binding domains, which are typical of other CPRs and necessary for enzyme activity. By truncation of the N-terminal transmembrane anchor and addition of a 6× His-tag, BbCPR was heterologously produced in Escherichia coli and purified by Ni-NTA affinity chromatography. The purified recombinant enzyme showed optimal reducing activity of cytochrome c at around a neutral pH at a temperature range of 30-37°C. For steady state kinetic parameters, the recombinant enzyme had a k m for cytochrome c and NADPH of 11.7±1.6 and 9.4±1.4 μM, and a k cat for cytochrome c and NADPH of 2.78±0.09 and 3.66±0.11 μmol/min/mg protein, respectively. This is the first study to perform the functional characterization of a CPR from eukaryotic microalgae. Copyright © 2017 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

  9. The Study of NADPH-Dependent Flavoenzyme-Catalyzed Reduction of Benzo[1,2-c]1,2,5-oxadiazole N-Oxides (Benzofuroxans

    Directory of Open Access Journals (Sweden)

    Jonas Šarlauskas

    2014-12-01

    Full Text Available The enzymatic reactivity of a series of benzo[1,2-c]1,2,5-oxadiazole N-oxides (benzofuroxans; BFXs towards mammalian single-electron transferring NADPH:cytochrome P-450 reductase (P-450R and two-electron (hydride transferring NAD(PH:quinone oxidoreductase (NQO1 was examined in this work. Since the =N+ (→OO− moiety of furoxan fragments of BFXs bears some similarity to the aromatic nitro-group, the reactivity of BFXs was compared to that of nitro-aromatic compounds (NACs whose reduction mechanisms by these and other related flavoenzymes have been extensively investigated. The reduction of BFXs by both P-450R and NQO1 was accompanied by O2 uptake, which was much lower than the NADPH oxidation rate; except for annelated BFXs, whose reduction was followed by the production of peroxide. In order to analyze the possible quantitative structure-activity relationships (QSARs of the enzymatic reactivity of the compounds, their electron-accepting potency and other reactivity indices were assessed by quantum mechanical methods. In P-450R-catalyzed reactions, both BFXs and NACs showed the same reactivity dependence on their electron-accepting potency which might be consistent with an “outer sphere” electron transfer mechanism. In NQO1-catalyzed two-electron (hydride transferring reactions, BFXs acted as more efficient substrates than NACs, and the reduction efficacy of BFXs by NQO1 was in general higher than by single-electron transferring P-450R. In NQO1-catalyzed reactions, QSARs obtained showed that the reduction efficacy of BFXs, as well as that of NACs, was determined by their electron-accepting potency and could be influenced by their binding mode in the active center of NQO1 and by their global softness as their electronic characteristic. The reductive conversion of benzofuroxan by both flavoenzymes yielded the same reduction product of benzofuroxan, 2,3-diaminophenazine, with the formation of o-benzoquinone dioxime as a putative primary

  10. Carnitine palmityl transferase I deficiency

    NARCIS (Netherlands)

    Al-Aqeel, A. I.; Rashed, M. S.; Ruiter, J. P.; Al-Husseini, H. F.; Al-Amoudi, M. S.; Wanders, R. J.

    2001-01-01

    Carnitine palmityl transferase I is the key enzyme in the carnitine dependent transport of long chain fatty acids across the mitochondrial inner membrane and its deficiency results in a decrease rate of fatty acids beta-oxidation with decreased energy production. We reported a family of 3 affected

  11. Hibiscus cannabinus feruloyl-coa:monolignol transferase

    Science.gov (United States)

    Wilkerson, Curtis; Ralph, John; Withers, Saunia; Mansfield, Shawn D.

    2016-11-15

    The invention relates to isolated nucleic acids encoding a feruloyl-CoA:monolignol transferase and feruloyl-CoA:monolignol transferase enzymes. The isolated nucleic acids and/or the enzymes enable incorporation of monolignol ferulates into the lignin of plants, where such monolignol ferulates include, for example, p-coumaryl ferulate, coniferyl ferulate, and/or sinapyl ferulate. The invention also includes methods and plants that include nucleic acids encoding a feruloyl-CoA:monolignol transferase enzyme and/or feruloyl-CoA:monolignol transferase enzymes.

  12. The in vitro NADPH-dependent inhibition by CCl4 of the ATP-dependent calcium uptake of hepatic microsomes from male rats. Studies on the mechanism of the inactivation of the hepatic microsomal calcium pump by the CCl3 radical

    International Nuclear Information System (INIS)

    Srivastava, S.P.; Chen, N.Q.; Holtzman, J.L.

    1990-01-01

    The hepatotoxicity of CCl4 is mediated through its initial reduction by cytochrome P-450 to the CCl3 radical. This radical then damages important metabolic systems such as the ATP-dependent microsomal Ca2+ pump. Previous studies from our laboratory on isolated microsomes have shown that NADPH in the absence of toxic agents inhibits this pump. We have now found in in vitro incubations that CCl4 (0.5-2.5 mM) enhanced the NADPH-dependent inhibition of Ca2+ uptake from 28% without CCl4 to a maximum of 68%. These concentrations are in the range found in the livers and blood of lethally intoxicated animals and are toxic to cultured hepatocytes. The inhibition of Ca2+ uptake was due both to a decrease in the Ca2(+)-dependent ATPase and to an enhanced release of Ca2+ from the microsomes. The NADPH-dependent CCl4 inhibition was greater under N2 and was totally prevented by CO. GSH (1-10 mM) added during the incubation with CCl4 prevented the inhibition. This protection was also seen when the incubations were performed under nitrogen. When samples were preincubated with CCl4, the CCl4 metabolism was stopped, and then the Ca2+ uptake was determined; GSH reversed the CCl4 inhibition of Ca2+ uptake. This reversal showed saturation kinetics for GSH with two Km values of 0.315 and 93 microM when both the preincubation and the Ca2+ uptake were performed under air, and 0.512 and 31 microM when both were performed under nitrogen. Cysteine did not prevent the NADPH-dependent CCl4 inhibition of Ca2+ uptake. CCl4 increased lipid peroxidation in air, but no lipid peroxidation was seen under nitrogen. Lipid peroxidation was only modestly reversed by GSH. GSH did not remove 14C bound to samples preincubated with the 14CCl4

  13. Identification of the S-transferase like superfamily bacillithiol transferases encoded by Bacillus subtilis

    Science.gov (United States)

    Perera, Varahenage R.; Lapek, John D.; Newton, Gerald L.; Gonzalez, David J.; Pogliano, Kit

    2018-01-01

    Bacillithiol is a low molecular weight thiol found in Firmicutes that is analogous to glutathione, which is absent in these bacteria. Bacillithiol transferases catalyze the transfer of bacillithiol to various substrates. The S-transferase-like (STL) superfamily contains over 30,000 putative members, including bacillithiol transferases. Proteins in this family are extremely divergent and are related by structural rather than sequence similarity, leaving it unclear if all share the same biochemical activity. Bacillus subtilis encodes eight predicted STL superfamily members, only one of which has been shown to be a bacillithiol transferase. Here we find that the seven remaining proteins show varying levels of metal dependent bacillithiol transferase activity. We have renamed the eight enzymes BstA-H. Mass spectrometry and gene expression studies revealed that all of the enzymes are produced to varying levels during growth and sporulation, with BstB and BstE being the most abundant and BstF and BstH being the least abundant. Interestingly, several bacillithiol transferases are induced in the mother cell during sporulation. A strain lacking all eight bacillithiol transferases showed normal growth in the presence of stressors that adversely affect growth of bacillithiol-deficient strains, such as paraquat and CdCl2. Thus, the STL bacillithiol transferases represent a new group of proteins that play currently unknown, but potentially significant roles in bacillithiol-dependent reactions. We conclude that these enzymes are highly divergent, perhaps to cope with an equally diverse array of endogenous or exogenous toxic metabolites and oxidants. PMID:29451913

  14. SIKLODEKSTRIN GLIKOSIL TRANSFERASE DAN PEMANFAATANNYA DALAM INDUSTRI [Cyclodextrin Glycosyl Transferase and its application in industries

    Directory of Open Access Journals (Sweden)

    Budiasih Wahyuntari

    2005-12-01

    Full Text Available Cyclodextrin glycosyl transferase (CGT-ase is mainly produced by Bacilli. Systematical name of the enzyme is E.C. 2.4.1.19 a-1,4 glucan-4-glycosyl transferase. The enzyme catalyzes hydrolysis of starch intramolecular, and intermolecular transglycosylation of a-1,4, glucan chains. Cyclodextrins are a-1,4 linked cyclic oligosaccharides resulting from enzymatic degradation of starch by cyclodextrin glycosyl transferase through untramolecular transglycosylation. The major cyclodextrins are made up of 6, 7 and 8 glucopyranose units which are known as a-, b-, and y-cyclodextrin. All CGT-ase catalyze three kinds of cyclodextrins, the proportion of the cyclodextrins depends on the enzyme source and reaction conditions. The intermolecular transglycosylation ability of the enzyme has been applied in transfering glycosyl residues into suitable acceptor. Transglycosylation by the enzymes have been tested to improve solubility of some flavonoids and to favor precipitation ci some glycosides.

  15. Genomic organization of plant aminopropyl transferases.

    Science.gov (United States)

    Rodríguez-Kessler, Margarita; Delgado-Sánchez, Pablo; Rodríguez-Kessler, Gabriela Theresia; Moriguchi, Takaya; Jiménez-Bremont, Juan Francisco

    2010-07-01

    Aminopropyl transferases like spermidine synthase (SPDS; EC 2.5.1.16), spermine synthase and thermospermine synthase (SPMS, tSPMS; EC 2.5.1.22) belong to a class of widely distributed enzymes that use decarboxylated S-adenosylmethionine as an aminopropyl donor and putrescine or spermidine as an amino acceptor to form in that order spermidine, spermine or thermospermine. We describe the analysis of plant genomic sequences encoding SPDS, SPMS, tSPMS and PMT (putrescine N-methyltransferase; EC 2.1.1.53). Genome organization (including exon size, gain and loss, as well as intron number, size, loss, retention, placement and phase, and the presence of transposons) of plant aminopropyl transferase genes were compared between the genomic sequences of SPDS, SPMS and tSPMS from Zea mays, Oryza sativa, Malus x domestica, Populus trichocarpa, Arabidopsis thaliana and Physcomitrella patens. In addition, the genomic organization of plant PMT genes, proposed to be derived from SPDS during the evolution of alkaloid metabolism, is illustrated. Herein, a particular conservation and arrangement of exon and intron sequences between plant SPDS, SPMS and PMT genes that clearly differs with that of ACL5 genes, is shown. The possible acquisition of the plant SPMS exon II and, in particular exon XI in the monocot SPMS genes, is a remarkable feature that allows their differentiation from SPDS genes. In accordance with our in silico analysis, functional complementation experiments of the maize ZmSPMS1 enzyme (previously considered to be SPDS) in yeast demonstrated its spermine synthase activity. Another significant aspect is the conservation of intron sequences among SPDS and PMT paralogs. In addition the existence of microsynteny among some SPDS paralogs, especially in P. trichocarpa and A. thaliana, supports duplication events of plant SPDS genes. Based in our analysis, we hypothesize that SPMS genes appeared with the divergence of vascular plants by a processes of gene duplication and the

  16. The Genetic Architecture of Murine Glutathione Transferases.

    Directory of Open Access Journals (Sweden)

    Lu Lu

    Full Text Available Glutathione S-transferase (GST genes play a protective role against oxidative stress and may influence disease risk and drug pharmacokinetics. In this study, massive multiscalar trait profiling across a large population of mice derived from a cross between C57BL/6J (B6 and DBA2/J (D2--the BXD family--was combined with linkage and bioinformatic analyses to characterize mechanisms controlling GST expression and to identify downstream consequences of this variation. Similar to humans, mice show a wide range in expression of GST family members. Variation in the expression of Gsta4, Gstt2, Gstz1, Gsto1, and Mgst3 is modulated by local expression QTLs (eQTLs in several tissues. Higher expression of Gsto1 in brain and liver of BXD strains is strongly associated (P < 0.01 with inheritance of the B6 parental allele whereas higher expression of Gsta4 and Mgst3 in brain and liver, and Gstt2 and Gstz1 in brain is strongly associated with inheritance of the D2 parental allele. Allele-specific assays confirmed that expression of Gsto1, Gsta4, and Mgst3 are modulated by sequence variants within or near each gene locus. We exploited this endogenous variation to identify coexpression networks and downstream targets in mouse and human. Through a combined systems genetics approach, we provide new insight into the biological role of naturally occurring variants in GST genes.

  17. Glutathione transferase mimics : Micellar catalysis of an enzymic reaction

    NARCIS (Netherlands)

    Lindkvist, Björn; Weinander, Rolf; Engman, Lars; Koetse, Marc; Engberts, Jan B.F.N.; Morgenstern, Ralf

    1997-01-01

    Substances that mimic the enzyme action of glutathione transferases (which serve in detoxification) are described. These micellar catalysts enhance the reaction rate between thiols and activated halogenated nitroarenes as well as alpha,beta-unsaturated carbonyls. The nucleophilic aromatic

  18. The association between glutathione S-transferase P1 ...

    African Journals Online (AJOL)

    Mahmoud I. Mahmoud

    2011-08-10

    Aug 10, 2011 ... B-adrenergic receptor polymorphisms and response to salmeterol. Am J Respir Crit Care ... transferase Pi locus and association with susceptibility to bladder, testicular and prostate cancer. Carcinogenesis 1997;18(4):641–4.

  19. Homogentisate solanesyl transferase (HST) cDNA’s in maize

    Science.gov (United States)

    Maize white seedling 3 (w3) has served as a model albino-seedling mutant since its discovery in 1923. We show that the w3 phenotype is caused by disruptions in homogentisate solanesyl transferase (HST), an enzyme that catalyzes the committed step in plastoquinone-9 (PQ9) biosynthesis. This reaction ...

  20. Insecticide resistance and glutathione S-transferases in mosquitoes ...

    African Journals Online (AJOL)

    Mosquito glutathione S-transferases (GSTs) have received considerable attention in the last 20 years because of their role in insecticide metabolism producing resistance. Many different compounds, including toxic xenobiotics and reactive products of intracellular processes such as lipid peroxidation, act as GST substrates.

  1. Steroid sulfatase and sulfuryl transferase activities in human brain tumors

    Czech Academy of Sciences Publication Activity Database

    Kříž, L.; Bičíková, M.; Mohapl, M.; Hill, M.; Černý, Ivan; Hampl, R.

    2008-01-01

    Roč. 109, č. 1 (2008), s. 31-39 ISSN 0960-0760 Institutional research plan: CEZ:AV0Z40550506 Keywords : dehydroepiandrosterone * steroid sulfatase * steroid sulfuryl transferase * brain Subject RIV: CC - Organic Chemistry Impact factor: 2.827, year: 2008

  2. 21 CFR 862.1030 - Alanine amino transferase (ALT/SGPT) test system.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Alanine amino transferase (ALT/SGPT) test system... Test Systems § 862.1030 Alanine amino transferase (ALT/SGPT) test system. (a) Identification. An alanine amino transferase (ALT/SGPT) test system is a device intended to measure the activity of the...

  3. Changes in amino transferases and muscle proteins when treating pigmeat with ionising rays

    Energy Technology Data Exchange (ETDEWEB)

    Hamm, R; Hofmann, K; Gruenewald, T; Partmann, W [Bundesanstalt fuer Fleischforschung, Kulmbach (F.R. Germany). Inst. fuer Chemie und Physik; Bundesforschungsanstalt fuer Ernaehrung, Karlsruhe [F.R. Germany

    1975-01-01

    Slices of lean pigmeat were treated with electron beams doses of 0.2, 1.0 and 5.0 Mrad. Low irradiation doses led to an increase in the activity of aspartate amino transferase (GOT) and alanin amino transferase (GPT) in the tissue generally and in the sarcoplasm (juice expressed from the muscle). 5 Mrad caused a great reduction in the activity of GOT and GPT in the tissue and the sarcoplasm. It seems doubtful whether this inactivation is due to a destruction of enzyme sulphhydryl groups. Irradiating with 5 Mrad resulted in partial release of the mitochondrial GOT isozyme (GOTsub(M)) into the sarcoplasm. This indicates damage to the mitochondrial membranes by ionising radiation. Irradiating the pigmeat increased the pH of the tissue and lowered its water binding ability (increase in drip). Up to a dose of 1 Mrad the solubility of the sarcoplasmic proteins was not definitely affected, but 5 Mrad caused a considerable drop in protein solubility. Surprisingly a dose of even 5 Mrad did not change the total number of sulphhydryl groups present in the tissue. Sephadex thin layer electrophoresis showed that at 0.2 Mrad there was a drastic decrease in the myosin band and an increase in peptide fragments of low molecular weight, whilst actin was little changed.

  4. Changes in amino transferases and muscle proteins when treating pigmeat with ionising rays

    International Nuclear Information System (INIS)

    Hamm, R.; Hofmann, K.; Gruenewald, T.; Partmann, W.

    1975-01-01

    Slices of lean pigmeat were treated with electron beams doses of 0.2, 1.0 and 5.0 Mrad. Low irradiation doses led to an increase in the activity of aspartate amino transferase (GOT) and alanin amino transferase (GPT) in the tissue generally and in the sarcoplasm (juice expressed from the muscle). 5 Mrad caused a great reduction in the activity of GOT and GPT in the tissue and the sarcoplasm. It seems doubtful whether this inactivation is due to a destruction of enzyme sulphhydryl groups. Irradiating with 5 Mrad resulted in partial release of the mitochondrial GOT isozyme (GOTsub(M)) into the sarcoplasm. This indicates damage to the mitochondrial membranes by ionising radiation. Irradiating the pigmeat increased the pH of the tissue and lowered its water binding ability (increase in drip). Up to a dose of 1 Mrad the solubility of the sarcoplasmic proteins was not definitely affected, but 5 Mrad caused a considerable drop in protein solubility. Surprisingly a dose of even 5 Mrad did not change the total number of sulphhydryl groups present in the tissue. Sephadex thin layer electrophoresis showed that at 0.2 Mrad there was a drastic decrease in the myosin band and an increase in peptide fragments of low molecular weight, whilst actin was little changed. (orig.) [de

  5. Interaction of pleuromutilin derivatives with the ribosomal peptidyl transferase center

    DEFF Research Database (Denmark)

    Long, K. S.; Hansen, L. K.; Jakobsen, L.

    2006-01-01

    Tiamulin is a pleuromutilin antibiotic that is used in veterinary medicine. The recently published crystal structure of a tiamulin-50S ribosomal subunit complex provides detailed information about how this drug targets the peptidyl transferase center of the ribosome. To promote rational design...... mutant strain is resistant to tiamulin and pleuromutilin, but not valnemulin, implying that valnemulin is better able to withstand an altered rRNA binding surface around the mutilin core. This is likely due to additional interactions made between the valnemulin side chain extension and the rRNA binding...

  6. A glutathione s-transferase confers herbicide tolerance in rice

    Directory of Open Access Journals (Sweden)

    Tingzhang Hu

    2014-07-01

    Full Text Available Plant glutathione S-transferases (GSTs have been a focus of attention due to their role in herbicide detoxification. OsGSTL2 is a glutathione S-transferase, lambda class gene from rice (Oryza sativa L.. Transgenic rice plants over-expressing OsGSTL2 were generated from rice calli by the use of an Agrobacterium transformation system, and were screened by a combination of hygromycin resistance, PCR and Southern blot analysis. In the vegetative tissues of transgenic rice plants, the over-expression of OsGSTL2 not only increased levels of OsGSTL2 transcripts, but also GST and GPX expression, while reduced superoxide. Transgenic rice plants also showed higher tolerance to glyphosate and chlorsulfuron, which often contaminate agricultural fields. The findings demonstrate the detoxification role of OsGSTL2 in the growth and development of rice plants. It should be possible to apply the present results to crops for developing herbicide tolerance and for limiting herbicide contamination in the food chain.

  7. Interaction of Pleuromutilin Derivatives with the Ribosomal Peptidyl Transferase Center

    Science.gov (United States)

    Long, Katherine S.; Hansen, Lykke H.; Jakobsen, Lene; Vester, Birte

    2006-01-01

    Tiamulin is a pleuromutilin antibiotic that is used in veterinary medicine. The recently published crystal structure of a tiamulin-50S ribosomal subunit complex provides detailed information about how this drug targets the peptidyl transferase center of the ribosome. To promote rational design of pleuromutilin-based drugs, the binding of the antibiotic pleuromutilin and three semisynthetic derivatives with different side chain extensions has been investigated using chemical footprinting. The nucleotides A2058, A2059, G2505, and U2506 are affected in all of the footprints, suggesting that the drugs are similarly anchored in the binding pocket by the common tricyclic mutilin core. However, varying effects are observed at U2584 and U2585, indicating that the side chain extensions adopt distinct conformations within the cavity and thereby affect the rRNA conformation differently. An Escherichia coli L3 mutant strain is resistant to tiamulin and pleuromutilin, but not valnemulin, implying that valnemulin is better able to withstand an altered rRNA binding surface around the mutilin core. This is likely due to additional interactions made between the valnemulin side chain extension and the rRNA binding site. The data suggest that pleuromutilin drugs with enhanced antimicrobial activity may be obtained by maximizing the number of interactions between the side chain moiety and the peptidyl transferase cavity. PMID:16569865

  8. A novel method for screening the glutathione transferase inhibitors

    Directory of Open Access Journals (Sweden)

    Węgrzyn Grzegorz

    2009-03-01

    Full Text Available Abstract Background Glutathione transferases (GSTs belong to the family of Phase II detoxification enzymes. GSTs catalyze the conjugation of glutathione to different endogenous and exogenous electrophilic compounds. Over-expression of GSTs was demonstrated in a number of different human cancer cells. It has been found that the resistance to many anticancer chemotherapeutics is directly correlated with the over-expression of GSTs. Therefore, it appears to be important to find new GST inhibitors to prevent the resistance of cells to anticancer drugs. In order to search for glutathione transferase (GST inhibitors, a novel method was designed. Results Our results showed that two fragments of GST, named F1 peptide (GYWKIKGLV and F2 peptide (KWRNKKFELGLEFPNL, can significantly inhibit the GST activity. When these two fragments were compared with several known potent GST inhibitors, the order of inhibition efficiency (measured in reactions with 2,4-dinitrochlorobenzene (CDNB and glutathione as substrates was determined as follows: tannic acid > cibacron blue > F2 peptide > hematin > F1 peptide > ethacrynic acid. Moreover, the F1 peptide appeared to be a noncompetitive inhibitor of the GST-catalyzed reaction, while the F2 peptide was determined as a competitive inhibitor of this reaction. Conclusion It appears that the F2 peptide can be used as a new potent specific GST inhibitor. It is proposed that the novel method, described in this report, might be useful for screening the inhibitors of not only GST but also other enzymes.

  9. Acrolein-detoxifying isozymes of glutathione transferase in plants.

    Science.gov (United States)

    Mano, Jun'ichi; Ishibashi, Asami; Muneuchi, Hitoshi; Morita, Chihiro; Sakai, Hiroki; Biswas, Md Sanaullah; Koeduka, Takao; Kitajima, Sakihito

    2017-02-01

    Acrolein is a lipid-derived highly reactive aldehyde, mediating oxidative signal and damage in plants. We found acrolein-scavenging glutathione transferase activity in plants and purified a low K M isozyme from spinach. Various environmental stressors on plants cause the generation of acrolein, a highly toxic aldehyde produced from lipid peroxides, via the promotion of the formation of reactive oxygen species, which oxidize membrane lipids. In mammals, acrolein is scavenged by glutathione transferase (GST; EC 2.5.1.18) isozymes of Alpha, Pi, and Mu classes, but plants lack these GST classes. We detected the acrolein-scavenging GST activity in four species of plants, and purified an isozyme showing this activity from spinach (Spinacia oleracea L.) leaves. The isozyme (GST-Acr), obtained after an affinity chromatography and two ion exchange chromatography steps, showed the K M value for acrolein 93 μM, the smallest value known for acrolein-detoxifying enzymes in plants. Peptide sequence homology search revealed that GST-Acr belongs to the GST Tau, a plant-specific class. The Arabidopsis thaliana GST Tau19, which has the closest sequence similar to spinach GST-Acr, also showed a high catalytic efficiency for acrolein. These results suggest that GST plays as a scavenger for acrolein in plants.

  10. Analysis of glutathione S-transferase (M1, T1 and P1) gene ...

    African Journals Online (AJOL)

    Glutathione S-transferase enzymes are active in detoxifying a wide number of endogenous and exogenous chemical carcinogens and subsequently, are crucial in protecting the DNA. Several studies show some differences in association of glutathione S-transferase M1, T1 and P1 genetic polymorphisms with the risk of ...

  11. From glutathione transferase to pore in a CLIC

    CERN Document Server

    Cromer, B A; Morton, C J; Parker, M W; 10.1007/s00249-002-0219-1

    2002-01-01

    Many plasma membrane chloride channels have been cloned and characterized in great detail. In contrast, very little is known about intracellular chloride channels. Members of a novel class of such channels, called the CLICs (chloride intracellular channels), have been identified over the last few years. A striking feature of the CLIC family of ion channels is that they can exist in a water- soluble state as well as a membrane-bound state. A major step forward in understanding the functioning of these channels has been the recent crystal structure determination of one family member, CLIC1. The structure confirms that CLICs are members of the glutathione S- transferase superfamily and provides clues as to how CLICs can insert into membranes to form chloride channels. (69 refs).

  12. Electronics

    Science.gov (United States)

    2001-01-01

    International Acer Incorporated, Hsin Chu, Taiwan Aerospace Industrial Development Corporation, Taichung, Taiwan American Institute of Taiwan, Taipei, Taiwan...Singapore and Malaysia .5 - 4 - The largest market for semiconductor products is the high technology consumer electronics industry that consumes up...Singapore, and Malaysia . A new semiconductor facility costs around $3 billion to build and takes about two years to become operational

  13. Functional analysis and localisation of a delta-class glutathione S-transferase from Sarcoptes scabiei.

    Science.gov (United States)

    Pettersson, Eva U; Ljunggren, Erland L; Morrison, David A; Mattsson, Jens G

    2005-01-01

    The mite Sarcoptes scabiei causes sarcoptic mange, or scabies, a disease that affects both animals and humans worldwide. Our interest in S. scabiei led us to further characterise a glutathione S-transferase. This multifunctional enzyme is a target for vaccine and drug development in several parasitic diseases. The S. scabiei glutathione S-transferase open reading frame reported here is 684 nucleotides long and yields a protein with a predicted molecular mass of 26 kDa. Through phylogenetic analysis the enzyme was classified as a delta-class glutathione S-transferase, and our paper is the first to report that delta-class glutathione S-transferases occur in organisms other than insects. The recombinant S. scabiei glutathione S-transferase was expressed in Escherichia coli via three different constructs and purified for biochemical analysis. The S. scabiei glutathione S-transferase was active towards the substrate 1-chloro-2,4-dinitrobenzene, though the positioning of fusion partners influenced the kinetic activity of the enzyme. Polyclonal antibodies raised against S. scabiei glutathione S-transferase specifically localised the enzyme to the integument of the epidermis and cavities surrounding internal organs in adult parasites. However, some minor staining of parasite intestines was observed. No staining was seen in host tissues, nor could we detect any antibody response against S. scabiei glutathione S-transferase in sera from naturally S. scabiei infected dogs or pigs. Additionally, the polyclonal sera raised against recombinant S. scabiei glutathione S-transferase readily detected a protein from mites, corresponding to the predicted size of native glutathione S-transferase.

  14. The C-type Arabidopsis thioredoxin reductase ANTR-C acts as an electron donor to 2-Cys peroxiredoxins in chloroplasts

    International Nuclear Information System (INIS)

    Moon, Jeong Chan; Jang, Ho Hee; Chae, Ho Byoung; Lee, Jung Ro; Lee, Sun Yong; Jung, Young Jun; Shin, Mi Rim; Lim, Hye Song; Chung, Woo Sik; Yun, Dae-Jin; Lee, Kyun Oh; Lee, Sang Yeol

    2006-01-01

    2-Cys peroxiredoxins (Prxs) play important roles in the antioxidative defense systems of plant chloroplasts. In order to determine the interaction partner for these proteins in Arabidopsis, we used a yeast two-hybrid screening procedure with a C175S-mutant of Arabidopsis 2-Cys Prx-A as bait. A cDNA encoding an NADPH-dependent thioredoxin reductase (NTR) isotype C was identified and designated ANTR-C. We demonstrated that this protein effected efficient transfer of electrons from NADPH to the 2-Cys Prxs of chloroplasts. Interaction between 2-Cys Prx-A and ANTR-C was confirmed by a pull-down experiment. ANTR-C contained N-terminal TR and C-terminal Trx domains. It exhibited both TR and Trx activities and co-localized with 2-Cys Prx-A in chloroplasts. These results suggest that ANTR-C functions as an electron donor for plastidial 2-Cys Prxs and represents the NADPH-dependent TR/Trx system in chloroplasts

  15. The Fusarium oxysporum gnt2, Encoding a Putative N-Acetylglucosamine Transferase, Is Involved in Cell Wall Architecture and Virulence

    Science.gov (United States)

    López-Fernández, Loida; Ruiz-Roldán, Carmen; Pareja-Jaime, Yolanda; Prieto, Alicia; Khraiwesh, Husam; Roncero, M. Isabel G.

    2013-01-01

    With the aim to decipher the molecular dialogue and cross talk between Fusarium oxysporum f.sp. lycopersci and its host during infection and to understand the molecular bases that govern fungal pathogenicity, we analysed genes presumably encoding N-acetylglucosaminyl transferases, involved in glycosylation of glycoproteins, glycolipids, proteoglycans or small molecule acceptors in other microorganisms. In silico analysis revealed the existence of seven putative N-glycosyl transferase encoding genes (named gnt) in F. oxysporum f.sp. lycopersici genome. gnt2 deletion mutants showed a dramatic reduction in virulence on both plant and animal hosts. Δgnt2 mutants had αalterations in cell wall properties related to terminal αor β-linked N-acetyl glucosamine. Mutant conidia and germlings also showed differences in structure and physicochemical surface properties. Conidial and hyphal aggregation differed between the mutant and wild type strains, in a pH independent manner. Transmission electron micrographs of germlings showed strong cell-to-cell adherence and the presence of an extracellular chemical matrix. Δgnt2 cell walls presented a significant reduction in N-linked oligosaccharides, suggesting the involvement of Gnt2 in N-glycosylation of cell wall proteins. Gnt2 was localized in Golgi-like sub-cellular compartments as determined by fluorescence microscopy of GFP::Gnt2 fusion protein after treatment with the antibiotic brefeldin A or by staining with fluorescent sphingolipid BODIPY-TR ceramide. Furthermore, density gradient ultracentrifugation allowed co-localization of GFP::Gnt2 fusion protein and Vps10p in subcellular fractions enriched in Golgi specific enzymatic activities. Our results suggest that N-acetylglucosaminyl transferases are key components for cell wall structure and influence interactions of F. oxysporum with both plant and animal hosts during pathogenicity. PMID:24416097

  16. The Fusarium oxysporum gnt2, encoding a putative N-acetylglucosamine transferase, is involved in cell wall architecture and virulence.

    Directory of Open Access Journals (Sweden)

    Loida López-Fernández

    Full Text Available With the aim to decipher the molecular dialogue and cross talk between Fusarium oxysporum f.sp. lycopersci and its host during infection and to understand the molecular bases that govern fungal pathogenicity, we analysed genes presumably encoding N-acetylglucosaminyl transferases, involved in glycosylation of glycoproteins, glycolipids, proteoglycans or small molecule acceptors in other microorganisms. In silico analysis revealed the existence of seven putative N-glycosyl transferase encoding genes (named gnt in F. oxysporum f.sp. lycopersici genome. gnt2 deletion mutants showed a dramatic reduction in virulence on both plant and animal hosts. Δgnt2 mutants had αalterations in cell wall properties related to terminal αor β-linked N-acetyl glucosamine. Mutant conidia and germlings also showed differences in structure and physicochemical surface properties. Conidial and hyphal aggregation differed between the mutant and wild type strains, in a pH independent manner. Transmission electron micrographs of germlings showed strong cell-to-cell adherence and the presence of an extracellular chemical matrix. Δgnt2 cell walls presented a significant reduction in N-linked oligosaccharides, suggesting the involvement of Gnt2 in N-glycosylation of cell wall proteins. Gnt2 was localized in Golgi-like sub-cellular compartments as determined by fluorescence microscopy of GFP::Gnt2 fusion protein after treatment with the antibiotic brefeldin A or by staining with fluorescent sphingolipid BODIPY-TR ceramide. Furthermore, density gradient ultracentrifugation allowed co-localization of GFP::Gnt2 fusion protein and Vps10p in subcellular fractions enriched in Golgi specific enzymatic activities. Our results suggest that N-acetylglucosaminyl transferases are key components for cell wall structure and influence interactions of F. oxysporum with both plant and animal hosts during pathogenicity.

  17. The role of glutathione transferases in renal cell carcinoma

    Directory of Open Access Journals (Sweden)

    Ćorić Vesna

    2016-01-01

    Full Text Available Mounting evidence suggest that members of the subfamily of cytosolic glutathione S-transferases (GSTs possess roles far beyond the classical glutathione-dependent enzymatic conjugation of electrophilic metabolites and xenobiotics. Namely, monomeric forms of certain GSTs are capable of forming protein: protein interactions with protein kinases and regulate cell apoptotic pathways. Due to this dual functionality of cytosolic GSTs, they might be implicated in both the development and the progression of renal cell carcinoma (RCC. Prominent genetic heterogeneity, resulting from the gene deletions, as well as from SNPs in the coding and non-coding regions of GST genes, might affect GST isoenzyme profiles in renal parenchyma and therefore serve as a valuable indicator for predicting the risk of cancer development. Namely, GSTs are involved in the biotransformation of several compounds recognized as risk factors for RCC. The most potent carcinogen of polycyclic aromatic hydrocarbon diol epoxides, present in cigarette smoke, is of benzo(apyrene (BPDE, detoxified by GSTs. So far, the relationship between GST genotype and BPDE-DNA adduct formation, in determining the risk for RCC, has not been evaluated in patients with RCC. Although the association between certain individual and combined GST genotypes and RCC risk has been debated in a the literature, the data on the prognostic value of GST polymorphism in patients with RCC are scarce, probably due to the fact that the molecular mechanism supporting the role of GSTs in RCC progression has not been clarified as yet.

  18. Glutathione transferase-mediated benzimidazole-resistance in Fusarium graminearum.

    Science.gov (United States)

    Sevastos, A; Labrou, N E; Flouri, F; Malandrakis, A

    2017-09-01

    Fusarium graminearum laboratory mutants moderately (MR) and highly (HR) benzimidazole-resistant, carrying or not target-site mutations at the β 2 -tubulin gene were utilized in an attempt to elucidate the biochemical mechanism(s) underlying the unique BZM-resistance paradigm of this fungal plant pathogen. Relative expression analysis in the presence or absence of carbendazim (methyl-2-benzimidazole carbamate) using a quantitative Real Time qPCR (RT-qPCR) revealed differences between resistant and the wild-type parental strain although no differences in expression levels of either β 1 - or β 2 -tubulin homologue genes were able to fully account for two of the highly resistant phenotypes. Glutathione transferase (GST)-mediated detoxification was shown to be -at least partly- responsible for the elevated resistance levels of a HR isolate bearing the β 2 -tubulin Phe200Tyr resistance mutation compared with another MR isolate carrying the same mutation. This benzimidazole-resistance mechanism is reported for the first time in F. graminearum. No indications of detoxification involved in benzimidazole resistance were found for the rest of the isolates as revealed by GST and glutathione peroxidase (GPx) activities and bioassays using monoxygenase and hydrolase detoxification enzyme inhibiting synergists. Interestingly, besides the Phe200Tyr mutation-carrying HR isolate, the remaining highly-carbendazim resistant phenotypes could not be associated with any of the target site modification/overproduction, detoxification or reduced uptake-increased efflux mechanisms. Copyright © 2016 Elsevier B.V. All rights reserved.

  19. Glutathione Transferase GSTπ In Breast Tumors Evaluated By Three Techniques

    Directory of Open Access Journals (Sweden)

    Rafael Molina

    1993-01-01

    Full Text Available The glutathione transferases are involved in intracellular detoxification reactions. One of these, GSTπ, is elevated in some breast cancer cells, particularly cells selected for resistance to anticancer agents. We evaluated GSTπ expression in 60 human breast tumors by three techniques, immunohistochemistry, Northern hybridization, and Western blot analysis. There was a significant positive correlation between the three methods, with complete concordance seen in 64% of the tumors. There was strong, inverse relationship between GSTπ expression and steroid receptor status with all of the techniques utili zed. [n addition, there was a trend toward higher GSTπ expression in poorly differentiated tumors, but no correlation was found between tumor GSTπ content and DNA ploidy or %S-phase. GSTπ expression was also detected in adjacent benign breast tissue as well as infiltrating lymphocytes; this expression may contribute to GSTπ measurements using either Northern hybridization or Western blot analysis. These re sults suggest that immunohistochemistry is the method of choice for measuring GSTπ in breast tumors.

  20. Glycoproteins and sialyl transferase of human B lymphoblastoid cell lines

    International Nuclear Information System (INIS)

    Lui, S.W.L.; Ng, M.H.

    1980-01-01

    We used two radiolabeling methods to study glycoproteins on the surface of lymphoblastoid cells. One of the methods affects tritiation of residues which are oxidized with galactose oxidase and the other causes tritiation of neuraminic acid residues. This approach was shown to allow a better resolution of cell surface glycoproteins than if either method were used alone. Glycoproteins of B 1 - 19 cells which harbor the Epstein-Barr virus genomes were compared with those of its parental cell line, BJAB, which does not harbor the viral genomes. These studies did not reveal a unique viral protein. A 28,000 mol. wt. glycoprotein was found to be the most prominent neuraminic acidlabeled product of B 1 - 19 cells and also of the two other cell lines, Raji and Ly38, which harbor the EBV genomes. A similar molecular weight species from BJAB cells identified by galactose oxidase labeling might be deficient in neuraminic acid residues as it was poorly labeled by the periodate oxidation method. The neuraminic acid content and level of sialyl transferase of BJAB cells were found to be lower than those of the other cell lines studied. (auth.)

  1. S-Nitrosation destabilizes glutathione transferase P1-1.

    Science.gov (United States)

    Balchin, David; Stoychev, Stoyan H; Dirr, Heini W

    2013-12-23

    Protein S-nitrosation is a post-translational modification that regulates the function of more than 500 human proteins. Despite its apparent physiological significance, S-nitrosation is poorly understood at a molecular level. Here, we investigated the effect of S-nitrosation on the activity, structure, stability, and dynamics of human glutathione transferase P1-1 (GSTP1-1), an important detoxification enzyme ubiquitous in aerobes. S-Nitrosation at Cys47 and Cys101 reduces the activity of the enzyme by 94%. Circular dichroism spectroscopy, acrylamide quenching, and amide hydrogen-deuterium exchange mass spectrometry experiments indicate that the loss of activity is caused by the introduction of local disorder at the active site of GSTP1-1. Furthermore, the modification destabilizes domain 1 of GSTP1-1 against denaturation, smoothing the unfolding energy landscape of the protein and introducing a refolding defect. In contrast, S-nitrosation at Cys101 alone introduces a refolding defect in domain 1 but compensates by stabilizing the domain kinetically. These data elucidate the physical basis for the regulation of GSTP1-1 by S-nitrosation and provide general insight into the consequences of S-nitrosation on protein stability and dynamics.

  2. Acetate:succinate CoA-transferase in the hydrogenosomes of Trichomonas vaginalis: Identification and characterization

    NARCIS (Netherlands)

    K.W.A. Grinsven; S. Rosnowsky (Silke); S.W.H. van Weelden (Susanne); S. Pütz (Simone); M. van der Giezen (Mark); W. Martin (William); J.J. van Hellemond (Jaap); A.G.M. Tielens (Aloysius); K. Henze (Katrin)

    2008-01-01

    textabstractAcetate:succinate CoA-transferases (ASCT) are acetate-producing enzymes in hydrogenosomes, anaerobically functioning mitochondria and in the aerobically functioning mitochondria of trypanosomatids. Although acetate is produced in the hydrogenosomes of a number of anaerobic microbial

  3. Glutathione S-transferase gene polymorphisms in presbycusis.

    Science.gov (United States)

    Ateş, Nurcan Aras; Unal, Murat; Tamer, Lülüfer; Derici, Ebru; Karakaş, Sevim; Ercan, Bahadir; Pata, Yavuz Selim; Akbaş, Yücel; Vayisoğlu, Yusuf; Camdeviren, Handan

    2005-05-01

    Glutathione and glutathione-related antioxidant enzymes are involved in the metabolism and detoxification of cytotoxic and carcinogenic compounds as well as reactive oxygen species. Reactive oxygen species generation occurs in prolonged relative hypoperfusion conditions such as in aging. The etiology of presbycusis is much less certain; however, a complex genetic cause is most likely. The effect of aging shows a wide interindividual range; we aimed to investigate whether profiles of (glutathione S-transferase (GST) M1, T1 and P1 genotypes may be associated with the risk of age-related hearing loss. We examined 68 adults with presbycusis and 69 healthy controls. DNA was extracted from whole blood, and the GSTM1, GSTT1 and GSTP1 polymorphisms were determined using a real-time polymerase chain reaction and fluorescence resonance energy transfer with a Light-Cycler Instrument. Associations between specific genotypes and the development of presbycusis were examined by use of logistic regression analyses to calculate odds ratios and 95% confidence intervals. Gene polymorphisms at GSTM1, GSTT1, and GSTP1 in subjects with presbycusis were not significantly different than in the controls (p > 0.05). Also, the combinations of different GSTM1, GSTT1, and GSTP1 genotypes were not an increased risk of presbycusis (p > 0.05). We could not demonstrate any significant association between the GSTM1, GSTT1, and GSTP1 polymorphism and age-related hearing loss in this population. This may be because of our sample size, and further studies need to investigate the exact role of GST gene polymorphisms in the etiopathogenesis of the presbycusis.

  4. Human glutathione transferases catalyzing the bioactivation of anticancer thiopurine prodrugs.

    Science.gov (United States)

    Eklund, Birgitta I; Gunnarsdottir, Sjofn; Elfarra, Adnan A; Mannervik, Bengt

    2007-06-01

    cis-6-(2-Acetylvinylthio)purine (cAVTP) and trans-6-(2-acetylvinylthio)guanine (tAVTG) are thiopurine prodrugs provisionally inactivated by an alpha,beta-unsaturated substituent on the sulfur of the parental thiopurines 6-mercaptopurine (6-MP) and 6-thioguanine (6-TG). The active thiopurines are liberated intracellularly by glutathione (GSH) in reactions catalyzed by glutathione transferases (GSTs) (EC 2.5.1.18). Catalytic activities of 13 human GSTs representing seven distinct classes of soluble GSTs have been determined. The bioactivation of cAVTP and tAVTG occurs via a transient addition of GSH to the activated double bond of the S-substituent of the prodrug, followed by elimination of the thiopurine. The first of these consecutive reactions is rate-limiting for thiopurine release, but GST-activation of this first addition is shifting the rate limitation to the subsequent elimination. Highly active GSTs reveal the transient intermediate, which is detectable by UV spectroscopy and HPLC analysis. LC/MS analysis of the reaction products demonstrates that the primary GSH conjugate, 4-glutathionylbuten-2-one, can react with a second GSH molecule to form the 4-(bis-glutathionyl)butan-2-one. GST M1-1 and GST A4-4 were the most efficient enzymes with tAVTG, and GST M1-1 and GST M2-2 had highest activity with cAVTP. The highly efficient GST M1-1 is polymorphic and is absent in approximately half of the human population. GST P1-1, which is overexpressed in many cancer cells, had no detectable activity with cAVTP and only minor activity with tAVTG. Other GST-activated prodrugs have targeted GST P1-1-expressing cancer cells. Tumors expressing high levels of GST M1-1 or GST A4-4 can be predicted to be particularly vulnerable to chemotherapy with cAVTP or tAVTG.

  5. The Role of Mitochondrial NADPH-Dependent Isocitrate Dehydrogenase in Cancer Cells

    Directory of Open Access Journals (Sweden)

    Katarína Smolková

    2012-01-01

    Full Text Available Isocitrate dehydrogenase 2 (IDH2 is located in the mitochondrial matrix. IDH2 acts in the forward Krebs cycle as an NADP+-consuming enzyme, providing NADPH for maintenance of the reduced glutathione and peroxiredoxin systems and for self-maintenance by reactivation of cystine-inactivated IDH2 by glutaredoxin 2. In highly respiring cells, the resulting NAD+ accumulation then induces sirtuin-3-mediated activating IDH2 deacetylation, thus increasing its protective function. Reductive carboxylation of 2-oxoglutarate by IDH2 (in the reverse Krebs cycle direction, which consumes NADPH, may follow glutaminolysis of glutamine to 2-oxoglutarate in cancer cells. When the reverse aconitase reaction and citrate efflux are added, this overall “anoxic” glutaminolysis mode may help highly malignant tumors survive aglycemia during hypoxia. Intermittent glycolysis would hypothetically be required to provide ATP. When oxidative phosphorylation is dormant, this mode causes substantial oxidative stress. Arg172 mutants of human IDH2—frequently found with similar mutants of cytosolic IDH1 in grade 2 and 3 gliomas, secondary glioblastomas, and acute myeloid leukemia—catalyze reductive carboxylation of 2-oxoglutarate and reduction to D-2-hydroxyglutarate, which strengthens the neoplastic phenotype by competitive inhibition of histone demethylation and 5-methylcytosine hydroxylation, leading to genome-wide histone and DNA methylation alternations. D-2-hydroxyglutarate also interferes with proline hydroxylation and thus may stabilize hypoxia-induced factor α.

  6. NADPH-dependent D-aldose reductases and xylose fermentation in Fusarium oxysporum

    DEFF Research Database (Denmark)

    Panagiotou, Gianni; Christakopoulos, P.

    2004-01-01

    Two aldose (xylose) reductases (ARI and ARII) from Fusarium oxysporum were purified and characterized. The native ARI was a monomer with M-r 41000, pI 5.2 and showed a 52-fold preference for NADPH over NADH, while ARII was homodimeric with a subunit of M-r 37000, pI 3.6 and a 60-fold preference...

  7. Structure of Hordeum vulgare NADPH-dependent thioredoxin reductase 2. Unwinding the reaction mechanism

    DEFF Research Database (Denmark)

    Kirkensgaard, Kristine Groth; Hägglund, Per; Finnie, Christine

    2009-01-01

    to the active form. Here, the first crystal structure of a cereal NTR, HvNTR2 from Hordeum vulgare (barley), is presented, which is also the first structure of a monocot plant NTR. The structure was determined at 2.6 A resolution and refined to an R (cryst) of 19.0% and an R (free) of 23.8%. The dimeric protein...

  8. The Role of Mitochondrial NADPH-Dependent Isocitrate Dehydrogenase in Cancer Cells

    Czech Academy of Sciences Publication Activity Database

    Smolková, Katarína; Ježek, Petr

    2012-01-01

    Roč. 2012, č. 2012 (2012), ID273947 ISSN 1687-8876 R&D Projects: GA ČR GPP301/12/P381; GA ČR(CZ) GAP302/10/0346 Institutional research plan: CEZ:AV0Z50110509 Institutional support: RVO:67985823 Keywords : isocitrate dehydrogenase 2 * Krebs cycle * cancer cells Subject RIV: ED - Physiology

  9. Presence of a Ca2+-sensitive CDPdiglyceride-inositol transferase in canine cardiac sarcoplasmic reticulum

    International Nuclear Information System (INIS)

    Kasinathan, C.; Kirchberger, M.A.

    1988-01-01

    Sarcoplasmic reticulum (SR) and plasma membranes from canine left ventricle were used to evaluate the presence of the enzyme CDPdiglyceride-inositol transferase in these membranes. (K + ,-Ca 2+ )-ATPase activity, a marker for SR, was 79.2 +/- 5.0 (SE) and 11.2 +/- 2.0 μmol x mg -1 x h -1 in SR and plasma membrane preparations, respectively, and (Na + , K + )-ATPase activity, a marker for plasma membranes, was 5.6 +/- 1.2 and 99.2 +/- 8.0 μmol x mg -1 x h -1 , respectively. Contamination of SR and plasma membrane preparations by mitochondria was estimated to be 2% and 8%, respectively, and by Golgi membranes, 0.9% and 1.8%, respectively. The transferase activity detected in the plasma membrane preparation could be accounted for largely, but not entirely, by contaminating SR membranes. The pH optimum for the SR transferase activity was between 8.0 and 9.0. Ca 2+ inhibited the enzyme, half-maximal inhibition occurring at about 10 μM Ca 2+ . No loss of [ 3 H]PtdIns could be detected when membranes were incubated in the presence or absence of Ca 2+ . The Ca 2+ inhibition of the transferase was noncompetitive with respect to CDP-dipalmitin while that with respect to myo-inositol was slightly noncompetitive at low [Ca 2+ ] and became uncompetitive at higher [Ca 2+ ]. It is concluded that CDPdiglyceride-inositol transferase is present on SR membranes and is sensitive to micromolar Ca 2+ . The data are consistent with a putative role for the inhibition of the SR transferase by Ca 2+ and acidic pH in the protection of the SR against calcium overload in ischemic myocardium

  10. Epidermal growth factor regulation of glutathione S-transferase gene expression in the rat is mediated by class Pi glutathione S-transferase enhancer I.

    OpenAIRE

    Matsumoto, M; Imagawa, M; Aoki, Y

    2000-01-01

    Using chloramphenicol acetyltransferase assays we showed that epidermal growth factor (EGF), transforming growth factor alpha (TGF alpha), and 3,3',4,4',5-pentachlorobiphenyl (PenCB) induce class Pi glutathione S-transferase (GSTP1) in primary cultured rat liver parenchymal cells. GSTP1 enhancer I (GPEI), which is required for the stimulation of GSTP1 expression by PenCB, also mediates EGF and TGF alpha stimulation of GSTP1 gene expression. However, hepatocyte growth factor and insulin did no...

  11. Purification of human hepatic glutathione S-transferases and the development of a radioimmunoassay for their measurement in plasma

    International Nuclear Information System (INIS)

    Hayes, J.D.; Gilligan, D.; Beckett, G.J.

    1983-01-01

    A purification scheme is described for six human hepatic glutathione S-transferases from a single liver. Five of the transferases comprised Ya monomers and had a molecular mass of 44000. The remaining enzyme comprised Yb monomers and had a molecular mass of 47000. Data are presented demonstrating that there are at least two distinct Ya monomers. A radioimmunoassay has been developed that has sufficient precision and sensitivity to allow direct measurement of glutathione S-transferase concentrations in unextracted plasma. A comparison of aminotransferase and glutathione S-transferase levels, in three patients who had taken a paracetamol overdose, indicated that glutathione S-transferase measurements provided a far more sensitive index of hepatocellular integrity than the more conventional aminotransferase measurements. (Auth.)

  12. Purification of human hepatic glutathione S-transferases and the development of a radioimmunoassay for their measurement in plasma

    Energy Technology Data Exchange (ETDEWEB)

    Hayes, J.D.; Gilligan, D.; Beckett, G.J. (Edinburgh Univ. (UK). Dept. of Clinical Chemistry); Chapman, B.J. (Royal Infirmary, Edinburgh (UK))

    1983-10-31

    A purification scheme is described for six human hepatic glutathione S-transferases from a single liver. Five of the transferases comprised Ya monomers and had a molecular mass of 44000. The remaining enzyme comprised Yb monomers and had a molecular mass of 47000. Data are presented demonstrating that there are at least two distinct Ya monomers. A radioimmunoassay has been developed that has sufficient precision and sensitivity to allow direct measurement of glutathione S-transferase concentrations in unextracted plasma. A comparison of aminotransferase and glutathione S-transferase levels, in three patients who had taken a paracetamol overdose, indicated that glutathione S-transferase measurements provided a far more sensitive index of hepatocellular integrity than the more conventional aminotransferase measurements.

  13. Glutathione s-transferase isoenzymes in relation to their role in detoxification of xenobiotics

    NARCIS (Netherlands)

    Vos, R.M.E.

    1989-01-01

    The glutathione S-transferases (GST) are a family of isoenzymes serving a major part in the biotransformation of many reactive compounds. The isoenzymes from rat, man and mouse are divided into three classes, alpha, mu and pi, on the basis of similar structural and enzymatic

  14. Isolation and characterization of an auxin-inducible glutathione S-transferase gene of Arabidopsis thaliana

    NARCIS (Netherlands)

    Kop, D.A.M. van der; Schuyer, M.; Scheres, B.J.G.; Zaal, B.J. van der; Hooykaas, P.J.J.

    1996-01-01

    Genes homologous to the auxin-inducible Nt103 glutathione S-transferase (GST) gene of tobacco, were isolated from a genomic library of Arabidopsis thaliana. We isolated a λ clone containing an auxin-inducible gene, At103-1a, and part of a constitutively expressed gene, At103-1b. The coding regions

  15. Global deletion of glutathione S-Transferase A4 exacerbates developmental nonalcoholic steatohepatitis

    Science.gov (United States)

    We established a mouse model of developmental nonalcoholic steatohepatitis (NASH) by feeding a high polyunsaturated fat liquid diet to female glutathione-S-transferase 4-4 (Gsta4-/-)/peroxisome proliferator activated receptor a (Ppara-/-) double knockout 129/SvJ mice for 12 weeks from weaning. We us...

  16. Involvement of human glutathione S-transferase isoenzymes in the conjugation of cyclophosphamide metabolites with glutathione

    NARCIS (Netherlands)

    Dirven, H.A.A.M.; Ommen, B. van; Bladeren, P.J. van

    1994-01-01

    Alkylating agents can be detoxified by conjugation with glutathione (GSH). One of the physiological significances of this lies in the observation that cancer cells resistant to the cytotoxic effects of alkylating agents have higher levels of GSH and high glutathione S-transferase (GST) activity.

  17. The role of glutathione S-transferase and claudin-1 gene polymorphisms in contact sensitization

    DEFF Research Database (Denmark)

    Ross-Hansen, K; Linneberg, A; Johansen, J D

    2013-01-01

    BACKGROUND: Contact sensitization is frequent in the general population and arises from excessive or repeated skin exposure to chemicals and metals. However, little is known about its genetic susceptibility. OBJECTIVES: To determine the role of polymorphisms of glutathione S-transferase (GST) genes...

  18. Effects of curcumin on cytochrome P450 and glutathione S-transferase activities in rat liver.

    NARCIS (Netherlands)

    Oetari, S.; Sudibyo, M.; Commandeur, J.N.M.; Samhoedi, R.; Vermeulen, N.P.E.

    1996-01-01

    The stability of curcumin, as well as the interactions between curcumin and cytochrome P450s (P450s) and glutathione S-transferases (GSTs) in rat liver, were studied. Curcumin is relatively unstable in phosphate buffer at pH 7.4. The stability of curcumin was strongly improved by lowering the pH or

  19. Bisubstrate Kinetics of Glutathione S-Transferase: A Colorimetric Experiment for the Introductory Biochemistry Laboratory

    Science.gov (United States)

    Stefanidis, Lazaros; Scinto, Krystal V.; Strada, Monica I.; Alper, Benjamin J.

    2018-01-01

    Most biochemical transformations involve more than one substrate. Bisubstrate enzymes catalyze multiple chemical reactions in living systems and include members of the transferase, oxidoreductase, and ligase enzyme classes. Working knowledge of bisubstrate enzyme kinetic models is thus of clear importance to the practicing biochemist. However,…

  20. Association study on glutathione S-transferase omega 1 and 2 and familial ALS

    NARCIS (Netherlands)

    van de Giessen, Elsmarieke; Fogh, Isabella; Gopinath, Sumana; Smith, Bradley; Hu, Xun; Powell, John; Andersen, Peter; Nicholson, Garth; Al Chalabi, Ammar; Shaw, Christopher E.

    2008-01-01

    Glutathione S-transferase omega 1 and 2 (GSTO1 and 2) protect from oxidative stress, a possible pathogenic mechanism underlying the pathogenesis of neurodegenerative diseases such as amyotrophic lateral sclerosis (ALS) and Alzheimer's disease. Significant association of age of onset in Alzheimer's

  1. Primary overproduction of urate caused by a partial deficiency of hypoxanthine-guanine phosphoribosyl transferase

    International Nuclear Information System (INIS)

    Cassidy, M.; Gregory, M.C.; Harley, E.H.

    1980-01-01

    Inherited enzyme deficiencies are found in a small proportion of patients with gout who produce an excess of uric acid. The clinical, biochemical and therapeutic aspects of a case of hyperuricaemia caused by an atypical mutant hypoxanthine-guanine phophoribosyl transferase are presented. Urate overproduction was moderate and controlled by allopurinol therapy

  2. Glutathione S-transferase genotype and p53 mutations in adenocarcinoma of the small intestine

    DEFF Research Database (Denmark)

    Pedersen, Lisbeth Nørum; Kaerlev, L; Stubbe Teglbjaerg, P

    2003-01-01

    Adenocarcinoma of the small intestine (ASI) is a rare disease of unknown aetiology. The glutathione S-transferase M1 (GSTM1) enzyme catalyses the detoxification of compounds involved in carcinogenesis of adenocarcinoma of the stomach, colon and lung, including constituents of tobacco smoke. We in...

  3. Antibiotic inhibition of the movement of tRNA substrates through a peptidyl transferase cavity

    DEFF Research Database (Denmark)

    Porse, B T; Rodriguez-Fonseca, C; Leviev, I

    1996-01-01

    The present review attempts to deal with movement of tRNA substrates through the peptidyl transferase centre on the large ribosomal subunit and to explain how this movement is interrupted by antibiotics. It builds on the concept of hybrid tRNA states forming on ribosomes and on the observed movem...

  4. Inhibition of rat, mouse, and human glutathione S-transferase by eugenol and its oxidation products

    NARCIS (Netherlands)

    Rompelberg, C.J.M.; Ploemen, J.H.T.M.; Jespersen, S.; Greef, J. van der; Verhagen, H.; Bladeren, P.J. van

    1996-01-01

    The irreversible and reversible inhibition of glutathione S-transferases (GSTs) by eugenol was studied in rat, mouse and man. Using liver cytosol of human, rat and mouse, species differences were found in the rate of irreversible inhibition of GSTs by eugenol in the presence of the enzyme

  5. Functional characterization of glutathione S-transferases associated with insecticide resistance in Tetranychus urticae

    NARCIS (Netherlands)

    Pavlidi, N.; Tseliou, V.; Riga, M.; Nauen, R.; Van Leeuwen, T.; Labrou, N.E.; Vontas, J.

    2015-01-01

    The two-spotted spider mite Tetranychus urticae is one of the most important agricultural pests world-wide. It is extremely polyphagous and develops resistance to acaricides. The overexpression of several glutathione S-transferases (GSTs) has been associated with insecticide resistance. Here, we

  6. Photoaffinity labelling of the active site of the rat glutathione transferases 3-3 and 1-1 and human glutathione transferase A1-1.

    OpenAIRE

    Cooke, R J; Björnestedt, R; Douglas, K T; McKie, J H; King, M D; Coles, B; Ketterer, B; Mannervik, B

    1994-01-01

    The glutathione transferases (GSTs) form a group of enzymes responsible for a wide range of molecular detoxications. The photoaffinity label S-(2-nitro-4-azidophenyl)glutathione was used to study the hydrophobic region of the active site of the rat liver GST 1-1 and 2-2 isoenzymes (class Alpha) as well as the rat class-Mu GST 3-3. Photoaffinity labelling was carried out using a version of S-(2-nitro-4-azidophenyl)glutathione tritiated in the arylazido ring. The labelling occurred with higher ...

  7. A novel plant glutathione S-transferase/peroxidase suppresses Bax lethality in yeast

    DEFF Research Database (Denmark)

    Kampranis, S C; Damianova, R; Atallah, M

    2000-01-01

    The mammalian inducer of apoptosis Bax is lethal when expressed in yeast and plant cells. To identify potential inhibitors of Bax in plants we transformed yeast cells expressing Bax with a tomato cDNA library and we selected for cells surviving after the induction of Bax. This genetic screen allows...... for the identification of plant genes, which inhibit either directly or indirectly the lethal phenotype of Bax. Using this method a number of cDNA clones were isolated, the more potent of which encodes a protein homologous to the class theta glutathione S-transferases. This Bax-inhibiting (BI) protein was expressed...... in Escherichia coli and found to possess glutathione S-transferase (GST) and weak glutathione peroxidase (GPX) activity. Expression of Bax in yeast decreases the intracellular levels of total glutathione, causes a substantial reduction of total cellular phospholipids, diminishes the mitochondrial membrane...

  8. Lectin Domains of Polypeptide GalNAc Transferases Exhibit Glycopeptide Binding Specificity

    DEFF Research Database (Denmark)

    Pedersen, Johannes W; Bennett, Eric P; Schjoldager, Katrine T-B G

    2011-01-01

    UDP-GalNAc:polypeptide a-N-acetylgalactosaminyltransferases (GalNAc-Ts) constitute a family of up to 20 transferases that initiate mucin-type O-glycosylation. The transferases are structurally composed of catalytic and lectin domains. Two modes have been identified for the selection...... of glycosylation sites by GalNAc-Ts: confined sequence recognition by the catalytic domain alone, and concerted recognition of acceptor sites and adjacent GalNAc-glycosylated sites by the catalytic and lectin domains, respectively. Thus far, only the catalytic domain has been shown to have peptide sequence...... on sequences of mucins MUC1, MUC2, MUC4, MUC5AC, MUC6, and MUC7 as well as a random glycopeptide bead library, we examined the binding properties of four different lectin domains. The lectin domains of GalNAc-T1, -T2, -T3, and -T4 bound different subsets of small glycopeptides. These results indicate...

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

    DEFF Research Database (Denmark)

    Song, Lina; Bachert, Collin; Schjoldager, Katrine T

    2014-01-01

    sequence influenced their activity and required modification, which we carried out based on previous in vitro work. Significantly, the modified T2 and T3 sensors were activated only in cells lacking their corresponding isozymes. Thus, we have developed T2- and T3-specific sensors that will be valuable......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....... Surprisingly, molecular probes to monitor GalNAc-transferase activity are lacking and there exist no effective global or isoform-specific inhibitors. Here we describe the development of T2- and T3-isoform specific fluorescence sensors that traffic in the secretory pathway. Each sensor yielded little signal...

  10. Caractérisation biochimique et fonctionnelle de glutathion-S-transferases (GSTs) chez Phanerochaete chrysosporium

    OpenAIRE

    Anak Ngadin , Andrew

    2011-01-01

    Phanerochaete chrysosporium is a ligninolytic fungus widely studied because of its capacities to degrade wood and xenobiotics through an extracellular enzymatic system. Its genome has been sequenced and has provided researchers with a complete inventory of the predicted proteins produced by this organism. This has allowed the description of many protein superfamilies. Among them, Glutathione S-transferases (GSTs) constitute a complex and widespread superfamily classified as enzymes of seconda...

  11. Imidazopyridine and Pyrazolopiperidine Derivatives as Novel Inhibitors of Serine Palmitoyl Transferase.

    Science.gov (United States)

    Genin, Michael J; Gonzalez Valcarcel, Isabel C; Holloway, William G; Lamar, Jason; Mosior, Marian; Hawkins, Eric; Estridge, Thomas; Weidner, Jeffrey; Seng, Thomas; Yurek, David; Adams, Lisa A; Weller, Jennifer; Reynolds, Vincent L; Brozinick, Joseph T

    2016-06-23

    To develop novel treatments for type 2 diabetes and dyslipidemia, we pursued inhibitors of serine palmitoyl transferase (SPT). To this end compounds 1 and 2 were developed as potent SPT inhibitors in vitro. 1 and 2 reduce plasma ceramides in rodents, have a slight trend toward enhanced insulin sensitization in DIO mice, and reduce triglycerides and raise HDL in cholesterol/cholic acid fed rats. Unfortunately these molecules cause a gastric enteropathy after chronic dosing in rats.

  12. The phosphopantetheinyl transferases: catalysis of a post-translational modification crucial for life

    DEFF Research Database (Denmark)

    Beld, Joris; Sonnenschein, Eva; Vickery, Christopher R.

    2013-01-01

    Covering: up to 2013 Although holo-acyl carrier protein synthase, AcpS, a phosphopantetheinyl transferase (PPTase), was characterized in the 1960s, it was not until the publication of the landmark paper by Lambalot et al. in 1996 that PPTases garnered wide-spread attention being classified...... knowledge on this class of enzymes that post-translationally install a 4′-phosphopantetheine arm on various carrier proteins....

  13. A 4'-phosphopantetheinyl transferase mediates non-ribosomal peptide synthetase activation in Aspergillus fumigatus.

    Science.gov (United States)

    Neville, Claire; Murphy, Alan; Kavanagh, Kevin; Doyle, Sean

    2005-04-01

    Aspergillus fumigatus is a significant human pathogen. Non-ribosomal peptide (NRP) synthesis is thought to be responsible for a significant proportion of toxin and siderophore production in the organism. Furthermore, it has been shown that 4'-phosphopantetheinylation is required for the activation of key enzymes involved in non-ribosomal peptide synthesis in other species. Here we report the cloning, recombinant expression and functional characterisation of a 4'-phosphopantetheinyl transferase from A. fumigatus and the identification of an atypical NRP synthetase (Afpes1), spanning 14.3 kb. Phylogenetic analysis has shown that the NRP synthetase exhibits greatest identity to NRP synthetases from Metarhizium anisolpiae (PesA) and Alternaria brassicae (AbrePsy1). Northern hybridisation and RT-PCR analysis have confirmed that both genes are expressed in A. fumigatus. A 120 kDa fragment of the A. fumigatus NRP synthetase, containing a putative thiolation domain, was cloned and expressed in the baculovirus expression system. Detection of a 4'-phosphopantetheinylated peptide (SFSAMK) from this protein, by MALDI-TOF mass spectrometric analysis after coincubation of the 4'-phosphopantetheinyl transferase with the recombinant NRP synthetase fragment and acetyl CoA, confirms that it is competent to play a role in NRP synthetase activation in A. fumigatus. The 4'-phosphopantetheinyl transferase also activates, by 4'-phosphopantetheinylation, recombinant alpha-aminoadipate reductase (Lys2p) from Candida albicans, a key enzyme involved in lysine biosynthesis.

  14. Interactions of photoactive DNAs with terminal deoxynucleotidyl transferase: Identification of peptides in the DNA binding domain

    International Nuclear Information System (INIS)

    Farrar, Y.J.K.; Evans, R.K.; Beach, C.M.; Coleman, M.S.

    1991-01-01

    Terminal deoxynucleotidyl transferase (terminal transferase) was specifically modified in the DNA binding site by a photoactive DNA substrate (hetero-40-mer duplex containing eight 5-azido-dUMP residues at one 3' end). Under optimal photolabeling conditions, 27-40% of the DNA was covalently cross-linked to terminal transferase. The specificity of the DNA and protein interaction was demonstrated by protection of photolabeling at the DNA binding domain with natural DNA substrates. In order to recover high yields of modified peptides from limited amounts of starting material, protein modified with 32 P-labeled photoactive DNA and digested with trypsin was extracted 4 times with phenol followed by gel filtration chromatography. All peptides not cross-linked to DNA were extracted into the phenol phase while the photolyzed DNA and the covalently cross-linked peptides remained in the aqueous phase. The 32 P-containing peptide-DNA fraction was subjected to amino acid sequence analysis. Two sequences, Asp 221 -Lys 231 (peptide B8) and Cys 234 -Lys 249 (peptide B10), present in similar yield, were identified. Structure predictions placed the two peptides in an α-helical array of 39 angstrom which would accommodate a DNA helix span of 11 nucleotides. These peptides share sequence similarity with a region in DNA polymerase β that has been implicated in the binding of DNA template

  15. Glutathione S - transferases class Pi and Mi and their significance in oncology

    Directory of Open Access Journals (Sweden)

    Zofia Marchewka

    2017-06-01

    Full Text Available In this article the current data, which shows that glutathione S-transferases (GST class Pi and Mi are interesting and promising biomarkers in acute and chronic inflammatory processes as well as in the oncology, were presented based on the review of the latest experimental and clinical studies. The article shows their characteristics, functions and participation (direct - GST Pi, indirect - GST Mi in the regulation of signaling pathways of JNK kinases, which are involved in cell differentiation. Overexpression of glutathione S-transferases class Pi and Mi in many cancer cells plays a key role in cancer treatment, making them resistant to chemotherapy. GST isoenzymes are involved in the metabolism of various types of xenobiotics and endogenous substrates, so their altered expression in cancer tissues as well as in serum and urine could be an important potential marker of the cancer and an indicator of oxidative stress. The study shows the role of glutathione S-transferases in redox homeostasis of tumor cells and in the mechanism of resistance to anticancer drugs.

  16. Glutathione S - transferases class Pi and Mi and their significance in oncology.

    Science.gov (United States)

    Marchewka, Zofia; Piwowar, Agnieszka; Ruzik, Sylwia; Długosz, Anna

    2017-06-19

    In this article the current data, which shows that glutathione S-transferases (GST) class Pi and Mi are interesting and promising biomarkers in acute and chronic inflammatory processes as well as in the oncology, were presented based on the review of the latest experimental and clinical studies. The article shows their characteristics, functions and participation (direct - GST Pi, indirect - GST Mi) in the regulation of signaling pathways of JNK kinases, which are involved in cell differentiation. Overexpression of glutathione S-transferases class Pi and Mi in many cancer cells plays a key role in cancer treatment, making them resistant to chemotherapy. GST isoenzymes are involved in the metabolism of various types of xenobiotics and endogenous substrates, so their altered expression in cancer tissues as well as in serum and urine could be an important potential marker of the cancer and an indicator of oxidative stress. The study shows the role of glutathione S-transferases in redox homeostasis of tumor cells and in the mechanism of resistance to anticancer drugs.

  17. Characterization of Affinity-Purified Isoforms of Acinetobacter calcoaceticus Y1 Glutathione Transferases

    Directory of Open Access Journals (Sweden)

    Chin-Soon Chee

    2014-01-01

    Full Text Available Glutathione transferases (GST were purified from locally isolated bacteria, Acinetobacter calcoaceticus Y1, by glutathione-affinity chromatography and anion exchange, and their substrate specificities were investigated. SDS-polyacrylamide gel electrophoresis revealed that the purified GST resolved into a single band with a molecular weight (MW of 23 kDa. 2-dimensional (2-D gel electrophoresis showed the presence of two isoforms, GST1 (pI 4.5 and GST2 (pI 6.2 with identical MW. GST1 was reactive towards ethacrynic acid, hydrogen peroxide, 1-chloro-2,4-dinitrobenzene, and trans,trans-hepta-2,4-dienal while GST2 was active towards all substrates except hydrogen peroxide. This demonstrated that GST1 possessed peroxidase activity which was absent in GST2. This study also showed that only GST2 was able to conjugate GSH to isoproturon, a herbicide. GST1 and GST2 were suggested to be similar to F0KLY9 (putative glutathione S-transferase and F0KKB0 (glutathione S-transferase III of Acinetobacter calcoaceticus strain PHEA-2, respectively.

  18. Characterization of Affinity-Purified Isoforms of Acinetobacter calcoaceticus Y1 Glutathione Transferases

    Science.gov (United States)

    Chee, Chin-Soon; Tan, Irene Kit-Ping; Alias, Zazali

    2014-01-01

    Glutathione transferases (GST) were purified from locally isolated bacteria, Acinetobacter calcoaceticus Y1, by glutathione-affinity chromatography and anion exchange, and their substrate specificities were investigated. SDS-polyacrylamide gel electrophoresis revealed that the purified GST resolved into a single band with a molecular weight (MW) of 23 kDa. 2-dimensional (2-D) gel electrophoresis showed the presence of two isoforms, GST1 (pI 4.5) and GST2 (pI 6.2) with identical MW. GST1 was reactive towards ethacrynic acid, hydrogen peroxide, 1-chloro-2,4-dinitrobenzene, and trans,trans-hepta-2,4-dienal while GST2 was active towards all substrates except hydrogen peroxide. This demonstrated that GST1 possessed peroxidase activity which was absent in GST2. This study also showed that only GST2 was able to conjugate GSH to isoproturon, a herbicide. GST1 and GST2 were suggested to be similar to F0KLY9 (putative glutathione S-transferase) and F0KKB0 (glutathione S-transferase III) of Acinetobacter calcoaceticus strain PHEA-2, respectively. PMID:24892084

  19. The interaction domain of the redox protein adrenodoxin is mandatory for binding of the electron acceptor CYP11A1, but is not required for binding of the electron donor adrenodoxin reductase

    International Nuclear Information System (INIS)

    Heinz, Achim; Hannemann, Frank; Mueller, Juergen J.; Heinemann, Udo; Bernhardt, Rita

    2005-01-01

    Adrenodoxin (Adx) is a [2Fe-2S] ferredoxin involved in electron transfer reactions in the steroid hormone biosynthesis of mammals. In this study, we deleted the sequence coding for the complete interaction domain in the Adx cDNA. The expressed recombinant protein consists of the amino acids 1-60, followed by the residues 89-128, and represents only the core domain of Adx (Adx-cd) but still incorporates the [2Fe-2S] cluster. Adx-cd accepts electrons from its natural redox partner, adrenodoxin reductase (AdR), and forms an individual complex with this NADPH-dependent flavoprotein. In contrast, formation of a complex with the natural electron acceptor, CYP11A1, as well as electron transfer to this steroid hydroxylase is prevented. By an electrostatic and van der Waals energy minimization procedure, complexes between AdR and Adx-cd have been proposed which have binding areas different from the native complex. Electron transport remains possible, despite longer electron transfer pathways

  20. Catalysis of Silver catfish Major Hepatic Glutathione Transferase proceeds via rapid equilibrium sequential random Mechanism

    Directory of Open Access Journals (Sweden)

    Ayodele O. Kolawole

    Full Text Available Fish hepatic glutathione transferases are connected with the elimination of intracellular pollutants and detoxification of organic micro-pollutants in their aquatic ecosystem. The two-substrate steady state kinetic mechanism of Silver catfish (Synodontis eupterus major hepatic glutathione transferases purified to apparent homogeneity was explored. The enzyme was dimeric enzyme with a monomeric size of 25.6 kDa. Initial-velocity studies and Product inhibition patterns by methyl glutathione and chloride with respect to GSH-CDNB; GSH-ρ-nitrophenylacetate; and GSH-Ethacrynic acid all conforms to a rapid equilibrium sequential random Bi Bi kinetic mechanism rather than steady state sequential random Bi Bi kinetic. α was 2.96 ± 0.35 for the model. The pH profile of Vmax/KM (with saturating 1-chloro-2,4-dinitrobenzene and variable GSH concentrations showed apparent pKa value of 6.88 and 9.86. Inhibition studies as a function of inhibitor concentration show that the enzyme is a homodimer and near neutral GST. The enzyme poorly conjugates 4-hydroxylnonenal and cumene hydroperoxide and may not be involved in oxidative stress protection. The seGST is unique and overwhelmingly shows characteristics similar to those of homodimeric class Pi GSTs, as was indicated by its kinetic mechanism, substrate specificity and inhibition studies. The rate- limiting step, probably the product release, of the reaction is viscosity-dependent and is consequential if macro-viscosogen or micro-viscosogen. Keywords: Silver catfish, Glutathione transferase, Steady-state, Kinetic mechanism, Inhibition

  1. Characterization of glutathione transferases involved in the pathogenicity of Alternaria brassicicola.

    Science.gov (United States)

    Calmes, Benoit; Morel-Rouhier, Mélanie; Bataillé-Simoneau, Nelly; Gelhaye, Eric; Guillemette, Thomas; Simoneau, Philippe

    2015-06-18

    Glutathione transferases (GSTs) represent an extended family of multifunctional proteins involved in detoxification processes and tolerance to oxidative stress. We thus anticipated that some GSTs could play an essential role in the protection of fungal necrotrophs against plant-derived toxic metabolites and reactive oxygen species that accumulate at the host-pathogen interface during infection. Mining the genome of the necrotrophic Brassica pathogen Alternaria brassicicola for glutathione transferase revealed 23 sequences, 17 of which could be clustered into the main classes previously defined for fungal GSTs and six were 'orphans'. Five isothiocyanate-inducible GSTs from five different classes were more thoroughly investigated. Analysis of their catalytic properties revealed that two GSTs, belonging to the GSTFuA and GTT1 classes, exhibited GSH transferase activity with isothiocyanates (ITC) and peroxidase activity with cumene hydroperoxide, respectively. Mutant deficient for these two GSTs were however neither more susceptible to ITC nor less aggressive than the wild-type parental strain. By contrast mutants deficient for two other GSTs, belonging to the Ure2pB and GSTO classes, were distinguished by their hyper-susceptibility to ITC and low aggressiveness against Brassica oleracea. In particular AbGSTO1 could participate in cell tolerance to ITC due to its glutathione-dependent thioltransferase activity. The fifth ITC-inducible GST belonged to the MAPEG class and although it was not possible to produce the soluble active form of this protein in a bacterial expression system, the corresponding deficient mutant failed to develop normal symptoms on host plant tissues. Among the five ITC-inducible GSTs analyzed in this study, three were found essential for full aggressiveness of A. brassicicola on host plant. This, to our knowledge is the first evidence that GSTs might be essential virulence factors for fungal necrotrophs.

  2. Glutathione Transferase from Trichoderma virens Enhances Cadmium Tolerance without Enhancing Its Accumulation in Transgenic Nicotiana tabacum

    Science.gov (United States)

    Dixit, Prachy; Mukherjee, Prasun K.; Ramachandran, V.; Eapen, Susan

    2011-01-01

    Background Cadmium (Cd) is a major heavy metal pollutant which is highly toxic to plants and animals. Vast agricultural areas worldwide are contaminated with Cd. Plants take up Cd and through the food chain it reaches humans and causes toxicity. It is ideal to develop plants tolerant to Cd, without enhanced accumulation in the edible parts for human consumption. Glutathione transferases (GST) are a family of multifunctional enzymes known to have important roles in combating oxidative stresses induced by various heavy metals including Cd. Some GSTs are also known to function as glutathione peroxidases. Overexpression/heterologous expression of GSTs is expected to result in plants tolerant to heavy metals such as Cd. Results Here, we report cloning of a glutathione transferase gene from Trichoderma virens, a biocontrol fungus and introducing it into Nicotiana tabacum plants by Agrobacterium-mediated gene transfer. Transgenic nature of the plants was confirmed by Southern blot hybridization and expression by reverse transcription PCR. Transgene (TvGST) showed single gene Mendelian inheritance. When transgenic plants expressing TvGST gene were exposed to different concentrations of Cd, they were found to be more tolerant compared to wild type plants, with transgenic plants showing lower levels of lipid peroxidation. Levels of different antioxidant enzymes such as glutathione transferase, superoxide dismutase, ascorbate peroxidase, guiacol peroxidase and catalase showed enhanced levels in transgenic plants expressing TvGST compared to control plants, when exposed to Cd. Cadmium accumulation in the plant biomass in transgenic plants were similar or lower than wild-type plants. Conclusion The results of the present study suggest that transgenic tobacco plants expressing a Trichoderma virens GST are more tolerant to Cd, without enhancing its accumulation in the plant biomass. It should be possible to extend the present results to crop plants for developing Cd tolerance and

  3. Glutathione transferase from Trichoderma virens enhances cadmium tolerance without enhancing its accumulation in transgenic Nicotiana tabacum.

    Directory of Open Access Journals (Sweden)

    Prachy Dixit

    Full Text Available BACKGROUND: Cadmium (Cd is a major heavy metal pollutant which is highly toxic to plants and animals. Vast agricultural areas worldwide are contaminated with Cd. Plants take up Cd and through the food chain it reaches humans and causes toxicity. It is ideal to develop plants tolerant to Cd, without enhanced accumulation in the edible parts for human consumption. Glutathione transferases (GST are a family of multifunctional enzymes known to have important roles in combating oxidative stresses induced by various heavy metals including Cd. Some GSTs are also known to function as glutathione peroxidases. Overexpression/heterologous expression of GSTs is expected to result in plants tolerant to heavy metals such as Cd. RESULTS: Here, we report cloning of a glutathione transferase gene from Trichoderma virens, a biocontrol fungus and introducing it into Nicotiana tabacum plants by Agrobacterium-mediated gene transfer. Transgenic nature of the plants was confirmed by Southern blot hybridization and expression by reverse transcription PCR. Transgene (TvGST showed single gene Mendelian inheritance. When transgenic plants expressing TvGST gene were exposed to different concentrations of Cd, they were found to be more tolerant compared to wild type plants, with transgenic plants showing lower levels of lipid peroxidation. Levels of different antioxidant enzymes such as glutathione transferase, superoxide dismutase, ascorbate peroxidase, guiacol peroxidase and catalase showed enhanced levels in transgenic plants expressing TvGST compared to control plants, when exposed to Cd. Cadmium accumulation in the plant biomass in transgenic plants were similar or lower than wild-type plants. CONCLUSION: The results of the present study suggest that transgenic tobacco plants expressing a Trichoderma virens GST are more tolerant to Cd, without enhancing its accumulation in the plant biomass. It should be possible to extend the present results to crop plants for

  4. The dyad palindromic glutathione transferase P enhancer binds multiple factors including AP1.

    OpenAIRE

    Diccianni, M B; Imagawa, M; Muramatsu, M

    1992-01-01

    Glutathione Transferase P (GST-P) gene expression is dominantly regulated by an upstream enhancer (GPEI) consisting of a dyad of palindromically oriented imperfect TPA (12-O-tetradecanoyl-phorbol-13-acetate)-responsive elements (TRE). GPEI is active in AP1-lacking F9 cells as well in AP1-containing HeLa cells. Despite GPEI's similarity to a TRE, c-jun co-transfection has only a minimal effect on transactivation. Antisense c-jun and c-fos co-transfection experiments further demonstrate the lac...

  5. Resistance to the Peptidyl Transferase Inhibitor Tiamulin Caused by Mutation of Ribosomal Protein L3

    OpenAIRE

    Bøsling, Jacob; Poulsen, Susan M.; Vester, Birte; Long, Katherine S.

    2003-01-01

    The antibiotic tiamulin targets the 50S subunit of the bacterial ribosome and interacts at the peptidyl transferase center. Tiamulin-resistant Escherichia coli mutants were isolated in order to elucidate mechanisms of resistance to the drug. No mutations in the rRNA were selected as resistance determinants using a strain expressing only a plasmid-encoded rRNA operon. Selection in a strain with all seven chromosomal rRNA operons yielded a mutant with an A445G mutation in the gene coding for ri...

  6. Generation of Active Bovine Terminal Deoxynucleotidyl Transferase (TdT in E.coli

    Directory of Open Access Journals (Sweden)

    Wee Liang Kuan

    2010-08-01

    Full Text Available A synthetic gene encoding bovine terminal deoxynucleotidyl transferase (TdT was generated, cloned into an expression vector and expressed in E.coli. The effects of altering culture and induction conditions on the nature of recombinant protein production were investigated. This led to the expression of active recombinant bovine TdT in E.coli. After purification and characterisation, the activity of the enzyme was assessed in a biological assay for apoptosis. The process described in this report enables the economical production of TdT for high throughput applications.

  7. Generation of Active Bovine Terminal Deoxynucleotidyl Transferase (TdT in E.coli

    Directory of Open Access Journals (Sweden)

    Wee Liang Kuan

    2010-01-01

    Full Text Available A synthetic gene encoding bovine terminal deoxynucleotidyl transferase (TdT was generated, cloned into an expression vector and expressed in E.coli. The effects of altering culture and induction conditions on the nature of recombinant protein production were investigated. This led to the expression of active recombinant bovine TdT in E.coli. After purification and characterisation, the activity of the enzyme was assessed in a biological assay for apoptosis. The process described in this report enables the economical production of TdT for high throughput applications.

  8. The O-GlcNAc Transferase Intellectual Disability Mutation L254F Distorts the TPR Helix.

    Science.gov (United States)

    Gundogdu, Mehmet; Llabrés, Salomé; Gorelik, Andrii; Ferenbach, Andrew T; Zachariae, Ulrich; van Aalten, Daan M F

    2018-05-17

    O-linked β-N-acetyl- D -glucosamine (O-GlcNAc) transferase (OGT) regulates protein O-GlcNAcylation, an essential post-translational modification that is abundant in the brain. Recently, OGT mutations have been associated with intellectual disability, although it is not understood how they affect OGT structure and function. Using a multi-disciplinary approach we show that the L254F OGT mutation leads to conformational changes of the tetratricopeptide repeats and reduced activity, revealing the molecular mechanisms contributing to pathogenesis. Copyright © 2018 The Authors. Published by Elsevier Ltd.. All rights reserved.

  9. Characterisation of the Candida albicans Phosphopantetheinyl Transferase Ppt2 as a Potential Antifungal Drug Target.

    Directory of Open Access Journals (Sweden)

    Katharine S Dobb

    Full Text Available Antifungal drugs acting via new mechanisms of action are urgently needed to combat the increasing numbers of severe fungal infections caused by pathogens such as Candida albicans. The phosphopantetheinyl transferase of Aspergillus fumigatus, encoded by the essential gene pptB, has previously been identified as a potential antifungal target. This study investigated the function of its orthologue in C. albicans, PPT2/C1_09480W by placing one allele under the control of the regulatable MET3 promoter, and deleting the remaining allele. The phenotypes of this conditional null mutant showed that, as in A. fumigatus, the gene PPT2 is essential for growth in C. albicans, thus fulfilling one aspect of an efficient antifungal target. The catalytic activity of Ppt2 as a phosphopantetheinyl transferase and the acyl carrier protein Acp1 as a substrate were demonstrated in a fluorescence transfer assay, using recombinant Ppt2 and Acp1 produced and purified from E.coli. A fluorescence polarisation assay amenable to high-throughput screening was also developed. Therefore we have identified Ppt2 as a broad-spectrum novel antifungal target and developed tools to identify inhibitors as potentially new antifungal compounds.

  10. Resistance to the peptidyl transferase inhibitor tiamulin caused by mutation of ribosomal protein l3.

    Science.gov (United States)

    Bøsling, Jacob; Poulsen, Susan M; Vester, Birte; Long, Katherine S

    2003-09-01

    The antibiotic tiamulin targets the 50S subunit of the bacterial ribosome and interacts at the peptidyl transferase center. Tiamulin-resistant Escherichia coli mutants were isolated in order to elucidate mechanisms of resistance to the drug. No mutations in the rRNA were selected as resistance determinants using a strain expressing only a plasmid-encoded rRNA operon. Selection in a strain with all seven chromosomal rRNA operons yielded a mutant with an A445G mutation in the gene coding for ribosomal protein L3, resulting in an Asn149Asp alteration. Complementation experiments and sequencing of transductants demonstrate that the mutation is responsible for the resistance phenotype. Chemical footprinting experiments show a reduced binding of tiamulin to mutant ribosomes. It is inferred that the L3 mutation, which points into the peptidyl transferase cleft, causes tiamulin resistance by alteration of the drug-binding site. This is the first report of a mechanism of resistance to tiamulin unveiled in molecular detail.

  11. Madumycin II inhibits peptide bond formation by forcing the peptidyl transferase center into an inactive state

    Energy Technology Data Exchange (ETDEWEB)

    Osterman, Ilya A.; Khabibullina, Nelli F.; Komarova, Ekaterina S.; Kasatsky, Pavel; Kartsev, Victor G.; Bogdanov, Alexey A.; Dontsova, Olga A.; Konevega, Andrey L.; Sergiev, Petr V.; Polikanov, Yury S. (InterBioScreen); (UIC); (MSU-Russia); (Kurchatov)

    2017-05-13

    The emergence of multi-drug resistant bacteria is limiting the effectiveness of commonly used antibiotics, which spurs a renewed interest in revisiting older and poorly studied drugs. Streptogramins A is a class of protein synthesis inhibitors that target the peptidyl transferase center (PTC) on the large subunit of the ribosome. In this work, we have revealed the mode of action of the PTC inhibitor madumycin II, an alanine-containing streptogramin A antibiotic, in the context of a functional 70S ribosome containing tRNA substrates. Madumycin II inhibits the ribosome prior to the first cycle of peptide bond formation. It allows binding of the tRNAs to the ribosomal A and P sites, but prevents correct positioning of their CCA-ends into the PTC thus making peptide bond formation impossible. We also revealed a previously unseen drug-induced rearrangement of nucleotides U2506 and U2585 of the 23S rRNA resulting in the formation of the U2506•G2583 wobble pair that was attributed to a catalytically inactive state of the PTC. The structural and biochemical data reported here expand our knowledge on the fundamental mechanisms by which peptidyl transferase inhibitors modulate the catalytic activity of the ribosome.

  12. Glutathione Transferases Superfamily: Cold-Inducible Expression of Distinct GST Genes in Brassica oleracea

    Directory of Open Access Journals (Sweden)

    Harshavardhanan Vijayakumar

    2016-07-01

    Full Text Available Plants, as sessile organisms, can suffer serious growth and developmental consequences under cold stress conditions. Glutathione transferases (GSTs, EC 2.5.1.18 are ubiquitous and multifunctional conjugating proteins, which play a major role in stress responses by preventing oxidative damage by reactive oxygen species (ROS. Currently, understanding of their function(s during different biochemical and signaling pathways under cold stress condition remain unclear. In this study, using combined computational strategy, we identified 65 Brassica oleracea glutathione transferases (BoGST and characterized them based on evolutionary analysis into 11 classes. Inter-species and intra-species duplication was evident between BoGSTs and Arabidopsis GSTs. Based on localization analyses, we propose possible pathways in which GST genes are involved during cold stress. Further, expression analysis of the predicted putative functions for GST genes were investigated in two cold contrasting genotypes (cold tolerance and susceptible under cold condition, most of these genes were highly expressed at 6 h and 1 h in the cold tolerant (CT and cold susceptible (CS lines, respectively. Overall, BoGSTU19, BoGSTU24, BoGSTF10 are candidate genes highly expressed in B. oleracea. Further investigation of GST superfamily in B. oleracea will aid in understanding complex mechanism underlying cold tolerance in plants.

  13. The role of a topologically conserved isoleucine in glutathione transferase structure, stability and function

    International Nuclear Information System (INIS)

    Achilonu, Ikechukwu; Gildenhuys, Samantha; Fisher, Loren; Burke, Jonathan; Fanucchi, Sylvia; Sewell, B. Trevor; Fernandes, Manuel; Dirr, Heini W.

    2010-01-01

    The role of a topologically conserved isoleucine in the structure of glutathione transferase was investigated by replacing the Ile71 residue in human GSTA1-1 by alanine or valine. The common fold shared by members of the glutathione-transferase (GST) family has a topologically conserved isoleucine residue at the N-terminus of helix 3 which is involved in the packing of helix 3 against two β-strands in domain 1. The role of the isoleucine residue in the structure, function and stability of GST was investigated by replacing the Ile71 residue in human GSTA1-1 by alanine or valine. The X-ray structures of the I71A and I71V mutants resolved at 1.75 and 2.51 Å, respectively, revealed that the mutations do not alter the overall structure of the protein compared with the wild type. Urea-induced equilibrium unfolding studies using circular dichroism and tryptophan fluorescence suggest that the mutation of Ile71 to alanine or valine reduces the stability of the protein. A functional assay with 1-chloro-2,4-dinitrobenzene shows that the mutation does not significantly alter the function of the protein relative to the wild type. Overall, the results suggest that conservation of the topologically conserved Ile71 maintains the structural stability of the protein but does not play a significant role in catalysis and substrate binding

  14. Epidermal growth factor regulation of glutathione S-transferase gene expression in the rat is mediated by class Pi glutathione S-transferase enhancer I.

    Science.gov (United States)

    Matsumoto, M; Imagawa, M; Aoki, Y

    2000-07-01

    Using chloramphenicol acetyltransferase assays we showed that epidermal growth factor (EGF), transforming growth factor alpha (TGF alpha), and 3,3',4,4',5-pentachlorobiphenyl (PenCB) induce class Pi glutathione S-transferase (GSTP1) in primary cultured rat liver parenchymal cells. GSTP1 enhancer I (GPEI), which is required for the stimulation of GSTP1 expression by PenCB, also mediates EGF and TGF alpha stimulation of GSTP1 gene expression. However, hepatocyte growth factor and insulin did not stimulate GPEI-mediated gene expression. On the other hand, the antioxidant reagents butylhydroxyanisole and t-butylhydroquinone, stimulated GPEI-mediated gene expression, but the level of GSTP1 mRNA was not elevated. Our observations suggest that EGF and TGF alpha induce GSTP1 by the same signal transduction pathway as PenCB. Since the sequence of GPEI is similar to that of the antioxidant responsive element (ARE), some factors which bind to ARE might play a role in GPEI-mediated gene expression.

  15. Reviewing Hit Discovery Literature for Difficult Targets: Glutathione Transferase Omega-1 as an Example.

    Science.gov (United States)

    Xie, Yiyue; Dahlin, Jayme L; Oakley, Aaron J; Casarotto, Marco G; Board, Philip G; Baell, Jonathan B

    2018-05-10

    Early stage drug discovery reporting on relatively new or difficult targets is often associated with insufficient hit triage. Literature reviews of such targets seldom delve into the detail required to critically analyze the associated screening hits reported. Here we take the enzyme glutathione transferase omega-1 (GSTO1-1) as an example of a relatively difficult target and review the associated literature involving small-molecule inhibitors. As part of this process we deliberately pay closer-than-usual attention to assay interference and hit quality aspects. We believe this Perspective will be a useful guide for future development of GSTO1-1 inhibitors, as well serving as a template for future review formats of new or difficult targets.

  16. Response of Glutathione and Glutathione S-transferase in Rice Seedlings Exposed to Cadmium Stress

    Directory of Open Access Journals (Sweden)

    Chun-hua ZHANG

    2008-03-01

    Full Text Available A hydroponic culture experiment was done to investigate the effect of Cd stress on glutathione content (GSH and glutathione S-transferase (GST, EC 2.5.1.18 activity in rice seedlings. The rice growth was severely inhibited when Cd level in the solution was higher than 10 mg/L. In rice shoots, GSH content and GST activity increased with the increasing Cd level, while in roots, GST was obviously inhibited by Cd treatments. Compared with shoots, the rice roots had higher GSH content and GST activity, indicating the ability of Cd detoxification was much higher in roots than in shoots. There was a significant correlation between Cd level and GSH content or GST activity, suggesting that both parameters may be used as biomarkers of Cd stress in rice.

  17. Differential roles of tau class glutathione S-transferases in oxidative stress

    DEFF Research Database (Denmark)

    Kilili, Kimiti G; Atanassova, Neli; Vardanyan, Alla

    2004-01-01

    The plant glutathione S-transferase BI-GST has been identified as a potent inhibitor of Bax lethality in yeast, a phenotype associated with oxidative stress and disruption of mitochondrial functions. Screening of a tomato two-hybrid library for BI-GST interacting proteins identified five homologous...... Tau class GSTs, which readily form heterodimers between them and BI-GST. All six LeGSTUs were found to be able to protect yeast cells from prooxidant-induced cell death. The efficiency of each LeGSTU was prooxidant-specific, indicating a different role for each LeGSTU in the oxidative stress......-response mechanism. The prooxidant protective effect of all six proteins was suppressed in the absence of YAP1, a transcription factor that regulates hydroperoxide homeostasis in Saccharomyces cerevisiae, suggesting a role for the LeGSTUs in the context of the YAP1-dependent stress-responsive machinery...

  18. Phi Class of Glutathione S-transferase Gene Superfamily Widely Exists in Nonplant Taxonomic Groups.

    Science.gov (United States)

    Munyampundu, Jean-Pierre; Xu, You-Ping; Cai, Xin-Zhong

    2016-01-01

    Glutathione S-transferases (GSTs) constitute a superfamily of enzymes involved in detoxification of noxious compounds and protection against oxidative damage. GST class Phi (GSTF), one of the important classes of plant GSTs, has long been considered as plant specific but was recently found in basidiomycete fungi. However, the range of nonplant taxonomic groups containing GSTFs remains unknown. In this study, the distribution and phylogenetic relationships of nonplant GSTFs were investigated. We identified GSTFs in ascomycete fungi, myxobacteria, and protists Naegleria gruberi and Aureococcus anophagefferens. GSTF occurrence in these bacteria and protists correlated with their genome sizes and habitats. While this link was missing across ascomycetes, the distribution and abundance of GSTFs among ascomycete genomes could be associated with their lifestyles to some extent. Sequence comparison, gene structure, and phylogenetic analyses indicated divergence among nonplant GSTFs, suggesting polyphyletic origins during evolution. Furthermore, in silico prediction of functional partners suggested functional diversification among nonplant GSTFs.

  19. Echinococcus granulosus: Evidence of a heterodimeric glutathione transferase built up by phylogenetically distant subunits.

    Science.gov (United States)

    Arbildi, Paula; La-Rocca, Silvana; Lopez, Veronica; Da-Costa, Natalia; Fernandez, Veronica

    2017-01-01

    In the cestode parasite Echinococcus granulosus, three phylogenetically distant cytosolic glutathione transferases (GSTs) (EgGST1, 2 and 3) were identified. Interestingly, the C-terminal domains of EgGST3 and EgGST2 but not EgGST1, exhibit all amino acids involved in Sigma-class GST dimerization. Here, we provide evidence indicating that EgGST2 and EgGST3 naturally form a heterodimeric structure (EgGST2-3), and also we report the enzymatic activity of the recombinant heterodimer. EgGST2-3 might display novel properties able to influence the infection establishment. This is the first report of a stable heterodimeric GST built up by phylogenetically distant subunits. Copyright © 2016 Elsevier B.V. All rights reserved.

  20. Isolation and purification of glutathione S-transferases from Brachionus plicatilis and B. calyciflorus (Rotifera).

    Science.gov (United States)

    Bowman, B P; Snell, T W; Cochrane, B J

    1990-01-01

    1. The enzyme glutathione S-transferase (GST), a critical element in xenobiotic metabolism, was isolated from the marine rotifer Brachionus plicatilis and its freshwater congener B. calyciflorus. 2. In B. plicatilis, GST comprised 4.2% of cytosolic protein and was present as three separate isozymes with mol. wts 30,000, 31,400 and 33,700. Specific activity of crude homogenates was 56 nmol min-1 mg-1 protein, while that of affinity chromatography purified GST was 1850. 3. In B. calyciflorus, GST was present as two isozymes with mol. wts of 26,300 and 28,500, representing 1.0% of cytosolic protein. Crude GST specific activity was 1750 nmol min-1 mg-1 protein and purified was 72,400. 4. Rotifer GSTs are unusual because they are monomers whereas all other animals thus far investigated posses dimeric GSTs.

  1. Nuclear translocation of glutathione transferase omega is a progression marker in Barrett's esophagus

    DEFF Research Database (Denmark)

    Piaggi, Simona; Marchi, Santino; Ciancia, Eugenio

    2009-01-01

    Barrett's esophagus (BE) represents a major risk factor for esophageal adenocarcinoma (AC). For this reason, patients with BE are subjected to a systematic endoscopic surveillance to detect initial evolution towards non-invasive neoplasia (NiN) and cancer, that eventually occurs only in a small......-S-transferase-omega 1 could be involved in the stress response of human cells playing a role in the cancer progression of Barrett's esophagus. Its immunohistochemical detection could represent a useful tool in the grading of Barrett's disease.......N in BE and to understand the mechanisms of the progression from BE to AC. We investigated the expression and subcellular localization of GSTO1 in biopsies from patients with BE and in human cancer cell lines subjected to heath shock treatment. A selective nuclear localisation of GSTO1 was found in 16/16 biopsies with low...

  2. Glutathione transferases are structural and functional outliers in the thioredoxin fold.

    Science.gov (United States)

    Atkinson, Holly J; Babbitt, Patricia C

    2009-11-24

    Glutathione transferases (GSTs) are ubiquitous scavengers of toxic compounds that fall, structurally and functionally, within the thioredoxin fold suprafamily. The fundamental catalytic capability of GSTs is catalysis of the nucleophilic addition or substitution of glutathione at electrophilic centers in a wide range of small electrophilic compounds. While specific GSTs have been studied in detail, little else is known about the structural and functional relationships between different groupings of GSTs. Through a global analysis of sequence and structural similarity, it was determined that variation in the binding of glutathione between the two major subgroups of cytosolic (soluble) GSTs results in a different mode of glutathione activation. Additionally, the convergent features of glutathione binding between cytosolic GSTs and mitochondrial GST kappa are described. The identification of these structural and functional themes helps to illuminate some of the fundamental contributions of the thioredoxin fold to catalysis in the GSTs and clarify how the thioredoxin fold can be modified to enable new functions.

  3. Glutathione-binding site of a bombyx mori theta-class glutathione transferase.

    Directory of Open Access Journals (Sweden)

    M D Tofazzal Hossain

    Full Text Available The glutathione transferase (GST superfamily plays key roles in the detoxification of various xenobiotics. Here, we report the isolation and characterization of a silkworm protein belonging to a previously reported theta-class GST family. The enzyme (bmGSTT catalyzes the reaction of glutathione with 1-chloro-2,4-dinitrobenzene, 1,2-epoxy-3-(4-nitrophenoxy-propane, and 4-nitrophenethyl bromide. Mutagenesis of highly conserved residues in the catalytic site revealed that Glu66 and Ser67 are important for enzymatic function. These results provide insights into the catalysis of glutathione conjugation in silkworm by bmGSTT and into the metabolism of exogenous chemical agents.

  4. Expression of polypeptide GalNAc-transferases in stratified epithelia and squamous cell carcinomas

    DEFF Research Database (Denmark)

    Mandel, U; Hassan, H; Therkildsen, M H

    1999-01-01

    GalNAc-T1, -T2, and -T3. Application of this panel of novel antibodies revealed that GalNAc- transferases are differentially expressed in different cell lines, in spermatozoa, and in oral mucosa and carcinomas. For example, GalNAc-T1 and -T2 but not -T3 were highly expressed in WI38 cells, and Gal......NAc-T3 but not GalNAc-T1 or -T2 was expressed in spermatozoa. The expression patterns in normal oral mucosa were found to vary with cell differentiation, and for GalNAc-T2 and -T3 this was reflected in oral squamous cell carcinomas. The expression pattern of GalNAc-T1 was on the other hand changed...

  5. Identification of Protein-Protein Interactions with Glutathione-S-Transferase (GST) Fusion Proteins.

    Science.gov (United States)

    Einarson, Margret B; Pugacheva, Elena N; Orlinick, Jason R

    2007-08-01

    INTRODUCTIONGlutathione-S-transferase (GST) fusion proteins have had a wide range of applications since their introduction as tools for synthesis of recombinant proteins in bacteria. GST was originally selected as a fusion moiety because of several desirable properties. First and foremost, when expressed in bacteria alone, or as a fusion, GST is not sequestered in inclusion bodies (in contrast to previous fusion protein systems). Second, GST can be affinity-purified without denaturation because it binds to immobilized glutathione, which provides the basis for simple purification. Consequently, GST fusion proteins are routinely used for antibody generation and purification, protein-protein interaction studies, and biochemical analysis. This article describes the use of GST fusion proteins as probes for the identification of protein-protein interactions.

  6. Serum glutathione transferase does not respond to indole-3-carbinol: A pilot study

    Directory of Open Access Journals (Sweden)

    Daniel R McGrath

    2010-05-01

    Full Text Available Daniel R McGrath1, Hamid Frydoonfar2, Joshua J Hunt3, Chris J Dunkley3, Allan D Spigelman41Ipswich Hospital, Ipswich, UK; 2Hunter Pathology Service, New South Wales; 3Royal Newcastle Centre, Newcastle; 4St Vincent’s Clinical School, Sydney, AustraliaBackground: Despite the well recognized protective effect of cruciferous vegetables against various cancers, including human colorectal cancers, little is known about how this effect is conferred. It is thought that some phytochemicals found only in these vegetables confer the protection. These compounds include the glucosinolates, of which indole-3-carbinol is one. They are known to induce carcinogen-metabolizing (phase II enzymes, including the glutathione S-transferase (GST family. Other effects in humans are not well documented. We wished to assess the effect of indole-3-carbinol on GST enzymes.Methods: We carried out a placebo-controlled human volunteer study. All patients were given 400 mg daily of indole-3-carbinol for three months, followed by placebo. Serum samples were tested for the GSTM1 genotype by polymerase chain reaction. Serum GST levels were assessed using enzyme-linked immunosorbent assay and Western Blot methodologies.Results: Forty-nine volunteers completed the study. GSTM1 genotypes were obtained for all but two volunteers. A slightly greater proportion of volunteers were GSTM1-positive, in keeping with the general population. GST was detected in all patients. Total GST level was not affected by indole-3-carbinol dosing compared with placebo. Although not statistically significant, the GSTM1 genotype affected the serum GST level response to indole-3-carbinol.Conclusion: Indole-3-carbinol does not alter total serum GST levels during prolonged dosing.Keywords: pilot study, colorectal cancer, glutathione transferase, human, indole-3-carbinol

  7. Loci Affecting Gamma-Glutamyl Transferase in Adults and Adolescents Show Age X SNP interaction and Cardiometabolic Disease Associations

    NARCIS (Netherlands)

    Middelberg, R.P.S.; Benyamin, B.; de Moor, M.H.M.; Warrington, N.M.; Gordon, S.; Henders, A.K.; Medland, S.E.; Nyholt, DR; de Geus, E.J.C.; Hottenga, J.J.; Willemsen, G.; Beilin, L.J.; Mori, T.A.; Wright, M.J.; Heath, A.C.; Madden, P.A.F.; Boomsma, D.I.; Pennell, C.E.; Montgomery, G.W.; Martin, N.G.; Whitfield, J.B.

    2012-01-01

    Serum gamma-glutamyl transferase (GGT) activity is a marker of liver disease which is also prospectively associated with the risk of all-cause mortality, cardiovascular disease, type 2 diabetes and cancers. We have discovered novel loci affecting GGT in a genome-wide association study (rs1497406 in

  8. Systemic catechol-O-methyl transferase inhibition enables the D1 agonist radiotracer R-[11C]SKF 82957

    DEFF Research Database (Denmark)

    Palner, Mikael; McCormick, Patrick; Parkes, Jun

    2010-01-01

    R-[(11)C]-SKF 82957 is a high-affinity and potent dopamine D(1) receptor agonist radioligand, which gives rise to a brain-penetrant lipophilic metabolite. In this study, we demonstrate that systemic administration of catechol-O-methyl transferase (COMT) inhibitors blocks this metabolic pathway, f...

  9. Glutathione S-transferase M1 and T1, CYP1A2-2467T/delT ...

    African Journals Online (AJOL)

    The present study investigated the impact of metabolic gene polymorphisms in modulating lung cancer risk susceptibility. Gene polymorphisms encoding Cytochrome 1A2 (CYP1A2) and Glutathione-S-transferases (GSTT1 and GSTM1) are involved in the bioactivation and detoxification of tobacco carcinogens and may ...

  10. Mapping important nucleotides in the peptidyl transferase centre of 23 S rRNA using a random mutagenesis approach

    DEFF Research Database (Denmark)

    Porse, B T; Garrett, R A

    1995-01-01

    Random mutations were generated in the lower half of the peptidyl transferase loop in domain V of 23 S rRNA from Escherichia coli using a polymerase chain reaction (PCR) approach, a rapid procedure for identifying mutants and a plasmid-based expression system. The effects of 21 single-site mutati...

  11. Cloning and characterization of a glutathione S-transferase homologue from the plant pathogenic fungus Botrytis cinerea

    NARCIS (Netherlands)

    Prins, T.W.; Wagemakers, L.; Schouten, A.; Kan, van J.A.L.

    2000-01-01

    A gene was cloned from Botrytis cinerea that encodes a protein homologous to glutathione S-transferase (GST). The gene, denominated Bcgst1, is present in a single copy and represents the first example of such a gene from a filamentous fungus. The biochemical function of GSTs is to conjugate toxic

  12. The pleuromutilin drugs tiamulin and valnemulin bind to the RNA at the peptidyl transferase centre on the ribosome.

    Science.gov (United States)

    Poulsen, S M; Karlsson, M; Johansson, L B; Vester, B

    2001-09-01

    The pleuromutilin antibiotic derivatives, tiamulin and valnemulin, inhibit protein synthesis by binding to the 50S ribosomal subunit of bacteria. The action and binding site of tiamulin and valnemulin was further characterized on Escherichia coli ribosomes. It was revealed that these drugs are strong inhibitors of peptidyl transferase and interact with domain V of 23S RNA, giving clear chemical footprints at nucleotides A2058-9, U2506 and U2584-5. Most of these nucleotides are highly conserved phylogenetically and functionally important, and all of them are at or near the peptidyl transferase centre and have been associated with binding of several antibiotics. Competitive footprinting shows that tiamulin and valnemulin can bind concurrently with the macrolide erythromycin but compete with the macrolide carbomycin, which is a peptidyl transferase inhibitor. We infer from these and previous results that tiamulin and valnemulin interact with the rRNA in the peptidyl transferase slot on the ribosomes in which they prevent the correct positioning of the CCA-ends of tRNAs for peptide transfer.

  13. The interdomain flexible linker of the polypeptide GalNAc transferases dictates their long-range glycosylation preferences

    DEFF Research Database (Denmark)

    Rivas, Matilde De Las; Lira-Navarrete, Erandi; Daniel, Earnest James Paul

    2017-01-01

    The polypeptide GalNAc-transferases (GalNAc-Ts), that initiate mucin-type O-glycosylation, consist of a catalytic and a lectin domain connected by a flexible linker. In addition to recognizing polypeptide sequence, the GalNAc-Ts exhibit unique long-range N- A nd/or C-terminal prior glycosylation ...

  14. Transmutation of human glutathione transferase A2-2 with peroxidase activity into an efficient steroid isomerase.

    Science.gov (United States)

    Pettersson, Par L; Johansson, Ann-Sofie; Mannervik, Bengt

    2002-08-16

    A major goal in protein engineering is the tailor-making of enzymes for specified chemical reactions. Successful attempts have frequently been based on directed molecular evolution involving libraries of random mutants in which variants with desired properties were identified. For the engineering of enzymes with novel functions, it would be of great value if the necessary changes of the active site could be predicted and implemented. Such attempts based on the comparison of similar structures with different substrate selectivities have previously met with limited success. However, the present work shows that the knowledge-based redesign restricted to substrate-binding residues in human glutathione transferase A2-2 can introduce high steroid double-bond isomerase activity into the enzyme originally characterized by glutathione peroxidase activity. Both the catalytic center activity (k(cat)) and catalytic efficiency (k(cat)/K(m)) match the values of the naturally evolved glutathione transferase A3-3, the most active steroid isomerase known in human tissues. The substrate selectivity of the mutated glutathione transferase was changed 7000-fold by five point mutations. This example demonstrates the functional plasticity of the glutathione transferase scaffold as well as the potential of rational active-site directed mutagenesis as a complement to DNA shuffling and other stochastic methods for the redesign of proteins with novel functions.

  15. The association of alcohol intake with gamma-glutamyl transferase (GGT) levels: Evidence for correlated genetic effects

    NARCIS (Netherlands)

    van Beek, J.H.D.A.; de Moor, M.H.M.; Geels, L.M.; Sinke, M.R.T.; de Geus, E.J.C.; Lubke, G.H.; Kluft, C.; Neuteboom, J.; Vink, J.M.; Willemsen, G.; Boomsma, D.I.

    2014-01-01

    Background: Blood levels of gamma-glutamyl transferase (GGT) are used as a marker for (heavy) alcohol use. The role of GGT in the anti-oxidant defense mechanism that is part of normal metabolism supposes a causal effect of alcohol intake on GGT. However, there is variability in the response of GGT

  16. Cooperation of terminal deoxynucleotidyl transferase with DNA polymerase α in the replication of ultraviolet-irradiated DNA

    International Nuclear Information System (INIS)

    Yoshida, S.; Masaki, S.; Nakamura, H.; Morita, T.

    1981-01-01

    The amount of DNA synthesis in vitro with the ultraviolet-irradiated poly(dT).oligo(rA) template initiators catalysed by DNA polymerase α (Masaki, S. and Yoshida, S., Biochim. Biophys. Acta 521, 74-88) decreased with the dose of ultraviolet-irradiation. The ultraviolet irradiation to the template, however, did not affect the rate of incorporation of incorrect deoxynucleotides into the newly synthesized poly(dA). The addition of terminal deoxynucleotidyl transferase to this system enhanced the DNA synthesis to a level which is comparable to that of the control and it concomitantly increased the incorporation of the mismatched deoxynucleotide into the newly synthesized poly(dA) strands. On the other hand, with an unirradiated template initiator, the misincorporation was only slightly enhanced by the addition of terminal deoxynucleotidyl transferase. The sizes of newly synthesized DNA measured by the sedimentation velocities were found to be smaller with the ultraviolet-irradiated templates but they increased to the control level with the addition of terminal deoxynucleotidyl transferase to the systems. These results suggest that terminal deoxynucleotidyl transferase can help DNA polymerase α to bypass thymine dimers in vitro by the formation of mismatched regions at the positions opposite to pyrimidine dimers on the template. (Auth.)

  17. Mutations in N-acetylglucosamine (O-GlcNAc) transferase in patients with X-linked intellectual disability

    NARCIS (Netherlands)

    Willems, A.P.; Gundogdu, M.; Kempers, M.J.E.; Giltay, J.C.; Pfundt, R.P.; Elferink, M.; Loza, B.F.; Fuijkschot, J.; Ferenbach, A.T.; Gassen, K.L. van; Aalten, D.M.F. van; Lefeber, D.J.

    2017-01-01

    N-Acetylglucosamine (O-GlcNAc) transferase (OGT) regulates protein O-GlcNAcylation, an essential and dynamic post-translational modification. The O-GlcNAc modification is present on numerous nuclear and cytosolic proteins and has been implicated in essential cellular functions such as signaling and

  18. Function and phylogeny of bacterial butyryl-CoA:acetate transferases and their diversity in the proximal colon of swine

    Science.gov (United States)

    Studying the host-associated butyrate-producing bacterial community is important because butyrate is essential for colonic homeostasis and gut health. Previous research has identified the butyryl-coA:acetate transferase (2.3.8.3) as a the main gene for butyrate production in intestinal ecosystems; h...

  19. Disruption of the NADPH-dependent glutamate dehydrogenase affects the morphology of two industrial strains of Penicillium chrysogenum

    DEFF Research Database (Denmark)

    Thykær, Jette; Kildegaard, Kanchana Rueksomtawin; Noorman, H.

    2009-01-01

    recipient strains, was conducted. The reference strains showed a compact structure with highly branched hyphal elements whereas the morphology of the Delta gdhA strains consisting of long elongated hyphal elements with few branches. On solid medium, the hyphal growth unit (length) increased from an average...... of 47 mu m tip(-1) in the reference strains to 117 mu m tip(-1) in the Delta gdhA strains and in submerged cultures a decrease of 18% in branching frequency was measured due to the gdhA deletion. P. chrysogenum Wis 54-1255, the ancestor of most production strains was also characterised and this strain...

  20. Contribution of liver mitochondrial membrane-bound glutathione transferase to mitochondrial permeability transition pores

    International Nuclear Information System (INIS)

    Hossain, Quazi Sohel; Ulziikhishig, Enkhbaatar; Lee, Kang Kwang; Yamamoto, Hideyuki; Aniya, Yoko

    2009-01-01

    We recently reported that the glutathione transferase in rat liver mitochondrial membranes (mtMGST1) is activated by S-glutathionylation and the activated mtMGST1 contributes to the mitochondrial permeability transition (MPT) pore and cytochrome c release from mitochondria [Lee, K.K., Shimoji, M., Quazi, S.H., Sunakawa, H., Aniya, Y., 2008. Novel function of glutathione transferase in rat liver mitochondrial membrane: role for cytochrome c release from mitochondria. Toxcol. Appl. Pharmacol. 232, 109-118]. In the present study we investigated the effect of reactive oxygen species (ROS), generator gallic acid (GA) and GST inhibitors on mtMGST1 and the MPT. When rat liver mitochondria were incubated with GA, mtMGST1 activity was increased to about 3 fold and the increase was inhibited with antioxidant enzymes and singlet oxygen quenchers including 1,4-diazabicyclo [2,2,2] octane (DABCO). GA-mediated mtMGST1 activation was prevented by GST inhibitors such as tannic acid, hematin, and cibacron blue and also by cyclosporin A (CsA). In addition, GA induced the mitochondrial swelling which was also inhibited by GST inhibitors, but not by MPT inhibitors CsA, ADP, and bongkrekic acid. GA also released cytochrome c from the mitochondria which was inhibited completely by DABCO, moderately by GST inhibitors, and somewhat by CsA. Ca 2+ -mediated mitochondrial swelling and cytochrome c release were inhibited by MPT inhibitors but not by GST inhibitors. When the outer mitochondrial membrane was isolated after treatment of mitochondria with GA, mtMGST1 activity was markedly increased and oligomer/aggregate of mtMGST1 was observed. These results indicate that mtMGST1 in the outer mitochondrial membrane is activated by GA through thiol oxidation leading to protein oligomerization/aggregation, which may contribute to the formation of ROS-mediated, CsA-insensitive MPT pore, suggesting a novel mechanism for regulation of the MPT by mtMGST1

  1. Inhibition of the recombinant cattle tick Rhipicephalus (Boophilus) annulatus glutathione S-transferase.

    Science.gov (United States)

    Guneidy, Rasha A; Shahein, Yasser E; Abouelella, Amira M K; Zaki, Eman R; Hamed, Ragaa R

    2014-09-01

    Rhipicephalus (Boophilus) annulatus is a bloodsucking ectoparasite that causes severe production losses in the cattle industry. This study aims to evaluate the in vitro effects of tannic acid, hematin (GST inhibitors) and different plant extracts (rich in tannic acid) on the activity of the recombinant glutathione S-transferase enzyme of the Egyptian cattle tick R. annulatus (rRaGST), in order to confirm their ability to inhibit the parasitic essential detoxification enzyme glutathione S-transferase. Extraction with 70% ethanol of Hibiscus cannabinus (kenaf flowers), Punica granatum (red and white pomegranate peel), Musa acuminata (banana peel) (Musaceae), Medicago sativa (alfalfa seeds), Tamarindus indicus (seed) and Cuminum cyminum (cumin seed) were used to assess: (i) inhibitory capacities of rRaGST and (ii) their phenolic and flavonoid contents. Ethanol extraction of red pomegranate peel contained the highest content of phenolic compounds (29.95mg gallic acid/g dry tissue) compared to the other studied plant extracts. The highest inhibition activities of rRaGST were obtained with kenaf and red pomegranate peel (P. granatum) extracts with IC50 values of 0.123 and 0.136mg dry tissue/ml, respectively. Tannic acid was the more effective inhibitor of rRaGST with an IC50 value equal to 4.57μM compared to delphinidine-HCl (IC50=14.9±3.1μM). Gossypol had a weak inhibitory effect (IC50=43.7μM), and caffeic acid had almost no effect on tick GST activity. The IC50 values qualify ethacrynic acid as a potent inhibitor of rRaGST activity (IC50=0.034μM). Cibacron blue and hematin showed a considerable inhibition effect on rRaGST activity, and their IC50 values were 0.13μM and 7.5μM, respectively. The activity of rRaGST was highest for CDNB (30.2μmol/min/mg protein). The enzyme had also a peroxidatic activity (the specific activity equals 26.5μmol/min/mg protein). Both tannic acid and hematin inhibited rRaGST activity non-competitively with respect to GSH and

  2. Structural snapshots along the reaction pathway of Yersinia pestis RipA, a putative butyryl-CoA transferase

    Energy Technology Data Exchange (ETDEWEB)

    Torres, Rodrigo; Lan, Benson; Latif, Yama; Chim, Nicholas [UC Irvine, 2212 Natural Sciences I, Irvine, CA 92697 (United States); Goulding, Celia W., E-mail: celia.goulding@uci.edu [UC Irvine, 2212 Natural Sciences I, Irvine, CA 92697 (United States); UC Irvine, 2302 Natural Sciences I, Irvine, CA 92697 (United States)

    2014-04-01

    The crystal structures of Y. pestis RipA mutants were determined to provide insights into the CoA transferase reaction pathway. Yersinia pestis, the causative agent of bubonic plague, is able to survive in both extracellular and intracellular environments within the human host, although its intracellular survival within macrophages is poorly understood. A novel Y. pestis three-gene rip (required for intracellular proliferation) operon, and in particular ripA, has been shown to be essential for survival and replication in interferon γ-induced macrophages. RipA was previously characterized as a putative butyryl-CoA transferase proposed to yield butyrate, a known anti-inflammatory shown to lower macrophage-produced NO levels. RipA belongs to the family I CoA transferases, which share structural homology, a conserved catalytic glutamate which forms a covalent CoA-thioester intermediate and a flexible loop adjacent to the active site known as the G(V/I)G loop. Here, functional and structural analyses of several RipA mutants are presented in an effort to dissect the CoA transferase mechanism of RipA. In particular, E61V, M31G and F60M RipA mutants show increased butyryl-CoA transferase activities when compared with wild-type RipA. Furthermore, the X-ray crystal structures of E61V, M31G and F60M RipA mutants, when compared with the wild-type RipA structure, reveal important conformational changes orchestrated by a conserved acyl-group binding-pocket phenylalanine, Phe85, and the G(V/I)G loop. Binary structures of M31G RipA and F60M RipA with two distinct CoA substrate conformations are also presented. Taken together, these data provide CoA transferase reaction snapshots of an open apo RipA, a closed glutamyl-anhydride intermediate and an open CoA-thioester intermediate. Furthermore, biochemical analyses support essential roles for both the catalytic glutamate and the flexible G(V/I)G loop along the reaction pathway, although further research is required to fully

  3. Structural snapshots along the reaction pathway of Yersinia pestis RipA, a putative butyryl-CoA transferase

    International Nuclear Information System (INIS)

    Torres, Rodrigo; Lan, Benson; Latif, Yama; Chim, Nicholas; Goulding, Celia W.

    2014-01-01

    The crystal structures of Y. pestis RipA mutants were determined to provide insights into the CoA transferase reaction pathway. Yersinia pestis, the causative agent of bubonic plague, is able to survive in both extracellular and intracellular environments within the human host, although its intracellular survival within macrophages is poorly understood. A novel Y. pestis three-gene rip (required for intracellular proliferation) operon, and in particular ripA, has been shown to be essential for survival and replication in interferon γ-induced macrophages. RipA was previously characterized as a putative butyryl-CoA transferase proposed to yield butyrate, a known anti-inflammatory shown to lower macrophage-produced NO levels. RipA belongs to the family I CoA transferases, which share structural homology, a conserved catalytic glutamate which forms a covalent CoA-thioester intermediate and a flexible loop adjacent to the active site known as the G(V/I)G loop. Here, functional and structural analyses of several RipA mutants are presented in an effort to dissect the CoA transferase mechanism of RipA. In particular, E61V, M31G and F60M RipA mutants show increased butyryl-CoA transferase activities when compared with wild-type RipA. Furthermore, the X-ray crystal structures of E61V, M31G and F60M RipA mutants, when compared with the wild-type RipA structure, reveal important conformational changes orchestrated by a conserved acyl-group binding-pocket phenylalanine, Phe85, and the G(V/I)G loop. Binary structures of M31G RipA and F60M RipA with two distinct CoA substrate conformations are also presented. Taken together, these data provide CoA transferase reaction snapshots of an open apo RipA, a closed glutamyl-anhydride intermediate and an open CoA-thioester intermediate. Furthermore, biochemical analyses support essential roles for both the catalytic glutamate and the flexible G(V/I)G loop along the reaction pathway, although further research is required to fully

  4. Expression profiling of selected glutathione transferase genes in Zea mays (L.) seedlings infested with cereal aphids.

    Science.gov (United States)

    Sytykiewicz, Hubert; Chrzanowski, Grzegorz; Czerniewicz, Paweł; Sprawka, Iwona; Łukasik, Iwona; Goławska, Sylwia; Sempruch, Cezary

    2014-01-01

    The purpose of this report was to evaluate the expression patterns of selected glutathione transferase genes (gst1, gst18, gst23 and gst24) in the tissues of two maize (Zea mays L.) varieties (relatively resistant Ambrozja and susceptible Tasty Sweet) that were colonized with oligophagous bird cherry-oat aphid (Rhopalosiphum padi L.) or monophagous grain aphid (Sitobion avenae L.). Simultaneously, insect-triggered generation of superoxide anion radicals (O2•-) in infested Z. mays plants was monitored. Quantified parameters were measured at 1, 2, 4, 8, 24, 48 and 72 h post-initial aphid infestation (hpi) in relation to the non-infested control seedlings. Significant increases in gst transcript amounts were recorded in aphid-stressed plants in comparison to the control seedlings. Maximal enhancement in the expression of the gst genes in aphid-attacked maize plants was found at 8 hpi (gst23) or 24 hpi (gst1, gst18 and gst24) compared to the control. Investigated Z. mays cultivars formed excessive superoxide anion radicals in response to insect treatments, and the highest overproduction of O2•- was noted 4 or 8 h after infestation, depending on the aphid treatment and maize genotype. Importantly, the Ambrozja variety could be characterized as having more profound increments in the levels of gst transcript abundance and O2•- generation in comparison with the Tasty Sweet genotype.

  5. In-vitro effect of flavonoids from Solidago canadensis extract on glutathione S-transferase.

    Science.gov (United States)

    Apáti, Pál; Houghton, Peter J; Kite, Geoffrey; Steventon, Glyn B; Kéry, Agnes

    2006-02-01

    Solidago canadensis is typical of a flavonoid-rich herb and the effect of an aqueous ethanol extract on glutathione-S-transferase (GST) activity using HepG2 cells was compared with those of the flavonol quercetin and its glycosides quercitrin and rutin, found as major constituents. The composition of the extract was determined by HPLC and rutin was found to be the major flavonoidal component of the extract. Total GST activity was assessed using 1-chloro-2,4-dinitrobenzene as a substrate. The glycosides rutin and quercitrin gave dose-dependent increases in GST activity, with a 50% and 24.5% increase at 250 mM, respectively, while the aglycone quercetin inhibited the enzyme by 30% at 250 mM. The total extract of the herb gave an overall dose-dependent increase, the fractions corresponding to the flavonoids showed activating effects while those containing caffeic acid derivatives were inhibitory. The activity observed corresponds to that reported for similar compounds in-vivo using rats, thus the HepG2 cell line could serve as a more satisfactory method of assessing the effects of extracts and compounds on GST.

  6. Terminal deoxynucleotidyl transferase in the diagnosis of leukemia and malignant lymphoma.

    Science.gov (United States)

    Kung, P C; Long, J C; McCaffrey, R P; Ratliff, R L; Harrison, T A; Baltimore, D

    1978-05-01

    Neoplastic cells from 253 patients with leukemia and 46 patients with malignant lymphoma were studied for the presence of terminal deoxynucleotidyl transferase (TdT) by biochemical and fluorescent antibody technics. TdT was detected in circulating blast cells from 73 of 77 patients with acute lymphoblastic leukemia, 24 of 72 patients with chronic myelogenous leukemia examined during the blastic phase of the disorder and in cell suspensions of lymph nodes from nine of nine patients with diffuse lymphoblastic lymphoma. Blast cells from six of 10 patients with acute undifferentiated leukemia were TdT positive, but the enzyme was found in only two of 55 patients with acute myeloblastic leukemia. TdT was not detected in other lymphocytic or granulocytic leukemias or in other types of malignant lymphomas. The fluorescent antibody assay for TdT permits rapid and specific identification of the enzyme in single cells. The TdT assay is clinically useful in confirming the diagnosis of acute lymphoblastic leukemia, evaluating patients with blastic chronic myelogenous leukemia, and distinguishing patients with lymphoblastic lymphoma, whose natural history includes rapid extranodal dissemination, from patients with other poorly differentiated malignant lymphomas.

  7. Lack of Ach1 CoA-Transferase Triggers Apoptosis and Decreases Chronological Lifespan in Yeast

    International Nuclear Information System (INIS)

    Orlandi, Ivan; Casatta, Nadia; Vai, Marina

    2012-01-01

    ACH1 encodes a mitochondrial enzyme of Saccharomyces cerevisiae endowed with CoA-transferase activity. It catalyzes the CoASH transfer from succinyl-CoA to acetate generating acetyl-CoA. It is known that ACH1 inactivation results in growth defects on media containing acetate as a sole carbon and energy source which are particularly severe at low pH. Here, we show that chronological aging ach1Δ cells which accumulate a high amount of extracellular acetic acid display a reduced chronological lifespan. The faster drop of cell survival is completely abrogated by alleviating the acid stress either by a calorie restricted regimen that prevents acetic acid production or by transferring chronologically aging mutant cells to water. Moreover, the short-lived phenotype of ach1Δ cells is accompanied by reactive oxygen species accumulation, severe mitochondrial damage, and an early insurgence of apoptosis. A similar pattern of endogenous severe oxidative stress is observed when ach1Δ cells are cultured using acetic acid as a carbon source under acidic conditions. On the whole, our data provide further evidence of the role of acetic acid as cell-extrinsic mediator of cell death during chronological aging and highlight a primary role of Ach1 enzymatic activity in acetic acid detoxification which is important for mitochondrial functionality.

  8. Serum γ-Glutamyl Transferase Is Inversely Associated with Bone Mineral Density Independently of Alcohol Consumption

    Directory of Open Access Journals (Sweden)

    Han Seok Choi

    2016-03-01

    Full Text Available Backgroundγ-Glutamyl transferase (GGT is a well-known marker of chronic alcohol consumption or hepatobiliary diseases. A number of studies have demonstrated that serum levels of GGT are independently associated with cardiovascular and metabolic disorders. The purpose of this study was to test if serum GGT levels are associated with bone mineral density (BMD in Korean adults.MethodsA total of 462 subjects (289 men and 173 women, who visited Severance Hospital for medical checkup, were included in this study. BMD was measured using dual energy X-ray absorptiometry. Cross-sectional association between serum GGT and BMD was evaluated.ResultsAs serum GGT levels increased from the lowest tertile (tertile 1 to the highest tertile (tertile 3, BMD decreased after adjusting for confounders such as age, body mass index, amount of alcohol consumed, smoking, regular exercise, postmenopausal state (in women, hypertension, diabetes mellitus, and hypercholesterolemia. A multiple linear regression analysis showed a negative association between log-transformed serum GGT levels and BMD. In a multiple logistic regression analysis, tertile 3 of serum GGT level was associated with an increased risk for low bone mass compared to tertile 1 (odds ratio, 2.271; 95% confidence interval, 1.340 to 3.850; P=0.002.ConclusionSerum GGT level was inversely associated with BMD in Korean adults. Further study is necessary to fully elucidate the mechanism of the inverse relationship.

  9. The dyad palindromic glutathione transferase P enhancer binds multiple factors including AP1.

    Science.gov (United States)

    Diccianni, M B; Imagawa, M; Muramatsu, M

    1992-10-11

    Glutathione Transferase P (GST-P) gene expression is dominantly regulated by an upstream enhancer (GPEI) consisting of a dyad of palindromically oriented imperfect TPA (12-O-tetradecanoyl-phorbol-13-acetate)-responsive elements (TRE). GPEI is active in AP1-lacking F9 cells as well in AP1-containing HeLa cells. Despite GPEI's similarity to a TRE, c-jun co-transfection has only a minimal effect on transactivation. Antisense c-jun and c-fos co-transfection experiments further demonstrate the lack of a role for AP1 in GPEI mediated trans-activation in F9 cells, although endogenously present AP1 can influence GPEI in HeLa cells. Co-transfection of delta fosB with c-jun, which forms an inactive c-Jun/delta FosB heterodimer that binds TRE sequences, inhibits GPEI-mediated transcription in AP1-lacking F9 cells as well as AP1-containing HeLa cells. These data suggest novel factor(s) other than AP1 are influencing GPEI. Binding studies reveal multiple nucleoproteins bind to GPEI. These factors are likely responsible for the high level of GPEI-mediated transcription observed in the absence of AP1 and during hepatocarcinogenesis.

  10. Expression Profiling of Selected Glutathione Transferase Genes in Zea mays (L.) Seedlings Infested with Cereal Aphids

    Science.gov (United States)

    Sytykiewicz, Hubert; Chrzanowski, Grzegorz; Czerniewicz, Paweł; Sprawka, Iwona; Łukasik, Iwona; Goławska, Sylwia; Sempruch, Cezary

    2014-01-01

    The purpose of this report was to evaluate the expression patterns of selected glutathione transferase genes (gst1, gst18, gst23 and gst24) in the tissues of two maize (Zea mays L.) varieties (relatively resistant Ambrozja and susceptible Tasty Sweet) that were colonized with oligophagous bird cherry-oat aphid (Rhopalosiphum padi L.) or monophagous grain aphid (Sitobion avenae L.). Simultaneously, insect-triggered generation of superoxide anion radicals (O2 •−) in infested Z. mays plants was monitored. Quantified parameters were measured at 1, 2, 4, 8, 24, 48 and 72 h post-initial aphid infestation (hpi) in relation to the non-infested control seedlings. Significant increases in gst transcript amounts were recorded in aphid-stressed plants in comparison to the control seedlings. Maximal enhancement in the expression of the gst genes in aphid-attacked maize plants was found at 8 hpi (gst23) or 24 hpi (gst1, gst18 and gst24) compared to the control. Investigated Z. mays cultivars formed excessive superoxide anion radicals in response to insect treatments, and the highest overproduction of O2 •− was noted 4 or 8 h after infestation, depending on the aphid treatment and maize genotype. Importantly, the Ambrozja variety could be characterized as having more profound increments in the levels of gst transcript abundance and O2 •− generation in comparison with the Tasty Sweet genotype. PMID:25365518

  11. Prevalence of glutathione S-transferase gene deletions and their effect on sickle cell patients

    Directory of Open Access Journals (Sweden)

    Pandey Sanjay

    2012-01-01

    Full Text Available BACKGROUND: Glutathione S-transferase gene deletions are known detoxification agents and cause oxidative damage. Due to the different pathophysiology of anemia in thalassemia and sickle cell disease, there are significant differences in the pathophysiology of iron overload and iron-related complications in these disorders. OBJECTIVE: The aim of this study was to estimate the frequency of the GSTM1 and GSTT1 genotypes in sickle cell disease patients and their effect on iron status. METHODS: Forty sickle cell anemia and sixty sickle ß-thalassemia patients and 100 controls were evaluated to determine the frequency of GST gene deletions. Complete blood counts were performed by an automated cell analyzer. Hemoglobin F, hemoglobin A, hemoglobin A2 and hemoglobin S were measured and diagnosis of patients was achieved by high performance liquid chromatography with DNA extraction by the phenol-chloroform method. The GST null genotype was determined using multiplex polymerase chain reaction and serum ferritin was measured using an ELISA kit. Statistical analysis was by EpiInfo and GraphPad statistics software. RESULTS: An increased frequency of the GSTT1 null genotype (p-value = 0.05 was seen in the patients. The mean serum ferritin level was higher in patients with the GST genotypes than in controls; this was statistically significant for all genotypes except GSTM1, however the higher levels of serum ferritin were due to blood transfusions in patients. CONCLUSION: GST deletions do not play a direct role in iron overload of sickle cell patients.

  12. Characterization and evolutionary implications of the triad Asp-Xxx-Glu in group II phosphopantetheinyl transferases.

    Science.gov (United States)

    Wang, Yue-Yue; Li, Yu-Dong; Liu, Jian-Bo; Ran, Xin-Xin; Guo, Yuan-Yang; Ren, Ni-Ni; Chen, Xin; Jiang, Hui; Li, Yong-Quan

    2014-01-01

    Phosphopantetheinyl transferases (PPTases), which play an essential role in both primary and secondary metabolism, are magnesium binding enzymes. In this study, we characterized the magnesium binding residues of all known group II PPTases by biochemical and evolutionary analysis. Our results suggested that group II PPTases could be classified into two subgroups, two-magnesium-binding-residue-PPTases containing the triad Asp-Xxx-Glu and three-magnesium-binding-residue-PPTases containing the triad Asp-Glu-Glu. Mutations of two three-magnesium-binding-residue-PPTases and one two-magnesium-binding-residue-PPTase indicate that the first and the third residues in the triads are essential to activities; the second residues in the triads are non-essential. Although variations of the second residues in the triad Asp-Xxx-Glu exist throughout the whole phylogenetic tree, the second residues are conserved in animals, plants, algae, and most prokaryotes, respectively. Evolutionary analysis suggests that: the animal group II PPTases may originate from one common ancestor; the plant two-magnesium-binding-residue-PPTases may originate from one common ancestor; the plant three-magnesium-binding-residue-PPTases may derive from horizontal gene transfer from prokaryotes.

  13. Characterization of glutathione S-transferase and its immunodiagnostic potential for detecting Taenia multiceps.

    Science.gov (United States)

    Sun, Ying; Wang, Yu; Huang, Xing; Gu, Xiaobing; Lai, Weimin; Peng, Xuerong; Yang, Guangyou

    2017-08-15

    Taenia multiceps is a widespread zoonotic tapeworm parasite which infects cloven-hoofed animals around the world. Animal infection with Coenurus cerebralis, the coenurus larvae of T. multiceps (Tm), is often fatal, which is a major cause of economic losses in stockbreeding. This study amplified the glutathione S-transferase (GST) gene from the total RNA of C. cerebralis. The resulting protein, Tm-GST, consisted of 201 amino acids, and had a predicted molecular mass of 23.1kDa. Its amino acid sequence shares 77.61% similarity with Echinococcus granulosus GST. Recombinant Tm-GST (rTm-GST) was expressed in Escherichia coli. The protein reacted with serum from goats infected with T. multiceps. Immunofluorescence signals indicated that Tm-GST was largely localized in the parenchymatous area of adult T. multiceps; in addition, it was also apparent in the coenurus. An enzyme-linked immunosorbent assay based on rTm-GST showed specificity of 92.8% (13/14) and sensitivity of 90% (18/20) in detecting anti-GST antibodies in serum from naturally infected animals. This study suggests that Tm-GST has the potential to be used as a diagnostic antigen for Coenurosis. Copyright © 2017. Published by Elsevier B.V.

  14. A glutathione S-transferase gene associated with antioxidant properties isolated from Apis cerana cerana

    Science.gov (United States)

    Liu, Shuchang; Liu, Feng; Jia, Haihong; Yan, Yan; Wang, Hongfang; Guo, Xingqi; Xu, Baohua

    2016-06-01

    Glutathione S-transferases (GSTs) are an important family of multifunctional enzymes in aerobic organisms. They play a crucial role in the detoxification of exogenous compounds, especially insecticides, and protection against oxidative stress. Most previous studies of GSTs in insects have largely focused on their role in insecticide resistance. Here, we isolated a theta class GST gene designated AccGSTT1 from Apis cerana cerana and aimed to explore its antioxidant and antibacterial attributes. Analyses of homology and phylogenetic relationships suggested that the predicted amino acid sequence of AccGSTT1 shares a high level of identity with the other hymenopteran GSTs and that it was conserved during evolution. Quantitative real-time PCR showed that AccGSTT1 is most highly expressed in adult stages and that the expression profile of this gene is significantly altered in response to various abiotic stresses. These results were confirmed using western blot analysis. Additionally, a disc diffusion assay showed that a recombinant AccGSTT1 protein may be roughly capable of inhibiting bacterial growth and that it reduces the resistance of Escherichia coli cells to multiple adverse stresses. Taken together, these data indicate that AccGSTT1 may play an important role in antioxidant processes under adverse stress conditions.

  15. Glutathione S-Transferases: Role in Combating Abiotic Stresses Including Arsenic Detoxification in Plants

    Directory of Open Access Journals (Sweden)

    Smita Kumar

    2018-06-01

    Full Text Available Arsenic (As, naturally occurring metalloid and a potential hazardous material, is found in low concentrations in the environment and emerges from natural sources and anthropogenic activities. The presence of As in ground water, which is used for irrigation, is a matter of great concern since it affects crop productivity and contaminates food chain. In plants, As alters various metabolic pathways in cells including the interaction of substrates/enzymes with the sulfhydryl groups of proteins and the replacement of phosphate in ATP for energy. In addition, As stimulates the generation of free radicals and reactive oxygen species (ROS, resulting in oxidative stress. Glutathione S-transferases (GSTs quench reactive molecules with the addition of glutathione (GSH and protect the cell from oxidative damage. GSTs are a multigene family of isozymes, known to catalyze the conjugation of GSH to miscellany of electrophilic and hydrophobic substrates. GSTs have been reported to be associated with plant developmental processes and are responsive to multitude of stressors. In past, several studies suggested involvement of plant GST gene family in As response due to the requirement of sulfur and GSH in the detoxification of this toxic metalloid. This review provides updated information about the role of GSTs in abiotic and biotic stresses with an emphasis on As uptake, metabolism, and detoxification in plants. Further, the genetic manipulations that helped in enhancing the understanding of the function of GSTs in abiotic stress response and heavy metal detoxification has been reviewed.

  16. The Biochemistry of O-GlcNAc Transferase: Which Functions Make It Essential in Mammalian Cells?

    Science.gov (United States)

    Levine, Zebulon G; Walker, Suzanne

    2016-06-02

    O-linked N-acetylglucosamine transferase (OGT) is found in all metazoans and plays an important role in development but at the single-cell level is only essential in dividing mammalian cells. Postmitotic mammalian cells and cells of invertebrates such as Caenorhabditis elegans and Drosophila can survive without copies of OGT. Why OGT is required in dividing mammalian cells but not in other cells remains unknown. OGT has multiple biochemical activities. Beyond its well-known role in adding β-O-GlcNAc to serine and threonine residues of nuclear and cytoplasmic proteins, OGT also acts as a protease in the maturation of the cell cycle regulator host cell factor 1 (HCF-1) and serves as an integral member of several protein complexes, many of them linked to gene expression. In this review, we summarize current understanding of the mechanisms underlying OGT's biochemical activities and address whether known functions of OGT could be related to its essential role in dividing mammalian cells.

  17. Solution Structural Studies of GTP:Adenosylcobinamide-Phosphateguanylyl Transferase (CobY from Methanocaldococcus jannaschii.

    Directory of Open Access Journals (Sweden)

    Kiran K Singarapu

    Full Text Available GTP:adenosylcobinamide-phosphate (AdoCbi-P guanylyl transferase (CobY is an enzyme that transfers the GMP moiety of GTP to AdoCbi yielding AdoCbi-GDP in the late steps of the assembly of Ado-cobamides in archaea. The failure of repeated attempts to crystallize ligand-free (apo CobY prompted us to explore its 3D structure by solution NMR spectroscopy. As reported here, the solution structure has a mixed α/β fold consisting of seven β-strands and five α-helices, which is very similar to a Rossmann fold. Titration of apo-CobY with GTP resulted in large changes in amide proton chemical shifts that indicated major structural perturbations upon complex formation. However, the CobY:GTP complex as followed by 1H-15N HSQC spectra was found to be unstable over time: GTP hydrolyzed and the protein converted slowly to a species with an NMR spectrum similar to that of apo-CobY. The variant CobYG153D, whose GTP complex was studied by X-ray crystallography, yielded NMR spectra similar to those of wild-type CobY in both its apo- state and in complex with GTP. The CobYG153D:GTP complex was also found to be unstable over time.

  18. [Synthesis of vitamin K2 by isopentenyl transferase NovA in Pichia pastoris Gpn12].

    Science.gov (United States)

    Wu, Xihua; Li, Zhemin; Liu, Hui; Wang, Peng; Wang, Li; Fang, Xue; Sun, Xiaowen; Ni, Wenfeng; Yang, Qiang; Zheng, Zhiming; Zhao, Genhai

    2018-01-25

    The effect of methanol addition on the heterologous expression of isoprenyl transferase NovQ was studied in Pichia pastoris Gpn12, with menadione and isopentenol as precursors to catalyze vitamin K2 (MK-3) synthesis. The expression of NovQ increased by 36% when 2% methanol was added every 24 h. The influence of initial pH, temperature, methanol addition, precursors (menadione, isopentenol) addition, catalytic time and cetyltrimethyl-ammonium bromide (CTAB) addition were explored in the P. pastoris whole-cell catalytic synthesis process of MK-3 in shaking flask. Three significant factors were then studied by response surface method. The optimal catalytic conditions obtained were as follows: catalytic temperature 31.56 ℃, menadione 295.54 mg/L, catalytic time 15.87 h. Consistent with the response surface prediction results, the optimized yield of MK-3 reached 98.47 mg/L in shaking flask, 35% higher than that of the control group. On this basis, the production in a 30-L fermenter reached 189.67 mg/L when the cell catalyst of 220 g/L (dry weight) was used to catalyze the synthesis for 24 h. This method laid the foundation for the large-scale production of MK-3 by P. pastoris Gpn12.

  19. The TCA cycle transferase DLST is important for MYC-mediated leukemogenesis.

    Science.gov (United States)

    Anderson, N M; Li, D; Peng, H L; Laroche, F J F; Mansour, M R; Gjini, E; Aioub, M; Helman, D J; Roderick, J E; Cheng, T; Harrold, I; Samaha, Y; Meng, L; Amsterdam, A; Neuberg, D S; Denton, T T; Sanda, T; Kelliher, M A; Singh, A; Look, A T; Feng, H

    2016-06-01

    Despite the pivotal role of MYC in the pathogenesis of T-cell acute lymphoblastic leukemia (T-ALL) and many other cancers, the mechanisms underlying MYC-mediated tumorigenesis remain inadequately understood. Here we utilized a well-characterized zebrafish model of Myc-induced T-ALL for genetic studies to identify novel genes contributing to disease onset. We found that heterozygous inactivation of a tricarboxylic acid (TCA) cycle enzyme, dihydrolipoamide S-succinyltransferase (Dlst), significantly delayed tumor onset in zebrafish without detectable effects on fish development. DLST is the E2 transferase of the α-ketoglutarate (α-KG) dehydrogenase complex (KGDHC), which converts α-KG to succinyl-CoA in the TCA cycle. RNAi knockdown of DLST led to decreased cell viability and induction of apoptosis in human T-ALL cell lines. Polar metabolomics profiling revealed that the TCA cycle was disrupted by DLST knockdown in human T-ALL cells, as demonstrated by an accumulation of α-KG and a decrease of succinyl-CoA. Addition of succinate, the downstream TCA cycle intermediate, to human T-ALL cells was sufficient to rescue defects in cell viability caused by DLST inactivation. Together, our studies uncovered an important role for DLST in MYC-mediated leukemogenesis and demonstrated the metabolic dependence of T-lymphoblasts on the TCA cycle, thus providing implications for targeted therapy.

  20. Corneal aldehyde dehydrogenase and glutathione S-transferase activity after excimer laser keratectomy in guinea pigs.

    Science.gov (United States)

    Bilgihan, K; Bilgihan, A; Hasanreisoğlu, B; Turkozkan, N

    1998-03-01

    The free radical balance of the eye may be changed by excimer laser keratectomy. Previous studies have demonstrated that excimer laser keratectomy increases the corneal temperature, decreases the superoxide dismutase activity of the aqueous, and induces lipid peroxidation in the superficial corneal stroma. Aldehyde dehydrogenase (ALDH) and glutathione S-transferase (GST) are known to play an important role in corneal metabolism, particularly in detoxification of aldehydes, which are generated from free radical reactions. In three groups of guinea pigs mechanical corneal de-epithelialisation was performed in group I, superficial corneal photoablation in group II, and deep corneal photoablation in group III, and the corneal ALDH and GST activities measured after 48 hours. The mean ALDH and GST activities of group I and II showed no differences compared with the controls (p > 0.05). The corneal ALDH activities were found to be significantly decreased (p < 0.05) and GST activities increased (p < 0.05) in group III. These results suggest that excimer laser treatment of high myopia may change the ALDH and GST activities, metabolism, and free radical balance of the cornea.

  1. Isozyme-specific fluorescent inhibitor of glutathione s-transferase omega 1.

    Science.gov (United States)

    Son, Junghyun; Lee, Jae-Jung; Lee, Jun-Seok; Schüller, Andreas; Chang, Young-Tae

    2010-05-21

    Recently, the glutathione S-transferase omega 1 (GSTO1) is suspected to be involved in certain cancers and neurodegenerative diseases. However, profound investigation on the pathological roles of GSTO1 has been hampered by the lack of specific methods to determine or modulate its activity in biological systems containing other isoforms with similar catalytic function. Here, we report a fluorescent compound that is able to inhibit and monitor the activity of GSTO1. We screened 43 fluorescent chemicals and found a compound (6) that binds specifically to the active site of GSTO1. We observed that compound 6 inhibits GSTO1 by covalent modification but spares other isoforms in HEK293 cells and demonstrated that compound 6 could report the activity of GSTO1 in NIH/3T3 or HEK293 cells by measuring the fluorescence intensity of the labeled amount of GSTO1 in SDS-PAGE. Compound 6 is a useful tool to study GSTO1, applicable as a specific inhibitor and an activity reporter.

  2. Identification and characterisation of multiple glutathione S-transferase genes from the diamondback moth, Plutella xylostella.

    Science.gov (United States)

    Chen, Xi'en; Zhang, Ya-lin

    2015-04-01

    The diamondback moth (DBM), Plutella xylostella, is one of the most harmful insect pests on crucifer crops worldwide. In this study, 19 cDNAs encoding glutathione S-transferases (GSTs) were identified from the genomic and transcriptomic database for DBM (KONAGAbase) and further characterized. Phylogenetic analysis showed that the 19 GSTs were classified into six different cytosolic classes, including four in delta, six in epsilon, three in omega, two in sigma, one in theta and one in zeta. Two GSTs were unclassified. RT-PCR analysis revealed that most GST genes were expressed in all developmental stages, with higher expression in the larval stages. Six DBM GSTs were expressed at the highest levels in the midgut tissue. Twelve purified recombinant GSTs showed varied enzymatic properties towards 1-chloro-2,4-dinitrobenzene and glutathione, whereas rPxGSTo2, rPxGSTz1 and rPxGSTu2 had no activity. Real-time quantitative PCR revealed that expression levels of the 19 DBM GST genes were varied and changed after exposure to acephate, indoxacarb, beta-cypermethrin and spinosad. PxGSTd3 was significantly overexpressed, while PxGSTe3 and PxGSTs2 were significantly downregulated by all four insecticide exposures. The changes in DBM GST gene expression levels exposed to different insecticides indicate that they may play individual roles in tolerance to insecticides and xenobiotics. © 2014 Society of Chemical Industry.

  3. Novel derivatives of aclacinomycin A block cancer cell migration through inhibition of farnesyl transferase.

    Science.gov (United States)

    Magi, Shigeyuki; Shitara, Tetsuo; Takemoto, Yasushi; Sawada, Masato; Kitagawa, Mitsuhiro; Tashiro, Etsu; Takahashi, Yoshikazu; Imoto, Masaya

    2013-03-01

    In the course of screening for an inhibitor of farnesyl transferase (FTase), we identified two compounds, N-benzyl-aclacinomycin A (ACM) and N-allyl-ACM, which are new derivatives of ACM. N-benzyl-ACM and N-allyl-ACM inhibited FTase activity with IC50 values of 0.86 and 2.93 μM, respectively. Not only ACM but also C-10 epimers of each ACM derivative failed to inhibit FTase. The inhibition of FTase by N-benzyl-ACM and N-allyl-ACM seems to be specific, because these two compounds did not inhibit geranylgeranyltransferase or geranylgeranyl pyrophosphate (GGPP) synthase up to 100 μM. In cultured A431 cells, N-benzyl-ACM and N-allyl-ACM also blocked both the membrane localization of H-Ras and activation of the H-Ras-dependent PI3K/Akt pathway. In addition, they inhibited epidermal growth factor (EGF)-induced migration of A431 cells. Thus, N-benzyl-ACM and N-allyl-ACM inhibited EGF-induced migration of A431 cells by inhibiting the farnesylation of H-Ras and subsequent H-Ras-dependent activation of the PI3K/Akt pathway.

  4. Electrochemical DNA biosensor based on grafting-to mode of terminal deoxynucleoside transferase-mediated extension.

    Science.gov (United States)

    Chen, Jinyuan; Liu, Zhoujie; Peng, Huaping; Zheng, Yanjie; Lin, Zhen; Liu, Ailin; Chen, Wei; Lin, Xinhua

    2017-12-15

    Previously reported electrochemical DNA biosensors based on in-situ polymerization approach reveal that terminal deoxynucleoside transferase (TdTase) has good amplifying performance and promising application in the design of electrochemical DNA biosensor. However, this method, in which the background is significantly affected by the amount of TdTase, suffers from being easy to produce false positive result and poor stability. Herein, we firstly present a novel electrochemical DNA biosensor based on grafting-to mode of TdTase-mediated extension, in which DNA targets are polymerized in homogeneous solution and then hybridized with DNA probes on BSA-based DNA carrier platform. It is surprising to find that the background in the grafting-to mode of TdTase-based electrochemical DNA biosensor have little interference from the employed TdTase. Most importantly, the proposed electrochemical DNA biosensor shows greatly improved detection performance over the in-situ polymerization approach-based electrochemical DNA biosensor. Copyright © 2017 Elsevier B.V. All rights reserved.

  5. Relation of serum γ-glutamyl transferase activity with copper in an adult population.

    Science.gov (United States)

    Peng, You-Fan; Wang, Chun-Fang; Pan, Guo-Gang

    2017-10-26

    The aim of this study was to evaluate the relationship between serum γ-glutamyl transferase (γ-GGT) activity and serum copper in an adult population. We analyzed 281 adult subjects who regularly attended the physical examination center at the Affiliated Hospital of Youjiang Medical University for Nationalities. The demographic and laboratory data of the participants were divided into two groups according to the median of serum γ-GGT activity. Serum copper concentrations in individuals with higher γ-GGT levels were significantly increased compared with those with lower γ-GGT concentrations (9.9±2.41 vs. 11.2±3.36 μmol/L, pcopper in all eligible subjects (r=0.198, p=0.001). Further, serum γ-GGT maintained a positive correlation with serum copper in both males and females (r=0.322, pcopper after adjusting for multiple potential confounders (b=0.464, p=0.001). This study suggests that serum γ-GGT activity is correlated with copper in the study population, indicating that serum γ-GGT may be a biomarker to evaluate serum copper levels in an adult population.

  6. Legionella shows a diverse secondary metabolism dependent on a broad spectrum Sfp-type phosphopantetheinyl transferase

    Directory of Open Access Journals (Sweden)

    Nicholas J. Tobias

    2016-11-01

    Full Text Available Several members of the genus Legionella cause Legionnaires’ disease, a potentially debilitating form of pneumonia. Studies frequently focus on the abundant number of virulence factors present in this genus. However, what is often overlooked is the role of secondary metabolites from Legionella. Following whole genome sequencing, we assembled and annotated the Legionella parisiensis DSM 19216 genome. Together with 14 other members of the Legionella, we performed comparative genomics and analysed the secondary metabolite potential of each strain. We found that Legionella contains a huge variety of biosynthetic gene clusters (BGCs that are potentially making a significant number of novel natural products with undefined function. Surprisingly, only a single Sfp-like phosphopantetheinyl transferase is found in all Legionella strains analyzed that might be responsible for the activation of all carrier proteins in primary (fatty acid biosynthesis and secondary metabolism (polyketide and non-ribosomal peptide synthesis. Using conserved active site motifs, we predict some novel compounds that are probably involved in cell-cell communication, differing to known communication systems. We identify several gene clusters, which may represent novel signaling mechanisms and demonstrate the natural product potential of Legionella.

  7. Glutathione S-transferase P influences redox and migration pathways in bone marrow.

    Directory of Open Access Journals (Sweden)

    Jie Zhang

    Full Text Available To interrogate why redox homeostasis and glutathione S-transferase P (GSTP are important in regulating bone marrow cell proliferation and migration, we isolated crude bone marrow, lineage negative and bone marrow derived-dendritic cells (BMDDCs from both wild type (WT and knockout (Gstp1/p2(-/- mice. Comparison of the two strains showed distinct thiol expression patterns. WT had higher baseline and reactive oxygen species-induced levels of S-glutathionylated proteins, some of which (sarco-endoplasmic reticulum Ca2(+-ATPase regulate Ca(2+ fluxes and subsequently influence proliferation and migration. Redox status is also a crucial determinant in the regulation of the chemokine system. CXCL12 chemotactic response was stronger in WT cells, with commensurate alterations in plasma membrane polarization/permeability and intracellular calcium fluxes; activities of the downstream kinases, ERK and Akt were also higher in WT. In addition, expression levels of the chemokine receptor CXCR4 and its associated phosphatase, SHP-2, were higher in WT. Inhibition of CXCR4 or SHP2 decreased the extent of CXCL12-induced migration in WT BMDDCs. The differential surface densities of CXCR4, SHP-2 and inositol trisphosphate receptor in WT and Gstp1/p2(-/- cells correlated with the differential CXCR4 functional activities, as measured by the extent of chemokine-induced directional migration and differences in intracellular signaling. These observed differences contribute to our understanding of how genetic ablation of GSTP causes different levels of myeloproliferation and migration [corrected

  8. Recognition and Detoxification of the Insecticide DDT by Drosophila melanogaster Glutathione S-Transferase D1

    Energy Technology Data Exchange (ETDEWEB)

    Low, Wai Yee; Feil, Susanne C.; Ng, Hooi Ling; Gorman, Michael A.; Morton, Craig J.; Pyke, James; McConville, Malcolm J.; Bieri, Michael; Mok, Yee-Foong; Robin, Charles; Gooley, Paul R.; Parker, Michael W.; Batterham, Philip (SVIMR-A); (Melbourne)

    2010-06-14

    GSTD1 is one of several insect glutathione S-transferases capable of metabolizing the insecticide DDT. Here we use crystallography and NMR to elucidate the binding of DDT and glutathione to GSTD1. The crystal structure of Drosophila melanogaster GSTD1 has been determined to 1.1 {angstrom} resolution, which reveals that the enzyme adopts the canonical GST fold but with a partially occluded active site caused by the packing of a C-terminal helix against one wall of the binding site for substrates. This helix would need to unwind or be displaced to enable catalysis. When the C-terminal helix is removed from the model of the crystal structure, DDT can be computationally docked into the active site in an orientation favoring catalysis. Two-dimensional {sup 1}H,{sup 15}N heteronuclear single-quantum coherence NMR experiments of GSTD1 indicate that conformational changes occur upon glutathione and DDT binding and the residues that broaden upon DDT binding support the predicted binding site. We also show that the ancestral GSTD1 is likely to have possessed DDT dehydrochlorinase activity because both GSTD1 from D. melanogaster and its sibling species, Drosophila simulans, have this activity.

  9. Targeting Glutathione-S Transferase Enzymes in Musculoskeletal Sarcomas: A Promising Therapeutic Strategy

    Directory of Open Access Journals (Sweden)

    Michela Pasello

    2011-01-01

    Full Text Available Recent studies have indicated that targeting glutathione-S-transferase (GST isoenzymes may be a promising novel strategy to improve the efficacy of conventional chemotherapy in the three most common musculoskeletal tumours: osteosarcoma, Ewing's sarcoma, and rhabdomyosarcoma. By using a panel of 15 drug-sensitive and drug-resistant human osteosarcoma, Ewing's sarcoma, and rhabdomyosarcoma cell lines, the efficay of the GST-targeting agent 6-(7-nitro-2,1,3-benzoxadiazol-4-ylthiohexanol (NBDHEX has been assessed and related to GST isoenzymes expression (namely GSTP1, GSTA1, GSTM1, and MGST. NBDHEX showed a relevant in vitro activity on all cell lines, including the drug-resistant ones and those with higher GSTs levels. The in vitro activity of NBDHEX was mostly related to cytostatic effects, with a less evident apoptotic induction. NBDHEX positively interacted with doxorubicin, vincristine, cisplatin but showed antagonistic effects with methotrexate. In vivo studies confirmed the cytostatic efficay of NBDHEX and its positive interaction with vincristine in Ewing's sarcoma cells, and also indicated a positive effect against the metastatisation of osteosarcoma cells. The whole body of evidence found in this study indicated that targeting GSTs in osteosarcoma, Ewing's sarcoma and rhabdomyosarcoma may be an interesting new therapeutic option, which can be considered for patients who are scarcely responsive to conventional regimens.

  10. Terminal deoxynucleotidyl transferase activity in the regenerating thymus of X-irradiated mice

    International Nuclear Information System (INIS)

    Daculsi, R.; Astier, T.; Legrand, E.; Duplan, J.F.

    1982-01-01

    The distribution of terminal deoyxnucleotidyl transferase (TdT) enzyme activity (EU per 10 8 cells) between peaks I and II was followed for a period of 42 days in regenerating thymus of lethally irradiated (9 Gy) C3H mice restored with 10 6 (C3H x AKR) F1 bone marrow cells. The detection of Thy-1.1 and Thy-1.2 surface antigens allowed for the discrimination between host and donor cells, and the main subpopulations of thymic cells were characterized by their sensitivity to H-2sup(k) antiserum and to dexamethazone. Two peaks of TdT activity could be detected on phosphocellulose chromatographic separation. The distribution of TdT activity between these two peaks was followed during the two periods of thymic endo- and exoregeneration. Peak I TdT activity was closely correlated with the variation in the percentage of the high H-2 population. Peak II activity was mostly related to low H-2 cells. The per cell content of both peak I and peak II activities exceeded the norm in rapidly expanding populations. Finally between days 10 and 14 the TdT activity of the endoregenerating population was apparently not different from that of the exoregenerating population between days 14 and 22. (Auth.)

  11. The pleuromutilin drugs tiamulin and valnemulin bind to the RNA at the peptidyl transferase centre on the ribosome

    DEFF Research Database (Denmark)

    Poulsen, S M; Karlsson, M; Johansson, L B

    2001-01-01

    The pleuromutilin antibiotic derivatives, tiamulin and valnemulin, inhibit protein synthesis by binding to the 50S ribosomal subunit of bacteria. The action and binding site of tiamulin and valnemulin was further characterized on Escherichia coli ribosomes. It was revealed that these drugs...... centre and have been associated with binding of several antibiotics. Competitive footprinting shows that tiamulin and valnemulin can bind concurrently with the macrolide erythromycin but compete with the macrolide carbomycin, which is a peptidyl transferase inhibitor. We infer from these and previous...... results that tiamulin and valnemulin interact with the rRNA in the peptidyl transferase slot on the ribosomes in which they prevent the correct positioning of the CCA-ends of tRNAs for peptide transfer....

  12. A model to environmental monitoring based on glutathione-S-transferase activity and branchial lesions in catfish

    Science.gov (United States)

    Neta, Raimunda Nonata Fortes Carvalho; Torres, Audalio Rebelo

    2017-11-01

    In this work, we validate the glutathione-S-transferase and branchial lesions as biomarkers in catfish Sciades herzbergii to obtain a predictive model of the environmental impact effects in a harbor of Brazil. The catfish were sampled from a port known to be contaminated with heavy metals and organic compounds and from a natural reserve in São Marcos Bay, Maranhão. Two biomarkers, hepatic glutathione S-transferase (GST) activity and branchial lesions were analyzed. The values for GST activity were modeled with the occurrence of branchial lesions by fitting a third order polynomial. Results from the mathematical model indicate that GST activity has a strong polynomial relationship with the occurrence of branchial lesions in both the wet and the dry seasons, but only at the polluted port site. Our mathematic model indicates that when the GST ceases to act, serious branchial lesions are observed in the catfish of the contaminated port area.

  13. Glutathione S-Transferase of Brown Planthoppers (Nilaparvata lugens) Is Essential for Their Adaptation to Gramine-Containing Host Plants

    OpenAIRE

    Sun, Xiao-Qin; Zhang, Mao-Xin; Yu, Jing-Ya; Jin, Yu; Ling, Bing; Du, Jin-Ping; Li, Gui-Hua; Qin, Qing-Ming; Cai, Qing-Nian

    2013-01-01

    Plants have evolved complex processes to ward off attacks by insects. In parallel, insects have evolved mechanisms to thwart these plant defenses. To gain insight into mechanisms that mediate this arms race between plants and herbivorous insects, we investigated the interactions between gramine, a toxin synthesized by plants of the family Gramineae, and glutathione S transferase (GST), an enzyme found in insects that is known to detoxify xenobiotics. Here, we demonstrate that rice (Oryza sati...

  14. The activity of glutathione S-transferase in hepatopancreas of Procambarus clarkii: seasonal variations and the influence of environmental pollutants.

    Science.gov (United States)

    Nies, E; Almar, M M; Hermenegildo, C; Monsalve, E; Romero, F J

    1991-01-01

    1. The glutathione S-transferase activity in hepatopancreas of the American red crayfish Procambarus clarkii after 15 days' acclimatization in tap water aquaria was measured in specimens collected monthly for a whole year, and shows seasonal variation. 2. Previous data on the environmental pollution of Lake Albufera suggest a possible correlation with the activity tested in the different seasons of the year considering the results of non-acclimatized animals.

  15. β(1,3-glucanosyl-transferase activity is essential for cell wall integrity and viability of Schizosaccharomyces pombe.

    Directory of Open Access Journals (Sweden)

    María de Medina-Redondo

    Full Text Available BACKGROUND: The formation of the cell wall in Schizosaccharomyces pombe requires the coordinated activity of enzymes involved in the biosynthesis and modification of β-glucans. The β(1,3-glucan synthase complex synthesizes linear β(1,3-glucans, which remain unorganized until they are cross-linked to other β(1,3-glucans and other cell wall components. Transferases of the GH72 family play important roles in cell wall assembly and its rearrangement in Saccharomyces cerevisiae and Aspergillus fumigatus. Four genes encoding β(1,3-glucanosyl-transferases -gas1(+, gas2(+, gas4(+ and gas5(+- are present in S. pombe, although their function has not been analyzed. METHODOLOGY/PRINCIPAL FINDINGS: Here, we report the characterization of the catalytic activity of gas1p, gas2p and gas5p together with studies directed to understand their function during vegetative growth. From the functional point of view, gas1p is essential for cell integrity and viability during vegetative growth, since gas1Δ mutants can only grow in osmotically supported media, while gas2p and gas5p play a minor role in cell wall construction. From the biochemical point of view, all of them display β(1,3-glucanosyl-transferase activity, although they differ in their specificity for substrate length, cleavage point and product size. In light of all the above, together with the differences in expression profiles during the life cycle, the S. pombe GH72 proteins may accomplish complementary, non-overlapping functions in fission yeast. CONCLUSIONS/SIGNIFICANCE: We conclude that β(1,3-glucanosyl-transferase activity is essential for viability in fission yeast, being required to maintain cell integrity during vegetative growth.

  16. Meat consumption, N-acetyl transferase 1 and 2 polymorphism and risk of breast cancer, in Danish postmenopausal women

    DEFF Research Database (Denmark)

    Egeberg, Rikke; Olsen, Anja; Autrup, Herman

    2008-01-01

    The aim of this study was to investigate whether polymorphisms in N-acetyl transferase 1 and 2 modify the association between meat consumption and risk of breast cancer. A nested case-control study was conducted among 24697 postmenopausal women included in the 'Diet, Cancer and Health' cohort study...... (1993-2000). Three hundred and seventy-eight breast cancer cases were identified and matched to 378 controls. The incidence rate ratio (95% confidence interval) for breast cancer was 1.09 (1.02-1.17) for total meat, 1.15 (1.01-1.31) for red meat and 1.23 (1.04-1.45) for processed meat per 25 g daily...... total meat intake and red meat intake and breast cancer risk were confined to intermediate/fast N-acetyl transferase 2 acetylators (P-interaction=0.03 and 0.04). Our findings support an association between meat consumption and breast cancer risk and that N-acetyl transferase 2 polymorphism has...

  17. Structures of a putative ζ-class glutathione S-transferase from the pathogenic fungus Coccidioides immitis

    International Nuclear Information System (INIS)

    Edwards, Thomas E.; Bryan, Cassie M.; Leibly, David J.; Dieterich, Shellie H.; Abendroth, Jan; Sankaran, Banumathi; Sivam, Dhileep; Staker, Bart L.; Van Voorhis, Wesley C.; Myler, Peter J.; Stewart, Lance J.

    2011-01-01

    The pathogenic fungus C. immitis causes coccidioidomycosis, a potentially fatal disease. Here, apo and glutathione-bound crystal structures of a previously uncharacterized protein from C. immitis that appears to be a ζ-class glutathione S-transferase are presented. Coccidioides immitis is a pathogenic fungus populating the southwestern United States and is a causative agent of coccidioidomycosis, sometimes referred to as Valley Fever. Although the genome of this fungus has been sequenced, many operons are not properly annotated. Crystal structures are presented for a putative uncharacterized protein that shares sequence similarity with ζ-class glutathione S-transferases (GSTs) in both apo and glutathione-bound forms. The apo structure reveals a nonsymmetric homodimer with each protomer comprising two subdomains: a C-terminal helical domain and an N-terminal thioredoxin-like domain that is common to all GSTs. Half-site binding is observed in the glutathione-bound form. Considerable movement of some components of the active site relative to the glutathione-free form was observed, indicating an induced-fit mechanism for cofactor binding. The sequence homology, structure and half-site occupancy imply that the protein is a ζ-class glutathione S-transferase, a maleylacetoacetate isomerase (MAAI)

  18. Neuroantibodies (NAB) in African-American Children: Associations with Gender, Glutathione-S-Transferase (GST)Pi Polymorphisms (SNP) and Heavy Metals

    Science.gov (United States)

    CONTACT (NAME ONLY): Hassan El-Fawal Abstract Details PRESENTATION TYPE: Platform or Poster CURRENT CATEGORY: Neurodegenerative Disease | Biomarkers | Neurotoxicity, Metals KEYWORDS: Autoantibodies, Glutathione-S-Transferase, DATE/TIME LAST MODIFIED: DATE/TIME SUBMITTED: Abs...

  19. Evaluation of antioxidant and antimicrobial activity of seaweed ( Sargassum sp.) extract: A study on inhibition of glutathione-S-transferase Activity

    Digital Repository Service at National Institute of Oceanography (India)

    Patra, J.K.; Rath, S.K.; Jena, K.B.; Rathod, V.K.; Thatoi, H.

    In the present study, the free radical scavenging potentials (DPPH radical and hydroxyl radical), inhibition of lipid peroxidation, and glutathione-S-transferase and antimicrobial properties of Sargassum sp. extract were investigated. The tested...

  20. Glutathione S-transferase P protects against cyclophosphamide-induced cardiotoxicity in mice

    Energy Technology Data Exchange (ETDEWEB)

    Conklin, Daniel J., E-mail: dj.conklin@louisville.edu [Diabetes and Obesity Center, University of Louisville, Louisville, KY 40292 (United States); Institute of Molecular Cardiology, University of Louisville, Louisville, KY 40292 (United States); Haberzettl, Petra; Jagatheesan, Ganapathy; Baba, Shahid [Diabetes and Obesity Center, University of Louisville, Louisville, KY 40292 (United States); Institute of Molecular Cardiology, University of Louisville, Louisville, KY 40292 (United States); Merchant, Michael L. [Diabetes and Obesity Center, University of Louisville, Louisville, KY 40292 (United States); Division of Nephrology, Department of Medicine, University of Louisville, Louisville, KY 40292 (United States); Prough, Russell A. [Diabetes and Obesity Center, University of Louisville, Louisville, KY 40292 (United States); Department of Biochemistry and Molecular Biology, University of Louisville, Louisville, KY 40292 (United States); Williams, Jessica D. [University of Cincinnati College of Medicine, Internal Medicine, Cincinnati, OH 45267 (United States); Prabhu, Sumanth D. [Division of Cardiovascular Disease, University of Alabama-Birmingham, Birmingham, AL 35294 (United States); Bhatnagar, Aruni [Diabetes and Obesity Center, University of Louisville, Louisville, KY 40292 (United States); Institute of Molecular Cardiology, University of Louisville, Louisville, KY 40292 (United States); Department of Biochemistry and Molecular Biology, University of Louisville, Louisville, KY 40292 (United States)

    2015-06-01

    High-dose chemotherapy regimens using cyclophosphamide (CY) are frequently associated with cardiotoxicity that could lead to myocyte damage and congestive heart failure. However, the mechanisms regulating the cardiotoxic effects of CY remain unclear. Because CY is converted to an unsaturated aldehyde acrolein, a toxic, reactive CY metabolite that induces extensive protein modification and myocardial injury, we examined the role of glutathione S-transferase P (GSTP), an acrolein-metabolizing enzyme, in CY cardiotoxicity in wild-type (WT) and GSTP-null mice. Treatment with CY (100–300 mg/kg) increased plasma levels of creatine kinase-MB isoform (CK·MB) and heart-to-body weight ratio to a significantly greater extent in GSTP-null than WT mice. In addition to modest yet significant echocardiographic changes following acute CY-treatment, GSTP insufficiency was associated with greater phosphorylation of c-Jun and p38 as well as greater accumulation of albumin and protein–acrolein adducts in the heart. Mass spectrometric analysis revealed likely prominent modification of albumin, kallikrein-1-related peptidase, myoglobin and transgelin-2 by acrolein in the hearts of CY-treated mice. Treatment with acrolein (low dose, 1–5 mg/kg) also led to increased heart-to-body weight ratio and myocardial contractility changes. Acrolein induced similar hypotension in GSTP-null and WT mice. GSTP-null mice also were more susceptible than WT mice to mortality associated with high-dose acrolein (10–20 mg/kg). Collectively, these results suggest that CY cardiotoxicity is regulated, in part, by GSTP, which prevents CY toxicity by detoxifying acrolein. Thus, humans with low cardiac GSTP levels or polymorphic forms of GSTP with low acrolein-metabolizing capacity may be more sensitive to CY toxicity. - Graphical abstract: Cyclophosphamide (CY) treatment results in P450-mediated metabolic formation of phosphoramide mustard and acrolein (3-propenal). Acrolein is either metabolized and

  1. Glutathione S-transferase P protects against cyclophosphamide-induced cardiotoxicity in mice

    International Nuclear Information System (INIS)

    Conklin, Daniel J.; Haberzettl, Petra; Jagatheesan, Ganapathy; Baba, Shahid; Merchant, Michael L.; Prough, Russell A.; Williams, Jessica D.; Prabhu, Sumanth D.; Bhatnagar, Aruni

    2015-01-01

    High-dose chemotherapy regimens using cyclophosphamide (CY) are frequently associated with cardiotoxicity that could lead to myocyte damage and congestive heart failure. However, the mechanisms regulating the cardiotoxic effects of CY remain unclear. Because CY is converted to an unsaturated aldehyde acrolein, a toxic, reactive CY metabolite that induces extensive protein modification and myocardial injury, we examined the role of glutathione S-transferase P (GSTP), an acrolein-metabolizing enzyme, in CY cardiotoxicity in wild-type (WT) and GSTP-null mice. Treatment with CY (100–300 mg/kg) increased plasma levels of creatine kinase-MB isoform (CK·MB) and heart-to-body weight ratio to a significantly greater extent in GSTP-null than WT mice. In addition to modest yet significant echocardiographic changes following acute CY-treatment, GSTP insufficiency was associated with greater phosphorylation of c-Jun and p38 as well as greater accumulation of albumin and protein–acrolein adducts in the heart. Mass spectrometric analysis revealed likely prominent modification of albumin, kallikrein-1-related peptidase, myoglobin and transgelin-2 by acrolein in the hearts of CY-treated mice. Treatment with acrolein (low dose, 1–5 mg/kg) also led to increased heart-to-body weight ratio and myocardial contractility changes. Acrolein induced similar hypotension in GSTP-null and WT mice. GSTP-null mice also were more susceptible than WT mice to mortality associated with high-dose acrolein (10–20 mg/kg). Collectively, these results suggest that CY cardiotoxicity is regulated, in part, by GSTP, which prevents CY toxicity by detoxifying acrolein. Thus, humans with low cardiac GSTP levels or polymorphic forms of GSTP with low acrolein-metabolizing capacity may be more sensitive to CY toxicity. - Graphical abstract: Cyclophosphamide (CY) treatment results in P450-mediated metabolic formation of phosphoramide mustard and acrolein (3-propenal). Acrolein is either metabolized and

  2. The poplar phi class glutathione transferase: expression, activity and structure of GSTF1

    Directory of Open Access Journals (Sweden)

    Henri ePégeot

    2014-12-01

    Full Text Available Glutathione transferases (GSTs constitute a superfamily of enzymes with essential roles in cellular detoxification and secondary metabolism in plants as in other organisms. Several plant GSTs, including those of the Phi class (GSTFs, require a conserved catalytic serine residue to perform glutathione (GSH-conjugation reactions. Genomic analyses revealed that terrestrial plants have around 10 GSTFs, 8 in the Populus trichocarpa genome, but their physiological functions and substrates are mostly unknown. Transcript expression analyses showed a predominant expression of all genes both in reproductive (female flowers, fruits, floral buds and vegetative organs (leaves, petioles. Here, we show that the recombinant poplar GSTF1 (PttGSTF1 possesses peroxidase activity towards cumene hydroperoxide and GSH-conjugation activity towards model substrates such as 2,4-dinitrochlorobenzene, benzyl and phenetyl isothiocyanate, 4-nitrophenyl butyrate and 4-hydroxy-2-nonenal but interestingly not on previously identified GSTF-class substrates. In accordance to analytical gel filtration data, crystal structure of PttGSTF1 showed a canonical dimeric organization with bound GSH or MES molecules. The structure of these protein-substrate complexes allowed delineating the residues contributing to both the G and H sites that form the active site cavity. In sum, the presence of GSTF1 transcripts and proteins in most poplar organs especially those rich in secondary metabolites such as flowers and fruits, together with its GSH-conjugation activity and its documented stress-responsive expression suggest that its function is associated with the catalytic transformation of metabolites and/or peroxide removal rather than with ligandin properties as previously reported for other GSTFs.

  3. Glutathione S Transferases Polymorphisms Are Independent Prognostic Factors in Lupus Nephritis Treated with Cyclophosphamide.

    Directory of Open Access Journals (Sweden)

    Alexandra Audemard-Verger

    Full Text Available To investigate association between genetic polymorphisms of GST, CYP and renal outcome or occurrence of adverse drug reactions (ADRs in lupus nephritis (LN treated with cyclophosphamide (CYC. CYC, as a pro-drug, requires bioactivation through multiple hepatic cytochrome P450s and glutathione S transferases (GST.We carried out a multicentric retrospective study including 70 patients with proliferative LN treated with CYC. Patients were genotyped for polymorphisms of the CYP2B6, CYP2C19, GSTP1, GSTM1 and GSTT1 genes. Complete remission (CR was defined as proteinuria ≤0.33g/day and serum creatinine ≤124 µmol/l. Partial remission (PR was defined as proteinuria ≤1.5g/day with a 50% decrease of the baseline proteinuria value and serum creatinine no greater than 25% above baseline.Most patients were women (84% and 77% were Caucasian. The mean age at LN diagnosis was 41 ± 10 years. The frequency of patients carrying the GST null genotype GSTT1-, GSTM1-, and the Ile→105Val GSTP1 genotype were respectively 38%, 60% and 44%. In multivariate analysis, the Ile→105Val GSTP1 genotype was an independent factor of poor renal outcome (achievement of CR or PR (OR = 5.01 95% CI [1.02-24.51] and the sole factor that influenced occurrence of ADRs was the GSTM1 null genotype (OR = 3.34 95% CI [1.064-10.58]. No association between polymorphisms of cytochrome P450s gene and efficacy or ADRs was observed.This study suggests that GST polymorphisms highly impact renal outcome and occurrence of ADRs related to CYC in LN patients.

  4. Glutathione-supported arsenate reduction coupled to arsenolysis catalyzed by ornithine carbamoyl transferase

    International Nuclear Information System (INIS)

    Nemeti, Balazs; Gregus, Zoltan

    2009-01-01

    Three cytosolic phosphorolytic/arsenolytic enzymes, (purine nucleoside phosphorylase [PNP], glycogen phosphorylase, glyceraldehyde-3-phosphate dehydrogenase) have been shown to mediate reduction of arsenate (AsV) to the more toxic arsenite (AsIII) in a thiol-dependent manner. With unknown mechanism, hepatic mitochondria also reduce AsV. Mitochondria possess ornithine carbamoyl transferase (OCT), which catalyzes phosphorolytic or arsenolytic citrulline cleavage; therefore, we examined if mitochondrial OCT facilitated AsV reduction in presence of glutathione. Isolated rat liver mitochondria were incubated with AsV, and AsIII formed was quantified. Glutathione-supplemented permeabilized or solubilized mitochondria reduced AsV. Citrulline (substrate for OCT-catalyzed arsenolysis) increased AsV reduction. The citrulline-stimulated AsV reduction was abolished by ornithine (OCT substrate inhibiting citrulline cleavage), phosphate (OCT substrate competing with AsV), and the OCT inhibitor norvaline or PALO, indicating that AsV reduction is coupled to OCT-catalyzed arsenolysis of citrulline. Corroborating this conclusion, purified bacterial OCT mediated AsV reduction in presence of citrulline and glutathione with similar responsiveness to these agents. In contrast, AsIII formation by intact mitochondria was unaffected by PALO and slightly stimulated by citrulline, ornithine, and norvaline, suggesting minimal role for OCT in AsV reduction in intact mitochondria. In addition to OCT, mitochondrial PNP can also mediate AsIII formation; however, its role in AsV reduction appears severely limited by purine nucleoside supply. Collectively, mitochondrial and bacterial OCT promote glutathione-dependent AsV reduction with coupled arsenolysis of citrulline, supporting the hypothesis that AsV reduction is mediated by phosphorolytic/arsenolytic enzymes. Nevertheless, because citrulline cleavage is disfavored physiologically, OCT may have little role in AsV reduction in vivo.

  5. The effect of chemical carcinogenesis on rat glutathione S-transferase P1 gene transcriptional regulation.

    Science.gov (United States)

    Liu, D; Liao, M; Zuo, J; Henner, W D; Fan, F

    2001-03-01

    To investigate mechanisms of rat glutathione S-transferase P1 gene (rGSTP1) expression regulation during chemical carcinogenesis. we studied enhancer elements located in the region between -2.5 kb to -2.2 kb. The region was upstream from the start site of transcription and was divided into two major fragments, GPEI and GPEII. The GPEII fragment was further divided into two smaller fragments, GPEII- I and GPEII-2. Using a luciferase reporter system, we identified a strong enhancer of GPEI and a weak enhancer of GPEII in HeLa and a rat hepatoma cell line CBRH79 19 cell. The enhancer of GPEII was located within the GPEII-I region. Chemical stimulation by glycidyl methatylate (GMA) and phorbol 12-o-tetradecanoate 13-acetate (TPA) analysis revealed that induction of rGSTP1 expression was mainly through GPEI. Although H2O2 could enhance GPEII enhancer activity, the enhancement is not mediated by the NF-kappaB factor that bound the NF-kappaB site in GPEII. Using electrophoretic mobility shift assays (EMSA) and the UV cross-linking assays, we found that HeLa and CBRH7919 cells had proteins that specifically bound GPEI core sequence and a 64 kDa protein that interacted with GPEII-1. The cells from normal rat liver did not express the binding proteins. Therefore, the trans-acting factors seem to be closely related to GPEI, GPEII enhancer activities and may play an important role in high expression of rGSTPI gene.

  6. Glutathione S-transferase expression and isoenzyme composition during cell differentiation of Caco-2 cells

    International Nuclear Information System (INIS)

    Scharmach, E.; Hessel, S.; Niemann, B.; Lampen, A.

    2009-01-01

    The human colon adenocarcinoma cell line Caco-2 is frequently used to study human intestinal metabolism and transport of xenobiotica. Previous studies have shown that both Caco-2 cells and human colon cells constitutively express the multigene family of detoxifying enzymes glutathione S-transferases (GSTs), particularly GST alpha and GST pi. GSTs may play a fundamental role in the molecular interplay between phase I, II enzymes and ABC-transporters. The gut fermentation product, butyrate, can modulate the potential for detoxification. The aim of this study was to investigate the basal expression of further cytosolic GSTs in Caco-2 cells during cell differentiation. In addition, a comparison was made with expression levels in MCF-7 and HepG2, two other cell types with barrier functions. Finally, the butyrate-mediated modulation of gene and protein expression was determined by real time PCR and western blot analysis. In Caco-2, gene and protein expression levels of GST alpha increased during cell differentiation. High levels of GSTO1 and GSTP1 were constantly expressed. No expression of GSTM5 and GSTT1 was detected. HepG2 expressed GSTO1 and MCF-7 GSTZ1 most intensively. No expression of GSTA5, GSTM5, or GSTP1 was detected in either cell. Incubation of Caco-2 cells with butyrate (5 mM) significantly induced GSTA1 and GSTM2 in proliferating Caco-2 cells. In differentiated cells, butyrate tended to increase GSTO1 and GSTP1. The results of this study show that a differentiation-dependent expression of GSTs in Caco-2 cells may reflect the in vivo situation and indicate the potential of butyrate to modify intestinal metabolism. GSTA1-A4 have been identified as good markers for cell differentiation. The Caco-2 cell line is a useful model for assessing the potential of food-related substances to modulate the GST expression pattern.

  7. Menadione stress in Saccharomyces cerevisiae strains deficient in the glutathione transferases.

    Science.gov (United States)

    Castro, F A V; Herdeiro, R S; Panek, A D; Eleutherio, E C A; Pereira, M D

    2007-02-01

    Using S. cerevisiae as a eukaryotic cell model we have analyzed the involvement of both glutathione transferase isoforms, Gtt1 and Gtt2, in constitutive resistance and adaptive response to menadione, a quinone which can exert its toxicity as redox cycling and/or electrophiles. The detoxification properties, of these enzymes, have also been analyzed by the appearance of S-conjugates in the media. Direct exposure to menadione (20 mM/60 min) showed to be lethal for cells deficient on both Gtt1 and Gtt2 isoforms. However, after pre-treatment with a low menadione concentration, cells deficient in Gtt2 displayed reduced ability to acquire tolerance when compared with the control and the Gtt1 deficient strains. Analyzing the toxic effects of menadione we observed that the gtt2 mutant showed no reduction in lipid peroxidation levels. Moreover, measuring the levels of intracellular oxidation during menadione stress we have shown that the increase of this oxidative stress parameter was due to the capacity menadione possesses in generating reactive oxygen species (ROS) and that both GSH and Gtt2 isoform were required to enhance ROS production. Furthermore, the efflux of the menadione-GSH conjugate, which is related with detoxification of xenobiotic pathways, was not detected in the gtt2 mutant. Taken together, these results suggest that acquisition of tolerance against stress generated by menadione and the process of detoxification through S-conjugates are dependent upon Gtt2 activity. This assessment was corroborated by the increase of GTT2 expression, and not of GTT1, after menadione treatment.

  8. Characterization and expression profiling of glutathione S-transferases in the diamondback moth, Plutella xylostella (L.).

    Science.gov (United States)

    You, Yanchun; Xie, Miao; Ren, Nana; Cheng, Xuemin; Li, Jianyu; Ma, Xiaoli; Zou, Minming; Vasseur, Liette; Gurr, Geoff M; You, Minsheng

    2015-03-05

    Glutathione S-transferases (GSTs) are multifunctional detoxification enzymes that play important roles in insects. The completion of several insect genome projects has enabled the identification and characterization of GST genes over recent years. This study presents a genome-wide investigation of the diamondback moth (DBM), Plutella xylostella, a species in which the GSTs are of special importance because this pest is highly resistant to many insecticides. A total of 22 putative cytosolic GSTs were identified from a published P. xylostella genome and grouped into 6 subclasses (with two unclassified). Delta, Epsilon and Omega GSTs were numerically superior with 5 genes for each of the subclasses. The resulting phylogenetic tree showed that the P. xylostella GSTs were all clustered into Lepidoptera-specific branches. Intron sites and phases as well as GSH binding sites were strongly conserved within each of the subclasses in the GSTs of P. xylostella. Transcriptome-, RNA-seq- and qRT-PCR-based analyses showed that the GST genes were developmental stage- and strain-specifically expressed. Most of the highly expressed genes in insecticide resistant strains were also predominantly expressed in the Malpighian tubules, midgut or epidermis. To date, this is the most comprehensive study on genome-wide identification, characterization and expression profiling of the GST family in P. xylostella. The diversified features and expression patterns of the GSTs are inferred to be associated with the capacity of this species to develop resistance to a wide range of pesticides and biological toxins. Our findings provide a base for functional research on specific GST genes, a better understanding of the evolution of insecticide resistance, and strategies for more sustainable management of the pest.

  9. Thermal- and urea-induced unfolding processes of glutathione S-transferase by molecular dynamics simulation.

    Science.gov (United States)

    Li, Jiahuang; Chen, Yuan; Yang, Jie; Hua, Zichun

    2015-05-01

    The Schistosoma juponicum 26 kDa glutathione S-transferase (sj26GST) consists of the N-terminal domain (N-domain), containing three alpha-helices (named H1-H3) and four anti-parallel beta-strands (S1-S4), and the C-terminal domain (C-domain), comprising five alpha-helices (named H4-H8). In present work, molecular dynamics simulations and fluorescence spectroscopic were used to gain insights into the unfolding process of sj26GST. The molecular dynamics simulations on sj26GST subunit both in water and in 8 M urea were carried out at 300 K, 400 K and 500 K, respectively. Spectroscopic measurements were employed to monitor structural changes. Molecular dynamics simulations of sj26GST subunit induced by urea and temperature showed that the initial unfolding step of sj26GST both in water and urea occurred on N-domain, involving the disruption of helices H2, H3 and strands S3 and S4, whereas H6 was the last region exposed to solution and was the last helix to unfold. Moreover, simulations analyses combining with fluorescence and circular dichroism spectra indicated that N-domain could not fold independent, suggesting that correct folding of N-domain depended on its interactions with C-domain. We further proposed that the folding of GSTs could begin with the hydrophobic collapse of C-domain whose H4, H5, H6 and H7 could move close to each other and form a hydrophobic core, especially H6 wrapped in the hydrophobic center and beginning spontaneous formation of the helix. S3, S4, H3, and H2 could form in the wake of the interaction between C-domain and N-domain. The paper can offer insights into the molecular mechanism of GSTs unfolding. © 2014 Wiley Periodicals, Inc.

  10. Glutathione S-transferase genotypes modify lung function decline in the general population: SAPALDIA cohort study

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    Ackermann-Liebrich Ursula

    2007-01-01

    Full Text Available Abstract Background Understanding the environmental and genetic risk factors of accelerated lung function decline in the general population is a first step in a prevention strategy against the worldwide increasing respiratory pathology of chronic obstructive pulmonary disease (COPD. Deficiency in antioxidative and detoxifying Glutathione S-transferase (GST gene has been associated with poorer lung function in children, smokers and patients with respiratory diseases. In the present study, we assessed whether low activity variants in GST genes are also associated with accelerated lung function decline in the general adult population. Methods We examined with multiple regression analysis the association of polymorphisms in GSTM1, GSTT1 and GSTP1 genes with annual decline in FEV1, FVC, and FEF25–75 during 11 years of follow-up in 4686 subjects of the prospective SAPALDIA cohort representative of the Swiss general population. Effect modification by smoking, gender, bronchial hyperresponisveness and age was studied. Results The associations of GST genotypes with FEV1, FVC, and FEF25–75 were comparable in direction, but most consistent for FEV1. GSTT1 homozygous gene deletion alone or in combination with GSTM1 homozygous gene deletion was associated with excess decline in FEV1 in men, but not women, irrespective of smoking status. The additional mean annual decline in FEV1 in men with GSTT1 and concurrent GSTM1 gene deletion was -8.3 ml/yr (95% confidence interval: -12.6 to -3.9 relative to men without these gene deletions. The GSTT1 effect on the FEV1 decline comparable to the observed difference in FEV1 decline between never and persistent smoking men. Effect modification by gender was statistically significant. Conclusion Our results suggest that genetic GSTT1 deficiency is a prevalent and strong determinant of accelerated lung function decline in the male general population.

  11. Farnesyl transferase inhibitors induce extended remissions in transgenic mice with mature B cell lymphomas

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    Refaeli Yosef

    2008-05-01

    Full Text Available Abstract Background We have used a mouse model based on overexpression of c-Myc in B cells genetically engineered to be self-reactive to test the hypothesis that farnesyl transferase inhibitors (FTIs can effectively treat mature B cell lymphomas. FTIs are undergoing clinical trials to treat both lymphoid and non-lymphoid malignancies and we wished to obtain evidence to support the inclusion of B cell lymphomas in future trials. Results We report that two FTIs, L-744,832 and SCH66336, blocked the growth of mature B cell lymphoma cells in vitro and in vivo. The FTI treatment affected the proliferation and survival of the transformed B cells to a greater extent than naïve B cells stimulated with antigen. In syngeneic mice transplanted with the transgenic lymphoma cells, L-744,832 treatment prevented the growth of the tumor cells and the morbidity associated with the resulting lymphoma progression. Tumors that arose from transplantation of the lymphoma cells regressed with as little as three days of treatment with L-744,832 or SCH66336. Treatment of these established lymphomas with L-744,832 for seven days led to long-term remission of the disease in approximately 25% of animals. Conclusion FTI treatment can block the proliferation and survival of self-reactive transformed B cells that overexpress Myc. In mice transplanted with mature B cell lymphomas, we found that FTI treatment led to regression of disease. FTIs warrant further consideration as therapeutic agents for mature B cell lymphomas and other lymphoid tumors.

  12. Enhanced tolerance and remediation of anthracene by transgenic tobacco plants expressing a fungal glutathione transferase gene

    Energy Technology Data Exchange (ETDEWEB)

    Dixit, Prachy; Mukherjee, Prasun K.; Sherkhane, Pramod D.; Kale, Sharad P. [Nuclear Agriculture and Biotechnology Division, Bhabha Atomic Research Centre, Mumbai 400085 (India); Eapen, Susan, E-mail: eapenhome@yahoo.com [Nuclear Agriculture and Biotechnology Division, Bhabha Atomic Research Centre, Mumbai 400085 (India)

    2011-08-15

    Highlights: {yields} Transgenic plants expressing a TvGST gene were tested for tolerance, uptake and degradation of anthracene. {yields} Transgenic plants were more tolerant to anthracene and take up more anthracene from soil and solutions compared to control plants. {yields} Using in vitro T{sub 1} seedlings, we showed that anthracene-a three fused benzene ring compound was phytodegraded to naphthalene derivatives, having two benzene rings. {yields} This is the first time that a transgenic plant was shown to have the potential to phytodegrade anthracene. - Abstract: Plants can be used for remediation of polyaromatic hydrocarbons, which are known to be a major concern for human health. Metabolism of xenobiotic compounds in plants occurs in three phases and glutathione transferases (GST) mediate phase II of xenobiotic transformation. Plants, although have GSTs, they are not very efficient for degradation of exogenous recalcitrant xenobiotics including polyaromatic hydrocarbons. Hence, heterologous expression of efficient GSTs in plants may improve their remediation and degradation potential of xenobiotics. In the present study, we investigated the potential of transgenic tobacco plants expressing a Trichoderma virens GST for tolerance, remediation and degradation of anthracene-a recalcitrant polyaromatic hydrocarbon. Transgenic plants with fungal GST showed enhanced tolerance to anthracene compared to control plants. Remediation of {sup 14}C uniformly labeled anthracene from solutions and soil by transgenic tobacco plants was higher compared to wild-type plants. Transgenic plants (T{sub 0} and T{sub 1}) degraded anthracene to naphthalene derivatives, while no such degradation was observed in wild-type plants. The present work has shown that in planta expression of a fungal GST in tobacco imparted enhanced tolerance as well as higher remediation potential of anthracene compared to wild-type plants.

  13. Determination of the serine palmitoyl transferase inhibitor myriocin by electrospray and Q-trap mass spectrometry.

    Science.gov (United States)

    Campisi, Giuseppe Matteo; Signorelli, Paola; Rizzo, Jessica; Ghilardi, Claudio; Antognetti, Jacopo; Caretti, Anna; Lazarević, Jelena S; Strettoi, Enrica; Novelli, Elena; Ghidoni, Riccardo; Rubino, Federico Maria; Paroni, Rita

    2017-12-01

    Myriocin is a potent inhibitor of serine-palmitoyl-transferase, the first and rate-determining enzyme in the sphingolipids biosynthetic pathway. This study developed, validated and applied a LC-MS/MS method to measure myriocin in minute specimens of animal tissue. The chemical analog 14-OH-myriocin was used as the internal standard. The two molecules were extracted from the tissue homogenate by solid-phase extraction, separated by gradient reversed-phase liquid chromatography and measured by negative ion electrospray mass spectrometry in the triple quadrupole. Detection was accomplished by multiple reaction monitoring, employing the most representative transitions, 400@104 and 402@104 for myriocin and 14-OH-myriocin, respectively. The typical limit of detection and lower limit of quantitation of the optimized method were 0.9 pmol/mL (~0.016 pmol injected) and 2.3 pmol/mL, respectively, and the method was linear up to 250 pmol/mL range (r 2  = 0.9996). The intra- and between-day repeatability afforded a coefficient of variation ≤7.0%. Applications included quantification of myriocin in mouse lungs after 24 h from administration of ~4 nmol by intra-tracheal delivery. Measured levels ranged from 4.11 (median; 2.3-7.4 IQR, n = 4) to 11.7 (median; 7.6-22.7 interquartile range (IQR), n = 6) pmol/lung depending on the different formulations used. Myriocin was also measured in retinas of mice treated by intravitreal injection and ranged from 0.045 (less than the limit of detection) to 0.35 pmol/retina. Copyright © 2017 John Wiley & Sons, Ltd.

  14. Cloning of a glutathione S-transferase decreasing during differentiation of HL60 cell line

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    Kim, Jae Chul; Park, In Kyu; Lee, Kyu Bo; Sohn, Sang Kyun; Kim, Moo Kyu; Kim, Jung Chul [College of Medicine, Kyungpook National Univ., Taegu (Korea, Republic of)

    1999-06-01

    By sequencing the Expressed Sequence Tags of human dermal papilla cDNA library, we identified a clone named K872 of which the expression decreased during differentiation of HL60 cell line. K872 plasmid DNA was isolated according to QIA plasmid extraction kit (Qiagen GmbH, Germany). The nucleotide sequencing was performed by Sanger's method with K872 plasmid DNA. The most updated GenBank EMBL necleic acid banks were searched through the internet by using BLAST (Basic Local Alignment Search Tools) program. Northern bots were performed using RNA isolated from various human tissues and cancer cell lines. The gene expression of the fusion protein was achieved by His-Patch Thiofusion expression system and the protein product was identified on SDS-PAGE. K872 clone is 1006 nucleotides long, and has a coding region of 675 nucleotides and a 3' non-coding region of 280 nucleotides. The presumed open reading frame starting at the 5' terminus of K872 encodes 226 amino acids, including the initiation methionine residue. The amino acid sequence deduced from the open reading frame of K872 shares 70% identity with that of rat glutathione S-transferase kappa 1 (rGSTK1). The transcripts were expressed inh a variety of human tissues and cancer cells. The levels of transcript were relatively high in those tissues such as heart, skeletal muscle, and peripheral blood leukocyte. It is noteworthy that K872 was found to be abundantly expressed in colorectal cancer and melanoma cell lines. Homology search result suggests that K872 clone is the human homolog of the rGSTK1 which is known to be involved in the resistance of cytotoxic therapy. We propose that meticulous functional analysis should be followed to confirm that.

  15. Glutathione-S-transferase profiles in the emerald ash borer, Agrilus planipennis.

    Science.gov (United States)

    Rajarapu, Swapna Priya; Mittapalli, Omprakash

    2013-05-01

    The emerald ash borer, Agrilus planipennis Fairmaire is a recently discovered invasive insect pest of ash, Fraxinus spp. in North America. Glutathione-S-transferases (GST) are a multifunctional superfamily of enzymes which function in conjugating toxic compounds to less toxic and excretable forms. In this study, we report the molecular characterization and expression patterns of different classes of GST genes in different tissues and developmental stages plus their specific activity. Multiple sequence alignment of all six A. planipennis GSTs (ApGST-E1, ApGST-E2, ApGST-E3, ApGST-O1, ApGST-S1 and ApGST-μ1) revealed conserved features of insect GSTs and a phylogenetic analysis grouped the GSTs within the epsilon, sigma, omega and microsomal classes of GSTs. Real time quantitative PCR was used to study field collected samples. In larval tissues high mRNA levels for ApGST-E1, ApGST-E3 and ApGST-O1 were obtained in the midgut and Malpighian tubules. On the other hand, ApGST-E2 and ApGST-S1 showed high mRNA levels in fat body and ApGST-μ1 showed constitutive levels in all the tissues assayed. During development, mRNA levels for ApGST-E2 were observed to be the highest in feeding instars, ApGST-S1 in prepupal instars; while the others showed constitutive patterns in all the developmental stages examined. At the enzyme level, total GST activity was similar in all the tissues and developmental stages assayed. Results obtained suggest that A. planipennis is potentially primed with GST-driven detoxification to metabolize ash allelochemicals. To our knowledge this study represents the first report of GSTs in A. planipennis and also in the family of wood boring beetles. Copyright © 2013 Elsevier Inc. All rights reserved.

  16. Enhanced tolerance and remediation of anthracene by transgenic tobacco plants expressing a fungal glutathione transferase gene

    International Nuclear Information System (INIS)

    Dixit, Prachy; Mukherjee, Prasun K.; Sherkhane, Pramod D.; Kale, Sharad P.; Eapen, Susan

    2011-01-01

    Highlights: → Transgenic plants expressing a TvGST gene were tested for tolerance, uptake and degradation of anthracene. → Transgenic plants were more tolerant to anthracene and take up more anthracene from soil and solutions compared to control plants. → Using in vitro T 1 seedlings, we showed that anthracene-a three fused benzene ring compound was phytodegraded to naphthalene derivatives, having two benzene rings. → This is the first time that a transgenic plant was shown to have the potential to phytodegrade anthracene. - Abstract: Plants can be used for remediation of polyaromatic hydrocarbons, which are known to be a major concern for human health. Metabolism of xenobiotic compounds in plants occurs in three phases and glutathione transferases (GST) mediate phase II of xenobiotic transformation. Plants, although have GSTs, they are not very efficient for degradation of exogenous recalcitrant xenobiotics including polyaromatic hydrocarbons. Hence, heterologous expression of efficient GSTs in plants may improve their remediation and degradation potential of xenobiotics. In the present study, we investigated the potential of transgenic tobacco plants expressing a Trichoderma virens GST for tolerance, remediation and degradation of anthracene-a recalcitrant polyaromatic hydrocarbon. Transgenic plants with fungal GST showed enhanced tolerance to anthracene compared to control plants. Remediation of 14 C uniformly labeled anthracene from solutions and soil by transgenic tobacco plants was higher compared to wild-type plants. Transgenic plants (T 0 and T 1 ) degraded anthracene to naphthalene derivatives, while no such degradation was observed in wild-type plants. The present work has shown that in planta expression of a fungal GST in tobacco imparted enhanced tolerance as well as higher remediation potential of anthracene compared to wild-type plants.

  17. Genetic polymorphism in three glutathione s-transferase genes and breast cancer risk

    Energy Technology Data Exchange (ETDEWEB)

    Woldegiorgis, S.; Ahmed, R.C.; Zhen, Y.; Erdmann, C.A.; Russell, M.L.; Goth-Goldstein, R.

    2002-04-01

    The role of the glutathione S-transferase (GST) enzyme family is to detoxify environmental toxins and carcinogens and to protect organisms from their adverse effects, including cancer. The genes GSTM1, GSTP1, and GSTT1 code for three GSTs involved in the detoxification of carcinogens, such as polycyclic aromatic hydrocarbons (PAHs) and benzene. In humans, GSTM1 is deleted in about 50% of the population, GSTT1 is absent in about 20%, whereas the GSTP1 gene has a single base polymorphism resulting in an enzyme with reduced activity. Epidemiological studies indicate that GST polymorphisms increase the level of carcinogen-induced DNA damage and several studies have found a correlation of polymorphisms in one of the GST genes and an increased risk for certain cancers. We examined the role of polymorphisms in genes coding for these three GST enzymes in breast cancer. A breast tissue collection consisting of specimens of breast cancer patients and non-cancer controls was analyzed by polymerase chain reaction (PCR) for the presence or absence of the GSTM1 and GSTT1 genes and for GSTP1 single base polymorphism by PCR/RFLP. We found that GSTM1 and GSTT1 deletions occurred more frequently in cases than in controls, and GSTP1 polymorphism was more frequent in controls. The effective detoxifier (putative low-risk) genotype (defined as presence of both GSTM1 and GSTT1 genes and GSTP1 wild type) was less frequent in cases than controls (16% vs. 23%, respectively). The poor detoxifier (putative high-risk) genotype was more frequent in cases than controls. However, the sample size of this study was too small to provide conclusive results.

  18. OGT (O-GlcNAc Transferase) Selectively Modifies Multiple Residues Unique to Lamin A.

    Science.gov (United States)

    Simon, Dan N; Wriston, Amanda; Fan, Qiong; Shabanowitz, Jeffrey; Florwick, Alyssa; Dharmaraj, Tejas; Peterson, Sherket B; Gruenbaum, Yosef; Carlson, Cathrine R; Grønning-Wang, Line M; Hunt, Donald F; Wilson, Katherine L

    2018-05-17

    The LMNA gene encodes lamins A and C with key roles in nuclear structure, signaling, gene regulation, and genome integrity. Mutations in LMNA cause over 12 diseases ('laminopathies'). Lamins A and C are identical for their first 566 residues. However, they form separate filaments in vivo, with apparently distinct roles. We report that lamin A is β- O -linked N -acetylglucosamine- (O -GlcNAc)-modified in human hepatoma (Huh7) cells and in mouse liver. In vitro assays with purified O -GlcNAc transferase (OGT) enzyme showed robust O -GlcNAcylation of recombinant mature lamin A tails (residues 385⁻646), with no detectable modification of lamin B1, lamin C, or 'progerin' (Δ50) tails. Using mass spectrometry, we identified 11 O -GlcNAc sites in a 'sweet spot' unique to lamin A, with up to seven sugars per peptide. Most sites were unpredicted by current algorithms. Double-mutant (S612A/T643A) lamin A tails were still robustly O -GlcNAc-modified at seven sites. By contrast, O -GlcNAcylation was undetectable on tails bearing deletion Δ50, which causes Hutchinson⁻Gilford progeria syndrome, and greatly reduced by deletion Δ35. We conclude that residues deleted in progeria are required for substrate recognition and/or modification by OGT in vitro. Interestingly, deletion Δ35, which does not remove the majority of identified O -GlcNAc sites, does remove potential OGT-association motifs (lamin A residues 622⁻625 and 639⁻645) homologous to that in mouse Tet1. These biochemical results are significant because they identify a novel molecular pathway that may profoundly influence lamin A function. The hypothesis that lamin A is selectively regulated by OGT warrants future testing in vivo, along with two predictions: genetic variants may contribute to disease by perturbing OGT-dependent regulation, and nutrient or other stresses might cause OGT to misregulate wildtype lamin A.

  19. O-GlcNAc transferase regulates transcriptional activity of human Oct4.

    Science.gov (United States)

    Constable, Sandii; Lim, Jae-Min; Vaidyanathan, Krithika; Wells, Lance

    2017-10-01

    O-linked β-N-acetylglucosamine (O-GlcNAc) is a single sugar modification found on many different classes of nuclear and cytoplasmic proteins. Addition of this modification, by the enzyme O-linked N-acetylglucosamine transferase (OGT), is dynamic and inducible. One major class of proteins modified by O-GlcNAc is transcription factors. O-GlcNAc regulates transcription factor properties through a variety of different mechanisms including localization, stability and transcriptional activation. Maintenance of embryonic stem (ES) cell pluripotency requires tight regulation of several key transcription factors, many of which are modified by O-GlcNAc. Octamer-binding protein 4 (Oct4) is one of the key transcription factors required for pluripotency of ES cells and more recently, the generation of induced pluripotent stem (iPS) cells. The action of Oct4 is modulated by the addition of several post-translational modifications, including O-GlcNAc. Previous studies in mice found a single site of O-GlcNAc addition responsible for transcriptional regulation. This study was designed to determine if this mechanism is conserved in humans. We mapped 10 novel sites of O-GlcNAc attachment on human Oct4, and confirmed a role for OGT in transcriptional activation of Oct4 at a site distinct from that found in mouse that allows distinction between different Oct4 target promoters. Additionally, we uncovered a potential new role for OGT that does not include its catalytic function. These results confirm that human Oct4 activity is being regulated by OGT by a mechanism that is distinct from mouse Oct4. © The Author 2017. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  20. Glutathione S-transferase expression and isoenzyme composition during cell differentiation of Caco-2 cells.

    Science.gov (United States)

    Scharmach, E; Hessel, S; Niemann, B; Lampen, A

    2009-11-30

    The human colon adenocarcinoma cell line Caco-2 is frequently used to study human intestinal metabolism and transport of xenobiotica. Previous studies have shown that both Caco-2 cells and human colon cells constitutively express the multigene family of detoxifying enzymes glutathione S-transferases (GSTs), particularly GST alpha and GST pi. GSTs may play a fundamental role in the molecular interplay between phase I, II enzymes and ABC-transporters. The gut fermentation product, butyrate, can modulate the potential for detoxification. The aim of this study was to investigate the basal expression of further cytosolic GSTs in Caco-2 cells during cell differentiation. In addition, a comparison was made with expression levels in MCF-7 and HepG2, two other cell types with barrier functions. Finally, the butyrate-mediated modulation of gene and protein expression was determined by real time PCR and western blot analysis. In Caco-2, gene and protein expression levels of GST alpha increased during cell differentiation. High levels of GSTO1 and GSTP1 were constantly expressed. No expression of GSTM5 and GSTT1 was detected. HepG2 expressed GSTO1 and MCF-7 GSTZ1 most intensively. No expression of GSTA5, GSTM5, or GSTP1 was detected in either cell. Incubation of Caco-2 cells with butyrate (5 mM) significantly induced GSTA1 and GSTM2 in proliferating Caco-2 cells. In differentiated cells, butyrate tended to increase GSTO1 and GSTP1. The results of this study show that a differentiation-dependent expression of GSTs in Caco-2 cells may reflect the in vivo situation and indicate the potential of butyrate to modify intestinal metabolism. GSTA1-A4 have been identified as good markers for cell differentiation. The Caco-2 cell line is a useful model for assessing the potential of food-related substances to modulate the GST expression pattern.

  1. Primary and secondary structural analyses of glutathione S-transferase pi from human placenta.

    Science.gov (United States)

    Ahmad, H; Wilson, D E; Fritz, R R; Singh, S V; Medh, R D; Nagle, G T; Awasthi, Y C; Kurosky, A

    1990-05-01

    The primary structure of glutathione S-transferase (GST) pi from a single human placenta was determined. The structure was established by chemical characterization of tryptic and cyanogen bromide peptides as well as automated sequence analysis of the intact enzyme. The structural analysis indicated that the protein is comprised of 209 amino acid residues and gave no evidence of post-translational modifications. The amino acid sequence differed from that of the deduced amino acid sequence determined by nucleotide sequence analysis of a cDNA clone (Kano, T., Sakai, M., and Muramatsu, M., 1987, Cancer Res. 47, 5626-5630) at position 104 which contained both valine and isoleucine whereas the deduced sequence from nucleotide sequence analysis identified only isoleucine at this position. These results demonstrated that in the one individual placenta studied at least two GST pi genes are coexpressed, probably as a result of allelomorphism. Computer assisted consensus sequence evaluation identified a hydrophobic region in GST pi (residues 155-181) that was predicted to be either a buried transmembrane helical region or a signal sequence region. The significance of this hydrophobic region was interpreted in relation to the mode of action of the enzyme especially in regard to the potential involvement of a histidine in the active site mechanism. A comparison of the chemical similarity of five known human GST complete enzyme structures, one of pi, one of mu, two of alpha, and one microsomal, gave evidence that all five enzymes have evolved by a divergent evolutionary process after gene duplication, with the microsomal enzyme representing the most divergent form.

  2. Electronics and electronic systems

    CERN Document Server

    Olsen, George H

    1987-01-01

    Electronics and Electronic Systems explores the significant developments in the field of electronics and electronic devices. This book is organized into three parts encompassing 11 chapters that discuss the fundamental circuit theory and the principles of analog and digital electronics. This book deals first with the passive components of electronic systems, such as resistors, capacitors, and inductors. These topics are followed by a discussion on the analysis of electronic circuits, which involves three ways, namely, the actual circuit, graphical techniques, and rule of thumb. The remaining p

  3. The NADPH thioredoxin reductase C functions as an electron donor to 2-Cys peroxiredoxin in a thermophilic cyanobacterium Thermosynechococcus elongatus BP-1

    International Nuclear Information System (INIS)

    Sueoka, Keigo; Yamazaki, Teruaki; Hiyama, Tetsuo; Nakamoto, Hitoshi

    2009-01-01

    An NADPH thioredoxin reductase C was co-purified with a 2-Cys peroxiredoxin by the combination of anion exchange chromatography and electroelution from gel slices after native PAGE from a thermophilic cyanobacterium Thermosynechococcus elongatus as an NAD(P)H oxidase complex induced by oxidative stress. The result provided a strong evidence that the NADPH thioredoxin reductase C interacts with the 2-Cys peroxiredoxin in vivo. An in vitro reconstitution assay with purified recombinant proteins revealed that both proteins were essential for an NADPH-dependent reduction of H 2 O 2 . These results suggest that the reductase transfers the reducing power from NADPH to the peroxiredoxin, which reduces peroxides in the cyanobacterium under oxidative stress. In contrast with other NADPH thioredoxin reductases, the NADPH thioredoxin reductase C contains a thioredoxin-like domain in addition to an NADPH thioredoxin reductase domain in the same polypeptide. Each domain contains a conserved CXYC motif. A point mutation at the CXYC motif in the NADPH thioredoxin reductase domain resulted in loss of the NADPH oxidation activity, while a mutation at the CXYC motif in the thioredoxin-like domain did not affect the electron transfer, indicating that this motif is not essential in the electron transport from NADPH to the 2-Cys peroxiredoxin.

  4. In vivo induction of phase II detoxifying enzymes, glutathione transferase and quinone reductase by citrus triterpenoids

    Directory of Open Access Journals (Sweden)

    Ahmad Hassan

    2010-09-01

    Full Text Available Abstract Background Several cell culture and animal studies demonstrated that citrus bioactive compounds have protective effects against certain types of cancer. Among several classes of citrus bioactive compounds, limonoids were reported to prevent different types of cancer. Furthermore, the structures of citrus limonoids were reported to influence the activity of phase II detoxifying enzymes. The purpose of the study was to evaluate how variations in the structures of citrus limonoids (namely nomilin, deacetyl nomilin, and isoobacunoic acid and a mixture of limonoids would influence phase II enzyme activity in excised tissues from a mouse model. Methods In the current study, defatted sour orange seed powder was extracted with ethyl acetate and subjected to silica gel chromatography. The HPLC, NMR and mass spectra were used to elucidate the purity and structure of compounds. Female A/J mice were treated with three limonoids and a mixture in order to evaluate their effect on phase II enzymes in four different tissues. Assays for glutathione S-transferase and NAD(PH: quinone reductase (QR were used to evaluate induction of phase II enzymatic activity. Results The highest induction of GST against 1-chloro-2,4-dinitrobenzene (CDNB was observed in stomach (whole, 58% by nomilin, followed by 25% isoobacunoic acid and 19% deacetyl nomilin. Deacetyl nomilin in intestine (small as well as liver significantly reduced GST activity against CDNB. Additionally isoobacunoic acid and the limonoid mixture in liver demonstrated a significant reduction of GST activity against CDNB. Nomilin significantly induced GST activity against 4-nitroquinoline 1-oxide (4NQO, intestine (280% and stomach (75% while deacetyl nomilin showed significant induction only in intestine (73%. Induction of GST activity was also observed in intestine (93% and stomach (45% treated with the limonoid mixture. Finally, a significant induction of NAD(PH: quinone reductase (QR activity was

  5. Glutathione S-transferase activity in follicular fluid from women undergoing ovarian stimulation: role in maturation.

    Science.gov (United States)

    Meijide, Susana; Hernández, M Luisa; Navarro, Rosaura; Larreategui, Zaloa; Ferrando, Marcos; Ruiz-Sanz, José Ignacio; Ruiz-Larrea, M Begoña

    2014-10-01

    Female infertility involves an emotional impact for the woman, often leading to a state of anxiety and low self-esteem. The assisted reproduction techniques (ART) are used to overcome the problem of infertility. In a first step of the in vitro fertilization therapy women are subjected to an ovarian stimulation protocol to obtain mature oocytes, which will result in competent oocytes necessary for fertilization to occur. Ovarian stimulation, however, subjects the women to a high physical and psychological stress, thus being essential to improve ART and to find biomarkers of dysfunction and fertility. GSH is an important antioxidant, and is also used in detoxification reactions, catalysed by glutathione S-transferases (GST). In the present work, we have investigated the involvement of GST in follicular maturation. Patients with fertility problems and oocyte donors were recruited for the study. From each woman follicles at two stages of maturation were extracted at the preovulatory stage. Follicular fluid was separated from the oocyte by centrifugation and used as the enzyme source. GST activity was determined based on its conjugation with 3,4-dichloronitrobenzene and the assay was adapted to a 96-well microplate reader. The absorbance was represented against the incubation time and the curves were adjusted to linearity (R(2)>0.990). Results showed that in both donors and patients GST activity was significantly lower in mature oocytes compared to small ones. These results suggest that GST may play a role in the follicle maturation by detoxifying xenobiotics, thus contributing to the normal development of the oocyte. Supported by FIS/FEDER (PI11/02559), Gobierno Vasco (Dep. Educación, Universiades e Investigación, IT687-13), and UPV/EHU (CLUMBER UFI11/20 and PES13/58). The work was approved by the Ethics Committee of the UPV/EHU (CEISH/96/2011/RUIZLARREA), and performed according to the UPV/EHU and IVI-Bilbao agreement (Ref. 2012/01). Copyright © 2014. Published by

  6. Clonorchis sinensis omega-class glutathione transferases are reliable biomarkers for serodiagnosis of clonorchiasis and opisthorchiasis.

    Science.gov (United States)

    Kim, J-G; Ahn, C-S; Sripa, B; Eom, K S; Kang, I; Sohn, W-M; Nawa, Y; Kong, Y

    2018-04-09

    To determine the potential for immunodiagnostic application of two recombinant forms of Clonorchis sinensis omega-class glutathione transferases (rCsGSTo1 and rCsGSTo2) against human small liver-fluke C. sinensis and Opisthorchis viverrini infections. Specific antibody levels against rCsGSTo1 and rCsGSTo2 in patients' sera of egg-positive opisthorchiasis (n = 87) and clonorchiasis (n = 120), as well as those in sera from patients with other helminthic infections (n = 252) and healthy controls (n = 40) were retrospectively analysed by ELISA. We observed highly positive correlation coefficients between specific antibody levels against rCsGSTo1 and rCsGSTo2 and egg counts per gramme of faeces (EPG) of patients with opisthorchiasis (n = 87; r = 0.88 for rCsGSTo1 and r = 0.90 for rCsGSTo2). Sera from opisthorchiasis patients whose EPG counts >100 (n = 43) revealed high antibody titres against both antigens. Patients' sera with low EPG counts (<100, n = 44) also exhibited reliable sensitivities of 93.2% and 97.7% for rCsGSTo1 and rCsGSTo2, respectively. Sera from clonorchiasis patients showed sensitivities of 90% (108/120 samples) and 89.2% (107/120 sera) for rCsGSTo1 and rCsGSTo2. Overall diagnostic sensitivities for liver-fluke infections were 92.3% for rCsGSTo1 (191/207 samples) and 93.2% for rCsGSTo2 (193/207 samples). Specificities were 89.7% (rCsGSTo1) and 97.6% (rCsGSTo2). Detection of specific antibody levels against rCsGSTo1 or rCsGSTo2 might be promising for the serodiagnosis of patients infected with these two phylogenetically close carcinogenic liver-flukes. Copyright © 2018 European Society of Clinical Microbiology and Infectious Diseases. Published by Elsevier Ltd. All rights reserved.

  7. SERUM GAMMA-GLUTAMYL TRANSFERASE AS A BIOMARKER OF TYPE-2 DM AMONG CIGARETTE SMOKERS

    Directory of Open Access Journals (Sweden)

    K. Suganthy

    2017-03-01

    Full Text Available BACKGROUND Smoking is one of the most common addictions of modern times and needs to be studied in a community as a public health issue. Also, smoking is a modifiable risk factor for type-2 DM. The smoking-related diseases share common pathophysiologies of imbalance of systemic oxidants and antioxidant status, increased inflammatory reactions, insulin resistance and dyslipidaemia. Biochemical assay of serum Gamma-Glutamyl Transferase (GGT activity is a low cost and highly sensitive laboratory test. Studies have indicated GGT is moderately elevated before the onset of other traditional risk factors for type-2 DM. So, among hepatic markers, the baseline GGT analysis can be an early risk marker of type 2 diabetes in cigarette smokers has to be studied. MATERIALS AND METHODS This is a case-control study on male cigarette smokers. 57 smokers were studied clinically and biochemically for plasma insulin, glucose and liver enzymes including GGT using standard biochemical methods and compared with 42 age and sex matched non-smokers as controls. RESULTS The mean serum GGT in smokers (25.45 ± 10.8 was increased compared to non-smokers (18.8 ± 5.8. Smokers GGT (r=0.396 and HOMA-IR (r=0.352 showed significant positive association with duration of smoking (p24 IU/L. Regression analysis showed none of the diabetic risk factors were observed to be dependent on GGT including other liver enzymes. Regression analysis showed GGT is not an independent risk factor for DM. Although, the mean fasting blood glucose (91.4 ± 21.3, BMI (26.1 ± 9.3 and HOMA-IR (7.3 ± 2.3 was increased among cigarette smokers with GGT >24 IU/L. CONCLUSION The baseline GGT assay in cigarette smokers might be associated with the proinflammatory status or be a marker of oxidative stress of smoke toxins. Smokers with baseline GGT >24 IU/L develop insulin resistance should be investigated in future longitudinal studies for prediabetes to consider cigarette smoking as an important modifiable

  8. Effects of imidacloprid on detoxifying enzyme glutathione S-transferase on Folsomia candida (Collembola).

    Science.gov (United States)

    Sillapawattana, Panwad; Schäffer, Andreas

    2017-04-01

    Chemical analyses of the environment can document contamination by various xenobiotics, but it is also important to understand the effect of pollutants on living organisms. Thus, in the present work, we investigated the effect of the pesticide imidacloprid on the detoxifying enzyme glutathione S-transferase (GST) from Folsomia candida (Collembola), a standard test organism for estimating the effects of pesticides and environmental pollutants on non-target soil arthropods. Test animals were treated with different concentrations of imidacloprid for 48 h. Changes in steady-state levels of GST messenger RNA (mRNA) and GST enzyme activity were investigated. Extracted proteins were separated according to their sizes by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and the resolved protein bands were detected by silver staining. The size of the glutathione (GSH) pool in Collembola was also determined. A predicted protein sequence of putative GSTs was identified with animals from control group. A 3-fold up-regulation of GST steady-state mRNA levels was detected in the samples treated with 5.0 mg L -1 imidacloprid compared to the control, while a 2.5- and 2.0- fold up-regulation was found in organisms treated with 2.5 and 7.5 mg L -1 imidacloprid, respectively. GST activity increased with increasing imidacloprid amounts from an initial activity of 0.11 μmol min -1  mg -1 protein in the control group up to 0.25 μmol min -1  mg -1 protein in the sample treated with the 5.0 mg L -1 of pesticide. By contrast, the total amount of GSH decreased with increasing imidacloprid concentration. The results suggest that the alteration of GST activity, steady-state level of GST mRNA, and GSH level may be involved in the response of F. candida to the exposure of imidacloprid and can be used as biomarkers to monitor the toxic effects of imidacloprid and other environmental pollutants on Collembola.

  9. Cobalamins uncovered by modern electronic structure calculations

    DEFF Research Database (Denmark)

    Kepp, Kasper Planeta; Ryde, Ulf

    2009-01-01

    electronic-structure calculations, in particular density functional methods, the understanding of the molecular mechanism of cobalamins has changed dramatically, going from a dominating view of trans-steric strain effects to a much more complex view involving an arsenal of catalytic strategies. Among...... these are cis-steric distortions, electrostatic stabilization of radical products, the realization that nucleotide units can serve as polar handles, and the careful design of the active sites, with polar residues in the radical enzymes and non-polar residues in the transferases. Together, these strategies...

  10. O-linked-N-acetylglucosamine modification of mammalian Notch receptors by an atypical O-GlcNAc transferase Eogt1

    Energy Technology Data Exchange (ETDEWEB)

    Sakaidani, Yuta [Department of Biochemistry II, Nagoya University Graduate School of Medicine, 65 Tsurumai, Showa-ku, Nagoya 466-0065 (Japan); Ichiyanagi, Naoki [Department of Applied Molecular Biosciences, Nagoya University Graduate School of Bioagricultural Sciences, Furo-cho, Chikusa-ku, Nagoya 464-8601 (Japan); Saito, Chika; Nomura, Tomoko [Department of Biochemistry II, Nagoya University Graduate School of Medicine, 65 Tsurumai, Showa-ku, Nagoya 466-0065 (Japan); Ito, Makiko [Department of Applied Molecular Biosciences, Nagoya University Graduate School of Bioagricultural Sciences, Furo-cho, Chikusa-ku, Nagoya 464-8601 (Japan); Nishio, Yosuke [Department of Biochemistry II, Nagoya University Graduate School of Medicine, 65 Tsurumai, Showa-ku, Nagoya 466-0065 (Japan); Nadano, Daita; Matsuda, Tsukasa [Department of Applied Molecular Biosciences, Nagoya University Graduate School of Bioagricultural Sciences, Furo-cho, Chikusa-ku, Nagoya 464-8601 (Japan); Furukawa, Koichi [Department of Biochemistry II, Nagoya University Graduate School of Medicine, 65 Tsurumai, Showa-ku, Nagoya 466-0065 (Japan); Okajima, Tetsuya, E-mail: tokajima@med.nagoya-u.ac.jp [Department of Biochemistry II, Nagoya University Graduate School of Medicine, 65 Tsurumai, Showa-ku, Nagoya 466-0065 (Japan); Department of Applied Molecular Biosciences, Nagoya University Graduate School of Bioagricultural Sciences, Furo-cho, Chikusa-ku, Nagoya 464-8601 (Japan)

    2012-03-02

    Highlights: Black-Right-Pointing-Pointer We characterized A130022J15Rik (Eogt1)-a mouse gene homologous to Drosophila Eogt. Black-Right-Pointing-Pointer Eogt1 encodes EGF domain O-GlcNAc transferase. Black-Right-Pointing-Pointer Expression of Eogt1 in Drosophila rescued the cell-adhesion defect in the Eogt mutant. Black-Right-Pointing-Pointer O-GlcNAcylation reaction in the secretory pathway is conserved through evolution. -- Abstract: O-linked-{beta}-N-acetylglucosamine (O-GlcNAc) modification is a unique cytoplasmic and nuclear protein modification that is common in nearly all eukaryotes, including filamentous fungi, plants, and animals. We had recently reported that epidermal growth factor (EGF) repeats of Notch and Dumpy are O-GlcNAcylated by an atypical O-GlcNAc transferase, EOGT, in Drosophila. However, no study has yet shown whether O-GlcNAcylation of extracellular proteins is limited to insects such as Drosophila or whether it occurs in other organisms, including mammals. Here, we report the characterization of A130022J15Rik, a mouse gene homolog of Drosophila Eogt (Eogt 1). Enzymatic analysis revealed that Eogt1 has a substrate specificity similar to that of Drosophila EOGT, wherein the Thr residue located between the fifth and sixth conserved cysteines of the folded EGF-like domains is modified. This observation is supported by the fact that the expression of Eogt1 in Drosophila rescued the cell-adhesion defect caused by Eogt downregulation. In HEK293T cells, Eogt1 expression promoted modification of Notch1 EGF repeats by O-GlcNAc, which was further modified, at least in part, by galactose to generate a novel O-linked-N-acetyllactosamine structure. These results suggest that Eogt1 encodes EGF domain O-GlcNAc transferase and that O-GlcNAcylation reaction in the secretory pathway is a fundamental biochemical process conserved through evolution.

  11. O-linked-N-acetylglucosamine modification of mammalian Notch receptors by an atypical O-GlcNAc transferase Eogt1

    International Nuclear Information System (INIS)

    Sakaidani, Yuta; Ichiyanagi, Naoki; Saito, Chika; Nomura, Tomoko; Ito, Makiko; Nishio, Yosuke; Nadano, Daita; Matsuda, Tsukasa; Furukawa, Koichi; Okajima, Tetsuya

    2012-01-01

    Highlights: ► We characterized A130022J15Rik (Eogt1)—a mouse gene homologous to Drosophila Eogt. ► Eogt1 encodes EGF domain O-GlcNAc transferase. ► Expression of Eogt1 in Drosophila rescued the cell-adhesion defect in the Eogt mutant. ► O-GlcNAcylation reaction in the secretory pathway is conserved through evolution. -- Abstract: O-linked-β-N-acetylglucosamine (O-GlcNAc) modification is a unique cytoplasmic and nuclear protein modification that is common in nearly all eukaryotes, including filamentous fungi, plants, and animals. We had recently reported that epidermal growth factor (EGF) repeats of Notch and Dumpy are O-GlcNAcylated by an atypical O-GlcNAc transferase, EOGT, in Drosophila. However, no study has yet shown whether O-GlcNAcylation of extracellular proteins is limited to insects such as Drosophila or whether it occurs in other organisms, including mammals. Here, we report the characterization of A130022J15Rik, a mouse gene homolog of Drosophila Eogt (Eogt 1). Enzymatic analysis revealed that Eogt1 has a substrate specificity similar to that of Drosophila EOGT, wherein the Thr residue located between the fifth and sixth conserved cysteines of the folded EGF-like domains is modified. This observation is supported by the fact that the expression of Eogt1 in Drosophila rescued the cell-adhesion defect caused by Eogt downregulation. In HEK293T cells, Eogt1 expression promoted modification of Notch1 EGF repeats by O-GlcNAc, which was further modified, at least in part, by galactose to generate a novel O-linked-N-acetyllactosamine structure. These results suggest that Eogt1 encodes EGF domain O-GlcNAc transferase and that O-GlcNAcylation reaction in the secretory pathway is a fundamental biochemical process conserved through evolution.

  12. Combined glutathione S transferase M1/T1 null genotypes is associated with type 2 diabetes mellitus

    Science.gov (United States)

    POROJAN, MIHAI D.; BALA, CORNELIA; ILIES, ROXANA; CATANA, ANDREEA; POPP, RADU A.; DUMITRASCU, DAN L.

    2015-01-01

    Background Due to new genetic insights, a considerably large number of genes and polymorphic gene variants are screened and linked with the complex pathogenesis of type 2 diabetes (DM). Our study aimed to investigate the association between the two isoforms of the glutathione S-transferase genes (Glutathione S transferase isoemzyme type M1- GSTM1 and Glutathione S transferase isoemzyme type T1-GSTT1) and the prevalence of DM in the Northern Romanian population. Methods We conducted a cross-sectional, randomized, case-control study evaluating the frequency of GSTM1 and GSTT1 null alleles in patients diagnosed with DM. A total of 106 patients diagnosed with DM and 124 healthy controls were included in the study. GSTM1 and GSTT1 null alleles genotyping was carried out using Multiplex PCR amplification of relevant gene fragments, followed by gel electrophoresis analysis of the resulting amplicons. Results Molecular analysis did not reveal an increased frequency of the null GSTM1 and GSTT1 alleles (mutant genotypes) respectively in the DM group compared to controls (p=0.171, OR=1.444 CI=0.852–2.447; p=0.647, OR=0.854, CI=0.436–1.673). Nevertheless, the combined GSTM1/GSTT1 null genotypes were statistically significantly higher in DM patients compared to control subjects (p=0.0021, OR=0.313, CI=0.149–0.655) Conclusions The main finding of our study is that the combined, double GSTM1/GSTT1 null genotypes are to be considered among the polymorphic genetic risk factors for type 2 DM. PMID:26528065

  13. Alterations at the peptidyl transferase centre of the ribosome induced by the synergistic action of the streptogramins dalfopristin and quinupristin

    Directory of Open Access Journals (Sweden)

    Fucini Paola

    2004-04-01

    Full Text Available Abstract Background The bacterial ribosome is a primary target of several classes of antibiotics. Investigation of the structure of the ribosomal subunits in complex with different antibiotics can reveal the mode of inhibition of ribosomal protein synthesis. Analysis of the interactions between antibiotics and the ribosome permits investigation of the specific effect of modifications leading to antimicrobial resistances. Streptogramins are unique among the ribosome-targeting antibiotics because they consist of two components, streptogramins A and B, which act synergistically. Each compound alone exhibits a weak bacteriostatic activity, whereas the combination can act bactericidal. The streptogramins A display a prolonged activity that even persists after removal of the drug. However, the mode of activity of the streptogramins has not yet been fully elucidated, despite a plethora of biochemical and structural data. Results The investigation of the crystal structure of the 50S ribosomal subunit from Deinococcus radiodurans in complex with the clinically relevant streptogramins quinupristin and dalfopristin reveals their unique inhibitory mechanism. Quinupristin, a streptogramin B compound, binds in the ribosomal exit tunnel in a similar manner and position as the macrolides, suggesting a similar inhibitory mechanism, namely blockage of the ribosomal tunnel. Dalfopristin, the corresponding streptogramin A compound, binds close to quinupristin directly within the peptidyl transferase centre affecting both A- and P-site occupation by tRNA molecules. Conclusions The crystal structure indicates that the synergistic effect derives from direct interaction between both compounds and shared contacts with a single nucleotide, A2062. Upon binding of the streptogramins, the peptidyl transferase centre undergoes a significant conformational transition, which leads to a stable, non-productive orientation of the universally conserved U2585. Mutations of this r

  14. Expression of an enzymatically active Yb3 glutathione S-transferase in Escherichia coli and identification of its natural form in rat brain.

    Science.gov (United States)

    Abramovitz, M; Ishigaki, S; Felix, A M; Listowsky, I

    1988-11-25

    Glutathione S-transferases containing Yb3 subunits are relatively uncommon forms that are expressed in a tissue-specific manner and have not been identified unequivocally or characterized. A cDNA clone containing the entire coding sequence of Yb3 glutathione S-transferase mRNA was incorporated into a pIN-III expression vector used to transform Escherichia coli. A fusion Yb3-protein containing 14 additional amino acid residues at its N terminus was purified to homogeneity. Recombinant Yb3 was enzymatically active with both 1-chloro-2,4-dinitrobenzene and 1,2-dichloro-4-nitrobenzene as substrates but lacked glutathione peroxidase activity. Substrate specificity patterns of recombinant Yb3 were more limited than those of glutathione S-transferase isoenzymes containing Yb1- or Yb2-type subunits. Peptides corresponding to unique amino acid sequences of Yb3 as well as a peptide from a region of homology with Yb1 and Yb2 subunits were synthesized. These synthetic peptides were used to raise antibodies specific to Yb3 and others that cross-reacted with all Yb forms. Immunoblotting was utilized to identify the natural counterpart of recombinant Yb3 among rat glutathione transferases. Brain and testis glutathione S-transferases were rich in Yb3 subunits, but very little was found in liver or kidney. Physical properties, substrate specificities, and binding patterns of the recombinant protein paralleled properties of the natural isoenzyme isolated from brain.

  15. Does occupational exposure to solvents and pesticides in association with glutathione S-transferase A1, M1, P1, and T1 polymorphisms increase the risk of bladder cancer? The Belgrade case-control study.

    Directory of Open Access Journals (Sweden)

    Marija G Matic

    Full Text Available OBJECTIVE: We investigated the role of the glutathione S-transferase A1, M1, P1 and T1 gene polymorphisms and potential effect modification by occupational exposure to different chemicals in Serbian bladder cancer male patients. PATIENTS AND METHODS: A hospital-based case-control study of bladder cancer in men comprised 143 histologically confirmed cases and 114 age-matched male controls. Deletion polymorphism of glutathione S-transferase M1 and T1 was identified by polymerase chain reaction method. Single nucleotide polymorphism of glutathione S-transferase A1 and P1 was identified by restriction fragment length polymorphism method. As a measure of effect size, odds ratio (OR with corresponding 95% confidence interval (95%CI was calculated. RESULTS: The glutathione S-transferase A1, T1 and P1 genotypes did not contribute independently toward the risk of bladder cancer, while the glutathione S-transferase M1-null genotype was overrepresented among cases (OR = 2.1, 95% CI = 1.1-4.2, p = 0.032. The most pronounced effect regarding occupational exposure to solvents and glutathione S-transferase genotype on bladder cancer risk was observed for the low activity glutathione S-transferase A1 genotype (OR = 9.2, 95% CI = 2.4-34.7, p = 0.001. The glutathione S-transferase M1-null genotype also enhanced the risk of bladder cancer among subjects exposed to solvents (OR = 6,5, 95% CI = 2.1-19.7, p = 0.001. The risk of bladder cancer development was 5.3-fold elevated among glutathione S-transferase T1-active patients exposed to solvents in comparison with glutathione S-transferase T1-active unexposed patients (95% CI = 1.9-15.1, p = 0.002. Moreover, men with glutathione S-transferase T1-active genotype exposed to pesticides exhibited 4.5 times higher risk in comparison with unexposed glutathione S-transferase T1-active subjects (95% CI = 0.9-22.5, p = 0.067. CONCLUSION: Null or low-activity genotypes of the

  16. Crystallization and preliminary X-ray diffraction analysis of a glutathione S-transferase from Xylella fastidiosa

    International Nuclear Information System (INIS)

    Garcia, Wanius; Travensolo, Regiane F.; Rodrigues, Nathalia C.; Muniz, João R. C.; Caruso, Célia S.; Lemos, Eliana G. M.; Araujo, Ana Paula U.; Carrilho, Emanuel

    2008-01-01

    Glutathione S-transferase from X. fastidiosa (xfGST) has been overexpressed in E. coli, purified and crystallized. Diffraction data were collected to 2.23 Å. Glutathione S-transferases (GSTs) form a group of multifunctional isoenzymes that catalyze the glutathione-dependent conjugation and reduction reactions involved in the cellular detoxification of xenobiotic and endobiotic compounds. GST from Xylella fastidiosa (xfGST) was overexpressed in Escherichia coli and purified by conventional affinity chromatography. In this study, the crystallization and preliminary X-ray analysis of xfGST is described. The purified protein was crystallized by the vapour-diffusion method, producing crystals that belonged to the triclinic space group P1. The unit-cell parameters were a = 47.73, b = 87.73, c = 90.74 Å, α = 63.45, β = 80.66, γ = 94.55°. xfGST crystals diffracted to 2.23 Å resolution on a rotating-anode X-ray source

  17. Glutathione transferase (GST) as a candidate molecular-based biomarker for soil toxin exposure in the earthworm Lumbricus rubellus

    International Nuclear Information System (INIS)

    LaCourse, E. James; Hernandez-Viadel, Mariluz; Jefferies, James R.; Svendsen, Claus; Spurgeon, David J.; Barrett, John; John Morgan, A.; Kille, Peter; Brophy, Peter M.

    2009-01-01

    The earthworm Lumbricus rubellus (Hoffmeister, 1843) is a terrestrial pollution sentinel. Enzyme activity and transcription of phase II detoxification superfamily glutathione transferases (GST) is known to respond in earthworms after soil toxin exposure, suggesting GST as a candidate molecular-based pollution biomarker. This study combined sub-proteomics, bioinformatics and biochemical assay to characterise the L. rubellus GST complement as pre-requisite to initialise assessment of the applicability of GST as a biomarker. L. rubellus possesses a range of GSTs related to known classes, with evidence of tissue-specific synthesis. Two affinity-purified GSTs dominating GST protein synthesis (Sigma and Pi class) were cloned, expressed and characterised for enzyme activity with various substrates. Electrospray ionisation mass spectrometry (ESI-MS) and tandem mass spectrometry (MS/MS) following SDS-PAGE were superior in retaining subunit stability relative to two-dimensional gel electrophoresis (2-DE). This study provides greater understanding of Phase II detoxification GST superfamily status of an important environmental pollution sentinel organism. - This study currently provides the most comprehensive view of the Phase II detoxification enzyme superfamily of glutathione transferases within the important environmental pollution sentinel earthworm Lumbricus rubellus.

  18. Glutathione transferase (GST) as a candidate molecular-based biomarker for soil toxin exposure in the earthworm Lumbricus rubellus

    Energy Technology Data Exchange (ETDEWEB)

    LaCourse, E. James, E-mail: james.la-course@liverpool.ac.u [Institute of Biological, Environmental, and Rural Sciences, Aberystwyth University, Aberystwyth SY23 3DA (United Kingdom); Hernandez-Viadel, Mariluz; Jefferies, James R. [Institute of Biological, Environmental, and Rural Sciences, Aberystwyth University, Aberystwyth SY23 3DA (United Kingdom); Svendsen, Claus; Spurgeon, David J. [Centre for Ecology and Hydrology, Huntingdon PE28 2LS (United Kingdom); Barrett, John [Institute of Biological, Environmental, and Rural Sciences, Aberystwyth University, Aberystwyth SY23 3DA (United Kingdom); John Morgan, A.; Kille, Peter [Biosciences, University of Cardiff, Cardiff CF10 3TL (United Kingdom); Brophy, Peter M. [Institute of Biological, Environmental, and Rural Sciences, Aberystwyth University, Aberystwyth SY23 3DA (United Kingdom)

    2009-08-15

    The earthworm Lumbricus rubellus (Hoffmeister, 1843) is a terrestrial pollution sentinel. Enzyme activity and transcription of phase II detoxification superfamily glutathione transferases (GST) is known to respond in earthworms after soil toxin exposure, suggesting GST as a candidate molecular-based pollution biomarker. This study combined sub-proteomics, bioinformatics and biochemical assay to characterise the L. rubellus GST complement as pre-requisite to initialise assessment of the applicability of GST as a biomarker. L. rubellus possesses a range of GSTs related to known classes, with evidence of tissue-specific synthesis. Two affinity-purified GSTs dominating GST protein synthesis (Sigma and Pi class) were cloned, expressed and characterised for enzyme activity with various substrates. Electrospray ionisation mass spectrometry (ESI-MS) and tandem mass spectrometry (MS/MS) following SDS-PAGE were superior in retaining subunit stability relative to two-dimensional gel electrophoresis (2-DE). This study provides greater understanding of Phase II detoxification GST superfamily status of an important environmental pollution sentinel organism. - This study currently provides the most comprehensive view of the Phase II detoxification enzyme superfamily of glutathione transferases within the important environmental pollution sentinel earthworm Lumbricus rubellus.

  19. The role of human demographic history in determining the distribution and frequency of transferase-deficient galactosaemia mutations.

    LENUS (Irish Health Repository)

    Flanagan, J M

    2010-02-01

    Classical or transferase-deficient galactosaemia is an inherited metabolic disorder caused by mutation in the human Galactose-1-phosphate uridyl transferase (GALT) gene. Of some 170 causative mutations reported, fewer than 10% are observed in more than one geographic region or ethnic group. To better understand the population history of the common GALT mutations, we have established a haplotyping system for the GALT locus incorporating eight single nucleotide polymorphisms and three short tandem repeat markers. We analysed haplotypes associated with the three most frequent GALT gene mutations, Q188R, K285N and Duarte-2 (D2), and estimated their age. Haplotype diversity, in conjunction with measures of genetic diversity and of linkage disequilibrium, indicated that Q188R and K285N are European mutations. The Q188R mutation arose in central Europe within the last 20 000 years, with its observed east-west cline of increasing relative allele frequency possibly being due to population expansion during the re-colonization of Europe by Homo sapiens in the Mesolithic age. K285N was found to be a younger mutation that originated in Eastern Europe and is probably more geographically restricted as it arose after all major European population expansions. The D2 variant was found to be an ancient mutation that originated before the expansion of Homo sapiens out of Africa.

  20. Acute cadmium intoxication induces alpha-class glutathione S-transferase protein synthesis and enzyme activity in rat liver

    International Nuclear Information System (INIS)

    Casalino, Elisabetta; Sblano, Cesare; Calzaretti, Giovanna; Landriscina, Clemente

    2006-01-01

    Acute cadmium intoxication affects glutathione S-transferase (GST) in rat liver. It has been found that 24 h after i.p. cadmium administration to rats, at a dose of 2.5 mg CdCl 2 kg -1 body weight, the activity of this enzyme in liver cytosol increased by 40%. A less stimulatory effect persisted till 48 h and thereafter the enzyme activity normalized. Since, GST isoenzymes belong to different classes in mammalian tissues, we used quantitative immunoassays to verify which family of GST isoenzymes is influenced by this intoxication. Only alpha-class glutathione S-transferase (α-GST) proteins were detected in rat liver cytosol and their level increased by about 25%, 24 h after cadmium treatment. No pi-GST isoforms were found in liver cytosol from either normal or cadmium-treated rats. Co-administration of actinomycin D with cadmium normalized both the protein level and the activity of α-GST, suggesting that some effect occurs on enzyme transcription of these isoenzymes by this metal. On the other hand, it seems unlikely that the stimulatory effect is due to the high level of peroxides caused by lipid peroxidation, since Vitamin E administration strongly reduced the TBARS level, but did not cause any GST activity decrease

  1. Crystallization and preliminary X-ray diffraction analysis of a glutathione S-transferase from Xylella fastidiosa

    Energy Technology Data Exchange (ETDEWEB)

    Garcia, Wanius, E-mail: wanius@if.sc.usp.br [Laboratório de Biofísica Molecular ‘Sérgio Mascarenhas’, Instituto de Física de São Carlos, Universidade de São Paulo (USP), São Carlos (Brazil); Travensolo, Regiane F. [Grupo de Bioanalítica, Microfabricação e Separações, Instituto de Química de São Carlos, Universidade de São Paulo (USP), São Carlos (Brazil); Rodrigues, Nathalia C.; Muniz, João R. C. [Laboratório de Biofísica Molecular ‘Sérgio Mascarenhas’, Instituto de Física de São Carlos, Universidade de São Paulo (USP), São Carlos (Brazil); Caruso, Célia S. [Grupo de Bioanalítica, Microfabricação e Separações, Instituto de Química de São Carlos, Universidade de São Paulo (USP), São Carlos (Brazil); Lemos, Eliana G. M. [Laboratório de Bioquímica de Microrganismos e de Plantas, Departamento de Tecnologia, UNESP, Jaboticabal (Brazil); Araujo, Ana Paula U. [Laboratório de Biofísica Molecular ‘Sérgio Mascarenhas’, Instituto de Física de São Carlos, Universidade de São Paulo (USP), São Carlos (Brazil); Carrilho, Emanuel, E-mail: wanius@if.sc.usp.br [Grupo de Bioanalítica, Microfabricação e Separações, Instituto de Química de São Carlos, Universidade de São Paulo (USP), São Carlos (Brazil); Laboratório de Biofísica Molecular ‘Sérgio Mascarenhas’, Instituto de Física de São Carlos, Universidade de São Paulo (USP), São Carlos (Brazil)

    2008-02-01

    Glutathione S-transferase from X. fastidiosa (xfGST) has been overexpressed in E. coli, purified and crystallized. Diffraction data were collected to 2.23 Å. Glutathione S-transferases (GSTs) form a group of multifunctional isoenzymes that catalyze the glutathione-dependent conjugation and reduction reactions involved in the cellular detoxification of xenobiotic and endobiotic compounds. GST from Xylella fastidiosa (xfGST) was overexpressed in Escherichia coli and purified by conventional affinity chromatography. In this study, the crystallization and preliminary X-ray analysis of xfGST is described. The purified protein was crystallized by the vapour-diffusion method, producing crystals that belonged to the triclinic space group P1. The unit-cell parameters were a = 47.73, b = 87.73, c = 90.74 Å, α = 63.45, β = 80.66, γ = 94.55°. xfGST crystals diffracted to 2.23 Å resolution on a rotating-anode X-ray source.

  2. Activity-Based Probes for Isoenzyme- and Site-Specific Functional Characterization of Glutathione S -Transferases

    Energy Technology Data Exchange (ETDEWEB)

    Stoddard, Ethan G. [Chemical Biology and Exposure; Killinger, Bryan J. [Chemical Biology and Exposure; Nair, Reji N. [Chemical Biology and Exposure; Sadler, Natalie C. [Chemical Biology and Exposure; Volk, Regan F. [Chemical Biology and Exposure; Purvine, Samuel O. [Chemical Biology and Exposure; Shukla, Anil K. [Chemical Biology and Exposure; Smith, Jordan N. [Chemical Biology and Exposure; Wright, Aaron T. [Chemical Biology and Exposure

    2017-11-01

    Glutathione S-transferases (GSTs) comprise a highly diverse family of phase II drug metabolizing enzymes whose shared function is the conjugation of reduced glutathione to various endo- and xenobiotics. Although the conglomerate activity of these enzymes can be measured by colorimetric assays, measurement of the individual contribution from specific isoforms and their contribution to the detoxification of xenobiotics in complex biological samples has not been possible. For this reason, we have developed two activity-based probes that characterize active glutathione transferases in mammalian tissues. The GST active site is comprised of a glutathione binding “G site” and a distinct substrate binding “H site”. Therefore, we developed (1) a glutathione-based photoaffinity probe (GSH-ABP) to target the “G site”, and (2) a probe designed to mimic a substrate molecule and show “H site” activity (GST-ABP). The GSH-ABP features a photoreactive moiety for UV-induced covalent binding to GSTs and glutathione-binding enzymes. The GST-ABP is a derivative of a known mechanism-based GST inhibitor that binds within the active site and inhibits GST activity. Validation of probe targets and “G” and “H” site specificity was carried out using a series of competitors in liver homogenates. Herein, we present robust tools for the novel characterization of enzyme- and active site-specific GST activity in mammalian model systems.

  3. Cantharidin Impedes Activity of Glutathione S-Transferase in the Midgut of Helicoverpa armigera Hübner

    Directory of Open Access Journals (Sweden)

    Ya Lin Zhang

    2013-03-01

    Full Text Available Previous investigations have implicated glutathione S-transferases (GSTs as one of the major reasons for insecticide resistance. Therefore, effectiveness of new candidate compounds depends on their ability to inhibit GSTs to prevent metabolic detoxification by insects. Cantharidin, a terpenoid compound of insect origin, has been developed as a bio-pesticide in China, and proves highly toxic to a wide range of insects, especially lepidopteran. In the present study, we test cantharidin as a model compound for its toxicity, effects on the mRNA transcription of a model Helicoverpa armigera glutathione S-transferase gene (HaGST and also for its putative inhibitory effect on the catalytic activity of GSTs, both in vivo and in vitro in Helicoverpa armigera, employing molecular and biochemical methods. Bioassay results showed that cantharidin was highly toxic to H. armigera. Real-time qPCR showed down-regulation of the HaGST at the mRNA transcript ranging from 2.5 to 12.5 folds while biochemical assays showed in vivo inhibition of GSTs in midgut and in vitro inhibition of rHaGST. Binding of cantharidin to HaGST was rationalized by homology and molecular docking simulations using a model GST (1PN9 as a template structure. Molecular docking simulations also confirmed accurate docking of the cantharidin molecule to the active site of HaGST impeding its catalytic activity.

  4. Glutathione S transferase polymorphisms influence on iron overload in β-thalassemia patients

    Directory of Open Access Journals (Sweden)

    Serena Sclafani

    2013-11-01

    Full Text Available In patients with β-thalassemia iron overload that leads to damage to vital organs is observed. Glutathione S transferase (GST enzymes have an antioxidant role in detoxification processes of toxic substances. This role is determined genetically. In this study, we correlated GSTT1 and GSTM1 genotypes with iron overload measured with direct and indirect non-invasive methods; in particular, we used serum ferritin and signal intensity of the magnetic resonance image (MRI in 42 patients with β-thalassemia, which were regularly subjected to chelation and transfusion therapy. Multiplex polymerase chain reaction was used to determine the genotype. The loss of both alleles leads to a decreased value of liver and heart MRI-signal intensity with a consequent iron accumulation in these organs; the loss of only one allele doesn’t lead to relevant overload. Serum ferritin doesn’t appear to be correlated to iron overload instead. 对于β-地中海贫血患者,由于铁过量而造成重要器官受损的情况也在观察之中。谷胱甘肽S转移酶(GST 酶类在对有毒物质进行解毒的过程中有着抗氧化剂的作用。该作用是由基因决定的。 在这份研究中,我们运用了直接和间接非侵入性的方法对基因型铁过量GSTT1 和GSTM1进行了相关性测量;特别地,我们对42位定期接受螯合和输血治疗的β-地中海贫血患者进行了血清铁蛋白和磁共振强度图像(MRI 的测试。 多重聚合酶链反应的测试也被运用来确定该基因型。 该两种等位基因的缺失,导致了肝功能减损及心脏磁共振强度的下降,并造成了在这些器官中铁含量的积累;其中一种等位基因的缺失并不会导致过度的铁含量。血清蛋白和铁过量之间,看起来并不存在相关性。

  5. The lectin domains of polypeptide GalNAc-transferases exhibit carbohydrate-binding specificity for GalNAc: lectin binding to GalNAc-glycopeptide substrates is required for high density GalNAc-O-glycosylation

    DEFF Research Database (Denmark)

    Wandall, Hans H; Irazoqui, Fernando; Tarp, Mads Agervig

    2007-01-01

    Initiation of mucin-type O-glycosylation is controlled by a large family of UDP GalNAc:polypeptide N-acetylgalactosaminyltransferases (GalNAc-transferases). Most GalNAc-transferases contain a ricin-like lectin domain in the C-terminal end, which may confer GalNAc-glycopeptide substrate specificit...

  6. Intracellular APP Domain Regulates Serine-Palmitoyl-CoA Transferase Expression and Is Affected in Alzheimer's Disease

    Science.gov (United States)

    Grimm, Marcus O. W.; Grösgen, Sven; Rothhaar, Tatjana L.; Burg, Verena K.; Hundsdörfer, Benjamin; Haupenthal, Viola J.; Friess, Petra; Müller, Ulrike; Fassbender, Klaus; Riemenschneider, Matthias; Grimm, Heike S.; Hartmann, Tobias

    2011-01-01

    Lipids play an important role as risk or protective factors in Alzheimer's disease (AD), a disease biochemically characterized by the accumulation of amyloid beta peptides (Aβ), released by proteolytic processing of the amyloid precursor protein (APP). Changes in sphingolipid metabolism have been associated to the development of AD. The key enzyme in sphingolipid de novo synthesis is serine-palmitoyl-CoA transferase (SPT). In the present study we identified a new physiological function of APP in sphingolipid synthesis. The APP intracellular domain (AICD) was found to decrease the expression of the SPT subunit SPTLC2, the catalytic subunit of the SPT heterodimer, resulting in that decreased SPT activity. AICD function was dependent on Fe65 and SPTLC2 levels are increased in APP knock-in mice missing a functional AICD domain. SPTLC2 levels are also increased in familial and sporadic AD postmortem brains, suggesting that SPT is involved in AD pathology. PMID:21660213

  7. Yeast one-hybrid system used to identify the binding proteins for rat glutathione S-transferase P enhancer I.

    Science.gov (United States)

    Liao, Ming-Xiang; Liu, Dong-Yuan; Zuo, Jin; Fang, Fu-De

    2002-03-01

    To detect the trans-factors specifically binding to the strong enhancer element (GPEI) in the upstream of rat glutathione S-transferase P (GST-P) gene. Yeast one-hybrid system was used to screen rat lung MATCHMAKER cDNA library to identify potential trans-factors that can interact with core sequence of GPEI(cGPEI). Electrophoresis mobility shift assay (EMSA) was used to analyze the binding of transfactors to cGPEI. cDNA fragments coding for the C-terminal part of the transcription factor c-Jun and rat adenine nucleotide translocator (ANT) were isolated. The binding of c-Jun and ANT to GPEI core sequence were confirmed. Rat c-jun transcriptional factor and ANT may interact with cGPEI. They could play an important role in the induced expression of GST-P gene.

  8. Glutathione S-transferase of brown planthoppers (Nilaparvata lugens) is essential for their adaptation to gramine-containing host plants.

    Science.gov (United States)

    Sun, Xiao-Qin; Zhang, Mao-Xin; Yu, Jing-Ya; Jin, Yu; Ling, Bing; Du, Jin-Ping; Li, Gui-Hua; Qin, Qing-Ming; Cai, Qing-Nian

    2013-01-01

    Plants have evolved complex processes to ward off attacks by insects. In parallel, insects have evolved mechanisms to thwart these plant defenses. To gain insight into mechanisms that mediate this arms race between plants and herbivorous insects, we investigated the interactions between gramine, a toxin synthesized by plants of the family Gramineae, and glutathione S transferase (GST), an enzyme found in insects that is known to detoxify xenobiotics. Here, we demonstrate that rice (Oryza sativa), a hydrophytic plant, also produces gramine and that rice resistance to brown planthoppers (Nilaparvata lugens, BPHs) is highly associated with in planta gramine content. We also show that gramine is a toxicant that causes BPH mortality in vivo and that knockdown of BPH GST gene nlgst1-1 results in increased sensitivity to diets containing gramine. These results suggest that the knockdown of key detoxification genes in sap-sucking insects may provide an avenue for increasing their sensitivity to natural plant-associated defense mechanisms.

  9. Dynamic interplay between catalytic and lectin domains of GalNAc-transferases modulates protein O-glycosylation

    DEFF Research Database (Denmark)

    Lira-Navarrete, Erandi; de Las Rivas, Matilde; Compañón, Ismael

    2015-01-01

    the first crystal structures of complexes of GalNAc-T2 with glycopeptides that together with enhanced sampling molecular dynamics simulations demonstrate a cooperative mechanism by which the lectin domain enables free acceptor sites binding of glycopeptides into the catalytic domain. Atomic force microscopy......Protein O-glycosylation is controlled by polypeptide GalNAc-transferases (GalNAc-Ts) that uniquely feature both a catalytic and lectin domain. The underlying molecular basis of how the lectin domains of GalNAc-Ts contribute to glycopeptide specificity and catalysis remains unclear. Here we present...... and small-angle X-ray scattering experiments further reveal a dynamic conformational landscape of GalNAc-T2 and a prominent role of compact structures that are both required for efficient catalysis. Our model indicates that the activity profile of GalNAc-T2 is dictated by conformational heterogeneity...

  10. Protective effect of lemongrass oil against dexamethasone induced hyperlipidemia in rats: possible role of decreased lecithin cholesterol acetyl transferase activity.

    Science.gov (United States)

    Kumar, V R Santhosh; Inamdar, Md Naseeruddin; Nayeemunnisa; Viswanatha, G L

    2011-08-01

    To evaluate the anti-hyperlipidemic activity of lemongrass oil against in dexamethasone induced hyperlipidemia in rats. Administration of dexamethasone was given at 10 mg/kg, sc. to the adult rats for 8 d induces hyperlipidemia characterized by marked increase in serum cholesterol and triglyceride levels along with increase in atherogenic index. Lemongrass oil (100 and 200 mg/kg, po.) treatment has showed significant inhibition against dexamethasone hyperlipidemia by maintaining the serum levels of cholesterol, triglycerides and atherogenic index near to the normal levels and the antihyperlipidemic effect of the lemongross oil was comparable with atorvastatin 10 mg/kg, po. The possible mechanism may be associated with decrease in lecithin cholesterol acetyl transferase (LCAT) activity. These results suggested that Lemon gross oil possess significant anti-hyperlipidemic activity. Copyright © 2011 Hainan Medical College. Published by Elsevier B.V. All rights reserved.

  11. A SERS protocol as a potential tool to access 6-mercaptopurine release accelerated by glutathione-S-transferase.

    Science.gov (United States)

    Wang, Ying; Sun, Jie; Yang, Qingran; Lu, Wenbo; Li, Yan; Dong, Jian; Qian, Weiping

    2015-11-21

    The developed method for monitoring GST, an important drug metabolic enzyme, could greatly facilitate researches on relative biological fields. In this work, we have developed a SERS technique to monitor the absorbance behaviour of 6-mercaptopurine (6-MP) and its glutathione-S-transferase (GST)-accelerated glutathione (GSH)-triggered release behaviour on the surface of gold nanoflowers (GNFs), using the GNFs as excellent SERS substrates. The SERS signal was used as an indicator of absorbance or release of 6-MP on the gold surface. We found that GST can accelerate GSH-triggered release behaviour of 6-MP from the gold surface. We speculated that GST catalyzes nucleophilic GSH to competitively bind with the electrophilic substance 6-MP. Experimental results have proved that the presented SERS protocol can be utilized as an effective tool for accessing the release of anticancer drugs.

  12. The identification and characterisation of a functional interaction between arginyl-tRNA-protein transferase and topoisomerase II

    International Nuclear Information System (INIS)

    Barker, Catherine R.; Mouchel, Nathalie A.P.; Jenkins, John R.

    2006-01-01

    Topoisomerase II is required for the viability of all eukaryotic cells. It plays important roles in DNA replication, recombination, chromosome segregation, and the maintenance of the nuclear scaffold. Proteins that interact with and regulate this essential enzyme are of great interest. To investigate the role of proteins interacting with the N-terminal domain of the Saccharomyces cerevisiae topoisomerase II, we used a yeast two-hybrid protein interaction screen. We identified an interaction between arginyl-tRNA-protein transferase (Ate1) and the N-terminal domain of the S. cerevisiae topoisomerase II, including the potential site of interaction. Ate1 is a component of the N-end rule protein degradation pathway which targets proteins for degradation. We also propose a previously unidentified role for Ate1 in modulating the level of topoisomerase II through the cell cycle

  13. Identification and suppression of the p-coumaroyl CoA:hydroxycinnamyl alcohol transferase in Zea mays L.

    Science.gov (United States)

    Marita, Jane M; Hatfield, Ronald D; Rancour, David M; Frost, Kenneth E

    2014-06-01

    Grasses, such as Zea mays L. (maize), contain relatively high levels of p-coumarates (pCA) within their cell walls. Incorporation of pCA into cell walls is believed to be due to a hydroxycinnamyl transferase that couples pCA to monolignols. To understand the role of pCA in maize development, the p-coumaroyl CoA:hydroxycinnamyl alcohol transferase (pCAT) was isolated and purified from maize stems. Purified pCAT was subjected to partial trypsin digestion, and peptides were sequenced by tandem mass spectrometry. TBLASTN analysis of the acquired peptide sequences identified a single full-length maize cDNA clone encoding all the peptide sequences obtained from the purified enzyme. The cDNA clone was obtained and used to generate an RNAi construct for suppressing pCAT expression in maize. Here we describe the effects of suppression of pCAT in maize. Primary screening of transgenic maize seedling leaves using a new rapid analytical platform was used to identify plants with decreased amounts of pCA. Using this screening method, mature leaves from fully developed plants were analyzed, confirming reduced pCA levels throughout plant development. Complete analysis of isolated cell walls from mature transgenic stems and leaves revealed that lignin levels did not change, but pCA levels decreased and the lignin composition was altered. Transgenic plants with the lowest levels of pCA had decreased levels of syringyl units in the lignin. Thus, altering the levels of pCAT expression in maize leads to altered lignin composition, but does not appear to alter the total amount of lignin present in the cell walls. © 2014 The Authors The Plant Journal © 2014 John Wiley & Sons Ltd.

  14. Synergistic and independent actions of multiple terminal nucleotidyl transferases in the 3' tailing of small RNAs in Arabidopsis.

    Directory of Open Access Journals (Sweden)

    Xiaoyan Wang

    2015-04-01

    Full Text Available All types of small RNAs in plants, piwi-interacting RNAs (piRNAs in animals and a subset of siRNAs in Drosophila and C. elegans are subject to HEN1 mediated 3' terminal 2'-O-methylation. This modification plays a pivotal role in protecting small RNAs from 3' uridylation, trimming and degradation. In Arabidopsis, HESO1 is a major enzyme that uridylates small RNAs to trigger their degradation. However, U-tail is still present in null hen1 heso1 mutants, suggesting the existence of (an enzymatic activities redundant with HESO1. Here, we report that UTP: RNA uridylyltransferase (URT1 is a functional paralog of HESO1. URT1 interacts with AGO1 and plays a predominant role in miRNA uridylation when HESO1 is absent. Uridylation of miRNA is globally abolished in a hen1 heso1 urt1 triple mutant, accompanied by an extensive increase of 3'-to-5' trimming. In contrast, disruption of URT1 appears not to affect the heterochromatic siRNA uridylation. This indicates the involvement of additional nucleotidyl transferases in the siRNA pathway. Analysis of miRNA tailings in the hen1 heso1 urt1 triple mutant also reveals the existence of previously unknown enzymatic activities that can add non-uridine nucleotides. Importantly, we show HESO1 may also act redundantly with URT1 in miRNA uridylation when HEN1 is fully competent. Taken together, our data not only reveal a synergistic action of HESO1 and URT1 in the 3' uridylation of miRNAs, but also independent activities of multiple terminal nucleotidyl transferases in the 3' tailing of small RNAs and an antagonistic relationship between uridylation and trimming. Our results may provide further insight into the mechanisms of small RNA 3' end modification and stability control.

  15. Phylogenetic characterization of Clonorchis sinensis proteins homologous to the sigma-class glutathione transferase and their differential expression profiles.

    Science.gov (United States)

    Bae, Young-An; Kim, Jeong-Geun; Kong, Yoon

    2016-01-01

    Glutathione transferase (GST) is one of the major antioxidant proteins with diverse supplemental activities including peroxidase, isomerase, and thiol transferase. GSTs are classified into multiple classes on the basis of their primary structures and substrate/inhibitor specificity. However, the evolutionary routes and physiological environments specific to each of the closely related bioactive enzymes remain elusive. The sigma-like GSTs exhibit amino acid conservation patterns similar to the prostaglandin D synthases (PGDSs). In this study, we analyzed the phylogenetic position of the GSTs of the biocarcinogenic liver fluke, Clonorchis sinensis. We also observed induction profile of the GSTs in association with the parasite's maturation and in response to exogenous oxidative stresses, with special attention to sigma-class GSTs and PGDSs. The C. sinensis genome encoded 12 GST protein species, which were separately assigned to cytosolic (two omega-, one zeta-, two mu-, and five sigma-class), mitochondrial (one kappa-class), and microsomal (one membrane-associated proteins in eicosanoid and glutathione metabolism-like protein) GST families. Multiple sigma GST (or PGDS) orthologs were also detected in Opisthorchis viverrini. Other trematode species possessed only a single sigma-like GST gene. A phylogenetic analysis demonstrated that one of the sigma GST lineages duplicated in the common ancestor of trematodes were specifically expanded in the opisthorchiids, but deleted in other trematodes. The induction profiles of these sigma GST genes along with the development and aging of C. sinensis, and against various exogenous chemical stimuli strongly suggest that the paralogous sigma GST genes might be undergone specialized evolution to cope with the diverse hostile biochemical environments within the mammalian hepatobiliary ductal system. Copyright © 2016 Elsevier B.V. All rights reserved.

  16. Maize w3 disrupts homogentisate solanesyl transferase (ZmHst) and reveals a plastoquinone-9 independent path for phytoene desaturation and tocopherol accumulation in kernels

    Science.gov (United States)

    Maize white seedling 3 (w3) has been used to study carotenoid deficiency for almost 100 years, although its genetic basis remained unknown. We show here that w3 phenotype is caused by disruption of homogentisate solanesyl transferase (HST), which catalyzes the first committed step in plastoquinone-9...

  17. Succinyl-CoA:acetoacetate transferase deficiency : identification of a new patient with a neonatal onset and review of the literature

    NARCIS (Netherlands)

    Niezen-Koning, K E; Wanders, R J; Ruiter, J P; Ijlst, L; Visser, G; Reitsma-Bierens, W C; Heijmans, Hugo; Reijngoud, D J; Smit, G P

    1997-01-01

    UNLABELLED: We describe the clinical symptoms and biochemical findings of a patient with succinyl-CoA:acetoacetate transferase deficiency who presented in the neonatal period and review the current literature on this subject. Our patient was initially suspected to have distal renal tubular acidosis,

  18. Effects of Deletion of the Streptococcus pneumoniae Lipoprotein Diacylglyceryl Transferase Gene lgt on ABC Transporter Function and on Growth In Vivo.

    NARCIS (Netherlands)

    Chimalapati, S.; Cohen, J.M.; Camberlein, E.; Macdonald, N.; Durmort, C.; Vernet, T.; Hermans, P.W.M.; Mitchell, T.; Brown, J.S.

    2012-01-01

    Lipoproteins are an important class of surface associated proteins that have diverse roles and frequently are involved in the virulence of bacterial pathogens. As prolipoproteins are attached to the cell membrane by a single enzyme, prolipoprotein diacylglyceryl transferase (Lgt), deletion of the

  19. Succinyl-CoA:acetoacetate transferase deficiency: identification of a new patient with a neonatal onset and review of the literature

    NARCIS (Netherlands)

    Niezen-Koning, K. E.; Wanders, R. J.; Ruiter, J. P.; IJlst, L.; Visser, G.; Reitsma-Bierens, W. C.; Heymans, H. S.; Reijngoud, D. J.; Smit, G. P.

    1997-01-01

    We describe the clinical symptoms and biochemical findings of a patient with succinyl-CoA:acetoacetate transferase deficiency who presented in the neonatal period and review the current literature on this subject. Our patient was initially suspected to have distal renal tubular acidosis, and

  20. The interaction of glutathione S-transferase M1-null variants with tobacco smoke exposure and the development of childhood asthma

    DEFF Research Database (Denmark)

    Rogers, A J; Brasch-Andersen, C; Ionita-Laza, I

    2009-01-01

    BACKGROUND: The glutathione S-transferase M1 (GSTM1)-null variant is a common copy number variant associated with adverse pulmonary outcomes, including asthma and airflow obstruction, with evidence of important gene-by-environment interactions with exposures to oxidative stress. OBJECTIVE: To exp...

  1. Quantum Mechanical/Molecular Mechanical Free Energy Simulations of the Glutathione S-Transferase (M1-1) Reaction with Phenanthrene 9,10-Oxide

    NARCIS (Netherlands)

    Ridder, L.; Rietjens, I.M.C.M.; Vervoort, J.J.M.; Mulholland, A.J.

    2002-01-01

    Glutathione S-transferases (GSTs) play an important role in the detoxification of xenobiotics in mammals. They catalyze the conjugation of glutathione to a wide range of electrophilic compounds. Phenanthrene 9,10-oxide is a model substrate for GSTs, representing an important group of epoxide

  2. Succinyl-CoA:acetoacetate transferase deficiency : identification of a new patient with a neonatal onset and review of the literature

    NARCIS (Netherlands)

    Niezen-Koning, K E; Wanders, R J; Ruiter, J P; Ijlst, L; Visser, G; Reitsma-Bierens, W C; Heijmans, Hugo; Reijngoud, D J; Smit, G P

    UNLABELLED: We describe the clinical symptoms and biochemical findings of a patient with succinyl-CoA:acetoacetate transferase deficiency who presented in the neonatal period and review the current literature on this subject. Our patient was initially suspected to have distal renal tubular acidosis,

  3. The role of the glutathione S-transferase genes GSTT1, GSTM1, and GSTP1 in acetaminophen-poisoned patients

    DEFF Research Database (Denmark)

    Buchard, Anders; Eefsen, Martin; Semb, Synne

    2012-01-01

    The aim of this study was to assess if genetic variants in the glutathione-S-transferase genes GST-T1, M1, and P1 reflect risk factors in acetaminophen (APAP)-poisoned patients assessed by investigation of the relation to prothrombin time (PT), which is a sensitive marker of survival in these pat...

  4. The association of glutathione S-transferase GSTT1 and GSTM1 gene polymorphism with pseudoexfoliative glaucoma in a Pakistani population.

    NARCIS (Netherlands)

    Khan, M.I.; Micheal, S.; Akhtar, F.; Ahmed, W.; Ijaz, B.; Ahmed, A.; Qamar, R.

    2010-01-01

    PURPOSE: The aim of the present study was to investigate the association of glutathione S-transferase GSTT1 and GSTM1 genotypes with pseudoexfoliative glaucoma (PEXG) in a group of Pakistani patients. METHODS: Multiplex polymerase chain reaction was used to study the GSTT1 and GSTM1 polymorphisms in

  5. Differential transcription of cytochrome P450s and glutathione S transferases in DDT-susceptible and resistant Drosophila melanogaster strains in response to DDT and oxidative stress

    Science.gov (United States)

    Metabolic DDT resistance in Drosophila melanogaster has previously been associated with constitutive over-transcription of cytochrome P450s. Increased P450 activity has also been associated with increased oxidative stress. In contrast, over-transcription of glutathione S transferases (GSTs) has been...

  6. Elevated gamma glutamyl transferase levels are associated with the location of acute pulmonary embolism. Cross-sectional evaluation in hospital setting

    Directory of Open Access Journals (Sweden)

    Ozge Korkmaz

    Full Text Available ABSTRACT CONTEXT AND OBJECTIVE: The location of embolism is associated with clinical findings and disease severity in cases of acute pulmonary embolism. The level of gamma-glutamyl transferase increases under oxidative stress-related conditions. In this study, we investigated whether gamma-glutamyl transferase levels could predict the location of pulmonary embolism. DESIGN AND SETTING: Hospital-based cross-sectional study at Cumhuriyet University, Sivas, Turkey. METHODS : 120 patients who were diagnosed with acute pulmonary embolism through computed tomography-assisted pulmonary angiography were evaluated. They were divided into two main groups (proximally and distally located, and subsequently into subgroups according to thrombus localization as follows: first group (thrombus in main pulmonary artery; n = 9; second group (thrombus in main pulmonary artery branches; n = 71; third group (thrombus in pulmonary artery segmental branches; n = 34; and fourth group (thrombus in pulmonary artery subsegmental branches; n = 8. RESULTS : Gamma-glutamyl transferase levels on admission, heart rate, oxygen saturation, right ventricular dilatation/hypokinesia, pulmonary artery systolic pressure and cardiopulmonary resuscitation requirement showed prognostic significance in univariate analysis. The multivariate logistic regression model showed that gamma-glutamyl transferase level on admission (odds ratio, OR = 1.044; 95% confidence interval, CI: 1.011-1.079; P = 0.009 and pulmonary artery systolic pressure (OR = 1.063; 95% CI: 1.005-1.124; P = 0.033 remained independently associated with proximally localized thrombus in pulmonary artery. CONCLUSIONS : The findings revealed a significant association between increased existing embolism load in the pulmonary artery and increased serum gamma-glutamyl transferase levels.

  7. Electronic technology

    International Nuclear Information System (INIS)

    Kim, Jin Su

    2010-07-01

    This book is composed of five chapters, which introduces electronic technology about understanding of electronic, electronic component, radio, electronic application, communication technology, semiconductor on its basic, free electron and hole, intrinsic semiconductor and semiconductor element, Diode such as PN junction diode, characteristic of junction diode, rectifier circuit and smoothing circuit, transistor on structure of transistor, characteristic of transistor and common emitter circuit, electronic application about electronic equipment, communication technology and education, robot technology and high electronic technology.

  8. The Electron

    Energy Technology Data Exchange (ETDEWEB)

    Thomson, George

    1972-01-01

    Electrons are elementary particles of atoms that revolve around and outside the nucleus and have a negative charge. This booklet discusses how electrons relate to electricity, some applications of electrons, electrons as waves, electrons in atoms and solids, the electron microscope, among other things.

  9. Hard electronics; Hard electronics

    Energy Technology Data Exchange (ETDEWEB)

    NONE

    1997-03-01

    Hard material technologies were surveyed to establish the hard electronic technology which offers superior characteristics under hard operational or environmental conditions as compared with conventional Si devices. The following technologies were separately surveyed: (1) The device and integration technologies of wide gap hard semiconductors such as SiC, diamond and nitride, (2) The technology of hard semiconductor devices for vacuum micro- electronics technology, and (3) The technology of hard new material devices for oxides. The formation technology of oxide thin films made remarkable progress after discovery of oxide superconductor materials, resulting in development of an atomic layer growth method and mist deposition method. This leading research is expected to solve such issues difficult to be easily realized by current Si technology as high-power, high-frequency and low-loss devices in power electronics, high temperature-proof and radiation-proof devices in ultimate electronics, and high-speed and dense- integrated devices in information electronics. 432 refs., 136 figs., 15 tabs.

  10. Electron radiography

    Science.gov (United States)

    Merrill, Frank E.; Morris, Christopher

    2005-05-17

    A system capable of performing radiography using a beam of electrons. Diffuser means receive a beam of electrons and diffuse the electrons before they enter first matching quadrupoles where the diffused electrons are focused prior to the diffused electrons entering an object. First imaging quadrupoles receive the focused diffused electrons after the focused diffused electrons have been scattered by the object for focusing the scattered electrons. Collimator means receive the scattered electrons and remove scattered electrons that have scattered to large angles. Second imaging quadrupoles receive the collimated scattered electrons and refocus the collimated scattered electrons and map the focused collimated scattered electrons to transverse locations on an image plane representative of the electrons' positions in the object.

  11. Antioxidant role of glutathione S-transferases: 4-Hydroxynonenal, a key molecule in stress-mediated signaling

    International Nuclear Information System (INIS)

    Singhal, Sharad S.; Singh, Sharda P.; Singhal, Preeti; Horne, David; Singhal, Jyotsana; Awasthi, Sanjay

    2015-01-01

    4-Hydroxy-2-trans-nonenal (4HNE), one of the major end products of lipid peroxidation (LPO), has been shown to induce apoptosis in a variety of cell lines. It appears to modulate signaling processes in more than one way because it has been suggested to have a role in signaling for differentiation and proliferation. It has been known that glutathione S-transferases (GSTs) can reduce lipid hydroperoxides through their Se-independent glutathione-peroxidase activity and that these enzymes can also detoxify LPO end-products such as 4HNE. Available evidence from earlier studies together with results of recent studies in our laboratories strongly suggests that LPO products, particularly hydroperoxides and 4HNE, are involved in the mechanisms of stress-mediated signaling and that it can be modulated by the alpha-class GSTs through the regulation of the intracellular concentrations of 4HNE. We demonstrate that 4HNE induced apoptosis in various cell lines is accompanied with c-Jun-N-terminal kinase (JNK) and caspase-3 activation. Cells exposed to mild, transient heat or oxidative stress acquire the capacity to exclude intracellular 4HNE at a faster rate by inducing GSTA4-4 which conjugates 4HNE to glutathione (GSH), and RLIP76 which mediates the ATP-dependent transport of the GSH-conjugate of 4HNE (GS-HNE). The balance between formation and exclusion promotes different cellular processes — higher concentrations of 4HNE promote apoptosis; whereas, lower concentrations promote proliferation. In this article, we provide a brief summary of the cellular effects of 4HNE, followed by a review of its GST-catalyzed detoxification, with an emphasis on the structural attributes that play an important role in the interactions with alpha-class GSTA4-4. Taken together, 4HNE is a key signaling molecule and that GSTs being determinants of its intracellular concentrations, can regulate stress-mediated signaling, are reviewed in this article. - Highlights: • GSTs are the major

  12. THE EFFECT OF POLYMORPHISM IN GLUTATHIONE S-TRANSFERASES ON THE DEVELOPING SECOND MALIGNANT NEOPLASMS AFTER LEUKEMIA TREATMENT IN CHILDHOOD

    Directory of Open Access Journals (Sweden)

    Janez Jazbec

    2004-12-01

    Full Text Available Background. Survivors of childhood leukemia have an increased risk of developing second malignant neoplasms and specific treatment factors such as alkylating agents, topoisomerase inhibitors and radiation have been associated with their occurrence. Genetic polymorphism in drug-metabolizing enzymes may result in impared detoxification of chemotherapeutics and may lead to increased risk for cancer.Methods. To test if polymorphism in glutathione S-transferases (GST genes is associated with occurrence of secondary malignant neoplasms, we compared GSTM1, GSTT1 and GSTP1 genotypes among 16 patients treated for childhood leukemia in whom second neoplasm occurred and matched the control group.Results. GSTM1 null genotype was found in 44% of patients with second neoplasms and in 50% in control group (p = 0.768, GSTT1 null genotype in 19% of cases and in 29% of controls (p = 0.729 and GSTP1 105 Ile/ile in 50% of cases and 37% of controls (p = 0.537. Differences in distribution of GST genotypes in patients with second neoplasms after childhood leukemia, compared to a matched control group of patients were not statistically significant.Conclusions. In our study we were not able to show relation between GST genotype and occurrence of second neoplasms after the childhood acute leukemia.

  13. Noninvasive evaluation of adult onset myopathy from carnitine palmitoyl transferase II deficiency using proton magnetic resonance spectroscopy.

    Science.gov (United States)

    Videen, J S; Haseler, L J; Karpinski, N C; Terkeltaub, R A

    1999-08-01

    The adult onset metabolic myopathy of carnitine palmitoyl transferase II (CPT II) deficiency is under-recognized, in part due to variable degrees of enzyme deficiency and symptomatology, as well as limitations in means for noninvasive evaluation. We describe a proton magnetic resonance spectroscopy (MRS) technique, using a standard clinical magnetic resonance imaging scanner, to diagnose and help monitor the response to therapy in adult CPT II deficiency. A 53-year-old woman presented with a long standing history of diffuse aching and fatigue provoked by high fat intake, fasting, or prolonged exertion. Muscle biopsy revealed myopathic features and a deficiency (33% of control) of CPT II activity with elevated palmitoyl carnitine. Proton MRS of the soleus muscle was performed using a 1.5 Tesla scanner before and during dietary therapy. Proton MRS revealed shortening of the transverse relaxation time (T2), consistent with increased acetylation of the carnitine pool. The symptoms resolved completely by treatment with frequent feedings of a high carbohydrate diet low in long chain fatty acids supplemented with medium chain triglycerides and L-carnitine. Recovery of normal muscle MRS and carnitine T2 relaxation was documented by the third month of therapy. Proton MRS is a novel, potentially useful, and readily available adjunct in the diagnosis and therapeutic monitoring of muscle CPT II deficiency.

  14. Purification and properties of an O-acetyl-transferase from Escherichia coli that can O-acetylate polysialic acid sequences

    International Nuclear Information System (INIS)

    Higa, H.; Varki, A.

    1986-01-01

    Certain strains of bacteria synthesize an outer polysialic acid (K1) capsule. Some strains of K1 + E.coli are also capable of adding O-acetyl-esters to the exocyclic hydroxyl groups of the sialic acid residues. Both the capsule and the O-acetyl modification have been correlated with differences in antigenicity and pathogenicity. The authors have developed an assay for an O-acetyl-transferase in E.coli that transfers O-[ 3 H]acetyl groups from [ 3 H]acetyl-Coenzyme A to colominic acid (fragments of the polysialic acid capsule). Using this assay, the enzyme was solubilized, and purified ∼ 600-fold using a single affinity chromatography step with Procion Red-A Agarose. The enzyme also binds to Coenzyme A Sepharose, and can be eluted with high salt or Coenzyme A. The partially purified enzyme has a pH optimum of 7.0 - 7.5, is unaffected by divalent cations, is inhibited by high salt concentrations, is inhibited by Coenzyme A (50% inhibition at 100 μM), and shows an apparent Km for colominic acid of 3.7 mM (sialic acid concentration). This enzyme could be involved in the O-acetyl +/- form variation seen in some strains of K1 + E.coli

  15. Differential substrate behaviours of ethylene oxide and propylene oxide towards human glutathione transferase theta hGSTT1-1.

    Science.gov (United States)

    Thier, R; Wiebel, F A; Bolt, H M

    1999-11-01

    The transformation of ethylene oxide (EO), propylene oxide (PO) and 1-butylene oxide (1-BuO) by human glutathione transferase theta (hGSTT1-1) was studied comparatively using 'conjugator' (GSTT1 + individuals) erythrocyte lysates. The relative sequence of velocity of enzymic transformation was PO > EO > 1-BuO. The faster transformation of PO compared to EO was corroborated in studies with human and rat GSTT1-1 (hGSTT1-1 and rGSTT1-1, respectively) expressed by Salmonella typhimurium TA1535. This sequence of reactivities of homologous epoxides towards GSTT1-1 contrasts to the sequence observed in homologous alkyl halides (methyl bromide, MBr; ethyl bromide, EtBr; n-propyl bromide, PrBr) where the relative sequence MeBr > EtBr > PrBr is observed. The higher reactivity towards GSTT1-1 of propylene oxide compared to ethylene oxide is consistent with a higher chemical reactivity. This is corroborated by experimental data of acid-catalysed hydrolysis of a number of aliphatic epoxides, including ethylene oxide and propylene oxide and consistent with semi-empirical molecular orbital modelings.

  16. A Turkish Patient With Succinyl-CoA:3-Oxoacid CoA Transferase Deficiency Mimicking Diabetic Ketoacidosis

    Directory of Open Access Journals (Sweden)

    Sahin Erdol MD

    2016-05-01

    Full Text Available Succinyl-CoA:3-oxoacid CoA transferase (SCOT deficiency is an autosomal recessive disorder of ketone body utilization that is clinically characterized with intermittent ketoacidosis crises. We report here the second Turkish case with SCOT deficiency. She experienced 3 ketoacidotic episodes: The first ketoacidotic crisis mimicked diabetic ketoacidosis because of the associated hyperglycemia. Among patients with SCOT deficiency, the blood glucose levels at the first crises were variable, and this case had the highest ever reported blood glucose level. She is a compound heterozygote with 2 novel mutations, c.517A>G (K173E and c.1543A>G (M515V, in exons 5 and 17 of the OXCT1 gene, respectively. In patient’s fibroblasts, SCOT activity was deficient and, by immunoblot analysis, SCOT protein was much reduced. The patient attained normal development and had no permanent ketosis. The accurate diagnosis of SCOT deficiency in this case had a vital impact on the management strategy and outcome.

  17. Fractal binding and dissociation kinetics of lecithin cholesterol acyl transferase (LCAT), a heart-related compound, on biosensor surfaces

    Science.gov (United States)

    Doke, Atul M.; Sadana, Ajit

    2006-05-01

    A fractal analysis is presented for the binding and dissociation of different heart-related compounds in solution to receptors immobilized on biosensor surfaces. The data analyzed include LCAT (lecithin cholesterol acyl transferase) concentrations in solution to egg-white apoA-I rHDL immobilized on a biosensor chip surface.1 Single- and dual- fractal models were employed to fit the data. Values of the binding and the dissociation rate coefficient(s), affinity values, and the fractal dimensions were obtained from the regression analysis provided by Corel Quattro Pro 8.0 (Corel Corporation Limited).2 The binding rate coefficients are quite sensitive to the degree of heterogeneity on the sensor chip surface. Predictive equations are developed for the binding rate coefficient as a function of the degree of heterogeneity present on the sensor chip surface and on the LCAT concentration in solution, and for the affinity as a function of the ratio of fractal dimensions present in the binding and the dissociation phases. The analysis presented provided physical insights into these analyte-receptor reactions occurring on different biosensor surfaces.

  18. Proteomic Profiling of Cytosolic Glutathione Transferases from Three Bivalve Species: Corbicula fluminea, Mytilus galloprovincialis and Anodonta cygnea

    Directory of Open Access Journals (Sweden)

    José Carlos Martins

    2014-01-01

    Full Text Available Suspension-feeding bivalves are considered efficient toxin vectors with a relative insensitivity to toxicants compared to other aquatic organisms. This fact highlights the potential role of detoxification enzymes, such as glutathione transferases (GSTs, in this bivalve resistance. Nevertheless, the GST system has not been extensively described in these organisms. In the present study, cytosolic GSTs isoforms (cGST were surveyed in three bivalves with different habitats and life strategies: Corbicula fluminea, Anodonta cygnea and Mytilus galloprovincialis. GSTs were purified by glutathione-agarose affinity chromatography, and the collection of expressed cGST classes of each bivalve were identified using a proteomic approach. All the purified extracts were also characterized kinetically. Results reveal variations in cGST subunits collection (diversity and properties between the three tested bivalves. Using proteomics, four pi-class and two sigma-class GST subunits were identified in M. galloprovincialis. C. fluminea also yielded four pi-class and one sigma-class GST subunits. For A. cygnea, two mu-class and one pi-class GST subunits were identified, these being the first record of GSTs from these freshwater mussels. The affinity purified extracts also show differences regarding enzymatic behavior among species. The variations found in cGST collection and kinetics might justify diverse selective advantages for each bivalve organism.

  19. In silico genome-wide identification and characterization of the glutathione S-transferase gene family in Vigna radiata.

    Science.gov (United States)

    Vaish, Swati; Awasthi, Praveen; Tiwari, Siddharth; Tiwari, Shailesh Kumar; Gupta, Divya; Basantani, Mahesh Kumar

    2018-05-01

    Plant glutathione S-transferases (GSTs) are integral to normal plant metabolism and biotic and abiotic stress tolerance. The GST gene family has been characterized in diverse plant species using molecular biology and bioinformatics approaches. In the current study, in silico analysis identified 44 GSTs in Vigna radiata. Of the total 44 GSTs identified, chromosomal locations of 31 GSTs were confirmed. The pI value of GST proteins ranged from 5.10 to 9.40. The predicted molecular weights ranged from 13.12 to 50 kDa. Subcellular localization analysis revealed that all GSTs were predominantly localized in the cytoplasm. The active site amino acids were confirmed to be serine in tau, phi, theta, zeta, and TCHQD; cysteine in lambda, DHAR, and omega; and tyrosine in EF1G. The gene architecture conformed to the two-exon/one-intron and three-exon/two-intron organization in the case of tau and phi classes, respectively. MEME analysis identified 10 significantly conserved motifs with the width of 8-50 amino acids. The motifs identified were either specific to a specific GST class or were shared by multiple GST classes. The results of the current study will be of potential importance in the characterization of the GST gene family in V. radiata, an economically important leguminous crop.

  20. Crystal Structure of Saccharomyces cerevisiae ECM4, a Xi-Class Glutathione Transferase that Reacts with Glutathionyl-(hydro)quinones

    Science.gov (United States)

    Schwartz, Mathieu; Didierjean, Claude; Hecker, Arnaud; Girardet, Jean-Michel; Morel-Rouhier, Mélanie; Gelhaye, Eric; Favier, Frédérique

    2016-01-01

    Glutathionyl-hydroquinone reductases (GHRs) belong to the recently characterized Xi-class of glutathione transferases (GSTXs) according to unique structural properties and are present in all but animal kingdoms. The GHR ScECM4 from the yeast Saccharomyces cerevisiae has been studied since 1997 when it was found to be potentially involved in cell-wall biosynthesis. Up to now and in spite of biological studies made on this enzyme, its physiological role remains challenging. The work here reports its crystallographic study. In addition to exhibiting the general GSTX structural features, ScECM4 shows extensions including a huge loop which contributes to the quaternary assembly. These structural extensions are probably specific to Saccharomycetaceae. Soaking of ScECM4 crystals with GS-menadione results in a structure where glutathione forms a mixed disulfide bond with the cysteine 46. Solution studies confirm that ScECM4 has reductase activity for GS-menadione in presence of glutathione. Moreover, the high resolution structures allowed us to propose new roles of conserved residues of the active site to assist the cysteine 46 during the catalytic act. PMID:27736955

  1. Crystal Structure of Saccharomyces cerevisiae ECM4, a Xi-Class Glutathione Transferase that Reacts with Glutathionyl-(hydroquinones.

    Directory of Open Access Journals (Sweden)

    Mathieu Schwartz

    Full Text Available Glutathionyl-hydroquinone reductases (GHRs belong to the recently characterized Xi-class of glutathione transferases (GSTXs according to unique structural properties and are present in all but animal kingdoms. The GHR ScECM4 from the yeast Saccharomyces cerevisiae has been studied since 1997 when it was found to be potentially involved in cell-wall biosynthesis. Up to now and in spite of biological studies made on this enzyme, its physiological role remains challenging. The work here reports its crystallographic study. In addition to exhibiting the general GSTX structural features, ScECM4 shows extensions including a huge loop which contributes to the quaternary assembly. These structural extensions are probably specific to Saccharomycetaceae. Soaking of ScECM4 crystals with GS-menadione results in a structure where glutathione forms a mixed disulfide bond with the cysteine 46. Solution studies confirm that ScECM4 has reductase activity for GS-menadione in presence of glutathione. Moreover, the high resolution structures allowed us to propose new roles of conserved residues of the active site to assist the cysteine 46 during the catalytic act.

  2. Inhibition of carnitine-acyl transferase I by oxfenicine studied in vivo with [11C]-labeled fatty acids

    International Nuclear Information System (INIS)

    Angsten, Gertrud; Valind, Sven; Takalo, Reijo; Neu, Henrik; Meurling, Staffan; Langstroem, Bengt

    2005-01-01

    Methods: Anesthetized pigs were studied with [ 11 C]-labeled fatty acids (FAs) with carbon chain length ranging from 8 to 16 carbon atoms, during control conditions and during inhibition of carnitine-palmitoyl transferase I (CPT I) with oxfenicine. The myocardial uptake of [ 11 C]-FAs from blood was measured together with the relative distribution of [ 11 C]-acyl-CoA between rapid mitochondrial oxidation and incorporation into slow turnover lipid pools in the heart. Results: During baseline conditions, the fractional oxidative utilization of palmitate was almost as high as that of carnitine-independent short-chain FAs, unless the carnitine shuttle was inhibited by high levels of lactate. Inhibition of CPT I almost completely blocked the oxidative pathway for palmitic acid and reduced the fractional oxidative utilization, while the rate of oxidative metabolism of acyl-CoA was unaffected. Conclusions: [ 11 C]-Labeled FAs allow rapid oxidation to be well separated from esterification into slow turnover lipid pools in the heart of anaesthetized pigs. The fractional oxidative utilization of [ 11 C]-palmitate serves well to characterize, in vivo, the carnitine-dependent transfer of long-chain FAs

  3. Inhibition of carnitine-acyl transferase I by oxfenicine studied in vivo with [{sup 11}C]-labeled fatty acids

    Energy Technology Data Exchange (ETDEWEB)

    Angsten, Gertrud [Department of Pediatric Surgery, University Children' s Hospital, S-751 85 Uppsala (Sweden)]. E-mail: gertrud.angsten@surgsci.uu.se; Valind, Sven [Uppsala University PET Centre, Uppsala University, S-751 05 Uppsala (Sweden); Department of Clinical Physiology, University Hospital, S-751 85 Uppsala (Sweden); Takalo, Reijo [Uppsala University PET Centre, Uppsala University, S-751 05 Uppsala (Sweden); Department of Clinical Physiology, University Hospital, S-751 85 Uppsala (Sweden); Neu, Henrik [Uppsala University PET Centre, Uppsala University, S-751 05 Uppsala (Sweden); Department of Organic Chemistry, Uppsala University, S-751 24 Uppsala (Sweden); Meurling, Staffan [Department of Pediatric Surgery, University Children' s Hospital, S-751 85 Uppsala (Sweden); Langstroem, Bengt [Uppsala University PET Centre, Uppsala University, S-751 05 Uppsala (Sweden); Department of Organic Chemistry, Uppsala University, S-751 24 Uppsala (Sweden)

    2005-07-01

    Methods: Anesthetized pigs were studied with [{sup 11}C]-labeled fatty acids (FAs) with carbon chain length ranging from 8 to 16 carbon atoms, during control conditions and during inhibition of carnitine-palmitoyl transferase I (CPT I) with oxfenicine. The myocardial uptake of [{sup 11}C]-FAs from blood was measured together with the relative distribution of [{sup 11}C]-acyl-CoA between rapid mitochondrial oxidation and incorporation into slow turnover lipid pools in the heart. Results: During baseline conditions, the fractional oxidative utilization of palmitate was almost as high as that of carnitine-independent short-chain FAs, unless the carnitine shuttle was inhibited by high levels of lactate. Inhibition of CPT I almost completely blocked the oxidative pathway for palmitic acid and reduced the fractional oxidative utilization, while the rate of oxidative metabolism of acyl-CoA was unaffected. Conclusions: [{sup 11}C]-Labeled FAs allow rapid oxidation to be well separated from esterification into slow turnover lipid pools in the heart of anaesthetized pigs. The fractional oxidative utilization of [{sup 11}C]-palmitate serves well to characterize, in vivo, the carnitine-dependent transfer of long-chain FAs.

  4. Mechanistic insights into EgGST1, a Mu class glutathione S-transferase from the cestode parasite Echinococcus granulosus.

    Science.gov (United States)

    Arbildi, Paula; Turell, Lucía; López, Verónica; Alvarez, Beatriz; Fernández, Verónica

    2017-11-01

    Glutathione transferases (GSTs) comprise a major detoxification system in helminth parasites, displaying both catalytic and non-catalytic activities. The kinetic mechanism of these enzymes is complex and depends on the isoenzyme which is being analyzed. Here, we characterized the kinetic mechanism of rEgGST1, a recombinant form of a cytosolic GST from Echinococcus granulosus (EgGST1), which is related to the Mu-class of mammalian enzymes, using the canonical substrates glutathione (GSH) and 1-chloro-2,4-dinitrobenzene (CDNB). Initial rate and product inhibition studies were consistent with a steady-state random sequential mechanism, where both substrates are bound to the enzyme before the products are released. Kinetic constants were also determined (pH 6.5 and 30 °C). Moreover, rEgGST1 lowered the pK a of GSH from 8.71 ± 0.07 to 6.77 ± 0.08, and enzyme-bound GSH reacted with CDNB 1 × 10 5 times faster than free GSH at pH 7.4. Finally, the dissociation of the enzyme-GSH complex was studied by means of intrinsic fluorescence, as well as that of the complex with the anthelminth drug mebendazole. This is the first report on mechanistic issues related to a helminth parasitic GST. Copyright © 2017 Elsevier Inc. All rights reserved.

  5. Human T cell recognition of the blood stage antigen Plasmodium hypoxanthine guanine xanthine phosphoribosyl transferase (HGXPRT in acute malaria

    Directory of Open Access Journals (Sweden)

    Woodberry Tonia

    2009-06-01

    Full Text Available Abstract Background The Plasmodium purine salvage enzyme, hypoxanthine guanine xanthine phosphoribosyl transferase (HGXPRT can protect mice against Plasmodium yoelii pRBC challenge in a T cell-dependent manner and has, therefore, been proposed as a novel vaccine candidate. It is not known whether natural exposure to Plasmodium falciparum stimulates HGXPRT T cell reactivity in humans. Methods PBMC and plasma collected from malaria-exposed Indonesians during infection and 7–28 days after anti-malarial therapy, were assessed for HGXPRT recognition using CFSE proliferation, IFNγ ELISPOT assay and ELISA. Results HGXPRT-specific T cell proliferation was found in 44% of patients during acute infection; in 80% of responders both CD4+ and CD8+ T cell subsets proliferated. Antigen-specific T cell proliferation was largely lost within 28 days of parasite clearance. HGXPRT-specific IFN-γ production was more frequent 28 days after treatment than during acute infection. HGXPRT-specific plasma IgG was undetectable even in individuals exposed to malaria for at least two years. Conclusion The prevalence of acute proliferative and convalescent IFNγ responses to HGXPRT demonstrates cellular immunogenicity in humans. Further studies to determine minimal HGXPRT epitopes, the specificity of responses for Plasmodia and associations with protection are required. Frequent and robust T cell proliferation, high sequence conservation among Plasmodium species and absent IgG responses distinguish HGXPRT from other malaria antigens.

  6. γ-Glutamyl Transferase as a Risk Factor for All-Cause or Cardiovascular Disease Mortality Among 5912 Ischemic Stroke.

    Science.gov (United States)

    Tu, Wen-Jun; Liu, Qiang; Cao, Jian-Lei; Zhao, Sheng-Jie; Zeng, Xian-Wei; Deng, Ai-Jun

    2017-10-01

    The aim of the study was to evaluate the association of the measurement of serum γ-glutamyl transferase (GGT) concentrations at admission with 1-year all-cause or cardiovascular disease (CVD) mortality in patients with acute ischemic stroke. This prospective, multicenter cohort study was conducted in 4 stroke centers in China. Baseline GGT measurements were tested. The relationship of GGT to the risk of death from all-cause or CVD was examined among 1-year follow-up patients. We recorded results from 5912 patients with stroke. In those patients, 51.0% were men, and the median age was 61 years. In both men and women, high GGT was significantly associated with total mortality from all-cause or CVD ( P mortality from all-cause and CVD, respectively. With an area under the curve of 0.69 (95% confidence interval, 0.66-0.73), GGT showed a significantly greater discriminatory ability to predict all-cause mortality as compared with others factors. GGT improved the National Institutes of Health Stroke Scale score (area under the curve of the combined model, 0.75 [95% confidence interval, 0.73-0.78]; P mortality in patients with ischemic stroke. © 2017 American Heart Association, Inc.

  7. Glutathione S-transferase of brown planthoppers (Nilaparvata lugens is essential for their adaptation to gramine-containing host plants.

    Directory of Open Access Journals (Sweden)

    Xiao-Qin Sun

    Full Text Available Plants have evolved complex processes to ward off attacks by insects. In parallel, insects have evolved mechanisms to thwart these plant defenses. To gain insight into mechanisms that mediate this arms race between plants and herbivorous insects, we investigated the interactions between gramine, a toxin synthesized by plants of the family Gramineae, and glutathione S transferase (GST, an enzyme found in insects that is known to detoxify xenobiotics. Here, we demonstrate that rice (Oryza sativa, a hydrophytic plant, also produces gramine and that rice resistance to brown planthoppers (Nilaparvata lugens, BPHs is highly associated with in planta gramine content. We also show that gramine is a toxicant that causes BPH mortality in vivo and that knockdown of BPH GST gene nlgst1-1 results in increased sensitivity to diets containing gramine. These results suggest that the knockdown of key detoxification genes in sap-sucking insects may provide an avenue for increasing their sensitivity to natural plant-associated defense mechanisms.

  8. Design and production of various fusion proteins of the nicotinamide/nicotinate mononucleotide adenilil transferase (NMNAT of Plasmodium falciparum

    Directory of Open Access Journals (Sweden)

    Carlos Alfonso Nieto Clavijo

    2017-09-01

    Full Text Available Recombinant proteins have become useful tools in biochemistry research. During their production, however, inclusion bodies (IB appear, on the one hand, due to the high expression rate from the recombinant plasmids, which have high efficiency promoters, and, on the other hand, intrinsic characteristics of the expressed protein. Furhtermore, the nicotinamide/nicotinate mononucleotide adenilyl transferase (NMNAT is a central protein in NAD(H+ biosynthesis, an essential cofactor in cell metabolism, and in protozoon parasite has been studied. To study the NMNAT protein of these parasites, their recombinant version in E. coli has been expressed, getting a great quantity of IB as a by-product. To increase the solubility of the protein, the coding sequence of the NMNAT enzyme of Plasmodium falciparum was cloned in different expression plasmids which were subsequently transformed into E. coli BL21(DE3 expression strain. The solubility of the recombinant proteins was assessed and the one with the highest presence in the soluble fraction was subsequently purified and its enzyme activity was determined. The recombinant protein with a MBP (maltose-binding protein tag showed an increased solubility and purity.

  9. Purification and properties of the glutathione S-transferases from the anoxia-tolerant turtle, Trachemys scripta elegans.

    Science.gov (United States)

    Willmore, William G; Storey, Kenneth B

    2005-07-01

    Glutathione S-transferases (GSTs) play critical roles in detoxification, response to oxidative stress, regeneration of S-thiolated proteins, and catalysis of reactions in nondetoxification metabolic pathways. Liver GSTs were purified from the anoxia-tolerant turtle, Trachemys scripta elegans. Purification separated a homodimeric (subunit relative molecular mass =34 kDa) and a heterodimeric (subunit relative molecular mass = 32.6 and 36.8 kDa) form of GST. The enzymes were purified 23-69-fold and 156-174-fold for homodimeric and heterodimeric GSTs, respectively. Kinetic data gathered using a variety of substrates and inhibitors suggested that both homodimeric and heterodimeric GSTs were of the alpha class although they showed significant differences in substrate affinities and responses to inhibitors. For example, homodimeric GST showed activity with known alpha class substrates, cumene hydroperoxide and p-nitrobenzylchloride, whereas heterodimeric GST showed no activity with cumene hydroperoxide. The specific activity of liver GSTs with chlorodinitrobenzene (CDNB) as the substrate was reduced by 2.6- and 8.7-fold for homodimeric and heterodimeric GSTs isolated from liver of anoxic turtles as compared with aerobic controls, suggesting an anoxia-responsive stable modification of the protein that may alter its function during natural anaerobiosis.

  10. Designing Ligands for Leishmania, Plasmodium, and Aspergillus N-Myristoyl Transferase with Specificity and Anti-Target-Safe Virtual Libraries.

    Science.gov (United States)

    Garcia-Sosa, Alfonso T

    2018-01-01

    Leishmaniasis, malaria, and fungal diseases are burdens on individuals and populations and can present severe complications. Easily accessible chemical treatments for these diseases are increasingly sought-after. Targeting the parasite N-myristoyl transferase while avoiding the human enzyme and other anti-targets may allow the prospect of compounds with pan-activity against these diseases, which would simplify treatments and costs. Developing chemical libraries, both virtual and physical, that have been filtered and flagged early on in the drug discovery process (before virtual screening) could reduce attrition rates of compounds being developed and failing late in development stages due to problems of side-effects or toxicity. Chemical libraries have been screened against the anti-targets pregnane-X-receptor, sulfotransferase, cytochrome P450 2a6, 2c9, and 3a4 with three different docking programs. Statistically significant differences are observed in their interactions with these enzymes as compared to small molecule drugs and bioactive non-drug datasets. A series of compounds are proposed with the best predicted profiles for inhibition of all parasite targets while sparing the human form and anti-targets. Some of the topranked compounds have confirmed experimental activity against Leishmania, and highlighted are those compounds with best properties for further development. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

  11. A Conserved Splicing Silencer Dynamically Regulates O-GlcNAc Transferase Intron Retention and O-GlcNAc Homeostasis

    Directory of Open Access Journals (Sweden)

    Sung-Kyun Park

    2017-08-01

    Full Text Available Modification of nucleocytoplasmic proteins with O-GlcNAc regulates a wide variety of cellular processes and has been linked to human diseases. The enzymes O-GlcNAc transferase (OGT and O-GlcNAcase (OGA add and remove O-GlcNAc, but the mechanisms regulating their expression remain unclear. Here, we demonstrate that retention of the fourth intron of OGT is regulated in response to O-GlcNAc levels. We further define a conserved intronic splicing silencer (ISS that is necessary for OGT intron retention. Deletion of the ISS in colon cancer cells leads to increases in OGT, but O-GlcNAc homeostasis is maintained by concomitant increases in OGA protein. However, the ISS-deleted cells are hypersensitive to OGA inhibition in culture and in soft agar. Moreover, growth of xenograft tumors from ISS-deleted cells is compromised in mice treated with an OGA inhibitor. Thus, ISS-mediated regulation of OGT intron retention is a key component in OGT expression and maintaining O-GlcNAc homeostasis.

  12. Volatile Gas Production by Methyl Halide Transferase: An In Situ Reporter Of Microbial Gene Expression In Soil.

    Science.gov (United States)

    Cheng, Hsiao-Ying; Masiello, Caroline A; Bennett, George N; Silberg, Jonathan J

    2016-08-16

    Traditional visual reporters of gene expression have only very limited use in soils because their outputs are challenging to detect through the soil matrix. This severely restricts our ability to study time-dependent microbial gene expression in one of the Earth's largest, most complex habitats. Here we describe an approach to report on dynamic gene expression within a microbial population in a soil under natural water levels (at and below water holding capacity) via production of methyl halides using a methyl halide transferase. As a proof-of-concept application, we couple the expression of this gas reporter to the conjugative transfer of a bacterial plasmid in a soil matrix and show that gas released from the matrix displays a strong correlation with the number of transconjugant bacteria that formed. Gas reporting of gene expression will make possible dynamic studies of natural and engineered microbes within many hard-to-image environmental matrices (soils, sediments, sludge, and biomass) at sample scales exceeding those used for traditional visual reporting.

  13. Association of N-acetyltransferase-2 and glutathione S-transferase polymorphisms with idiopathic male infertility in Vietnam male subjects.

    Science.gov (United States)

    Trang, Nguyen Thi; Huyen, Vu Thi; Tuan, Nguyen Thanh; Phan, Tran Duc

    2018-04-25

    N-acetyltransferase-2 (NAT2) and Glutathione S-transferases (GSTs) are phase-II xenobiotic metabolizing enzymes participating in detoxification of toxic arylamines, aromatic amines, hydrazines and reactive oxygen species (ROS), which are produced under oxidative and electrophile stresses. The purpose of this research was to investigate whether two common single-nucleotide polymorphisms (SNP) of NAT2 (rs1799929, rs1799930) and GSTP1 (rs1138272, rs1695) associated with susceptibility to idiopathic male infertility. A total 300 DNA samples (150 infertile patients and 150 healthy control) were genotyped for the polymorphisms by ARMS - PCR. We revealed a significant association between the NAT2 variant genotypes (CT + TT (rs1799929), (OR: 3.74; p male infertility in subjects from Vietnam. This pilot study is the first (as far as we know) to reveal that polymorphisms of NAT2 (rs1799929, rs1799930) and GSTP1 (rs1138272, rs1695) are some novel genetic markers for susceptibility to idiopathic male infertility. Copyright © 2018 Elsevier B.V. All rights reserved.

  14. Effect of Arsenic and Chromium on the Serum Amino-Transferases Activity in Indian Major Carp, Labeo rohita

    Directory of Open Access Journals (Sweden)

    Anjaneyulu Yerramilli

    2007-09-01

    Full Text Available Arsenic and hexavalent chromium toxicity results from their ability to interact with sulfahydryl groups of proteins and enzymes, and to substitute phosphorus in a variety of biochemical reactions. Alanine aminotransferase (ALT; E.C: 2.6.1.2 and Aspartate amino transferase (AST; EC 2.6.1.1 play a crucial role in transamination reactions and can be used as potential biomarkers to indicate hepatotoxicity and cellular damage. While histopathological studies in liver tissue require more time and expertise, simple and reliable biochemical analysis of ALT and AST can be used for a rapid assessment of tissue and cellular damage within 96 h. The main objective of this study was to determine the acute effects of arsenic and hexavalent chromium on the activity of ALT and AST in the Indian major carp, Labeo rohita for 24 h and 96 h. Significant increase in the activity of ALT (P < 0.01 from controls in arsenic exposed fish indicates serious hepatic damage and distress condition to the fish. However, no such significant changes were observed in chromium-exposed fish suggesting that arsenic is more toxic to the fish. These findings indicate that ALT and AST are candidate biomarkers for arsenic-induced hepatotoxicity in Labeo rohita.

  15. Thymidylate Synthase, Thymidine Phosphorylase and Orotate Phosphoribosyl Transferase Levels as Predictive Factors of Chemotherapy in Oral Squamous Cell Carcinoma

    International Nuclear Information System (INIS)

    Ogiuchi, Yosuke; Maruoka, Yasubumi; Ando, Tomohiro; Kobayashi, Makio; Ogiuchi, Hideki

    2008-01-01

    We conducted a clinicopathologic study on protein and mRNA levels of thymidylate synthase (TS), thymidine phosphorylase (TP) and orotate phosphoribosyl transferase (OPRT) using biopsy tissue specimens before treatment. The mRNA levels have been measured in tumor cells microdissected from paraffin-embedded specimens (Danenberg Tumor Profile method: DTP method). We studied the mRNA and protein expression as effect predictive factors in chemotherapy. The subjects consisted of 20 cases of untreated oral squamous cell carcinoma who had undergone chemotherapy with TS-1 (16 males and 4 females, tongue in 8 cases, upper gingiva in 3 cases, lower gingiva in 3 cases, buccal mucosa in 5 cases and floor of the mouth in 1 case). TS gene expressions of the responders were lower than those for the nonresponders. Furthermore, regarding males who were less than 70 years of age, stage I and II, well differentiated type and tongue, TS mRNA expression of the responders were lower than that for the nonresponders. The mRNA expression of OPRT for the male responders was lower than that for the nonresponders. No remarkable difference was observed by immunohistochemistry. In this study, the measurement of the TS levels using the DTP method may potentially act as a predictive factor of antitumor effectiveness

  16. An acetylation site in lectin domain modulates the biological activity of polypeptide GalNAc-transferase-2

    DEFF Research Database (Denmark)

    Zlocowski, Natacha; Lorenz, Virginia; Bennett, Eric Paul

    2013-01-01

    Abstract Polypeptide GalNAc-transferases (ppGalNAc-Ts) are a family of enzymes that catalyze the initiation of mucin-type O-glycosylation. All ppGalNAc-T family members contain a common (QXW)3 motif which is present in R-type lectin group. Acetylation site K521 is part of the QKW motif of ß......-trefoil in the lectin domain of ppGalNAc-T2. We used a combination of acetylation and site-directed mutagenesis approaches to examine the functional role of K521 in ppGalNAc-T2. Binding assays of non-acetylated and acetylated forms of the mutant ppGalNAc-T2K521Q to various naked and aGalNAc-glycosylated mucin peptides...... indicated that degree of interaction of lectin domain with aGalNAc depends on the peptide sequence of mucin. Studies of inhibitory effect of various carbohydrates on interactions of ppGalNAc-T2 with MUC1aGalNAc indicate that point K521Q mutation enhance the carbohydrate specificity of lectin domain for aGalNAc...

  17. Electrons, Electronic Publishing, and Electronic Display.

    Science.gov (United States)

    Brownrigg, Edwin B.; Lynch, Clifford A.

    1985-01-01

    Provides a perspective on electronic publishing by distinguishing between "Newtonian" publishing and "quantum-mechanical" publishing. Highlights include media and publishing, works delivered through electronic media, electronic publishing and the printed word, management of intellectual property, and recent copyright-law issues…

  18. Evaluation of the precision-cut liver and lung slice systems for the study of induction of CYP1, epoxide hydrolase and glutathione S-transferase activities.

    Science.gov (United States)

    Pushparajah, Daphnee S; Umachandran, Meera; Plant, Kathryn E; Plant, Nick; Ioannides, Costas

    2007-02-28

    The principal objective was to ascertain whether precision-cut tissue slices can be used to evaluate the potential of chemicals to induce CYP1, epoxide hydrolase and glutathione S-transferase activities, all being important enzymes involved in the metabolism of polycyclic aromatic hydrocarbons. Precision-cut rat liver and lung slices were incubated with a range of benzo[a]pyrene concentrations for various time periods. A rise in the O-deethylation of ethoxyresorufin was seen in both liver and lung slices exposed to benzo[a]pyrene, which was accompanied by increased CYP1A apoprotein levels. Pulmonary CYP1B1 apoprotein levels and hepatic mRNA levels were similarly enhanced. Elevated epoxide hydrolase and glutathione S-transferase activities were also observed in liver slices following incubation for 24h; similarly, a rise in apoprotein levels of both enzymes was evident, peak levels occurring at the same time point. When mRNA levels were monitored, a rise in the levels of both enzymes was seen as early as 4h after incubation, but maximum levels were attained at 24 h. In lung slices, induction of epoxide hydrolase by benzo[a]pyrene was observed after a 24-h incubation, and at a concentration of 1 microM; a rise in apoprotein levels was seen at this time point. Glutathione S-transferase activity was not inducible in lung slices by benzo[a]pyrene but a modest increase was observed in hepatic slices. Collectively, these studies confirmed CYP1A induction in rat liver slices and established that CYP1B1 expression, and epoxide hydrolase and glutathione S-transferase activities are inducible in precision-cut tissue slices.

  19. Purification, molecular cloning and heterologous expression of a glutathione S-transferase involved in insecticide resistance from the rice brown planthopper, Nilaparvata lugens.

    OpenAIRE

    Vontas, John G; Small, Graham J; Nikou, Dimitra C; Ranson, Hilary; Hemingway, Janet

    2002-01-01

    A novel glutathione S-transferase (GST)-based pyrethroid resistance mechanism was recently identified in Nilaparvata lugens [Vontas, Small and Hemingway (2001) Biochem. J. 357, 65-72]. To determine the nature of GSTs involved in conferring this resistance, the GSTs from resistant and susceptible strains of N. lugens were partially purified by anion exchange and affinity chromatography. The majority of peroxidase activity, previously correlated with resistance, was confined to the fraction tha...

  20. An ethylene-responsive enhancer element is involved in the senescence-related expression of the carnation glutathione-S-transferase (GST1) gene.

    OpenAIRE

    Itzhaki, H; Maxson, J M; Woodson, W R

    1994-01-01

    The increased production of ethylene during carnation petal senescence regulates the transcription of the GST1 gene encoding a subunit of glutathione-S-transferase. We have investigated the molecular basis for this ethylene-responsive transcription by examining the cis elements and trans-acting factors involved in the expression of the GST1 gene. Transient expression assays following delivery of GST1 5' flanking DNA fused to a beta-glucuronidase receptor gene were used to functionally define ...

  1. Two pear glutathione S-transferases genes are regulated during fruit development and involved in response to salicylic acid, auxin, and glucose signaling.

    Directory of Open Access Journals (Sweden)

    Hai-Yan Shi

    Full Text Available Two genes encoding putative glutathione S-transferase proteins were isolated from pear (Pyrus pyrifolia and designated PpGST1 and PpGST2. The deduced PpGST1 and PpGST2 proteins contain conserved Glutathione S-transferase N-terminal domain (GST_N and Glutathione S-transferase, C-terminal domain (GST_C. Using PCR amplification technique, the genomic clones corresponding to PpGST1 and PpGST2 were isolated and shown to contain two introns and a singal intron respectively with typical GT/AG boundaries defining the splice junctions. Phylogenetic analysis clearly demonstrated that PpGST1 belonged to Phi class of GST superfamilies and had high homology with apple MdGST, while PpGST2 was classified into the Tau class of GST superfamilies. The expression of PpGST1 and PpGST2 genes was developmentally regulated in fruit. Further study demonstrated that PpGST1 and PpGST2 expression was remarkably induced by glucose, salicylic acid (SA and indole-3-aceticacid (IAA treatments in pear fruit, and in diseased fruit. These data suggested that PpGST1 and PpGST2 might be involved in response to sugar, SA, and IAA signaling during fruit development of pear.

  2. EFEK POLIMORFISME GENA GSTP-1 TERHADAP AKTIVITAS GLUTATION S-TRANSFERASE (GST PADA INDIVIDU TERPAPAR LOGAM BERAT TIMBAL (Effect of GSTP-1 Gene Polymorphismson Glutation S- Transferase (GST Activity in Heavy Metals Lead-Exposed Individual

    Directory of Open Access Journals (Sweden)

    Hernayanti Hernayanti

    2015-11-01

    Full Text Available ABSTRAK Gena GSTP-1 merupakan penghasil enzim glutation S- transferase (GST, yang berfungsi dalam proses detoksifikasi senyawa toksik di hati. Faktor keberadaan polimorfisme gena GSTP-1 akan menyebabkan penurunan ekspresi GST, sehingga proses detoksifikasi terhadap senyawa toksik akan terhambat. Kerentanan terhadap paparan senyawa toksik pada manusia akan meningkat apabila dijumpai polimorfisme gena. Salah satu senyawa toksik yang dapat menghambat aktivitas GST adalah timbal (Pb, terutama dalam bentuk tetra ethyl lead (TEL. Tujuan penelitian adalah untuk mengetahui pengaruh polimorfisme gena GSTP-1 terhadap aktivitas GST pada individu terpapar Pb, yang diwakili pekerja bengkel mobil. Faktor keberadaan polimorfisme gena individu ditentukan dengan metode PCR-RFLP dan enzim restriksi BsmA1. Parameter yang diukur adalah kadar Pb dan aktivitas GST. Analisis molekuler gena GSTP-1 dilakukan secara deskriptif. Data kadar Pb dan aktivitas GST dianalisis dengan uji t independent. Hasil analisis gena GSTP-1 dari 40 orang subyek kasus setelah dilakukan digesti dengan enzim BsmA1, ditemukan sebanyak 10 orang individu dengan polimorfisme Ile105Val gena GSTP 1 atau sekitar 25% dengan genotip Ile-Val, sedangkan 30 orang atau 75% ditemukan tanpa polimorfisme dengan genotip Ile-Ile. Pita DNA individu dengan polimorfisme terpotong menjadi 3 fragmen sepanjang 176, 91 dan 85 pp (mutan heterozygot, sedangkan tanpa polimorfisme terletak pada 176 bp. Subyek kasus dengan polimorfisme gena GSTP-1 memiliki kadar Pb lebih tinggi dan aktivitas GST lebih rendah dibandingkan individu non polimorfisme. Telah terbukti bahwa polimorfisme gena GSTP-1 menyebabkan penurunan ekspresi enzim GST. Pada individu terpapar Pb dengan polimorfisme gena GSTP-1 memiliki aktivitas GST lebih rendah dibandingkan individu tanpa polimorfisme. ABSTRACT GSTP-1 gene regulates the expression of gluthation S-transferase enzyme, which role in detoxification of toxicant on liver. If the polymorphisms

  3. Electron detector

    International Nuclear Information System (INIS)

    Hashimoto, H.; Mogami, A.

    1975-01-01

    A device for measuring electron densities at a given energy level in an electron beam or the like having strong background noise, for example, in the detection of Auger electric energy spectrums is described. An electron analyzer passes electrons at the given energy level and at the same time electrons of at least one adjacent energy level. Detecting means associated therewith produce signals indicative of the densities of the electrons at each energy level and combine these signals to produce a signal indicative of the density of the electrons of the given energy level absent background noise

  4. Replacing Escherichia coli NAD-dependent glyceraldehyde 3-phosphate dehydrogenase (GAPDH) with a NADP-dependent enzyme from Clostridium acetobutylicum facilitates NADPH dependent pathways.

    Science.gov (United States)

    Martínez, Irene; Zhu, Jiangfeng; Lin, Henry; Bennett, George N; San, Ka-Yiu

    2008-11-01

    Reactions requiring reducing equivalents, NAD(P)H, are of enormous importance for the synthesis of industrially valuable compounds such as carotenoids, polymers, antibiotics and chiral alcohols among others. The use of whole-cell biocatalysis can reduce process cost by acting as catalyst and cofactor regenerator at the same time; however, product yields might be limited by cofactor availability within the cell. Thus, our study focussed on the genetic manipulation of a whole-cell system by modifying metabolic pathways and enzymes to improve the overall production process. In the present work, we genetically engineered an Escherichia coli strain to increase NADPH availability to improve the productivity of products that require NADPH in its biosynthesis. The approach involved an alteration of the glycolysis step where glyceraldehyde-3-phosphate (GAP) is oxidized to 1,3 bisphophoglycerate (1,3-BPG). This reaction is catalyzed by NAD-dependent endogenous glyceraldehyde-3-phosphate dehydrogenase (GAPDH) encoded by the gapA gene. We constructed a recombinant E. coli strain by replacing the native NAD-dependent gapA gene with a NADP-dependent GAPDH from Clostridium acetobutylicum, encoded by the gene gapC. The beauty of this approach is that the recombinant E. coli strain produces 2 mol of NADPH, instead of NADH, per mole of glucose consumed. Metabolic flux analysis showed that the flux through the pentose phosphate (PP) pathway, one of the main pathways that produce NADPH, was reduced significantly in the recombinant strain when compared to that of the parent strain. The effectiveness of the NADPH enhancing system was tested using the production of lycopene and epsilon-caprolactone as model systems using two different background strains. The recombinant strains, with increased NADPH availability, consistently showed significant higher productivity than the parent strains.

  5. Identification and characterization of NADPH-dependent cytochrome P450 reductase gene and cytochrome b₅ gene from Plutella xylostella: possible involvement in resistance to beta-cypermethrin.

    Science.gov (United States)

    Chen, Xi'en; Zhang, Yalin

    2015-03-10

    NADPH-cytochrome P450 reductase (CPR) and cytochrome b5 (b5) are essential for cytochrome P450 mediated biological reactions. CPR and b5 in several insects have been found to be associated with insecticide resistance. However, CPR and b5 in the diamondback moth (DBM), Plutella xylostella, are not characterized and their roles remain undefined. A full-length cDNA of CPR encoding 678 amino acids and a full-length cDNA of b5 encoding 127 amino acids were cloned from DBM. Their deduced amino acid sequences shared high identities with those of other insects and showed characteristics of classical CPRs and b5s, respectively. The mRNAs of both genes were detectable in all developmental stages with the highest expression levels occurring in the 4th instar larvae. Tissue-specific expression analysis showed that their transcripts were most abundant in gut. Transcripts of CPR and b5 in the beta-cypermethrin resistant DBM strain were 13.2- and 2.84-fold higher than those in the beta-cypermethrin susceptible strain, respectively. The expression levels of CPR and b5 were enhanced by beta-cypermethrin at the concentration of 12 mg L(-1) (~LC10). The results indicate that CPR and b5 may play essential roles in the P450 mediated resistance of DBM to beta-cypermethrin or even other insecticides. Copyright © 2015 Elsevier B.V. All rights reserved.

  6. Differential activation of diverse glutathione transferases of Clonorchis sinensis in response to the host bile and oxidative stressors.

    Directory of Open Access Journals (Sweden)

    Young-An Bae

    Full Text Available BACKGROUND: Clonorchis sinensis causes chronic cumulative infections in the human hepatobiliary tract and is intimately associated with cholangiocarcinoma. Approximately 35 million people are infected and 600 million people are at risk of infections worldwide. C. sinensis excretory-secretory products (ESP constitute the first-line effector system affecting the host-parasite interrelationship by interacting with bile fluids and ductal epithelium. However, the secretory behavior of C. sinensis in an environment close to natural host conditions is unclear. C. sinensis differs from Fasciola hepatica in migration to, and maturation in, the hepatic bile duct, implying that protein profile of the ESP of these two trematodes might be different from each other. METHODOLOGY/PRINCIPAL FINDINGS: We conducted systemic approaches to analyze the C. sinensis ESP proteome and the biological reactivity of C. sinensis glutathione transferases (GSTs, such as global expression patterns and induction profiles under oxidative stress and host bile. When we observed ex host excretion behavior of C. sinensis in the presence of 10% host bile, the global proteome pattern was not significantly altered, but the amount of secretory proteins was increased by approximately 3.5-fold. Bioactive molecules secreted by C. sinensis revealed universal/unique features in relation to its intraluminal hydrophobic residing niche. A total of 38 protein spots identified abundantly included enzymes involved in glucose metabolism (11 spots, 28.9% and diverse-classes of glutathione transferases (GSTs; 10 spots, 26.3%. Cathepsin L/F (four spots, 10.5% and transporter molecules (three spots, 7.9% were also recognized. The universal secretory proteins found in other parasites, such as several enzymes involved in glucose metabolism and oxygen transporters, were commonly detected. C. sinensis secreted less cysteine proteases and fatty acid binding proteins compared to other tissue-invading or

  7. Differential Activation of Diverse Glutathione Transferases of Clonorchis sinensis in Response to the Host Bile and Oxidative Stressors

    Science.gov (United States)

    Bae, Young-An; Ahn, Do-Whan; Lee, Eung-Goo; Kim, Seon-Hee; Cai, Guo-Bin; Kang, Insug; Sohn, Woon-Mok; Kong, Yoon

    2013-01-01

    Background Clonorchis sinensis causes chronic cumulative infections in the human hepatobiliary tract and is intimately associated with cholangiocarcinoma. Approximately 35 million people are infected and 600 million people are at risk of infections worldwide. C. sinensis excretory-secretory products (ESP) constitute the first-line effector system affecting the host-parasite interrelationship by interacting with bile fluids and ductal epithelium. However, the secretory behavior of C. sinensis in an environment close to natural host conditions is unclear. C. sinensis differs from Fasciola hepatica in migration to, and maturation in, the hepatic bile duct, implying that protein profile of the ESP of these two trematodes might be different from each other. Methodology/Principal Findings We conducted systemic approaches to analyze the C. sinensis ESP proteome and the biological reactivity of C. sinensis glutathione transferases (GSTs), such as global expression patterns and induction profiles under oxidative stress and host bile. When we observed ex host excretion behavior of C. sinensis in the presence of 10% host bile, the global proteome pattern was not significantly altered, but the amount of secretory proteins was increased by approximately 3.5-fold. Bioactive molecules secreted by C. sinensis revealed universal/unique features in relation to its intraluminal hydrophobic residing niche. A total of 38 protein spots identified abundantly included enzymes involved in glucose metabolism (11 spots, 28.9%) and diverse-classes of glutathione transferases (GSTs; 10 spots, 26.3%). Cathepsin L/F (four spots, 10.5%) and transporter molecules (three spots, 7.9%) were also recognized. The universal secretory proteins found in other parasites, such as several enzymes involved in glucose metabolism and oxygen transporters, were commonly detected. C. sinensis secreted less cysteine proteases and fatty acid binding proteins compared to other tissue-invading or intravascular

  8. Are glutathione S transferases involved in DNA damage signalling? Interactions with DNA damage and repair revealed from molecular epidemiology studies

    Energy Technology Data Exchange (ETDEWEB)

    Dusinska, Maria, E-mail: Maria.DUSINSKA@nilu.no [CEE-Health Effects Group, NILU - Norwegian Institute for Air Research, Kjeller (Norway); Staruchova, Marta; Horska, Alexandra [Department of Experimental and Applied Genetics, Slovak Medical University, Bratislava (Slovakia); Smolkova, Bozena [Laboratory of Cancer Genetics, Cancer Research Institute of the Slovak Academy of Sciences, Bratislava (Slovakia); Collins, Andrew [Department of Nutrition, Faculty of Medicine, University of Oslo (Norway); Bonassi, Stefano [Unit of Clinical and Molecular Epidemiology, IRCCS San Raffaele Pisana, Rome (Italy); Volkovova, Katarina [Department of Experimental and Applied Genetics, Slovak Medical University, Bratislava (Slovakia)

    2012-08-01

    Glutathione S-transferases (GSTs) are members of a multigene family of isoenzymes that are important in the control of oxidative stress and in phase II metabolism. Acting non-enzymically, GSTs can modulate signalling pathways of cell proliferation, cell differentiation and apoptosis. Using a molecular epidemiology approach, we have investigated a potential involvement of GSTs in DNA damage processing, specifically the modulation of DNA repair in a group of 388 healthy adult volunteers; 239 with at least 5 years of occupational exposure to asbestos, stone wool or glass fibre, and 149 reference subjects. We measured DNA damage in lymphocytes using the comet assay (alkaline single cell gel electrophoresis): strand breaks (SBs) and alkali-labile sites, oxidised pyrimidines with endonuclease III, and oxidised purines with formamidopyrimidine DNA glycosylase. We also measured GST activity in erythrocytes, and the capacity for base excision repair (BER) in a lymphocyte extract. Polymorphisms in genes encoding three GST isoenzymes were determined, namely deletion of GSTM1 and GSTT1 and single nucleotide polymorphism Ile105Val in GSTP1. Consumption of vegetables and wine correlated negatively with DNA damage and modulated BER. GST activity correlated with oxidised bases and with BER capacity, and differed depending on polymorphisms in GSTP1, GSTT1 and GSTM1. A significantly lower BER rate was associated with the homozygous GSTT1 deletion in all asbestos site subjects and in the corresponding reference group. Multifactorial analysis revealed effects of sex and exposure in GSTP1 Ile/Val heterozygotes but not in Ile/Ile homozygotes. These variants affected also SBs levels, mainly by interactions of GSTP1 genotype with exposure, with sex, and with smoking habit; and by an interaction between sex and smoking. Our results show that GST polymorphisms and GST activity can apparently influence DNA stability and repair of oxidised bases, suggesting a potential new role for these

  9. Biochemical Characterization and Vaccine Potential of a Heme-Binding Glutathione Transferase from the Adult Hookworm Ancylostoma caninum

    Science.gov (United States)

    Zhan, Bin; Liu, Sen; Perally, Samirah; Xue, Jian; Fujiwara, Ricardo; Brophy, Peter; Xiao, Shuhua; Liu, Yueyuan; Feng, Jianjun; Williamson, Angela; Wang, Yan; Bueno, Lilian L.; Mendez, Susana; Goud, Gaddam; Bethony, Jeffrey M.; Hawdon, John M.; Loukas, Alex; Jones, Karen; Hotez, Peter J.

    2005-01-01

    We report the cloning and expression of Ac-GST-1, a novel glutathione S-transferase from the adult hookworm Ancylostoma caninum, and its possible role in parasite blood feeding and as a vaccine target. The predicted Ac-GST-1 open reading frame contains 207 amino acids (mass, 24 kDa) and exhibited up to 65% amino acid identity with other nematode GSTs. mRNA encoding Ac-GST-1 was detected in adults, eggs, and larval stages, but the protein was detected only in adult hookworm somatic extracts and excretory/secretory products. Using antiserum to the recombinant protein, Ac-GST-1 was immunolocalized to the parasite hypodermis and muscle tissue and weakly to the intestine. Recombinant Ac-GST-1 was enzymatically active, as determined by conjugation of glutathione to a model substrate, and exhibited a novel high-affinity binding site for hematin. The possible role of Ac-GST-1 in parasite heme detoxification during hemoglobin digestion or heme uptake prompted interest in evaluating it as a potential vaccine antigen. Vaccination of dogs with Ac-GST-1 resulted in a 39.4% reduction in the mean worm burden and 32.3% reduction in egg counts compared to control dogs following larval challenge, although the reductions were not statistically significant. However, hamsters vaccinated with Ac-GST-1 exhibited statistically significant worm reduction (53.7%) following challenge with heterologous Necator americanus larvae. These studies suggest that Ac-GST-1 is a possible drug and vaccine target for hookworm infection. PMID:16177370

  10. Nuclear translocation of glutathione S-transferase {pi} is mediated by a non-classical localization signal

    Energy Technology Data Exchange (ETDEWEB)

    Kawakatsu, Miho [Department of Stem Cell Biology, Atomic Bomb Disease Institute, Nagasaki University Graduate School of Biomedical Sciences, Nagasaki 852-8523 (Japan); Goto, Shinji, E-mail: sgoto@nagasaki-u.ac.jp [Department of Stem Cell Biology, Atomic Bomb Disease Institute, Nagasaki University Graduate School of Biomedical Sciences, Nagasaki 852-8523 (Japan); Yoshida, Takako; Urata, Yoshishige; Li, Tao-Sheng [Department of Stem Cell Biology, Atomic Bomb Disease Institute, Nagasaki University Graduate School of Biomedical Sciences, Nagasaki 852-8523 (Japan)

    2011-08-12

    Highlights: {yields} Nuclear translocation of GST{pi} is abrogated by the deletion of the last 16 amino acid residues in the carboxy-terminal region, indicating that residues 195-208 of GST{pi} are required for nuclear translocation. {yields} The lack of a contiguous stretch of positively charged amino acid residues within the carboxy-terminal region of GST{pi}, suggests that the nuclear translocation of GST{pi} is mediated by a non-classical nuclear localization signal. {yields} An in vitro transport assay shows that the nuclear translocation of GST{pi} is dependent on cytosolic factors and ATP. -- Abstract: Glutathione S-transferase {pi} (GST{pi}), a member of the GST family of multifunctional enzymes, is highly expressed in human placenta and involved in the protection of cellular components against electrophilic compounds or oxidative stress. We have recently found that GST{pi} is expressed in the cytoplasm, mitochondria, and nucleus in some cancer cells, and that the nuclear expression of GST{pi} appears to correlate with resistance to anti-cancer drugs. Although the mitochondrial targeting signal of GST{pi} was previously identified in the amino-terminal region, the mechanism of nuclear translocation remains completely unknown. In this study, we find that the region of GST{pi}195-208 is critical for nuclear translocation, which is mediated by a novel and non-classical nuclear localization signal. In addition, using an in vitro transport assay, we demonstrate that the nuclear translocation of GST{pi} depends on the cytosolic extract and ATP. Although further experiments are needed to understand in depth the precise mechanism of nuclear translocation of GST{pi}, our results may help to establish more efficient anti-cancer therapy, especially with respect to resistance to anti-cancer drugs.

  11. JS-K, a glutathione S-transferase-activated nitric oxide donor with antineoplastic activity in malignant gliomas.

    Science.gov (United States)

    Weyerbrock, Astrid; Osterberg, Nadja; Psarras, Nikolaos; Baumer, Brunhilde; Kogias, Evangelos; Werres, Anna; Bette, Stefanie; Saavedra, Joseph E; Keefer, Larry K; Papazoglou, Anna

    2012-02-01

    Glutathione S-transferases (GSTs) control multidrug resistance and are upregulated in many cancers, including malignant gliomas. The diazeniumdiolate JS-K generates nitric oxide (NO) on enzymatic activation by glutathione and GST, showing promising NO-based anticancer efficacy. To evaluate the role of NO-based antitumor therapy with JS-K in U87 gliomas in vitro and in vivo. U87 glioma cells and primary glioblastoma cell lines were exposed to JS-K and a variety of inhibitors to study cell death by necrosis, apoptosis, and other mechanisms. GST expression was evaluated by immunocytochemistry, polymerase chain reaction, and Western blot, and NO release from JS-K was studied with a NO assay. The growth-inhibitory effect of JS-K was studied in a U87 xenograft model in vivo. Dose-dependent inhibition of cell proliferation was observed in human U87 glioma cells and primary glioblastoma cells in vitro. Cell death was partially induced by caspase-dependent apoptosis, which could be blocked by Z-VAD-FMK and Q-VD-OPH. Inhibition of GST by sulfasalazine, cGMP inhibition by ODQ, and MEK1/2 inhibition by UO126 attenuated the antiproliferative effect of JS-K, suggesting the involvement of various intracellular death signaling pathways. Response to JS-K correlated with mRNA and protein expression of GST and the amount of NO released by the glioma cells. Growth of U87 xenografts was reduced significantly, with immunohistochemical evidence for increased necrosis and apoptosis and reduced proliferation. Our data show for the first time the potent antiproliferative effect of JS-K in gliomas in vitro and in vivo. These findings warrant further investigation of this novel NO-releasing prodrug in gliomas.

  12. Post-transcriptional generation of miRNA variants by multiple nucleotidyl transferases contributes to miRNA transcriptome complexity.

    Science.gov (United States)

    Wyman, Stacia K; Knouf, Emily C; Parkin, Rachael K; Fritz, Brian R; Lin, Daniel W; Dennis, Lucas M; Krouse, Michael A; Webster, Philippa J; Tewari, Muneesh

    2011-09-01

    Modification of microRNA sequences by the 3' addition of nucleotides to generate so-called "isomiRs" adds to the complexity of miRNA function, with recent reports showing that 3' modifications can influence miRNA stability and efficiency of target repression. Here, we show that the 3' modification of miRNAs is a physiological and common post-transcriptional event that shows selectivity for specific miRNAs and is observed across species ranging from C. elegans to human. The modifications result predominantly from adenylation and uridylation and are seen across tissue types, disease states, and developmental stages. To quantitatively profile 3' nucleotide additions, we developed and validated a novel assay based on NanoString Technologies' nCounter platform. For certain miRNAs, the frequency of modification was altered by processes such as cell differentiation, indicating that 3' modification is a biologically regulated process. To investigate the mechanism of 3' nucleotide additions, we used RNA interference to screen a panel of eight candidate miRNA nucleotidyl transferases for 3' miRNA modification activity in human cells. Multiple enzymes, including MTPAP, PAPD4, PAPD5, ZCCHC6, ZCCHC11, and TUT1, were found to govern 3' nucleotide addition to miRNAs in a miRNA-specific manner. Three of these enzymes-MTPAP, ZCCHC6, and TUT1-have not previously been known to modify miRNAs. Collectively, our results indicate that 3' modification observed in next-generation small RNA sequencing data is a biologically relevant process, and identify enzymatic mechanisms that may lead to new approaches for modulating miRNA activity in vivo.

  13. Glutathione S-Transferase (GST Gene Diversity in the Crustacean Calanus finmarchicus--Contributors to Cellular Detoxification.

    Directory of Open Access Journals (Sweden)

    Vittoria Roncalli

    Full Text Available Detoxification is a fundamental cellular stress defense mechanism, which allows an organism to survive or even thrive in the presence of environmental toxins and/or pollutants. The glutathione S-transferase (GST superfamily is a set of enzymes involved in the detoxification process. This highly diverse protein superfamily is characterized by multiple gene duplications, with over 40 GST genes reported in some insects. However, less is known about the GST superfamily in marine organisms, including crustaceans. The availability of two de novo transcriptomes for the copepod, Calanus finmarchicus, provided an opportunity for an in depth study of the GST superfamily in a marine crustacean. The transcriptomes were searched for putative GST-encoding transcripts using known GST proteins from three arthropods as queries. The identified transcripts were then translated into proteins, analyzed for structural domains, and annotated using reciprocal BLAST analysis. Mining the two transcriptomes yielded a total of 41 predicted GST proteins belonging to the cytosolic, mitochondrial or microsomal classes. Phylogenetic analysis of the cytosolic GSTs validated their annotation into six different subclasses. The predicted proteins are likely to represent the products of distinct genes, suggesting that the diversity of GSTs in C. finmarchicus exceeds or rivals that described for insects. Analysis of relative gene expression in different developmental stages indicated low levels of GST expression in embryos, and relatively high expression in late copepodites and adult females for several cytosolic GSTs. A diverse diet and complex life history are factors that might be driving the multiplicity of GSTs in C. finmarchicus, as this copepod is commonly exposed to a variety of natural toxins. Hence, diversity in detoxification pathway proteins may well be key to their survival.

  14. Effects of three pesticides on superoxide dismutase and glutathione-S-transferase activities and reproduction of Daphnia magna

    Directory of Open Access Journals (Sweden)

    Song Yuzhi

    2017-03-01

    Full Text Available Applying pesticides to crops is one of the causes of water pollution by surface runoff, and chlorpyrifos, trifluralin and chlorothalonil are used respectively as insecticide, herbicide and fungicide for crop plants widely. To explore effects of three pesticides on aquatic organisms, superoxide dismutase (SOD and glutathione S-transferase (GST activities were determined after 24 h and 48 h exposure of D. magna with ages of 6–24 h to several low concentrations of chlorpyrifos (0.36, 0.72, 1.43, 2.86, 5.72 μg∙L−1, trifluralin (0.17, 0.33, 0.66, 1.33, 2.65 mg∙L−1 and chlorothalonil (0.09, 0.18, 0.36, 0.72, 1.43 mg∙L−1 respectively. Main reproductive parameters including first pregnancy time, first brood time, the number of first brood and total fecundity after 21 d exposures at the same concentrations of pesticides as described above were also measured. The results showed that the activities of GST increased in lower concentrations and decreased in higher concentrations after 24 h exposure to three pesticides, respectively. The activities of SOD showed the same changes after 48 h exposure. With the time prolonged, the activities of GST decreased while the activities of SOD increased. After 21 d exposure, the first pregnancy time and first brood time were delayed, while the number of the first brood and total fecundity per female decreased with increasing concentrations. These results corroborated that GST activity was more sensitive to those pesticides than SOD activity, and there was a significant relationship between total fecundity and pesticides-dose(r>0.94, n=6, GST activity after 48 h exposure and total fecundity after 21 d exposure (r>0.92, n=6.

  15. Are glutathione S transferases involved in DNA damage signalling? Interactions with DNA damage and repair revealed from molecular epidemiology studies

    International Nuclear Information System (INIS)

    Dusinska, Maria; Staruchova, Marta; Horska, Alexandra; Smolkova, Bozena; Collins, Andrew; Bonassi, Stefano; Volkovova, Katarina

    2012-01-01

    Glutathione S-transferases (GSTs) are members of a multigene family of isoenzymes that are important in the control of oxidative stress and in phase II metabolism. Acting non-enzymically, GSTs can modulate signalling pathways of cell proliferation, cell differentiation and apoptosis. Using a molecular epidemiology approach, we have investigated a potential involvement of GSTs in DNA damage processing, specifically the modulation of DNA repair in a group of 388 healthy adult volunteers; 239 with at least 5 years of occupational exposure to asbestos, stone wool or glass fibre, and 149 reference subjects. We measured DNA damage in lymphocytes using the comet assay (alkaline single cell gel electrophoresis): strand breaks (SBs) and alkali-labile sites, oxidised pyrimidines with endonuclease III, and oxidised purines with formamidopyrimidine DNA glycosylase. We also measured GST activity in erythrocytes, and the capacity for base excision repair (BER) in a lymphocyte extract. Polymorphisms in genes encoding three GST isoenzymes were determined, namely deletion of GSTM1 and GSTT1 and single nucleotide polymorphism Ile105Val in GSTP1. Consumption of vegetables and wine correlated negatively with DNA damage and modulated BER. GST activity correlated with oxidised bases and with BER capacity, and differed depending on polymorphisms in GSTP1, GSTT1 and GSTM1. A significantly lower BER rate was associated with the homozygous GSTT1 deletion in all asbestos site subjects and in the corresponding reference group. Multifactorial analysis revealed effects of sex and exposure in GSTP1 Ile/Val heterozygotes but not in Ile/Ile homozygotes. These variants affected also SBs levels, mainly by interactions of GSTP1 genotype with exposure, with sex, and with smoking habit; and by an interaction between sex and smoking. Our results show that GST polymorphisms and GST activity can apparently influence DNA stability and repair of oxidised bases, suggesting a potential new role for these

  16. Overlapping protective roles for glutathione transferase gene family members in chemical and oxidative stress response in Agrobacterium tumefaciens.

    Science.gov (United States)

    Skopelitou, Katholiki; Muleta, Abdi W; Pavli, Ourania; Skaracis, Georgios N; Flemetakis, Emmanouil; Papageorgiou, Anastassios C; Labrou, Nikolaos E

    2012-03-01

    In the present work, we describe the characterisation of the glutathione transferase (GST) gene family from Agrobacterium tumefaciens C58. A genome survey revealed the presence of eight GST-like proteins in A. tumefaciens (AtuGSTs). Comparison by multiple sequence alignment generated a dendrogram revealing the phylogenetic relationships of AtuGSTs-like proteins. The beta and theta classes identified in other bacterial species are represented by five members in A. tumefaciens C58. In addition, there are three "orphan" sequences that do not fit into any previously recognised GST classes. The eight GST-like genes were cloned, expressed in Escherichia coli and their substrate specificity was determined towards 17 different substrates. The results showed that AtuGSTs catalyse a broad range of reactions, with different members of the family exhibiting quite varied substrate specificity. The 3D structures of AtuGSTs were predicted using molecular modelling. The use of comparative sequence and structural analysis of the AtuGST isoenzymes allowed us to identify local sequence and structural characteristics between different GST isoenzymes and classes. Gene expression profiling was conducted under normal culture conditions as well as under abiotic stress conditions (addition of xenobiotics, osmotic stress and cold and heat shock) to induce and monitor early stress-response mechanisms. The results reveal the constitutive expression of GSTs in A. tumefaciens and a modulation of GST activity after treatments, indicating that AtuGSTs presumably participate in a wide range of functions, many of which are important in counteracting stress conditions. These functions may be relevant to maintaining cellular homeostasis as well as in the direct detoxification of toxic compounds.

  17. Cys-X scanning for expansion of active-site residues and modulation of catalytic functions in a glutathione transferase.

    Science.gov (United States)

    Norrgård, Malena A; Hellman, Ulf; Mannervik, Bengt

    2011-05-13

    We propose Cys-X scanning as a semisynthetic approach to engineer the functional properties of recombinant proteins. As in the case of Ala scanning, key residues in the primary structure are identified, and one of them is replaced by Cys via site-directed mutagenesis. The thiol of the residue introduced is subsequently modified by alternative chemical reagents to yield diverse Cys-X mutants of the protein. This chemical approach is orthogonal to Ala or Cys scanning and allows the expansion of the repertoire of amino acid side chains far beyond those present in natural proteins. In its present application, we have introduced Cys-X residues in human glutathione transferase (GST) M2-2, replacing Met-212 in the substrate-binding site. To achieve selectivity of the modifications, the Cys residues in the wild-type enzyme were replaced by Ala. A suite of simple substitutions resulted in a set of homologous Met derivatives ranging from normethionine to S-heptyl-cysteine. The chemical modifications were validated by HPLC and mass spectrometry. The derivatized mutant enzymes were assayed with alternative GST substrates representing diverse chemical reactions: aromatic substitution, epoxide opening, transnitrosylation, and addition to an ortho-quinone. The Cys substitutions had different effects on the alternative substrates and differentially enhanced or suppressed catalytic activities depending on both the Cys-X substitution and the substrate assayed. As a consequence, the enzyme specificity profile could be changed among the alternative substrates. The procedure lends itself to large-scale production of Cys-X modified protein variants.

  18. Correlation between serum and peritoneal fluid glutathione S-transferases T1 concentration with different stages of endometriosis

    Directory of Open Access Journals (Sweden)

    Sohail Mashayekhi

    2018-03-01

    Full Text Available Endometriosis is a gynecological disease defined by the histological presence of endometrial glands and stroma outside the uterine cavity. Ectopic endometrial cell proliferation and chronic inflammation in endometriosis were shown to be associated with oxidative stress (OS induction. OS is a condition in which reactive oxygen species (ROS overproduction and antioxidant deficiency cause a shift in oxidant/antioxidant balance. Glutathione S-transferases (GSTs comprise a family of eukaryotic and prokaryotic phase II metabolic isozymes best known for their ability to catalyze the conjugation of the reduced form of glutathione (GSH to xenobiotic substrates for the purpose of detoxification. The aim of this project was to study the concentrations of GSTT1 in the serum and peritoneal fluid (PF of patients with different stages of endometriosis. Frothy two PF and serum from normal and 152 from different stages of patients with endometriosis (stage I: n = 30, stage II: n = 39, stage III: n = 43 and stage IV: n = 40 were included in this study. The level of GSTT1 in the serum was determined by enzyme linked immunosorbent assay (ELISA. The results showed the presence of GSTT1 in all serum and peritoneal fluid samples, while, starting from stages I to IV endometriosis, a significant decrease in GSTT1 concentration was seen as compared to controls. It is concluded that levels of GSTT1 is negatively correlated with advanced stages of endometriosis. It is also suggested that the detection of serum and/or peritoneal fluid GSTT1 concentration may be valuable in the classifying of endometriosis.

  19. SILENCING THE NUCLEOCYTOPLASMIC O-GLCNAC TRANSFERASE REDUCES PROLIFERATION, ADHESION AND MIGRATION OF CANCER AND FETAL HUMAN COLON CELL LINES

    Directory of Open Access Journals (Sweden)

    AGATA eSTEENACKERS

    2016-05-01

    Full Text Available The post-translational modification of proteins by O-linked β-N-acetylglucosamine (O-GlcNAc is regulated by a unique couple of enzymes. O-GlcNAc transferase (OGT transfers the GlcNAc residue from UDP-GlcNAc, the final product of the hexosamine biosynthetic pathway (HBP, whereas O-GlcNAcase (OGA removes it. This study and others show that OGT and O-GlcNAcylation levels are increased in cancer cell lines. In that context we studied the effect of OGT silencing in the colon cancer cell lines HT29 and HCT116 and the primary colon cell line CCD841CoN. Herein we report that OGT silencing diminished proliferation, in vitro cell survival and adhesion of primary and cancer cell lines. SiOGT dramatically de-creased HT29 and CCD841CoN migration, CCD841CoN harboring high capabilities of mi-gration in Boyden chamber system when compared to HT29 and HCT116. The expression levels of actin and tubulin were unaffected by OGT knockdown but siOGT seemed to disor-ganize microfilament, microtubule and vinculin networks in CCD841CoN. While cancer cell lines harbor higher levels of OGT and O-GlcNAcylation to fulfill their proliferative and migra-tory properties, in agreement with their higher consumption of HBP main substrates glucose and glutamine, our data demonstrate that OGT expression is not only necessary for the biolog-ical properties of cancer cell lines but also for normal cells.

  20. Novel function of glutathione transferase in rat liver mitochondrial membrane: Role for cytochrome c release from mitochondria

    International Nuclear Information System (INIS)

    Lee, Kang Kwang; Shimoji, Manami; Hossain, Quazi Sohel; Sunakawa, Hajime; Aniya, Yoko

    2008-01-01

    Microsomal glutathione transferase (MGST1) is activated by oxidative stress. Although MGST1 is found in mitochondrial membranes (mtMGST1), there is no information about the oxidative activation of mtMGST1. In the present study, we aimed to determine whether mtMGST1 also undergoes activation and about its function. When rats were treated with galactosamine/lipopolysaccharide (GalN/LPS), mtMGST1 activity was significantly increased, and the increased activity was reduced by the disulfide reducing agent dithiothreitol. In mitochondria from GalN/LPS-treated rats, disulfide-linked mtMGST1 dimer and mixed protein glutathione disulfides (glutathionylation) were detected. In addition, cytochrome c release from mitochondria isolated from GalN/LPS-treated rats was observed, and the release was inhibited by anti-MGST1 antibodies. Incubation of mitochondria from control rats with diamide and diamide plus GSH in vitro resulted in dimer- and mixed disulfide bond-mediated activation of mtMGST1, respectively. The activation of mtMGST1 by diamide plus GSH caused cytochrome c release from the mitochondria, and the release was prevented by treatment with anti-MGST1 antibodies. In addition, diamide plus GSH treatment caused mitochondrial swelling accompanied by cytochrome c release, which was inhibited by cyclosporin A (CsA) and bongkrekic acid (BKA), inhibitors of the mitochondrial permeability transition (MPT) pore. Furthermore, mtMGST1 activity was also inhibited by CsA and BKA. These results indicate that mtMGST1 is activated through mixed disulfide bond formation that contributes to cytochrome c release from mitochondria through the MPT pore

  1. Microsomal Glutathione Transferase 1 Protects Against Toxicity Induced by Silica Nanoparticles but Not by Zinc Oxide Nanoparticles

    Science.gov (United States)

    2012-01-01

    Microsomal glutathione transferase 1 (MGST1) is an antioxidant enzyme located predominantly in the mitochondrial outer membrane and endoplasmic reticulum and has been shown to protect cells from lipid peroxidation induced by a variety of cytostatic drugs and pro-oxidant stimuli. We hypothesized that MGST1 may also protect against nanomaterial-induced cytotoxicity through a specific effect on lipid peroxidation. We evaluated the induction of cytotoxicity and oxidative stress by TiO2, CeO2, SiO2, and ZnO in the human MCF-7 cell line with or without overexpression of MGST1. SiO2 and ZnO nanoparticles caused dose- and time-dependent toxicity, whereas no obvious cytotoxic effects were induced by nanoparticles of TiO2 and CeO2. We also noted pronounced cytotoxicity for three out of four additional SiO2 nanoparticles tested. Overexpression of MGST1 reversed the cytotoxicity of the main SiO2 nanoparticles tested and for one of the supplementary SiO2 nanoparticles but did not protect cells against ZnO-induced cytotoxic effects. The data point toward a role of lipid peroxidation in SiO2 nanoparticle-induced cell death. For ZnO nanoparticles, rapid dissolution was observed, and the subsequent interaction of Zn2+ with cellular targets is likely to contribute to the cytotoxic effects. A direct inhibition of MGST1 by Zn2+ could provide a possible explanation for the lack of protection against ZnO nanoparticles in this model. Our data also showed that SiO2 nanoparticle-induced cytotoxicity is mitigated in the presence of serum, potentially through masking of reactive surface groups by serum proteins, whereas ZnO nanoparticles were cytotoxic both in the presence and in the absence of serum. PMID:22303956

  2. Molecular cloning, heterologous expression and functional characterization of gamma tocopherol methyl transferase (γ-TMT) from Glycine max.

    Science.gov (United States)

    Tewari, Kalpana; Dahuja, Anil; Sachdev, Archana; Kumar, Vaibhav; Ali, Kishwar; Kumar, Amresh; Kumari, Sweta

    2017-12-01

    γ-Tocopherol methyltransferase (γ-TMT) (EC 2.1.1.95) is the last enzyme in the tocopherol biosynthetic pathway and it catalyzes the conversion of γ-tocopherol into α-tocopherol, the nutritionally significant and most bioactive form of vitamin E. Although the γ-TMT gene has been successfully overexpressed in many crops to enhance their α-tocopherol content but still only few attempts have been made to uncover its structural, functional and regulation aspects at protein level. In this study, we have cloned the complete 909bp coding sequence of Glycine max γ-TMT (Gm γ-TMT) gene that encodes the corresponding protein comprising of 302 amino acid residues. The deduced Gm γ-TMT protein showed 74-87% sequence identity with other characterized plant γ-TMTs. Gm γ-TMT belongs to Class I Methyl Transferases that have a Rossmann-like fold which consists of a seven-stranded β sheet joined by α helices. Heterologous expression of Gm γ-TMT in pET29a expression vector under the control of bacteriophage T7 promoter produced a 37.9 kDa recombinant Gm γ-TMT protein with histidine hexamer tag at its C-terminus. The expression of recombinant Gm γ-TMT protein was confirmed by western blotting using anti-His antibody. The recombinant protein was purified by Ni 2+ -NTA column chromatography. The purified protein showed SAM dependent methyltransferase activity. The α-tocopherol produced in the in-vitro reaction catalyzed by the purified enzyme was detected using reverse phase HPLC. This study has laid the foundation to unveil the biochemical understanding of Gm γ-TMT enzyme which can be further explored by studying its kinetic behaviour, substrate specificity and its interaction with other biomolecules. Copyright © 2017 Elsevier Inc. All rights reserved.

  3. Effect of recombinant glutathione S-transferase as vaccine antigen against Rhipicephalus appendiculatus and Rhipicephalus sanguineus infestation.

    Science.gov (United States)

    Sabadin, Gabriela Alves; Parizi, Luís Fernando; Kiio, Irene; Xavier, Marina Amaral; da Silva Matos, Renata; Camargo-Mathias, Maria Izabel; Githaka, Naftaly Wang'ombe; Nene, Vish; da Silva Vaz, Itabajara

    2017-12-04

    The ticks Rhipicephalus appendiculatus and Rhipicephalus sanguineus are the main vectors of Theileria parva and Babesia spp. in cattle and dogs, respectively. Due to their impact in veterinary care and industry, improved methods against R. appendiculatus and R. sanguineus parasitism are under development, including vaccines. We have previously demonstrated the induction of a cross-protective humoral response against Rhipicephalus microplus following vaccination with recombinant glutathione S-transferase from Haemaphysalis longicornis tick (rGST-Hl), suggesting that this protein could control tick infestations. In the present work, we investigated the effect of rGST-Hl vaccine against R. appendiculatus and R. sanguineus infestation in rabbits. In silico analysis revealed that GST from H. longicornis, R. appendiculatus and R. sanguineus have >80% protein sequence similarity, and multiple conserved antigenic sites. After the second vaccine dose, rGST-Hl-immunized rabbits showed elevated antibody levels which persisted until the end of experiment (75 and 60 days for R. appendiculatus and R. sanguineus, respectively). Western blot assays demonstrated cross-reactivity between anti-rGST-Hl antibodies and native R. appendiculatus and R. sanguineus GST extracts from ticks at different life stages. Vaccination with rGST-Hl decreased the number, weight, and fertility of engorged R. appendiculatus adults, leading to an overall vaccine efficacy of 67%. Interestingly, histological analysis of organ morphology showed damage to salivary glands and ovaries of R. appendiculatus adult females fed on vaccinated animals. In contrast, rGST-Hl vaccination did not affect R. appendiculatus nymphs, and it was ineffective against R. sanguineus across the stages of nymph and adult. Taken together, our results show the potential application of rGST-Hl as an antigen in anti-tick vaccine development, however indicating a broad difference in efficacy among tick species. Copyright © 2017 Elsevier

  4. Human glutathione S-transferase P1-1 functions as an estrogen receptor α signaling modulator

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Xiyuan [Department of Biological Science, Sookmyung Women’s University, Seoul (Korea, Republic of); An, Byoung Ha [Department of Food and Nutrition, College of Life Science, Sookmyung Women’s University, Seoul (Korea, Republic of); Kim, Min Jung; Park, Jong Hoon [Department of Biological Science, Sookmyung Women’s University, Seoul (Korea, Republic of); Kang, Young Sook [Department of Pharmacy, College of Pharmacy, Sookmyung Women’s University, Seoul (Korea, Republic of); Chang, Minsun, E-mail: minsunchang@sm.ac.kr [Department of Medical and Pharmaceutical Science, College of Science, Sookmyung Women’s University, Seoul (Korea, Republic of)

    2014-09-26

    Highlights: • GSTP induces the classical ERα signaling event. • The functional GSTP is a prerequisite for GSTP-induced ERα transcription activity. • The expression of RIP140, a transcription cofactor, was inhibited by GSTP protein. • We propose the novel non-enzymatic role of GSTP. - Abstract: Estrogen receptor α (ERα) plays a crucial role in estrogen-mediated signaling pathways and exerts its action as a nuclear transcription factor. Binding of the ligand-activated ERα to the estrogen response element (ERE) is a central part of ERα-associated signal transduction pathways and its aberrant modulation is associated with many disease conditions. Human glutathione S-transferase P1-1 (GSTP) functions as an enzyme in conjugation reactions in drug metabolism and as a regulator of kinase signaling pathways. It is overexpressed in tumors following chemotherapy and has been associated with a poor prognosis in breast cancer. In this study, a novel regulatory function of GSTP has been proposed in which GSTP modulates ERE-mediated ERα signaling events. Ectopic expression of GSTP was able to induce the ERα and ERE-mediated transcriptional activities in ERα-positive but GSTP-negative MCF7 human breast cancer cells. This inductive effect of GSTP on the ERE-transcription activity was diminished when the cells express a mutated form of the enzyme or are treated with a GSTP-specific chemical inhibitor. It was found that GSTP inhibited the expression of the receptor interacting protein 140 (RIP140), a negative regulator of ERα transcription, at both mRNA and protein levels. Our study suggests a novel non-enzymatic role of GSTP which plays a significant role in regulating the classical ERα signaling pathways via modification of transcription cofactors such as RIP140.

  5. Interaction of Ferulic Acid with Glutathione S-Transferase and Carboxylesterase Genes in the Brown Planthopper, Nilaparvata lugens.

    Science.gov (United States)

    Yang, Jun; Sun, Xiao-Qin; Yan, Shu-Ying; Pan, Wen-Jun; Zhang, Mao-Xin; Cai, Qing-Nian

    2017-07-01

    Plant phenolics are crucial defense phytochemicals against herbivores and glutathione S-transferase (GST) and carboxylesterase (CarE) in herbivorous insects are well-known detoxification enzymes for such xenobiotics. To understand relationship between a plant phenolic and herbivore GST or CarE genes, we evaluated the relationship between a rice phenolic ferulic acid and resistance to brown planthopper (BPH, Nilaparvata lugens), and investigated the interaction of ferulic acid with GST or CarE genes in BPH. The results indicate that ferulic acid content in tested rice varieties was highly associated with resistance to BPH. Bioassays using artificial diets show that the phenolic acid toxicity to BPH was dose dependent and the LC 25 and LC 50 were 5.81 and 23.30 μg/ml at 72 hr, respectively. Activities of the enzymes BPH GST and CarE were increased at concentrations below the LC 50 of ferulic acid. Moreover, low ferulic acid concentrations (gene silencing (DIGS) of GST or CarE, it was shown that suppressed expression levels of NlGSTD1, NlGSTE1 and NlCE were 14.6%-21.2%, 27.8%-34.2%, and 10.5%-19.8%, respectively. Combination of NlGSTD1, NlGSTE1 or NlCE knockdown with ferulic acid increased nymph mortality by 92.9%, 119.9%, or 124.6%, respectively. These results suggest that depletion of detoxification genes in herbivorous insects by plant-mediated RNAi technology might be a new potential resource for improving rice resistance to BPH.

  6. Systemic catechol-O-methyl transferase inhibition enables the D1 agonist radiotracer R-[11C]SKF 82957

    International Nuclear Information System (INIS)

    Palner, Mikael; McCormick, Patrick; Parkes, Jun; Knudsen, Gitte M.; Wilson, Alan A.

    2010-01-01

    Introduction: R-[ 11 C]-SKF 82957 is a high-affinity and potent dopamine D 1 receptor agonist radioligand, which gives rise to a brain-penetrant lipophilic metabolite. In this study, we demonstrate that systemic administration of catechol-O-methyl transferase (COMT) inhibitors blocks this metabolic pathway, facilitating the use of R-[ 11 C]-SKF 82957 to image the high-affinity state of the dopamine D 1 receptor with PET. Methods: R-[ 11 C]SKF 82957 was administered to untreated and COMT inhibitor-treated conscious rats, and the radioactive metabolites present in the brain and plasma were quantified by HPLC. Under optimal conditions, cerebral uptake and dopamine D 1 binding of R-[ 11 C]SKF 82957 were measured ex vivo. In addition, pharmacological challenges with the receptor antagonist SCH 23390, amphetamine, the dopamine reuptake inhibitor RTI-32 and the dopamine hydroxylase inhibitor α-methyl-p-tyrosine were performed to study the specificity and sensitivity of R-[ 11 C]-SKF 82957 dopamine D 1 binding in COMT-inhibited animals. Results: Treatment with the COMT inhibitor tolcapone was associated with a dose-dependent (EC 90 5.3±4.3 mg/kg) reduction in the lipophilic metabolite. Tolcapone treatment (20 mg/kg) also resulted in a significant increase in the striatum/cerebellum ratio of R-[ 11 C]SKF 82957, from 15 (controls) to 24. Treatment with the dopamine D 1 antagonist SCH 23390 reduced the striatal binding to the levels of the cerebellum, demonstrating a high specificity and selectivity of R-[ 11 C]SKF 82957 binding. Conclusions: Pre-treatment with the COMT inhibitor tolcapone inhibits formation of an interfering metabolite of R-[ 11 C]SKF 82957. Under such conditions, R-[ 11 C]SKF 82957 demonstrates high potential as the first agonist radiotracer for imaging the dopamine D 1 receptor by PET.

  7. Growth hormone alters the glutathione S-transferase and mitochondrial thioredoxin systems in long-living Ames dwarf mice.

    Science.gov (United States)

    Rojanathammanee, Lalida; Rakoczy, Sharlene; Brown-Borg, Holly M

    2014-10-01

    Ames dwarf mice are deficient in growth hormone (GH), prolactin, and thyroid-stimulating hormone and live significantly longer than their wild-type (WT) siblings. The lack of GH is associated with stress resistance and increased longevity. However, the mechanism underlying GH's actions on cellular stress defense have yet to be elucidated. In this study, WT or Ames dwarf mice were treated with saline or GH (WT saline, Dwarf saline, and Dwarf GH) two times daily for 7 days. The body and liver weights of Ames dwarf mice were significantly increased after 7 days of GH administration. Mitochondrial protein levels of the glutathione S-transferase (GST) isozymes, K1 and M4 (GSTK1 and GSTM4), were significantly higher in dwarf mice (Dwarf saline) when compared with WT mice (WT saline). GH administration downregulated the expression of GSTK1 proteins in dwarf mice. We further investigated GST activity from liver lysates using different substrates. Substrate-specific GST activity (bromosulfophthalein, dichloronitrobenzene, and 4-hydrox-ynonenal) was significantly reduced in GH-treated dwarf mice. In addition, GH treatment attenuated the activity of thioredoxin and glutaredoxin in liver mitochondria of Ames mice. Importantly, GH treatment suppressed Trx2 and TrxR2 mRNA expression. These data indicate that GH has a role in stress resistance by altering the functional capacity of the GST system through the regulation of specific GST family members in long-living Ames dwarf mice. It also affects the regulation of thioredoxin and glutaredoxin, factors that regulate posttranslational modification of proteins and redox balance, thereby further influencing stress resistance. © The Author 2013. Published by Oxford University Press on behalf of The Gerontological Society of America. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  8. Large-scale determination of sequence, structure, and function relationships in cytosolic glutathione transferases across the biosphere.

    Science.gov (United States)

    Mashiyama, Susan T; Malabanan, M Merced; Akiva, Eyal; Bhosle, Rahul; Branch, Megan C; Hillerich, Brandan; Jagessar, Kevin; Kim, Jungwook; Patskovsky, Yury; Seidel, Ronald D; Stead, Mark; Toro, Rafael; Vetting, Matthew W; Almo, Steven C; Armstrong, Richard N; Babbitt, Patricia C

    2014-04-01

    The cytosolic glutathione transferase (cytGST) superfamily comprises more than 13,000 nonredundant sequences found throughout the biosphere. Their key roles in metabolism and defense against oxidative damage have led to thousands of studies over several decades. Despite this attention, little is known about the physiological reactions they catalyze and most of the substrates used to assay cytGSTs are synthetic compounds. A deeper understanding of relationships across the superfamily could provide new clues about their functions. To establish a foundation for expanded classification of cytGSTs, we generated similarity-based subgroupings for the entire superfamily. Using the resulting sequence similarity networks, we chose targets that broadly covered unknown functions and report here experimental results confirming GST-like activity for 82 of them, along with 37 new 3D structures determined for 27 targets. These new data, along with experimentally known GST reactions and structures reported in the literature, were painted onto the networks to generate a global view of their sequence-structure-function relationships. The results show how proteins of both known and unknown function relate to each other across the entire superfamily and reveal that the great majority of cytGSTs have not been experimentally characterized or annotated by canonical class. A mapping of taxonomic classes across the superfamily indicates that many taxa are represented in each subgroup and highlights challenges for classification of superfamily sequences into functionally relevant classes. Experimental determination of disulfide bond reductase activity in many diverse subgroups illustrate a theme common for many reaction types. Finally, sequence comparison between an enzyme that catalyzes a reductive dechlorination reaction relevant to bioremediation efforts with some of its closest homologs reveals differences among them likely to be associated with evolution of this unusual reaction

  9. Large-scale determination of sequence, structure, and function relationships in cytosolic glutathione transferases across the biosphere.

    Directory of Open Access Journals (Sweden)

    Susan T Mashiyama

    2014-04-01

    Full Text Available The cytosolic glutathione transferase (cytGST superfamily comprises more than 13,000 nonredundant sequences found throughout the biosphere. Their key roles in metabolism and defense against oxidative damage have led to thousands of studies over several decades. Despite this attention, little is known about the physiological reactions they catalyze and most of the substrates used to assay cytGSTs are synthetic compounds. A deeper understanding of relationships across the superfamily could provide new clues about their functions. To establish a foundation for expanded classification of cytGSTs, we generated similarity-based subgroupings for the entire superfamily. Using the resulting sequence similarity networks, we chose targets that broadly covered unknown functions and report here experimental results confirming GST-like activity for 82 of them, along with 37 new 3D structures determined for 27 targets. These new data, along with experimentally known GST reactions and structures reported in the literature, were painted onto the networks to generate a global view of their sequence-structure-function relationships. The results show how proteins of both known and unknown function relate to each other across the entire superfamily and reveal that the great majority of cytGSTs have not been experimentally characterized or annotated by canonical class. A mapping of taxonomic classes across the superfamily indicates that many taxa are represented in each subgroup and highlights challenges for classification of superfamily sequences into functionally relevant classes. Experimental determination of disulfide bond reductase activity in many diverse subgroups illustrate a theme common for many reaction types. Finally, sequence comparison between an enzyme that catalyzes a reductive dechlorination reaction relevant to bioremediation efforts with some of its closest homologs reveals differences among them likely to be associated with evolution of this

  10. Role of induced glutathione-S-transferase from Helicoverpa armigera (Lepidoptera: Noctuidae) HaGST-8 in detoxification of pesticides.

    Science.gov (United States)

    Labade, Chaitali P; Jadhav, Abhilash R; Ahire, Mehul; Zinjarde, Smita S; Tamhane, Vaijayanti A

    2018-01-01

    The present study deals with glutathione-S-transferase (GST) based detoxification of pesticides in Helicoverpa armigera and its potential application in eliminating pesticides from the environment. Dietary exposure of a pesticide mixture (organophosphates - chlorpyrifos and dichlorvos, pyrethroid - cypermethrin; 2-15ppm each) to H. armigera larvae resulted in a dose dependant up-regulation of GST activity and gene expression. A variant GST from H. armigera (HaGST-8) was isolated from larvae fed with 10ppm pesticide mixture and it was recombinantly expressed in yeast (Pichia pastoris HaGST-8). HaGST-8 had a molecular mass of 29kDa and was most active at pH 9 at 30°C. GC-MS and LC-HRMS analysis validated that HaGST-8 was effective in eliminating organophosphate type of pesticides and partially reduced the cypermethrin content (53%) from aqueous solutions. Unlike the untransformed yeast, P. pastoris HaGST-8 grew efficiently in media supplemented with pesticide mixtures (200 and 400ppm each pesticide) signifying the detoxification ability of HaGST-8. The amino acid sequence of HaGST-8 and the already reported sequence of HaGST-7 had just 2 mismatches. The studies on molecular interaction strengths revealed that HaGST-8 had stronger binding affinities with organophosphate, pyrethroid, organochloride, carbamate and neonicotinoid type of pesticides. The abilities of recombinant HaGST-8 to eliminate pesticides and P. pastoris HaGST-8 to grow profusely in the presence of high level of pesticide content can be applied for removal of such residues from food, water resources and bioremediation. Copyright © 2017 Elsevier Inc. All rights reserved.

  11. Contribution of Arginine 13 to the Catalytic Activity of Human Class Pi Glutathione Transferase P1-1

    International Nuclear Information System (INIS)

    Kong, Ji Na; Jo, Dong Hyeon; Do, Hyun Dong; Lee, Jin Ju; Kong, Kwang Hoon

    2010-01-01

    Arg13 is a conserved active-site residue in all known Pi class glutathione S-transferases (GSTs) and in most Alpha class GSTs. To evaluate its contribution to substrate binding and catalysis of this residue, three mutants (R13A, R13K, and R13L) were expressed in Escherichia coli and purified by GSH affinity chromatography. The substitutions of Arg13 significantly affected GSH-conjugation activity, while scarcely affecting glutathione peroxidase or steroid isomerase activities. Mutation of Arg13 into Ala largely reduced the GSH-conjugation activity by approximately 85 - 95%, whereas substitutions by Lys and Leu barely affected activity. These results suggest that, in the GSH-conjugation activity of hGST P1-1, the contribution of Arg13 toward catalytic activity is highly dependent on substrate specificities and the size of the side chain at position 13. From the kinetic parameters, introduction of larger side chains at position 13 results in stronger affinity (Leu > Lys, Arg > Ala) towards GSH. The substitutions of Arg13 with alanine and leucine significantly affected k cat , whereas substitution with Lys was similar to that of the wild type, indicating the significance of a positively charged residue at position 13. From the plots of log (k cat /K m CDNB ) against pH, the pK a values of the thiol group of GSH bound in R13A, R13K, and R13L were estimated to be 1.8, 1.4, and 1.8 pK units higher than the pK a value of the wildtype enzyme, demonstrating the contribution of the Arg13 guanidinium group to the electrostatic field in the active site. From these results, we suggest that contribution of Arg13 in substrate binding is highly dependent on the nature of the electrophilic substrates, while in the catalytic mechanism, it stabilizes the GSH thiolate through hydrogen bonding

  12. Are glutathione S-transferase polymorphisms (GSTM1, GSTT1) associated with primary open angle glaucoma? A meta-analysis.

    Science.gov (United States)

    Lu, Yan; Shi, Yuhua; Yin, Jie; Huang, Zhenping

    2013-09-15

    Glutathione S-transferase (GST) variants have been considered as risk factors for the pathogenesis of primary open angle glaucoma (POAG). However, the results have been inconsistent. In this study, we performed a meta-analysis to assess the association between GSTM1 and GSTT1 null genotypes and the risk for POAG. Published literature from PubMed and EMBASE databases was retrieved. All studies evaluating the association between GSTM1/GSTT1 variants and POAG were included. Pooled odds ratio (OR) and 95% confidence interval (CI) were calculated using fixed- or random-effects model. 14 studies (1711 POAG cases and 1537 controls) were included in the meta-analysis of GSTM1 genotypes and 10 studies (1306 POAG cases and 1114 controls) were included in the meta-analysis of GSTT1 genotypes. The overall result showed that the association between GSTM1 and GSTT1 null genotypes and risk for POAG was not statistically significant (GSTM1: OR=1.19, 95% CI=0.82-1.73, p=0.361; GSTT1: OR=1.26, 95% CI=0.77-2.06, p=0.365). The results by ethnicity showed that the association between the GSTM1 null genotype and risk for POAG is statistically significant in East Asians (OR=1.41, 95% CI=1.04-1.90, p=0.026), but not in Caucasians (OR=1.13, 95% CI=0.69-1.84, p=0.638) and Latin-American (OR=1.09, 95% CI=0.62-1.92, p=0.767). In addition, there was no significant association of GSTT1 null genotype with risk for POAG in either ethnic population. The present meta-analysis suggested that there might be a significant association of GSTM1 null genotype with POAG risk in East Asians. © 2013 Elsevier B.V. All rights reserved.

  13. Effects of high-intensity intermittent training on carnitine palmitoyl transferase activity in the gastrocnemius muscle of rats

    Directory of Open Access Journals (Sweden)

    L.C. Carnevali Jr

    2012-08-01

    Full Text Available We examined the capacity of high-intensity intermittent training (HI-IT to facilitate the delivery of lipids to enzymes responsible for oxidation, a task performed by the carnitine palmitoyl transferase (CPT system in the rat gastrocnemius muscle. Male adult Wistar rats (160-250 g were randomly distributed into 3 groups: sedentary (Sed, N = 5, HI-IT (N = 10, and moderate-intensity continuous training (MI-CT, N = 10. The trained groups were exercised for 8 weeks with a 10% (HI-IT and a 5% (MI-CT overload. The HI-IT group presented 11.8% decreased weight gain compared to the Sed group. The maximal activities of CPT-I, CPT-II, and citrate synthase were all increased in the HI-IT group compared to the Sed group (P < 0.01, as also was gene expression, measured by RT-PCR, of fatty acid binding protein (FABP; P < 0.01 and lipoprotein lipase (LPL; P < 0.05. Lactate dehydrogenase also presented a higher maximal activity (nmol·min-1·mg protein-1 in HI-IT (around 83%. We suggest that 8 weeks of HI-IT enhance mitochondrial lipid transport capacity thus facilitating the oxidation process in the gastrocnemius muscle. This adaptation may also be associated with the decrease in weight gain observed in the animals and was concomitant to a higher gene expression of both FABP and LPL in HI-IT, suggesting that intermittent exercise is a "time-efficient" strategy inducing metabolic adaptation.

  14. Effects of high-intensity intermittent training on carnitine palmitoyl transferase activity in the gastrocnemius muscle of rats

    Energy Technology Data Exchange (ETDEWEB)

    Carnevali, L.C. Jr. [Grupo de Biologia Molecular da Célula, Instituto de Ciências Biomédicas, Universidade de São Paulo, São Paulo SP (Brazil); Centro Universitário Ítalo-Brasileiro (Unítalo), São Paulo SP (Brazil); Eder, R.; Lira, F.S. [Grupo de Biologia Molecular da Célula, Instituto de Ciências Biomédicas, Universidade de São Paulo, São Paulo SP (Brazil); Lima, W.P. [Grupo de Biologia Molecular da Célula, Instituto de Ciências Biomédicas, Universidade de São Paulo, São Paulo SP (Brazil); Instituto Federal de Educação,Ciência e Tecnologia de São Paulo, São Paulo SP (Brazil); Gonçalves, D.C. [Grupo de Biologia Molecular da Célula, Instituto de Ciências Biomédicas, Universidade de São Paulo, São Paulo SP (Brazil); Zanchi, N.E. [Laboratorio de Nutrição e Metabolismo Aplicado à Atividade Motora, Escola de Educação Física e Esporte, Universidade de São Paulo, São Paulo SP (Brazil); Centro de Pesquisa do Genoma Humano, Instituto de Ciências Biomédicas, Universidade de São Paulo, São Paulo SP (Brazil); Nicastro, H. [Laboratorio de Nutrição e Metabolismo Aplicado à Atividade Motora, Escola de Educação Física e Esporte, Universidade de São Paulo, São Paulo SP (Brazil); Lavoie, J.M. [Department of Kinesiology, University of Montreal, Montreal (Canada); Seelaender, M.C.L. [Grupo de Biologia Molecular da Célula, Instituto de Ciências Biomédicas, Universidade de São Paulo, São Paulo SP (Brazil)

    2012-06-29

    We examined the capacity of high-intensity intermittent training (HI-IT) to facilitate the delivery of lipids to enzymes responsible for oxidation, a task performed by the carnitine palmitoyl transferase (CPT) system in the rat gastrocnemius muscle. Male adult Wistar rats (160-250 g) were randomly distributed into 3 groups: sedentary (Sed, N = 5), HI-IT (N = 10), and moderate-intensity continuous training (MI-CT, N = 10). The trained groups were exercised for 8 weeks with a 10% (HI-IT) and a 5% (MI-CT) overload. The HI-IT group presented 11.8% decreased weight gain compared to the Sed group. The maximal activities of CPT-I, CPT-II, and citrate synthase were all increased in the HI-IT group compared to the Sed group (P < 0.01), as also was gene expression, measured by RT-PCR, of fatty acid binding protein (FABP; P < 0.01) and lipoprotein lipase (LPL; P < 0.05). Lactate dehydrogenase also presented a higher maximal activity (nmol·min{sup −1}·mg protein{sup −1}) in HI-IT (around 83%). We suggest that 8 weeks of HI-IT enhance mitochondrial lipid transport capacity thus facilitating the oxidation process in the gastrocnemius muscle. This adaptation may also be associated with the decrease in weight gain observed in the animals and was concomitant to a higher gene expression of both FABP and LPL in HI-IT, suggesting that intermittent exercise is a “time-efficient” strategy inducing metabolic adaptation.

  15. Comparative study of acetylcholinesterase and glutathione S-transferase activities of closely related cave and surface Asellus aquaticus (Isopoda: Crustacea.

    Directory of Open Access Journals (Sweden)

    Anita Jemec

    Full Text Available The freshwater isopod crustacean Asellus aquaticus has recently been developed as an emerging invertebrate cave model for studying evolutionary and developmental biology. Mostly morphological and genetic differences between cave and surface A. aquaticus populations have been described up to now, while scarce data are available on other aspects, including physiology. The purpose of this study was to advance our understanding of the physiological differences between cave A. aquaticus and its surface-dwelling counterparts. We sampled two surface populations from the surface section of the sinking Pivka River (central Slovenia, Europe, i.e. locality Pivka Polje, and locality Planina Polje, and one cave population from the subterranean section of the sinking Pivka River, i.e. locality Planina Cave. Animals were sampled in spring, summer and autumn. We measured the activities of acetylcholinesterase (AChE and glutathione S-transferase (GST in individuals snap-frozen in the field immediately after collection. Acetylcholinesterase is likely related to animals' locomotor activity, while GST activity is related to the metabolic activity of an organism. Our study shows significantly lower AChE and GST activities in the cave population in comparison to both surface A. aquaticus populations. This confirms the assumption that cave A. aquaticus have lower locomotor and metabolic activity than surface A. aquaticus in their respective natural environments. In surface A. aquaticus populations, seasonal fluctuations in GST activity were observed, while these were less pronounced in individuals from the more stable cave environment. On the other hand, AChE activity was generally season-independent in all populations. To our knowledge, this is the first study of its kind conducted in A. aquaticus. Our results show that among closely related cave and surface A. aquaticus populations also physiological differences are present besides the morphological and genetic

  16. Effects of high-intensity intermittent training on carnitine palmitoyl transferase activity in the gastrocnemius muscle of rats

    International Nuclear Information System (INIS)

    Carnevali, L.C. Jr.; Eder, R.; Lira, F.S.; Lima, W.P.; Gonçalves, D.C.; Zanchi, N.E.; Nicastro, H.; Lavoie, J.M.; Seelaender, M.C.L.

    2012-01-01

    We examined the capacity of high-intensity intermittent training (HI-IT) to facilitate the delivery of lipids to enzymes responsible for oxidation, a task performed by the carnitine palmitoyl transferase (CPT) system in the rat gastrocnemius muscle. Male adult Wistar rats (160-250 g) were randomly distributed into 3 groups: sedentary (Sed, N = 5), HI-IT (N = 10), and moderate-intensity continuous training (MI-CT, N = 10). The trained groups were exercised for 8 weeks with a 10% (HI-IT) and a 5% (MI-CT) overload. The HI-IT group presented 11.8% decreased weight gain compared to the Sed group. The maximal activities of CPT-I, CPT-II, and citrate synthase were all increased in the HI-IT group compared to the Sed group (P < 0.01), as also was gene expression, measured by RT-PCR, of fatty acid binding protein (FABP; P < 0.01) and lipoprotein lipase (LPL; P < 0.05). Lactate dehydrogenase also presented a higher maximal activity (nmol·min −1 ·mg protein −1 ) in HI-IT (around 83%). We suggest that 8 weeks of HI-IT enhance mitochondrial lipid transport capacity thus facilitating the oxidation process in the gastrocnemius muscle. This adaptation may also be associated with the decrease in weight gain observed in the animals and was concomitant to a higher gene expression of both FABP and LPL in HI-IT, suggesting that intermittent exercise is a “time-efficient” strategy inducing metabolic adaptation

  17. Molecular evolution and the role of oxidative stress in the expansion and functional diversification of cytosolic glutathione transferases

    Directory of Open Access Journals (Sweden)

    Vasconcelos Vítor

    2010-09-01

    Full Text Available Abstract Background Cytosolic glutathione transferases (cGST are a large group of ubiquitous enzymes involved in detoxification and are well known for their undesired side effects during chemotherapy. In this work we have performed thorough phylogenetic analyses to understand the various aspects of the evolution and functional diversification of cGSTs. Furthermore, we assessed plausible correlations between gene duplication and substrate specificity of gene paralogs in humans and selected species, notably in mammalian enzymes and their natural substrates. Results We present a molecular phylogeny of cytosolic GSTs that shows that several classes of cGSTs are more ubiquitous and thus have an older ancestry than previously thought. Furthermore, we found that positive selection is implicated in the diversification of cGSTs. The number of duplicate genes per class is generally higher for groups of enzymes that metabolize products of oxidative damage. Conclusions 1 Protection against oxidative stress seems to be the major driver of positive selection in mammalian cGSTs, explaining the overall expansion pattern of this subfamily; 2 Given the functional redundancy of GSTs that metabolize xenobiotic chemicals, we would expect the loss of gene duplicates, but by contrast we observed a gene expansion of this family, which likely has been favored by: i the diversification of endogenous substrates; ii differential tissue expression; and iii increased specificity for a particular molecule; 3 The increased availability of sequence data from diversified taxa is likely to continue to improve our understanding of the early origin of the different cGST classes.

  18. Palmitoylation of the Cysteine Residue in the DHHC Motif of a Palmitoyl Transferase Mediates Ca2+ Homeostasis in Aspergillus.

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    Yuanwei Zhang

    2016-04-01

    Full Text Available Finely tuned changes in cytosolic free calcium ([Ca2+]c mediate numerous intracellular functions resulting in the activation or inactivation of a series of target proteins. Palmitoylation is a reversible post-translational modification involved in membrane protein trafficking between membranes and in their functional modulation. However, studies on the relationship between palmitoylation and calcium signaling have been limited. Here, we demonstrate that the yeast palmitoyl transferase ScAkr1p homolog, AkrA in Aspergillus nidulans, regulates [Ca2+]c homeostasis. Deletion of akrA showed marked defects in hyphal growth and conidiation under low calcium conditions which were similar to the effects of deleting components of the high-affinity calcium uptake system (HACS. The [Ca2+]c dynamics in living cells expressing the calcium reporter aequorin in different akrA mutant backgrounds were defective in their [Ca2+]c responses to high extracellular Ca2+ stress or drugs that cause ER or plasma membrane stress. All of these effects on the [Ca2+]c responses mediated by AkrA were closely associated with the cysteine residue of the AkrA DHHC motif, which is required for palmitoylation by AkrA. Using the acyl-biotin exchange chemistry assay combined with proteomic mass spectrometry, we identified protein substrates palmitoylated by AkrA including two new putative P-type ATPases (Pmc1 and Spf1 homologs, a putative proton V-type proton ATPase (Vma5 homolog and three putative proteins in A. nidulans, the transcripts of which have previously been shown to be induced by extracellular calcium stress in a CrzA-dependent manner. Thus, our findings provide strong evidence that the AkrA protein regulates [Ca2+]c homeostasis by palmitoylating these protein candidates and give new insights the role of palmitoylation in the regulation of calcium-mediated responses to extracellular, ER or plasma membrane stress.

  19. Genetic polymorphisms in glutathione S-transferase (GST) superfamily and arsenic metabolism in residents of the Red River Delta, Vietnam

    International Nuclear Information System (INIS)

    Agusa, Tetsuro; Iwata, Hisato; Fujihara, Junko; Kunito, Takashi; Takeshita, Haruo; Tu Binh Minh; Pham Thi Kim Trang; Pham Hung Viet; Tanabe, Shinsuke

    2010-01-01

    To elucidate the role of genetic factors in arsenic metabolism, we investigated associations of genetic polymorphisms in the members of glutathione S-transferase (GST) superfamily with the arsenic concentrations in hair and urine, and urinary arsenic profile in residents in the Red River Delta, Vietnam. Genotyping was conducted for GST ω1 (GSTO1) Ala140Asp, Glu155del, Glu208Lys, Thr217Asn, and Ala236Val, GST ω2 (GSTO2) Asn142Asp, GST π1 (GSTP1) Ile105Val, GST μ1 (GSTM1) wild/null, and GST θ1 (GSTT1) wild/null. There were no mutation alleles for GSTO1 Glu208Lys, Thr217Asn, and Ala236Val in this population. GSTO1 Glu155del hetero type showed higher urinary concentration of As V than the wild homo type. Higher percentage of DMA V in urine of GSTM1 wild type was observed compared with that of the null type. Strong correlations between GSTP1 Ile105Val and arsenic exposure level and profile were observed in this study. Especially, heterozygote of GSTP1 Ile105Val had a higher metabolic capacity from inorganic arsenic to monomethyl arsenic, while the opposite trend was observed for ability of metabolism from As V to As III . Furthermore, other factors including sex, age, body mass index, arsenic level in drinking water, and genotypes of As (+ 3 oxidation state) methyltransferase (AS3MT) were also significantly co-associated with arsenic level and profile in the Vietnamese. To our knowledge, this is the first study indicating the associations of genetic factors of GST superfamily with arsenic metabolism in a Vietnamese population.

  20. A novel biomarker for marine environmental pollution of pi-class glutathione S-transferase from Mytilus coruscus.

    Science.gov (United States)

    Liu, Huihui; He, Jianyu; Zhao, Rongtao; Chi, Changfeng; Bao, Yongbo

    2015-08-01

    Glutathione S-transferases (GSTs) are the superfamily of phase II detoxification enzymes that play crucial roles in innate immunity. In this study, a pi-class GST homolog was identified from Mytilus coruscus (named as McGST1, KC525103). The full-length cDNA sequence of McGST1 was 621bp with a 5' untranslated region (UTR) of 70bp and a 3'-UTR of 201bp. The deduced amino acid sequence was 206 residues in length with theoretical pI/MW of 5.60/23.72kDa, containing the conserved G-site and diversiform H-site. BLASTn analysis and phylogenetic relationship strongly suggested that this cDNA sequence was a member of pi class GST family. The prediction of secondary structure displayed a preserved N-terminal and a C-terminal comprised with α-helixes. Quantitative real time RT-PCR showed that constitutive expression of McGST1 was occurred, with increasing order in mantle, muscle, gill, hemocyte, gonad and hepatopancreas. The stimulation of bacterial infection, heavy metals and 180CST could up-regulate McGST1 mRNA expression in hepatopancreas with time-dependent manners. The maximum expression appeared at 6h after pathogenic bacteria injected, with 10-fold in Vibrio alginolyticus and 16-fold in Vibrio harveyi higher than that of the control. The highest point of McGST1 mRNA appeared at different time for exposure to copper (10-fold at day 15), cadmium (9-fold at day10) and 180 CST (10-fold at day 15). These results suggested that McGST1 played a significant role in antioxidation and might potentially be used as indicators and biomarkers for detection of marine environmental pollution. Copyright © 2015 Elsevier Inc. All rights reserved.

  1. O-GlcNAc site-mapping of liver X receptor-α and O-GlcNAc transferase.

    Science.gov (United States)

    Fan, Qiong; Moen, Anders; Anonsen, Jan Haug; Bindesbøll, Christian; Sæther, Thomas; Carlson, Cathrine Rein; Grønning-Wang, Line M

    2018-05-05

    The Liver X Receptor α (LXRα) belongs to the nuclear receptor superfamily and plays an essential role in regulating cholesterol, lipid and glucose metabolism and inflammatory responses. We have previously shown that LXRα is post-translationally modified by O-linked β-N-acetyl-glucosamine (O-GlcNAc) with increased transcriptional activity. Moreover, we showed that LXRα associates with O-GlcNAc transferase (OGT) in vitro and in vivo in mouse liver. In this study, we report that human LXRα is O-GlcNAc modified in its N-terminal domain (NTD) by identifying a specific O-GlcNAc site S49 and a novel O-GlcNAc modified peptide 20 LWKPGAQDASSQAQGGSSCILRE 42 . However, O-GlcNAc site-mutations did not modulate LXRα transactivation of selected target gene promoters in vitro. Peptide array and co-immunoprecipitation assays demonstrate that LXRα interacts with OGT in its NTD and ligand-binding domain (LBD) in a ligand-independent fashion. Moreover, we map two new O-GlcNAc sites in the longest OGT isoform (ncOGT): S437 in the tetratricopeptide repeat (TPR) 13 domain and T1043 in the far C-terminus, and a new O-GlcNAc modified peptide (amino acids 826-832) in the intervening region (Int-D) within the catalytic domain. We also map four new O-GlcNAc sites in the short isoform sOGT: S391, T393, S399 and S437 in the TPRs 11-13 domain. Future studies will reveal the biological role of identified O-GlcNAc sites in LXRα and OGT. Copyright © 2018 Elsevier Inc. All rights reserved.

  2. Skeletal muscle O-GlcNAc transferase is important for muscle energy homeostasis and whole-body insulin sensitivity.

    Science.gov (United States)

    Shi, Hao; Munk, Alexander; Nielsen, Thomas S; Daughtry, Morgan R; Larsson, Louise; Li, Shize; Høyer, Kasper F; Geisler, Hannah W; Sulek, Karolina; Kjøbsted, Rasmus; Fisher, Taylor; Andersen, Marianne M; Shen, Zhengxing; Hansen, Ulrik K; England, Eric M; Cheng, Zhiyong; Højlund, Kurt; Wojtaszewski, Jørgen F P; Yang, Xiaoyong; Hulver, Matthew W; Helm, Richard F; Treebak, Jonas T; Gerrard, David E

    2018-05-01

    Given that cellular O-GlcNAcylation levels are thought to be real-time measures of cellular nutrient status and dysregulated O-GlcNAc signaling is associated with insulin resistance, we evaluated the role of O-GlcNAc transferase (OGT), the enzyme that mediates O-GlcNAcylation, in skeletal muscle. We assessed O-GlcNAcylation levels in skeletal muscle from obese, type 2 diabetic people, and we characterized muscle-specific OGT knockout (mKO) mice in metabolic cages and measured energy expenditure and substrate utilization pattern using indirect calorimetry. Whole body insulin sensitivity was assessed using the hyperinsulinemic euglycemic clamp technique and tissue-specific glucose uptake was subsequently evaluated. Tissues were used for histology, qPCR, Western blot, co-immunoprecipitation, and chromatin immunoprecipitation analyses. We found elevated levels of O-GlcNAc-modified proteins in obese, type 2 diabetic people compared with well-matched obese and lean controls. Muscle-specific OGT knockout mice were lean, and whole body energy expenditure and insulin sensitivity were increased in these mice, consistent with enhanced glucose uptake and elevated glycolytic enzyme activities in skeletal muscle. Moreover, enhanced glucose uptake was also observed in white adipose tissue that was browner than that of WT mice. Interestingly, mKO mice had elevated mRNA levels of Il15 in skeletal muscle and increased circulating IL-15 levels. We found that OGT in muscle mediates transcriptional repression of Il15 by O-GlcNAcylating Enhancer of Zeste Homolog 2 (EZH2). Elevated muscle O-GlcNAc levels paralleled insulin resistance and type 2 diabetes in humans. Moreover, OGT-mediated signaling is necessary for proper skeletal muscle metabolism and whole-body energy homeostasis, and our data highlight O-GlcNAcylation as a potential target for ameliorating metabolic disorders. Copyright © 2018 The Authors. Published by Elsevier GmbH.. All rights reserved.

  3. Glutathione S-transferases in human renal cortex and neoplastic tissue: enzymatic activity, isoenzyme profile and immunohistochemical localization.

    Science.gov (United States)

    Rodilla, V; Benzie, A A; Veitch, J M; Murray, G I; Rowe, J D; Hawksworth, G M

    1998-05-01

    1. Glutathione S-transferase (GST) activity in the cytosol of renal cortex and tumours from eight men and eight women was measured using 1-chloro-2,4-dinitrobenzene (CDNB) as a substrate. GST activities ranged from 685 to 2192 nmol/min/mg protein in cortex (median 1213) and from non-detectable (minimum 45) to 2424 nmol/min/mg protein in tumours (median 469). The activities in the tumours were lower than those in the normal cortices (p 0.05). 3. The age of the patients ranged from 42 to 81 years (median 62) and was not found to play a role in the levels of GST activity observed in cortex or in renal tumours from either sex. 4. Immunoblotting and immunohistochemical studies confirmed that GST-alpha was the predominant form expressed both in normal cortex and tumour and probably accounted for most of the GST activity present in these samples. GST-mu and GST-phi were expressed in both tumours and normal cortex and, while in some cases the level of expression in the cortices was higher than that found in the tumours, the reverse was also observed. Within the GST-mu class, GST M1/M2 was only detected in one sample (tumour), which showed the highest overall expression of GST-mu. GSTM3 was the predominant isoenzyme of the mu class in normal and tumour tissue, whereas GTM4 and GSTM5 were not detected. 5. These differences could have functional significance where xenobiotics or cytotoxic drugs are specific substrates for the different classes of GSTs.

  4. Electron/electron acoustic instability

    International Nuclear Information System (INIS)

    Gary, S.P.

    1987-01-01

    The electron acoustic wave becomes a normal mode of an unmagnetized collisionless plasma in the presence of two electron components with similar densities, but strongly disparate temperatures. The characteristic frequency of this mode is the plasma frequency of the cooler electron component. If these two electron components have a relative drift speed several times the thermal speed of the cooler component, the electron/electron acoustic instability may arise. This paper describes the parametric dependences of the threshold drift speed and maximum growth rate of this instability, and compares these with the same properties of the electron/ion acoustic instability. Under the condition of zero current, the electron/ion acoustic instability typically has the lower threshold drift speed, so that observation of the electron/electron acoustic instability is a strong indication of the presence of an electrical current in the plasma

  5. Electronic emission and electron guns

    International Nuclear Information System (INIS)

    Roy, Amitava

    2010-01-01

    This paper reviews the process of electron emission from metal surface. Although electrons move freely in conductors like metals, they normally do not leave the metal without some manipulation. In fact, heating and bombardment are the two primary ways in which electrons are emitted through the use of a heating element behind the cathode (termed thermionic emission) or as a result of bombardment with a beam of electrons, ions, or metastable atoms (termed secondary emission). Another important emission mechanism called Explosive Electron Emission (EEE) is also often used in various High Voltage Pulse Power Systems to generate very high current (few hundreds of kA) pulsed electron beams. The electron gun is the device in that it shoots off a continuous (or pulsed) stream of electrons. A brief idea about the evolution of the electron gun components and their basis of functioning are also discussed. (author)

  6. Sticker electronics

    KAUST Repository

    Hussain, Muhammad Mustafa; Torres Sevilla, Galo Andres; Diaz Cordero, Marlon Steven

    2017-01-01

    Electronic stickers may be manufactured on flexible substrates (110, 120, 130) as layers and packaged together. The package may then have an adhesive applied to one side to provide capability for sticking the electronic devices to surfaces

  7. ELECTRONIC SIGNATURES

    African Journals Online (AJOL)

    10332324

    "[to] promote the understanding and, acceptance of and growth in the number of electronic transactions .... Chapter III of the ECT Act is based on the UNCITRAL Model Law on Electronic. Commerce ... Communications Technology Law 146. 22.

  8. Electronic components

    CERN Document Server

    Colwell, Morris A

    1976-01-01

    Electronic Components provides a basic grounding in the practical aspects of using and selecting electronics components. The book describes the basic requirements needed to start practical work on electronic equipment, resistors and potentiometers, capacitance, and inductors and transformers. The text discusses semiconductor devices such as diodes, thyristors and triacs, transistors and heat sinks, logic and linear integrated circuits (I.C.s) and electromechanical devices. Common abbreviations applied to components are provided. Constructors and electronics engineers will find the book useful

  9. Understand electronics

    CERN Document Server

    Bishop, Owen

    2013-01-01

    Understand Electronics provides a readable introduction to the exciting world of electronics for the student or enthusiast with little previous knowledge. The subject is treated with the minimum of mathematics and the book is extensively illustrated.This is an essential guide for the newcomer to electronics, and replaces the author's best-selling Beginner's Guide to Electronics.The step-by-step approach makes this book ideal for introductory courses such as the Intermediate GNVQ.

  10. Electronic Commerce

    OpenAIRE

    Slavko Đerić

    2016-01-01

    Electronic commerce can be defined in different ways. Any definition helps to understand and explain that concept as better as possible.. Electronic commerce is a set of procedures and technologies that automate the tasks of financial transactions using electronic means. Also, according to some authors, electronic commerce is defined as a new concept, which is being developed and which includes process of buying and selling or exchanging products, services or information via computer networks...

  11. Electronic Publishing.

    Science.gov (United States)

    Lancaster, F. W.

    1989-01-01

    Describes various stages involved in the applications of electronic media to the publishing industry. Highlights include computer typesetting, or photocomposition; machine-readable databases; the distribution of publications in electronic form; computer conferencing and electronic mail; collaborative authorship; hypertext; hypermedia publications;…

  12. Epoxyalkane:Coenzyme M Transferase Gene Diversity and Distribution in Groundwater Samples from Chlorinated-Ethene-Contaminated Sites

    Science.gov (United States)

    Liu, Xikun

    2016-01-01

    ABSTRACT Epoxyalkane:coenzyme M transferase (EaCoMT) plays a critical role in the aerobic biodegradation and assimilation of alkenes, including ethene, propene, and the toxic chloroethene vinyl chloride (VC). To improve our understanding of the diversity and distribution of EaCoMT genes in the environment, novel EaCoMT-specific terminal-restriction fragment length polymorphism (T-RFLP) and nested-PCR methods were developed and applied to groundwater samples from six different contaminated sites. T-RFLP analysis revealed 192 different EaCoMT T-RFs. Using clone libraries, we retrieved 139 EaCoMT gene sequences from these samples. Phylogenetic analysis revealed that a majority of the sequences (78.4%) grouped with EaCoMT genes found in VC- and ethene-assimilating Mycobacterium strains and Nocardioides sp. strain JS614. The four most-abundant T-RFs were also matched with EaCoMT clone sequences related to Mycobacterium and Nocardioides strains. The remaining EaCoMT sequences clustered within two emergent EaCoMT gene subgroups represented by sequences found in propene-assimilating Gordonia rubripertincta strain B-276 and Xanthobacter autotrophicus strain Py2. EaCoMT gene abundance was positively correlated with VC and ethene concentrations at the sites studied. IMPORTANCE The EaCoMT gene plays a critical role in assimilation of short-chain alkenes, such as ethene, VC, and propene. An improved understanding of EaCoMT gene diversity and distribution is significant to the field of bioremediation in several ways. The expansion of the EaCoMT gene database and identification of incorrectly annotated EaCoMT genes currently in the database will facilitate improved design of environmental molecular diagnostic tools and high-throughput sequencing approaches for future bioremediation studies. Our results further suggest that potentially significant aerobic VC degraders in the environment are not well represented in pure culture. Future research should aim to isolate and

  13. Antioxidant role of glutathione S-transferases: 4-Hydroxynonenal, a key molecule in stress-mediated signaling.

    Science.gov (United States)

    Singhal, Sharad S; Singh, Sharda P; Singhal, Preeti; Horne, David; Singhal, Jyotsana; Awasthi, Sanjay

    2015-12-15

    4-Hydroxy-2-trans-nonenal (4HNE), one of the major end products of lipid peroxidation (LPO), has been shown to induce apoptosis in a variety of cell lines. It appears to modulate signaling processes in more than one way because it has been suggested to have a role in signaling for differentiation and proliferation. It has been known that glutathione S-transferases (GSTs) can reduce lipid hydroperoxides through their Se-independent glutathione-peroxidase activity and that these enzymes can also detoxify LPO end-products such as 4HNE. Available evidence from earlier studies together with results of recent studies in our laboratories strongly suggests that LPO products, particularly hydroperoxides and 4HNE, are involved in the mechanisms of stress-mediated signaling and that it can be modulated by the alpha-class GSTs through the regulation of the intracellular concentrations of 4HNE. We demonstrate that 4HNE induced apoptosis in various cell lines is accompanied with c-Jun-N-terminal kinase (JNK) and caspase-3 activation. Cells exposed to mild, transient heat or oxidative stress acquire the capacity to exclude intracellular 4HNE at a faster rate by inducing GSTA4-4 which conjugates 4HNE to glutathione (GSH), and RLIP76 which mediates the ATP-dependent transport of the GSH-conjugate of 4HNE (GS-HNE). The balance between formation and exclusion promotes different cellular processes - higher concentrations of 4HNE promote apoptosis; whereas, lower concentrations promote proliferation. In this article, we provide a brief summary of the cellular effects of 4HNE, followed by a review of its GST-catalyzed detoxification, with an emphasis on the structural attributes that play an important role in the interactions with alpha-class GSTA4-4. Taken together, 4HNE is a key signaling molecule and that GSTs being determinants of its intracellular concentrations, can regulate stress-mediated signaling, are reviewed in this article. Copyright © 2015 Elsevier Inc. All rights

  14. Skeletal muscle O-GlcNAc transferase is important for muscle energy homeostasis and whole-body insulin sensitivity

    Directory of Open Access Journals (Sweden)

    Hao Shi

    2018-05-01

    Full Text Available Objective: Given that cellular O-GlcNAcylation levels are thought to be real-time measures of cellular nutrient status and dysregulated O-GlcNAc signaling is associated with insulin resistance, we evaluated the role of O-GlcNAc transferase (OGT, the enzyme that mediates O-GlcNAcylation, in skeletal muscle. Methods: We assessed O-GlcNAcylation levels in skeletal muscle from obese, type 2 diabetic people, and we characterized muscle-specific OGT knockout (mKO mice in metabolic cages and measured energy expenditure and substrate utilization pattern using indirect calorimetry. Whole body insulin sensitivity was assessed using the hyperinsulinemic euglycemic clamp technique and tissue-specific glucose uptake was subsequently evaluated. Tissues were used for histology, qPCR, Western blot, co-immunoprecipitation, and chromatin immunoprecipitation analyses. Results: We found elevated levels of O-GlcNAc-modified proteins in obese, type 2 diabetic people compared with well-matched obese and lean controls. Muscle-specific OGT knockout mice were lean, and whole body energy expenditure and insulin sensitivity were increased in these mice, consistent with enhanced glucose uptake and elevated glycolytic enzyme activities in skeletal muscle. Moreover, enhanced glucose uptake was also observed in white adipose tissue that was browner than that of WT mice. Interestingly, mKO mice had elevated mRNA levels of Il15 in skeletal muscle and increased circulating IL-15 levels. We found that OGT in muscle mediates transcriptional repression of Il15 by O-GlcNAcylating Enhancer of Zeste Homolog 2 (EZH2. Conclusions: Elevated muscle O-GlcNAc levels paralleled insulin resistance and type 2 diabetes in humans. Moreover, OGT-mediated signaling is necessary for proper skeletal muscle metabolism and whole-body energy homeostasis, and our data highlight O-GlcNAcylation as a potential target for ameliorating metabolic disorders. Keywords: O-GlcNAc signaling, Type 2 diabetes, N

  15. Glutathione S-transferase M1 null genotype: lack of association with tumour characteristics and survival in advanced breast cancer

    International Nuclear Information System (INIS)

    Lizard-Nacol, Sarab; Coudert, Bruno; Colosetti, Pascal; Riedinger, Jean-Marc; Fargeot, Pierre; Brunet-Lecomte, Patrick

    1999-01-01

    Glutathione S-transferase (GST)M1, a member of the μ class GST gene family, has been shown to be polymorphic because of a partial gene deletion. This results in a failure to express the GSTM1 gene in 50-60% of individuals. Several studies have demonstrated a possible link with the GSTM1-null genotype and susceptibility to cancer. Furthermore, a GSTM1 isoenzyme has been positively associated with protective effect against mutagenic drugs, such as alkylating agents and anthracyclines. To determine whether GSTM1 polymorphisms are associated with tumour characteristics and survival in advanced breast cancer patients, and whether it may constitute a prognostic factor. We genotyped 92 patients receiving primary chemotherapy, which included cyclophosphamide, doxorubicine and 5-fluorouracil. The relationships between allelism at GSTM1 and clinicopathological parameters including age, menopausal status, tumour size, grade hormone receptors, involved nodes and p53 gene mutations were analysed. Of the patients with GSTM1-positive genotype, tissue samples obtained before and after treatment were available from 28 cases, allowing RNA extraction and GSTM1 expression by reverse transcription polymerase chain reaction. Relationships with clinical response to chemotherapy, and disease-free and overall survival were also evaluated. The data obtained was analysed using logistic regression to estimate the odds ratio and 95% confidence interval. Of 92 patients, 57.6% (n = 53) were classified as heritably GSTM1-deficient, and 42.4% (n = 39) were of the GSTM1-positive genotype. There were no statistically significant relationships between GSTM1-null genotype and the clinicopathological parameters analysed. No relationship was observed between GSTM1 RNA expression and objective clinical response to chemotherapy. Objective clinical response to chemotherapy was related only to clinical tumour size (P = 0.0177) and to the absence of intraductal carcinoma (P = 0.0013). GSTM1-null genotype

  16. Gene polymorphisms of glutathione S-transferase omega 1 and 2, urinary arsenic methylation profile and urothelial carcinoma

    Energy Technology Data Exchange (ETDEWEB)

    Chung, Chi-Jung [School of Public Health, College of Public Health and Nutrition, Taipei Medical University, Taipei, Taiwan (China); Pu, Yeong-Shiau [Department of Urology, National Taiwan University Hospital, Taipei, Taiwan (China); Su, Chien-Tien [Department of Family Medicine, Taipei Medical University Hospital, Taipei, Taiwan (China); Huang, Chao-Yuan [Department of Urology, National Taiwan University Hospital, Taipei, Taiwan (China); Graduate Institute of Clinical Medicine, College of Medicine, Taipei Medical University (China); Hsueh, Yu-Mei, E-mail: ymhsueh@tmu.edu.tw [School of Public Health, College of Public Health and Nutrition, Taipei Medical University, Taipei, Taiwan (China); Department of Public Health, School of Medicine, College of Medicine, Taipei Medical University, Taipei, Taiwan (China)

    2011-01-01

    Genetic polymorphisms in arsenic-metabolizing enzymes may be involved in the biotransformation of inorganic arsenic and may increase the risk of developing urothelial carcinoma (UC). The present study evaluated the roles of glutathione S-transferase omega 1 (GSTO1) and GSTO2 polymorphisms in UC carcinogenesis. A hospital-based case-control study was conducted. Questionnaire information and biological specimens were collected from 149 UC cases and 251 healthy controls in a non-obvious inorganic arsenic exposure area in Taipei, Taiwan. The urinary arsenic profile was determined using high-performance liquid chromatography and hydride generator-atomic absorption spectrometry. Genotyping for GSTO1 Ala140Asp and GSTO2 Asn142Asp was conducted using polymerase chain reaction-restriction fragment length polymerase. GSTO1 Glu208Lys genotyping was performed using high-throughput matrix-assisted laser desorption and ionization time-of-flight mass spectrometry. A significant positive association was found between total arsenic, inorganic arsenic percentage and monomethylarsonic acid percentage and UC, while dimethylarsinic acid percentage was significantly inversely associated with UC. The minor allele frequency of GSTO1 Ala140Asp, GSTO1 Glu208Lys and GSTO2 Asn142Asp was 18%, 1% and 26%, respectively. A significantly higher MMA% was found in people who carried the wild type of GSTO1 140 Ala/Ala compared to those who carried the GSTO1 140 Ala/Asp and Asp/Asp genotype (p = 0.02). The homogenous variant genotype of GSTO2 142 Asp/Asp was inversely associated with UC risk (OR = 0.17; 95% CI, 0.03 - 0.88; p = 0.03). Large-scale studies will be required to verify the association between the single nucleotide polymorphisms of arsenic-metabolism-related enzymes and UC risk. - Research Highlights: {yields} The homogenous variant genotype of GSTO2 was inversely associated with UC risk. {yields} A higher urinary MMA% was found in people carrying the wild type of GSTO1 Ala140Asp. {yields

  17. Association between glutathione S-transferase M1 and T1 polymorphisms and colorectal cancer risk in patients from Kazakhstan.

    Science.gov (United States)

    Zhunussova, Gulnur; Zhunusbekova, Benazir; Djansugurova, Leyla

    2015-01-01

    Colorectal cancer (CRC) is one of the most common malignancies worldwide and the incidence is increasing in developed as well as developing countries including Kazakhstan. Glutathione S-transferases (GSTs) are considered to be cancer susceptibility genes as they play a role in the detoxification of carcinogenic species. In this case-control study the influence of GSTM1 and GSTT1 polymorphisms on CRC risk in Kazakhstan population were evaluated. Blood samples were collected from patients diagnosed with rectal or colon cancer (300 individuals) as well as a control cohort of healthy volunteers (300 individuals), taking into account the age, gender, ethnicity, and smoking habits of the CRC patients. Deletion polymorphisms were genotyped employing a multiplex PCR amplification method. Association between polymorphisms and CRC susceptibility risk was calculated using multivariate analysis and logistic regression for odd ratio (OR). The homozygous GSTM1 null genotype was associated with significantly increased risk of CRC (OR = 2.01, 95% CI = 1.45-2.79, p = 0.0001) while the homozygous GSST1 null genotype was not associated with the risk of developing CRC (OR = 1.10, 95% CI = 0.78-1.55, p = 0.001), but the heterozygous genotype correlated with CRC susceptibility (OR = 1.98, 95% CI = 1.30-3.00, p = 0.001). Also, separate analyses of each of the main ethnic groups (Kazakh and Russian) showed a strong association of GSTM1 null genotype with CRC risk (for Kazakhs OR = 2.36, 95% CI = 1.35-4.10, p = 0.006 and for Russians OR = 1.84, 95% CI = 1.17-2.89, p = 0.003). The CRC risk of GSTM1 null genotype in smokers was considerably higher (OR = 3.37, 95% CI = 1.78-6.38, p = 0.0007). The combination of the GSTM1 and GSTT1 null genotypes in combined mixed population of Kazakhstan showed a trend to increasing the risk of developing CRC (OR = 1.60, 95% CI = 1.00-2.56), but it was not statistically significant. In conclusion, the results of this case-control study for sporadic cases of

  18. Pattern of γ-glutamyl transferase activity in cow milk throughout lactation and relationships with metabolic conditions and milk composition.

    Science.gov (United States)

    Calamari, L; Gobbi, L; Russo, F; Cappelli, F Piccioli

    2015-08-01

    The main objective of this experiment was to study the γ-glutamyl transferase (GGT) activity in milk during lactation and its relationship with metabolic status of dairy cows, milk yield, milk composition, and cheesemaking properties. The study was performed in a tied stall barn and involved 20 lactations from 12 healthy multiparous Italian Friesian dairy cows. During lactation starting at d 10, milk samples were collected weekly and analyzed for composition, somatic cells count, titratable acidity, and milk coagulation properties. The GGT activity was measured in defatted samples. Blood samples were collected weekly to assess biochemical indicators related to energy, protein, and mineral metabolism, markers of inflammation and some enzyme activities. The lactations of each cow were retrospectively categorized into 2 groups according to their milk GGT activity value through lactation. A median value of GGT activity in the milk of all lactations was calculated (3,045 U/L), and 10 lactations with lower GGT activity were classified as low while 10 lactations with greater GGT activity were classified as high. The average value of milk GGT activity during lactation was 3,863 and 3,024 U/L for high and low, respectively. The GGT activity decreased in early lactation and reached minimum values in the second month (3,289 and 2,355 U/L for high and low, respectively). Thereafter GGT activity increased progressively, reaching values in late lactation of 4,511 and 3,540 U/L in high and low, respectively. On average, milk yield was 40.81 and 42.76 kg/d in high and low, respectively, and a negative partial correlation with milk GGT activity was observed. A greater milk protein concentration was observed in high (3.39%) compared with low (3.18%), and a positive partial correlation with milk GGT activity was observed. Greater titratable acidity in high than that in low (3.75 vs. 3.45 degrees Soxhlet-Henkel/50 mL, respectively) was also observed. Plasma glucose was greater in

  19. Toxicological responses on cytochrome P450 and metabolic transferases in liver of goldfish (Carassius auratus) exposed to lead and paraquat.

    Science.gov (United States)

    Xu, Xiaoming; Cui, Zhaojie; Wang, Xinlei; Wang, Xixin; Zhang, Su

    2018-04-30

    As the producer of reactive oxygen species (ROS), both lead (Pb) and paraquat (PQ) can generate serious oxidative stress in target organs which result in irreversible toxic effects on organisms. They can disturb the normal catalytic activities of many enzymes by means of different toxicity mechanism. The changed responses of enzymes are frequently used as the biomarkers for indicating the relationship between toxicological effects and exposure levels. In this work, goldfish was exposed to a series of test groups containing lead and paraquat in the range of 0.05-10mg/L, respectively. Four hepatic enzyme activities, including 7-ethoxyresorufinO-deethylase (EROD), 7-benzyloxy-4-trifluoromethyl-coumarin-O-debenzyloxylase (BFCOD), glutathione S-transferase (GST) and UDP-glucuronosyltransferase (UGT) were determined after 1, 7, 14, 28 days exposure. The results showed that the activities of EROD and BFCOD in fish were significantly inhibited in response to paraquat at all exposure levels during the whole experiment. Similarly, the inhibitory effects of lead exposure on BFCOD activity were found in our study, while different responses of lead on EROD were observed. There were no significant differences on EROD activity under lower concentrations of lead (less than 0.1mg/L) before 14 days until an obvious increase was occurred for the 0.5mg/L lead treatment group at day 14. Furthermore, lead showed stronger inhibition on GST activity than paraquat when the concentrations of the two toxicants were more than 0.5mg/L. However, the similar dose and time-dependent manners of UGT activity were found under lead and paraquat exposure. Our results indicated that higher exposure levels and longer accumulations caused inhibitory effects on the four enzymes regardless of lead or paraquat stress. In addition, the responses of phase I enzymes were more sensitive than that of phase II enzymes and they may be served as the acceptable biomarkers for evaluating the toxicity effects of both

  20. Glutathione-S-transferase A3 knockout mice are sensitive to acute cytotoxic and genotoxic effects of aflatoxin B1

    International Nuclear Information System (INIS)

    Ilic, Zoran; Crawford, Dana; Egner, Patricia A.; Sell, Stewart

    2010-01-01

    Aflatoxin B1 (AFB1) is a major risk factor for hepatocellular carcinoma (HCC) in humans. However, mice, a major animal model for the study of AFB1 carcinogenesis, are resistant, due to high constitutive expression, in the mouse liver, of glutathione S-transferase A3 subunit (mGSTA3) that is lacking in humans. Our objective was to establish that a mouse model for AFB1 toxicity could be used to study mechanisms of toxicity that are relevant for human disease, i.e., an mGSTA3 knockout (KO) mouse that responds to toxicants such as AFB1 in a manner similar to humans. Exons 3-6 of the mGSTA3 were replaced with a neomycin cassette by homologous recombination. Southern blotting, RT-PCR, Western blotting, and measurement of AFB1-N 7 -DNA adduct formation were used to evaluate the mGSTA3 KO mice. The KO mice have deletion of exons 3-6 of the mGSTA3 gene, as expected, as well as a lack of mGSTA3 expression at the mRNA and protein levels. Three hours after injection of 5 mg/kg AFB1, mGSTA3 KO mice have more than 100-fold more AFB1-N 7 -DNA adducts in their livers than do similarly treated wild-type (WT) mice. In addition, the mGSTA3 KO mice die of massive hepatic necrosis, at AFB1 doses that have minimal toxic effects in WT mice. We conclude that mGSTA3 KO mice are sensitive to the acute cytotoxic and genotoxic effects of AFB1, confirming the crucial role of GSTA3 subunit in protection of normal mice against AFB1 toxicity. We propose the mGSTA3 KO mouse as a useful model with which to study the interplay of risk factors leading to HCC development in humans, as well as for testing of additional possible functions of mGSTA3.

  1. Protective role for ovarian glutathione S-transferase isoform pi during 7,12-dimethylbenz[a]anthracene-induced ovotoxicity

    Energy Technology Data Exchange (ETDEWEB)

    Bhattacharya, Poulomi, E-mail: poulomib@iastate.edu; Keating, Aileen F., E-mail: akeating@iastate.edu

    2012-04-15

    7,12-Dimethylbenz[a]anthracene (DMBA) destroys ovarian follicles at all developmental stages. This study investigated a role for the glutathione S-transferase (Gst) isoforms alpha (a), mu (m) and pi (p) and the transcription factors, Ahr and Nrf2, during DMBA-induced ovotoxicity, and their regulation by phosphatidylinositol-3 kinase (PI3K) signaling. Negative regulation of JNK by GSTP during DMBA exposure was also studied. Post-natal day (PND) 4 Fischer 344 rat ovaries were exposed to vehicle control (1% DMSO) ± DMBA (1 μM) or vehicle control (1% DMSO) ± LY294002 (PI3K inhibitor; 20 μM) for 1, 2, 4, or 6 days. Total RNA or protein was isolated, followed by RT-PCR or Western blotting to determine mRNA or protein level, respectively. Immunoprecipitation using an anti-GSTP antibody was performed to determine interaction between GSTP and JNK, followed by Western blotting to determine JNK and p-c-Jun protein level. DMBA had no impact on Gsta, Gstm or Nrf2 mRNA level, but increased Gstp mRNA and protein after 2 days. Ahr mRNA and protein increased after 2 and 4 days of DMBA exposure, respectively and DMBA increased NRF2 protein level after 4 days. JNK bound to GSTP was increased during DMBA exposure, with a concomitant decrease in unbound JNK and p-c-Jun. Ahr and Gstp mRNA were decreased (2 days) and increased (4 days) by PI3K inhibition, while Gstm mRNA increased (P < 0.05) after both time points, and there was no effect on Nrf2 mRNA. PI3K inhibition increased AHR, NRF2 and GSTP protein level. These findings support involvement of ovarian GSTP during DMBA exposure, and indicate a regulatory role for the PI3K signaling pathway on ovarian xenobiotic metabolism gene expression. -- Highlights: ► Ovarian GSTP is activated in response to DMBA exposure. ► AhR and Nrf2 transcription factors are up-regulated by DMBA. ► PI3K signaling regulates Ahr, Nrf2 and Gstp expression. ► GSTP negatively regulates ovarian JNK in response to DMBA exposure.

  2. Sticker electronics

    KAUST Repository

    Hussain, Muhammad Mustafa

    2017-09-08

    Electronic stickers may be manufactured on flexible substrates (110, 120, 130) as layers and packaged together. The package may then have an adhesive applied to one side to provide capability for sticking the electronic devices to surfaces. The stickers can be wrappable, placed on surfaces, glued on walls or mirrors or wood or stone, and have electronics (112, 122, 132) which may or may not be ultrathin. Packaging for the electronic sticker can use polymer on cellulose manufacturing and/or three dimensional (3-D) printing. The electronic stickers may provide lighting capability, sensing capability, and/or recharging capabilities.

  3. Basic electronics

    CERN Document Server

    Holbrook, Harold D

    1971-01-01

    Basic Electronics is an elementary text designed for basic instruction in electricity and electronics. It gives emphasis on electronic emission and the vacuum tube and shows transistor circuits in parallel with electron tube circuits. This book also demonstrates how the transistor merely replaces the tube, with proper change of circuit constants as required. Many problems are presented at the end of each chapter. This book is comprised of 17 chapters and opens with an overview of electron theory, followed by a discussion on resistance, inductance, and capacitance, along with their effects on t

  4. Electronic Commerce and Electronic Business

    Indian Academy of Sciences (India)

    R. Narasimhan (Krishtel eMaging) 1461 1996 Oct 15 13:05:22

    This special issue is motivated by the recent upsurge of research activity in the areas of electronic commerce and electronic business both in India and all over the world. The current ... Monte Carlo methods for pricing financial options are then.

  5. Electronic Government and Electronic Participation

    NARCIS (Netherlands)

    Tambouris, E; Scholl, H.J.; Janssen, M.F.W.H.A.; Wimmer, M.A.; Tarabanis, K; Gascó, M; Klievink, A.J.; Lindgren, I; Milano, M; Panagiotopoulos, P; Pardo, T.A.; Parycek, P; Sæbø, Ø

    2016-01-01

    Electronic government and electronic participation continue to transform the public sector and society worldwide and are constantly being transformed themselves by emerging information and communication technologies.This book presents papers from the 14th International Federation for Information

  6. Electronic Government and Electronic Participation

    NARCIS (Netherlands)

    Tambouris, E.; Scholl, H.J.; Janssen, M.F.W.H.A.; Wimmer, M.A.; Tarabanis, K.; Gascó, M.; Klievink, A.J.; Lindgren, I.; Milano, M.; Panagiotopoulos, P.; Pardo, T.A.; Parycek, P.; Sæbø, O.

    2015-01-01

    Electronic government and electronic participation continue to transform the public sector and society worldwide and are constantly being transformed themselves by emerging information and communication technologies. This book presents papers from the 14th International Federation for Information

  7. Miopatia por deficiência de carnitina-palmitil-transferase: relato de 2 casos com dosagens enzimáticas no tecido muscular

    Directory of Open Access Journals (Sweden)

    Lineu Cesar Werneck

    1983-12-01

    Full Text Available Relato dos casos de dois irmãos, que desde a infância apresentavam dores musculares após exercício prolongado ou exposição ao frio, com diminuição da força, à medida que o exercício continuava. Um deles desenvolveu mioglobinúria recorrente e, em um episódio apresentou insuficiência renal aguda, necessitando de diálise peritonial. A investigação laboratorial intercrise foi normal, mas durante o episódio de mioglobinúria, apresentou grande aumento da creatinafosfoquinase. A eletromiografia foi sugestiva de processo de denervação. Teste de produção de lactato durante isquemia foi normal. Biópsias musculares mostraram discreto aumento dos lipídios nas fibras musculares e maior atividade da desidrogenase succínica na histoquímica. O estudo bioquímico do tecido muscular dos dois pacientes, revelou importante redução da atividade da carnitina-palmitil-transferase, com atividade normal da carnitina-octanoil-transferase e carnitina-acetil-transferase. São discutidas as vias metabólicas, sua importância na manutenção da energia muscular durante o exercício prolongado e o papel dos ácidos graxos como fonte energética muscular durante condições normais e patológicas.

  8. Electron Tree

    DEFF Research Database (Denmark)

    Appelt, Ane L; Rønde, Heidi S

    2013-01-01

    The photo shows a close-up of a Lichtenberg figure – popularly called an “electron tree” – produced in a cylinder of polymethyl methacrylate (PMMA). Electron trees are created by irradiating a suitable insulating material, in this case PMMA, with an intense high energy electron beam. Upon discharge......, during dielectric breakdown in the material, the electrons generate branching chains of fractures on leaving the PMMA, producing the tree pattern seen. To be able to create electron trees with a clinical linear accelerator, one needs to access the primary electron beam used for photon treatments. We...... appropriated a linac that was being decommissioned in our department and dismantled the head to circumvent the target and ion chambers. This is one of 24 electron trees produced before we had to stop the fun and allow the rest of the accelerator to be disassembled....

  9. `Twisted' electrons

    Science.gov (United States)

    Larocque, Hugo; Kaminer, Ido; Grillo, Vincenzo; Leuchs, Gerd; Padgett, Miles J.; Boyd, Robert W.; Segev, Mordechai; Karimi, Ebrahim

    2018-04-01

    Electrons have played a significant role in the development of many fields of physics during the last century. The interest surrounding them mostly involved their wave-like features prescribed by the quantum theory. In particular, these features correctly predict the behaviour of electrons in various physical systems including atoms, molecules, solid-state materials, and even in free space. Ten years ago, new breakthroughs were made, arising from the new ability to bestow orbital angular momentum (OAM) to the wave function of electrons. This quantity, in conjunction with the electron's charge, results in an additional magnetic property. Owing to these features, OAM-carrying, or twisted, electrons can effectively interact with magnetic fields in unprecedented ways and have motivated materials scientists to find new methods for generating twisted electrons and measuring their OAM content. Here, we provide an overview of such techniques along with an introduction to the exciting dynamics of twisted electrons.

  10. Identification of an enhancer element of class Pi glutathione S-transferase gene required for expression by a co-planar polychlorinated biphenyl.

    OpenAIRE

    Matsumoto, M; Imagawa, M; Aoki, Y

    1999-01-01

    3,3',4,4',5-Pentachlorobiphenyl (PenCB), one of the most toxic co-planar polychlorinated biphenyl congeners, specifically induces class Pi glutathione S-transferase (GSTP1) as well as cytochrome P-450 1A1 in primary cultured rat liver parenchymal cells [Aoki, Matsumoto and Suzuki (1993) FEBS Lett. 333, 114-118]. However, the 5'-flanking sequence of the GSTP1 gene does not contain a xenobiotic responsive element, to which arylhydrocarbon receptor binds. Using a chloramphenicol acetyltransferas...

  11. Targeted cytosine deaminase-uracil phosphoribosyl transferase suicide gene therapy induces small cell lung cancer-specific cytotoxicity and tumor growth delay

    DEFF Research Database (Denmark)

    Christensen, Camilla L; Gjetting, Torben; Poulsen, Thomas Tuxen

    2010-01-01

    Small cell lung cancer (SCLC) is a highly malignant cancer for which there is no curable treatment. Novel therapies are therefore in great demand. In the present study we investigated the therapeutic effect of transcriptionally targeted suicide gene therapy for SCLC based on the yeast cytosine...... deaminase (YCD) gene alone or fused with the yeast uracil phosphoribosyl transferase (YUPRT) gene followed by administration of 5-fluorocytosine (5-FC) prodrug. Experimental design: The YCD gene or the YCD-YUPRT gene was placed under regulation of the SCLC-specific promoter insulinoma-associated 1 (INSM1...

  12. Possible gene dosage effect of glutathione-S-transferases on atopic asthma: using real-time PCR for quantification of GSTM1 and GSTT1 gene copy numbers

    DEFF Research Database (Denmark)

    Brasch-Andersen, Charlotte; Christiansen, L; Tan, Q

    2004-01-01

    -S-transferase (GST) involved in the antioxidant defense were tested for association to asthma using 246 Danish atopic families in a family-based transmission disequilibrium test (TDT) design. A real-time PCR assay for relative quantification of gene copy number of GSTM1 and GSTT1 was developed. The assay made......Asthma is a complex genetic disorder characterized by chronic inflammation in the airways. As oxidative stress is a key component of inflammation, variations in genes involved in antioxidant defense could therefore be likely candidates for asthma. Three enzymes from the superfamily glutathione...

  13. Electronic Commerce

    Directory of Open Access Journals (Sweden)

    Slavko Đerić

    2016-12-01

    Full Text Available Electronic commerce can be defined in different ways. Any definition helps to understand and explain that concept as better as possible.. Electronic commerce is a set of procedures and technologies that automate the tasks of financial transactions using electronic means. Also, according to some authors, electronic commerce is defined as a new concept, which is being developed and which includes process of buying and selling or exchanging products, services or information via computer networks, including the Internet. Electronic commerce is not limited just to buying and selling, but it also includes all pre-sales and after-sales ongoing activities along the supply chain. Introducing electronic commerce, using the Internet and Web services in business, realizes the way to a completely new type of economy - internet economy.

  14. Advanced Electronics

    Science.gov (United States)

    2017-07-21

    AFRL-RV-PS- AFRL-RV-PS- TR-2017-0114 TR-2017-0114 ADVANCED ELECTRONICS Ashwani Sharma 21 Jul 2017 Interim Report APPROVED FOR PUBLIC RELEASE...NUMBER Advanced Electronics 5b. GRANT NUMBER 5c. PROGRAM ELEMENT NUMBER 62601F 6. AUTHOR(S) 5d. PROJECT NUMBER 4846 Ashwani Sharma 5e. TASK NUMBER...Approved for public release; distribution is unlimited. (RDMX-17-14919 dtd 20 Mar 2018) 13. SUPPLEMENTARY NOTES 14. ABSTRACT The Space Electronics

  15. Electron spectroscopy

    International Nuclear Information System (INIS)

    Hegde, M.S.

    1979-01-01

    An introduction to the various techniques in electron spectroscopy is presented. These techniques include: (1) UV Photoelectron spectroscopy, (2) X-ray Photoelectron spectroscopy, (3) Auger electron spectroscopy, (4) Electron energy loss spectroscopy, (5) Penning ionization spectroscopy and (6) Ion neutralization spectroscopy. The radiations used in each technique, the basis of the technique and the special information obtained in structure determination in atoms and molecules by each technique are summarised. (A.K.)

  16. Polymer electronics

    CERN Document Server

    Hsin-Fei, Meng

    2013-01-01

    Polymer semiconductor is the only semiconductor that can be processed in solution. Electronics made by these flexible materials have many advantages such as large-area solution process, low cost, and high performance. Researchers and companies are increasingly dedicating time and money in polymer electronics. This book focuses on the fundamental materials and device physics of polymer electronics. It describes polymer light-emitting diodes, polymer field-effect transistors, organic vertical transistors, polymer solar cells, and many applications based on polymer electronics. The book also disc

  17. Electronics Industry

    National Research Council Canada - National Science Library

    Bell, Robert; Carroll-Garrison, Martina; Donovan, Daniel; Fisher, John; Guemmer, Paul; Harms, Robert; Kelly, Timothy; Love, Mattie; McReynolds, James; Ward, Ralph

    2006-01-01

    .... Government action to preserve strategic access to semiconductor producers is clearly needed to ensure DoD electronic systems can be built without compromising sensitive technology, though every...

  18. Microfluidic electronics.

    Science.gov (United States)

    Cheng, Shi; Wu, Zhigang

    2012-08-21

    Microfluidics, a field that has been well-established for several decades, has seen extensive applications in the areas of biology, chemistry, and medicine. However, it might be very hard to imagine how such soft microfluidic devices would be used in other areas, such as electronics, in which stiff, solid metals, insulators, and semiconductors have previously dominated. Very recently, things have radically changed. Taking advantage of native properties of microfluidics, advances in microfluidics-based electronics have shown great potential in numerous new appealing applications, e.g. bio-inspired devices, body-worn healthcare and medical sensing systems, and ergonomic units, in which conventional rigid, bulky electronics are facing insurmountable obstacles to fulfil the demand on comfortable user experience. Not only would the birth of microfluidic electronics contribute to both the microfluidics and electronics fields, but it may also shape the future of our daily life. Nevertheless, microfluidic electronics are still at a very early stage, and significant efforts in research and development are needed to advance this emerging field. The intention of this article is to review recent research outcomes in the field of microfluidic electronics, and address current technical challenges and issues. The outlook of future development in microfluidic electronic devices and systems, as well as new fabrication techniques, is also discussed. Moreover, the authors would like to inspire both the microfluidics and electronics communities to further exploit this newly-established field.

  19. Electron holography

    CERN Document Server

    Tonomura, Akira

    1993-01-01

    Holography was devised for breaking through the resolution limit of electron microscopes The advent of a "coherent" field emission electron beam has enabled the use of Electron Holography in various areas of magnetic domain structures observation, fluxon observation in superconductors, and fundamental experiments in physics which have been inaccessible using other techniques After examining the fundamentals of electron holography and its applications to the afore mentioned fields, a detailed discussion of the Aharonov-Bohm effect and the related experiments is presented Many photographs and illustrations are included to elucidate the text

  20. The EGF repeat-specific O-GlcNAc-transferase Eogt interacts with notch signaling and pyrimidine metabolism pathways in Drosophila.

    Directory of Open Access Journals (Sweden)

    Reto Müller

    Full Text Available The O-GlcNAc transferase Eogt modifies EGF repeats in proteins that transit the secretory pathway, including Dumpy and Notch. In this paper, we show that the Notch ligands Delta and Serrate are also substrates of Eogt, that mutation of a putative UDP-GlcNAc binding DXD motif greatly reduces enzyme activity, and that Eogt and the cytoplasmic O-GlcNAc transferase Ogt have distinct substrates in Drosophila larvae. Loss of Eogt is larval lethal and disrupts Dumpy functions, but does not obviously perturb Notch signaling. To identify novel genetic interactions with eogt, we investigated dominant modification of wing blister formation caused by knock-down of eogt. Unexpectedly, heterozygosity for several members of the canonical Notch signaling pathway suppressed wing blister formation. And importantly, extensive genetic interactions with mutants in pyrimidine metabolism were identified. Removal of pyrimidine synthesis alleles suppressed wing blister formation, while removal of uracil catabolism alleles was synthetic lethal with eogt knock-down. Therefore, Eogt may regulate protein functions by O-GlcNAc modification of their EGF repeats, and cellular metabolism by affecting pyrimidine synthesis and catabolism. We propose that eogt knock-down in the wing leads to metabolic and signaling perturbations that increase cytosolic uracil levels, thereby causing wing blister formation.

  1. Mutations in ribosomal protein L3 and 23S ribosomal RNA at the peptidyl transferase centre are associated with reduced susceptibility to tiamulin in Brachyspira spp. isolates.

    Science.gov (United States)

    Pringle, Märit; Poehlsgaard, Jacob; Vester, Birte; Long, Katherine S

    2004-12-01

    The pleuromutilin antibiotic tiamulin binds to the ribosomal peptidyl transferase centre. Three groups of Brachyspira spp. isolates with reduced tiamulin susceptibility were analysed to define resistance mechanisms to the drug. Mutations were identified in genes encoding ribosomal protein L3 and 23S rRNA at positions proximal to the peptidyl transferase centre. In two groups of laboratory-selected mutants, mutations were found at nucleotide positions 2032, 2055, 2447, 2499, 2504 and 2572 of 23S rRNA (Escherichia coli numbering) and at amino acid positions 148 and 149 of ribosomal protein L3 (Brachyspira pilosicoli numbering). In a third group of clinical B. hyodysenteriae isolates, only a single mutation at amino acid 148 of ribosomal protein L3 was detected. Chemical footprinting experiments show a reduced binding of tiamulin to ribosomal subunits from mutants with decreased susceptibility to the drug. This reduction in drug binding is likely the resistance mechanism for these strains. Hence, the identified mutations located near the tiamulin binding site are predicted to be responsible for the resistance phenotype. The positions of the mutated residues relative to the bound drug advocate a model where the mutations affect tiamulin binding indirectly through perturbation of nucleotide U2504.

  2. Track detection on the cells exposed to high Linear Energy Transfer heavy-ions by Cr-39 plastic and terminal deoxynucleotidyl transferase(Td T)

    International Nuclear Information System (INIS)

    Mehnati, P.; Keshtkar, A.; Mesbahi, A.; Sasaki, H.

    2006-01-01

    The fatal effect of ionizing radiation on cells depends on Linear Energy Transfer level. The distribution of ionizing radiation is sparse and homogeneous for low Linear Energy Transfer radiations such as X or y, but it is dense and concentrated for high Linear Energy Transfer radiation such as heavy-ions radiation. Materials and Methods: Chinese hamster ovary cells (CHO-K1) were exposed to 4 Gy Fe-ion 2000 keV/μm. The Cr-39 is a special and sensitive plastic used to verify exact position of heavy-ions traversal. Terminal deoxynucleotidyl transferase is an enzyme labeled with [3 H ] d ATP for detection of cellular DNA damage by autoradiography assay. Results: The track of heavy ions traversals presented by pit size was almost similar for all different doses of radiation. No pits to show the track of traversal were found in 20% of the cell nuclei of the irradiation. Apparently these fractions of cells wave not hit by heavy ions. Conclusion: This study indicated the possible usefulness of both the Cr-39 plastics and DNA labeling with Terminal deoxynucleotidyl transferase method for evaluating the biological effect of heavy-ions in comparison with low Linear Energy Transfer ionizing radiation

  3. Use of serum gamma glutamyl transferase as a biomarker of stress and metabolic dysfunctions in Rathi cattle of arid tract in India

    Directory of Open Access Journals (Sweden)

    Kataria N.

    2012-08-01

    Full Text Available The investigation was carried out to determine serum gamma glutamyl transferase enzyme as a biomarker of stress and metabolic dysfunctions in Rathi cattle of arid tract in India. Blood samples were collected to harvest serum from healthy male and female, drought affected, ketotic cows, recently aborted cows, cows with diarrhoea, cows with traumatic pericarditis, calves with urinary calculi, cows affected with urea poisoning and cows affected with acidosis. The mean values of γ glutamyl transferase showed significant variations (p≤0.05 according to sex and age in the healthy group of animals. The normal range in healthy animals was from 12 to 34 UL-1. In affected group an average 23.69 times rise in the value was observed from that of healthy group. Cows affected with urea poisoning and acidosis were having highest mean values whereas drought affected animals were having least value. It was concluded that present study attempted to provide a new insight about an old enzyme. As the number of animals in the present study was statistically sufficient therefore the mean value of healthy group can be used as reference value for γ GT in Rathi cattle and other cattle breeds which can help to interpret the variations of serum γ GT in various metabolic diseases of cattle.

  4. A chloroplast-localized and auxin-induced glutathione S-transferase from phreatophyte Prosopis juliflora confer drought tolerance on tobacco.

    Science.gov (United States)

    George, Suja; Venkataraman, Gayatri; Parida, Ajay

    2010-03-01

    Plant growth and productivity are adversely affected by various abiotic stress factors. In our previous study, we used Prosopis juliflora, a drought-tolerant tree species of Fabaceae, as a model plant system for mining genes functioning in abiotic stress tolerance. Large-scale random EST sequencing from a cDNA library obtained from drought-stressed leaves of 2-month-old P. juliflora plants resulted in identification of three different auxin-inducible glutathione S-transferases. In this paper, we report the cellular localization and the ability to confer drought tolerance in transgenic tobacco of one of these GSTs (PjGSTU1). PjGSTU1 was overexpressed in Escherichia coli and GST and GPX activities in total protein samples were assayed and compared with controls. The results indicated that PjGSTU1 protein forms a functional homo-dimer in recombinant bacteria with glutathione transferase as well as glutathione peroxidase activities. PjGSTU1 transgenic tobacco lines survived better under conditions of 15% PEG stress compared with control un-transformed plants. In vivo localization studies for PjGSTU1 using GFP fusion revealed protein localization in chloroplasts of transgenic plants. The peroxidase activity of PjGSTU1 and its localization in the chloroplast indicates a possible role for PjGSTU1 in ROS removal. Copyright 2009 Elsevier GmbH. All rights reserved.

  5. The relationship of glutathione-S-transferases copy number variation and indoor air pollution to symptoms and markers of respiratory disease

    DEFF Research Database (Denmark)

    Hersoug, Lars-Georg; Brasch-Andersen, Charlotte; Husemoen, Lise-Lotte

    2012-01-01

    Introduction: Exposure to particulate matter (PM) may induce inflammation and oxidative stress in the airways. Carriers of null polymorphisms of glutathione S-transferases (GSTs), which detoxify reactive oxygen species, may be particularly susceptible to the effects of PM. Objectives: To investig....... The relationship of glutathione-S-transferases copy number variation and indoor air pollution to symptoms and markers of respiratory disease. Clin Respir J 2011; DOI:10.1111/j.1752-699X.2011.00258.x.......: To investigate whether deletions of GSTM1 and GSTT1 modify the potential effects of exposure to indoor sources of PM on symptoms and objective markers of respiratory disease. Methods: We conducted a population-based, cross-sectional study of 3471 persons aged 18-69 years. Information about exposure to indoor......: We found that none of the symptoms and objective markers of respiratory disease were significantly associated with the GST null polymorphisms. An increasing number of positive alleles of the GSTM1 polymorphism tended to be associated lower prevalence of wheeze, cough, and high forced expiratory...

  6. The electron

    International Nuclear Information System (INIS)

    Hestenes, David; Weingartshofer, Antonio

    1991-01-01

    The stupendous successes of the Dirac equation and quantum electro-dynamics have established the electron as the best understood of the fundamental constituents of matter. Nevertheless, physicists agree that the electron still has secrets to reveal. Moreover, powerful new theoretical and experimental tools for probing those secrets have been sharpened during the last decade. This workshop was organized to bring theorists and experimentalists together to discuss their common goal of knowing the electron. Present state and future prospects for progress toward that goal are here described. The theoretical papers encompass a wide range of views on the electron. Several argue that the 'Zitter-bewegung' is more than a mathematical peculiarity of the Dirac equation, that it may well be a real physical phenomenon and worthy of serious study, theoretically and experimentally. Besides generating the electron spin and magnetic moment, the 'Zitterbewegung' may be a vital clue to electron structure and self-interaction. Some of the papers employ a radical new formulation of the Dirac theory which reveals a hidden geo-metric structure in the theory that supports a 'Zitterbewegung' inter-pretation. For the last half century the properties of electrons have been probed primarily by scattering experiments at ever higher energies. Recently, however, two powerful new experimental techniques have emerged capable of giving alternative experimental views of the electron. First, techniques for confining single electrons for long term study have led to the most accurate measurements of the electron magnetic moment. Second, the interaction of high intensity laser fields with atoms and electrons have revealed striking new phenomena such as multiphoton ionization. refs.; figs.; tabs

  7. Effect of cadmium on glutathione S-transferase and metallothionein gene expression in coho salmon liver, gill and olfactory tissues

    International Nuclear Information System (INIS)

    Espinoza, Herbert M.; Williams, Chase R.; Gallagher, Evan P.

    2012-01-01

    Highlights: ► Developed qPCR assays to distinguish closely related GST isoforms in salmon. ► Examined the effect of cadmium on GST and metallothionein genes in 3 tissues. ► Modulation of GST varied among isoforms, tissues, and included a loss of expression. ► Metallothionein outperformed, but generally complemented, GSTs as biomarkers. ► Salmon olfactory genes were among the most responsive to cadmium. - Abstract: The glutathione S-transferases (GSTs) are a multifunctional family of phase II enzymes that detoxify a variety of environmental chemicals, reactive intermediates, and secondary products of oxidative damage. GST mRNA expression and catalytic activity have been used as biomarkers of exposure to environmental chemicals. However, factors such as species differences in induction, partial analyses of multiple GST isoforms, and lack of understanding of fish GST gene regulation, have confounded the use of GSTs as markers of pollutant exposure. In the present study, we examined the effect of exposure to cadmium (Cd), a prototypical environmental contaminant and inducer of mammalian GST, on GST mRNA expression in coho salmon (Oncorhynchus kisutch) liver, gill, and olfactory tissues. GST expression data were compared to those for metallothionein (MT), a prototypical biomarker of metal exposure. Data mining of genomic databases led to the development of quantitative real-time PCR (qPCR) assays for salmon GST isoforms encompassing 9 subfamilies, including alpha, mu, pi, theta, omega, kappa, rho, zeta and microsomal GST. In vivo acute (8–48 h) exposures to low (3.7 ppb) and high (347 ppb) levels of Cd relevant to environmental scenarios elicited a variety of transient, albeit minor changes (<2.5-fold) in tissue GST profiles, including some reductions in GST mRNA expression. In general, olfactory GSTs were the earliest to respond to cadmium, whereas, more pronounced effects in olfactory and gill GST expression were observed at 48 h relative to earlier time

  8. Printed Electronics

    Science.gov (United States)

    Korkut, Sibel (Inventor); Chiang, Katherine S. (Inventor); Crain, John M. (Inventor); Aksay, Ilhan A. (Inventor); Lettow, John S. (Inventor); Chen, Chuan-Hua (Inventor); Prud'Homme, Robert K. (Inventor)

    2018-01-01

    Printed electronic device comprising a substrate onto at least one surface of which has been applied a layer of an electrically conductive ink comprising functionalized graphene sheets and at least one binder. A method of preparing printed electronic devices is further disclosed.

  9. O-Glycosylation Modulates Proprotein Convertase Activation of Angiopoietin-like Protein 3: POSSIBLE ROLE OF POLYPEPTIDE GalNAc-TRANSFERASE-2 IN REGULATION OF CONCENTRATIONS OF PLASMA LIPIDS

    DEFF Research Database (Denmark)

    Schjoldager, Katrine Ter-Borch Gram; Vester-Christensen, Malene B; Bennett, Eric Paul

    2010-01-01

    immediately C-terminal (TT(226)). We developed an in vivo model system in CHO ldlD cells that was used to show that O-glycosylation in the processing site blocked processing of ANGPTL3. Genome-wide SNP association studies have identified the polypeptide GalNAc-transferase gene, GALNT2, as a candidate gene...... for low HDL and high triglyceride blood levels. We hypothesized that the GalNAc-T2 transferase performed critical O-glycosylation of proteins involved in lipid metabolism. Screening of a panel of proteins known to affect lipid metabolism for potential sites glycosylated by GalNAc-T2 led to identification...

  10. Digital electronics

    CERN Document Server

    Morris, John

    2013-01-01

    An essential companion to John C Morris's 'Analogue Electronics', this clear and accessible text is designed for electronics students, teachers and enthusiasts who already have a basic understanding of electronics, and who wish to develop their knowledge of digital techniques and applications. Employing a discovery-based approach, the author covers fundamental theory before going on to develop an appreciation of logic networks, integrated circuit applications and analogue-digital conversion. A section on digital fault finding and useful ic data sheets completes th

  11. Electronic diagrams

    CERN Document Server

    Colwell, Morris A

    1976-01-01

    Electronic Diagrams is a ready reference and general guide to systems and circuit planning and in the preparation of diagrams for both newcomers and the more experienced. This book presents guidelines and logical procedures that the reader can follow and then be equipped to tackle large complex diagrams by recognition of characteristic 'building blocks' or 'black boxes'. The goal is to break down many of the barriers that often seem to deter students and laymen in learning the art of electronics, especially when they take up electronics as a spare time occupation. This text is comprised of nin

  12. Polymer electronics

    CERN Document Server

    Geoghegan, Mark

    2013-01-01

    Polymer electronics is the science behind many important new developments in technology, such as the flexible electronic display (e-ink) and many new developments in transistor technology. Solar cells, light-emitting diodes, and transistors are all areas where plastic electronics is likely to, or is already having, a serious impact on our daily lives. With polymer transistors and light-emitting diodes now being commercialised, there is a clear need for a pedagogic text thatdiscusses the subject in a clear and concise fashion suitable for senior undergraduate and graduate students. The content

  13. Starting electronics

    CERN Document Server

    Brindley, Keith

    2005-01-01

    Starting Electronics is unrivalled as a highly practical introduction for hobbyists, students and technicians. Keith Brindley introduces readers to the functions of the main component types, their uses, and the basic principles of building and designing electronic circuits. Breadboard layouts make this very much a ready-to-run book for the experimenter; and the use of multimeter, but not oscilloscopes, puts this practical exploration of electronics within reach of every home enthusiast's pocket. The third edition has kept the simplicity and clarity of the original. New material

  14. Stretchable electronics

    CERN Document Server

    Someya, Takao

    2012-01-01

    With its comprehensive coverage this handbook and ready reference brings together some of the most outstanding scientists in the field to lay down the undisputed knowledge on how to make electronics stretchable.As such, it focuses on gathering and evaluating the materials, designs, models and technologies that enable the fabrication of fully elastic electronic devices which can sustain high strain. Furthermore, it provides a review of those specific applications that directly benefit from highly compliant electronics, including transistors, photonic devices and sensors. In addition to stre

  15. Electron optics

    CERN Document Server

    Grivet, Pierre; Bertein, F; Castaing, R; Gauzit, M; Septier, Albert L

    1972-01-01

    Electron Optics, Second English Edition, Part I: Optics is a 10-chapter book that begins by elucidating the fundamental features and basic techniques of electron optics, as well as the distribution of potential and field in electrostatic lenses. This book then explains the field distribution in magnetic lenses; the optical properties of electrostatic and magnetic lenses; and the similarities and differences between glass optics and electron optics. Subsequent chapters focus on lens defects; some electrostatic lenses and triode guns; and magnetic lens models. The strong focusing lenses and pris

  16. Electronic identity

    CERN Document Server

    de Andrade, Norberto Nuno Gomes; Argles, David

    2014-01-01

    With the increasing availability of electronic services, security and a reliable means by which identity is verified is essential.Written by Norberto Andrade the first chapter of this book provides an overview of the main legal and regulatory aspects regarding electronic identity in Europe and assesses the importance of electronic identity for administration (public), business (private) and, above all, citizens. It also highlights the role of eID as a key enabler of the economy.In the second chapter Lisha Chen-Wilson, David Argles, Michele Schiano di Zenise and Gary Wills discuss the user-cent

  17. Power Electronics

    DEFF Research Database (Denmark)

    Iov, Florin; Ciobotaru, Mihai; Blaabjerg, Frede

    2008-01-01

    is to change the electrical power production sources from the conventional, fossil (and short term) based energy sources to renewable energy resources. The other is to use high efficient power electronics in power generation, power transmission/distribution and end-user application. This paper discuss the most...... emerging renewable energy sources, wind energy, which by means of power electronics are changing from being a minor energy source to be acting as an important power source in the energy system. Power electronics is the enabling technology and the presentation will cover the development in wind turbine...... technology from kW to MW, discuss which power electronic solutions are most feasible and used today....

  18. Paper electronics.

    Science.gov (United States)

    Tobjörk, Daniel; Österbacka, Ronald

    2011-05-03

    Paper is ubiquitous in everyday life and a truly low-cost substrate. The use of paper substrates could be extended even further, if electronic applications would be applied next to or below the printed graphics. However, applying electronics on paper is challenging. The paper surface is not only very rough compared to plastics, but is also porous. While this is detrimental for most electronic devices manufactured directly onto paper substrates, there are also approaches that are compatible with the rough and absorptive paper surface. In this review, recent advances and possibilities of these approaches are evaluated and the limitations of paper electronics are discussed. Copyright © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  19. Electron Microprobe

    Data.gov (United States)

    Federal Laboratory Consortium — The JEOL JXA-8600 is a conventional hairpin filament thermal emission electron microprobe that is more than 20 years old. It is capable of performing qualitative and...

  20. Electronic Aggression

    Centers for Disease Control (CDC) Podcasts

    Aggression is no longer limited to the school yard. New forms of electronic media, such as blogs, instant messaging, chat rooms, email, text messaging, and the internet are providing new arenas for youth violence to occur.

  1. Electron Emitters

    National Research Council Canada - National Science Library

    Tzeng, Yonhua

    2002-01-01

    When two carbon-nanotube coated electrodes are placed at a small distance from each other, electron emission from carbon nanotubes allows a DC or AC electrical current to flow between these two electrodes...

  2. Electronic plants

    Science.gov (United States)

    Stavrinidou, Eleni; Gabrielsson, Roger; Gomez, Eliot; Crispin, Xavier; Nilsson, Ove; Simon, Daniel T.; Berggren, Magnus

    2015-01-01

    The roots, stems, leaves, and vascular circuitry of higher plants are responsible for conveying the chemical signals that regulate growth and functions. From a certain perspective, these features are analogous to the contacts, interconnections, devices, and wires of discrete and integrated electronic circuits. Although many attempts have been made to augment plant function with electroactive materials, plants’ “circuitry” has never been directly merged with electronics. We report analog and digital organic electronic circuits and devices manufactured in living plants. The four key components of a circuit have been achieved using the xylem, leaves, veins, and signals of the plant as the template and integral part of the circuit elements and functions. With integrated and distributed electronics in plants, one can envisage a range of applications including precision recording and regulation of physiology, energy harvesting from photosynthesis, and alternatives to genetic modification for plant optimization. PMID:26702448

  3. Electronic Elections

    DEFF Research Database (Denmark)

    Schürmann, Carsten

    2009-01-01

    Electronic voting technology is a two edged sword. It comes with many risks but brings also many benefits. Instead of flat out rejecting the technology as uncontrollably dangerous, we advocate in this paper a different technological angle that renders electronic elections trustworthy beyond...... the usual levels of doubt. We exploit the trust that voters currently have into the democratic process and model our techniques around that observation accordingly. In particular, we propose a technique of trace emitting computations to record the individual steps of an electronic voting machine...... for a posteriori validation on an acceptably small trusted computing base. Our technology enables us to prove that an electronic elections preserves the voter’s intent, assuming that the voting machine and the trace verifier are independent....

  4. Electronic commerce

    OpenAIRE

    Zvolánková, Pavla

    2010-01-01

    The thesis deals with a description of electronic commerce from its beginning up to present situation in this area. It explains basic terms connected with electronic commerce and it summarizes the relevant legislation. Moreover it describes e-contracts and rights and duties of both contractual parties. The main view is the view of Internet retailer, which is reflected in the practical part focused on concrete problems of retailers.

  5. Printed Electronics

    Science.gov (United States)

    Wade, Jessica; Hollis, Joseph Razzell; Wood, Sebastian

    2018-04-01

    The combination of printing technology with manufacturing electronic devices enables a new paradigm of printable electronics, where 'smart' functionality can be readily incorporated into almost any product at low cost. Over recent decades, rapid progress has been made in this field, which is now emerging into the industrial andcommercial realm. However, successful development and commercialisation on a large scale presents some significant technical challenges. For fully-printable electronic systems, all the component parts must be deposited from solutions (inks), requiring the development of new inorganic, organic and hybrid materials.A variety of traditional printing techniques are being explored and adapted forprinting these new materials in ways that result in the best performing electronicdevices. Whilst printed electronics research has initially focused on traditional typesof electronic device such as light-emitting diodes, transistors, and photovoltaics, it is increasingly apparent that a much wider range of applications can be realised. The soft and stretchable nature of printable materials makes them perfect candidates forbioelectronics, resulting in a wealth of research looking at biocompatible printable inks and biosensors. Regardless of application, the properties of printed electronicmaterials depend on the chemical structures, processing conditions, device architecture,and operational conditions, the complex inter-relationships of which aredriving ongoing research. We focus on three particular 'hot topics', where attention is currently focused: novel materials, characterisation techniques, and device stability. With progress advancing very rapidly, printed electronics is expected to grow over the next decade into a key technology with an enormous economic and social impact.

  6. UV-induced modifications in the peptidyl transferase loop of 23S rRNA dependent on binding of the streptogramin B antibiotic, pristinamycin IA

    DEFF Research Database (Denmark)

    Porse, B T; Kirillov, S V; Awayez, M J

    1999-01-01

    The naturally occurring streptogramin B antibiotic, pristinamycin IA, which inhibits peptide elongation, can produce two modifications in 23S rRNA when bound to the Escherichia coli 70S ribosome and irradiated at 365 nm. Both drug-induced effects map to highly conserved nucleotides within...... in the latter modification to A2062/C2063. Pristinamycin IA can also produce a modification on binding to deproteinized, mature 23S rRNA, at position U2500/C2501. The same modification occurs on an approximately 37-nt fragment, encompassing positions approximately 2496-2532 of the peptidyl transferase loop...... the sequence Cm-C-U-C-G-m2A-psi-G2505 are important for pristinamycin IA binding and/or the antibiotic-dependent modification of 23S rRNA....

  7. Identification of an enhancer element of class Pi glutathione S-transferase gene required for expression by a co-planar polychlorinated biphenyl.

    Science.gov (United States)

    Matsumoto, M; Imagawa, M; Aoki, Y

    1999-01-01

    3,3',4,4',5-Pentachlorobiphenyl (PenCB), one of the most toxic co-planar polychlorinated biphenyl congeners, specifically induces class Pi glutathione S-transferase (GSTP1) as well as cytochrome P-450 1A1 in primary cultured rat liver parenchymal cells [Aoki, Matsumoto and Suzuki (1993) FEBS Lett. 333, 114-118]. However, the 5'-flanking sequence of the GSTP1 gene does not contain a xenobiotic responsive element, to which arylhydrocarbon receptor binds. Using a chloramphenicol acetyltransferase assay we demonstrate here that the enhancer termed GSTP1 enhancer I (GPEI) is necessary for the stimulation by PenCB of GSTP1 gene expression in primary cultured rat liver parenchymal cells. GPEI is already known to contain a dyad of PMA responsive element-like elements oriented palindromically. It is suggested that a novel signal transduction pathway activated by PenCB contributes to the stimulation of GSTP1 expression. PMID:10051428

  8. Liver Melanomacrophages and Gluthation S-Transferase Activity in Leptodactylus chaquensis (ANURA, LEPTODACTYLIDAE as Biomarkers of Oxidative Stress Due to Chlorpyrifos Exposition

    Directory of Open Access Journals (Sweden)

    Ivan Huespe

    2017-05-01

    Full Text Available We quantified and compared the hepatic melanomacrophage (MM and glutathione S-transferase (GST enzyme activity (two oxidative stress biomarkers in the liver of Leptodatylus chaquensis adults (Anura, Leptodactylidae collected in a rice field (CA in San Javier department, Santa Fe (Argentina, seven days after the application of chlorpyrifos and in a reference site (SR. The histological analysis revealed a significant amount (p = 0.028 and area occupied by MM (p = 0.017 in livers of CA compared to SR. Furthermore, a significant inhibition of GST activity was recorded in the CA frogs compared to the SR (p = 0.030. The histopathological and enzymatic effects provide evidences of ecotoxicological risk for anurans in rice field with CPF application.

  9. Mutations in 23S rRNA at the Peptidyl Transferase Center and Their Relationship to Linezolid Binding and Cross-Resistance

    DEFF Research Database (Denmark)

    Long, Katherine; Munck, Christian

    2010-01-01

    The oxazolidinone antibiotic linezolid targets the peptidyl transferase center (PTC) on the bacterial ribosome. Thirteen single and four double 23S rRNA mutations were introduced into a Mycobacterium smegmatis strain with a single rRNA operon. Converting bacterial base identity by single mutations...... at positions 2032, 2453, and 2499 to human cytosolic base identity did not confer significantly reduced susceptibility to linezolid. The largest decrease in linezolid susceptibility for any of the introduced single mutations was observed with the G2576U mutation at a position that is 7.9 Å from linezolid....... Smaller decreases were observed with the A2503G, U2504G, and G2505A mutations at nucleotides proximal to linezolid, showing that the degree of resistance conferred is not simply inversely proportional to the nucleotide-drug distance. The double mutations G2032A-C2499A, G2032A-U2504G, C2055A-U2504G, and C...

  10. Comparative structural analysis of a novel glutathioneS-transferase (ATU5508) from Agrobacterium tumefaciens at 2.0 A resolution.

    Science.gov (United States)

    Kosloff, Mickey; Han, Gye Won; Krishna, S Sri; Schwarzenbacher, Robert; Fasnacht, Marc; Elsliger, Marc-André; Abdubek, Polat; Agarwalla, Sanjay; Ambing, Eileen; Astakhova, Tamara; Axelrod, Herbert L; Canaves, Jaume M; Carlton, Dennis; Chiu, Hsiu-Ju; Clayton, Thomas; DiDonato, Michael; Duan, Lian; Feuerhelm, Julie; Grittini, Carina; Grzechnik, Slawomir K; Hale, Joanna; Hampton, Eric; Haugen, Justin; Jaroszewski, Lukasz; Jin, Kevin K; Johnson, Hope; Klock, Heath E; Knuth, Mark W; Koesema, Eric; Kreusch, Andreas; Kuhn, Peter; Levin, Inna; McMullan, Daniel; Miller, Mitchell D; Morse, Andrew T; Moy, Kin; Nigoghossian, Edward; Okach, Linda; Oommachen, Silvya; Page, Rebecca; Paulsen, Jessica; Quijano, Kevin; Reyes, Ron; Rife, Christopher L; Sims, Eric; Spraggon, Glen; Sridhar, Vandana; Stevens, Raymond C; van den Bedem, Henry; Velasquez, Jeff; White, Aprilfawn; Wolf, Guenter; Xu, Qingping; Hodgson, Keith O; Wooley, John; Deacon, Ashley M; Godzik, Adam; Lesley, Scott A; Wilson, Ian A

    2006-11-15

    Glutathione S-transferases (GSTs) comprise a diverse superfamily of enzymes found in organisms from all kingdoms of life. GSTs are involved in diverse processes, notably small-molecule biosynthesis or detoxification, and are frequently also used in protein engineering studies or as biotechnology tools. Here, we report the high-resolution X-ray structure of Atu5508 from the pathogenic soil bacterium Agrobacterium tumefaciens (atGST1). Through use of comparative sequence and structural analysis of the GST superfamily, we identified local sequence and structural signatures, which allowed us to distinguish between different GST classes. This approach enables GST classification based on structure, without requiring additional biochemical or immunological data. Consequently, analysis of the atGST1 crystal structure suggests a new GST class, distinct from previously characterized GSTs, which would make it an attractive target for further biochemical studies. (c) 2006 Wiley-Liss, Inc.

  11. Role of household exposure, dietary habits and glutathione S-Transferases M1, T1 polymorphisms in susceptibility to lung cancer among women in Mizoram India.

    Science.gov (United States)

    Phukan, Rup Kumar; Saikia, Bhaskar Jyoti; Borah, Prasanta Kumar; Zomawia, Eric; Sekhon, Gaganpreet Singh; Mahanta, Jagadish

    2014-01-01

    A case-control study was conducted to evaluate the effect of household exposure, dietary habits, smoking and Glutathione S-Transferases M1, T1 polymorphisms on lung cancer among women in Mizoram, India. We selected 230 newly diagnosed primary lung cases and 460 controls from women in Mizoram. Multivariate logistic regression analysis was performed to estimate adjusted odds ratio (OR). Exposure of cooking oil fumes (pkitchen inside living room (p=0.001), improper ventilated house (p=0.003), roasting of soda in kitchen (p=0.001), current smokers of tobacco (p=0.043), intake of smoked fish (p=0.006), smoked meat (p=0.001), Soda (poil emission and wood smoke, intake of smoked meat, smoked fish and soda (an alkali preparation used as food additives in Mizoram) and tobacco consumption for increase risk of lung cancer among Women in Mizoram.

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

    Directory of Open Access Journals (Sweden)

    Rekoslavskaya N.I.

    2012-08-01

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

  13. Caribbean yellow band disease compromises the activity of catalase and glutathione S-transferase in the reef-building coral Orbicella faveolata exposed to anthracene.

    Science.gov (United States)

    Montilla, Luis Miguel; Ramos, Ruth; García, Elia; Cróquer, Aldo

    2016-05-03

    Healthy and diseased corals are threatened by different anthropogenic sources, such as pollution, a problem expected to become more severe in the near future. Despite the fact that coastal pollution and coral diseases might represent a serious threat to coral reef health, there is a paucity of controlled experiments showing whether the response of diseased and healthy corals to xenobiotics differs. In this study, we exposed healthy and Caribbean yellow band disease (CYBD)-affected Orbicella faveolata colonies to 3 sublethal concentrations of anthracene to test if enzymatic responses to this hydrocarbon were compromised in CYBD-affected tissues. For this, a 2-factorial fully orthogonal design was used in a controlled laboratory bioassay, using tissue condition (2 levels: apparently healthy and diseased) and pollutant concentration (4 levels: experimental control, 10, 30 and 100 ppb concentration) as fixed factors. A permutation-based ANOVA (PERMANOVA) was used to test the effects of condition and concentration on the specific activity of 3 enzymatic biomarkers: catalase, glutathione S-transferase, and glutathione peroxidase. We found a significant interaction between the concentration of anthracene and the colony condition for catalase (Pseudo-F = 3.84, df = 3, p < 0.05) and glutathione S-transferase (Pseudo-F = 3.29, df = 3, p < 0.05). Moreover, our results indicated that the enzymatic response to anthracene in CYBD-affected tissues was compromised, as the activity of these enzymes decreased 3- to 4-fold compared to healthy tissues. These results suggest that under a potential scenario of increasing hydrocarbon coastal pollution, colonies of O. faveolata affected with CYBD might become more vulnerable to the deleterious effects of chemical pollution.

  14. Copy number variation in glutathione-S-transferase T1 and M1 predicts incidence and 5-year survival from prostate and bladder cancer, and incidence of corpus uteri cancer in the general population

    DEFF Research Database (Denmark)

    Nørskov, M S; Frikke-Schmidt, R; Bojesen, S E

    2011-01-01

    Glutathione-S-transferase T1 (GSTT1) and GSTM1 detoxify carcinogens and thus potentially contribute to inter-individual susceptibility to cancer. We determined the ability of GST copy number variation (CNV) to predict the risk of cancer in the general population. Exact copy numbers of GSTT1 and G...

  15. Succinyl-CoA:3-ketoacid CoA transferase (SCOT): cloning of the human SCOT gene, tertiary structural modeling of the human SCOT monomer, and characterization of three pathogenic mutations

    NARCIS (Netherlands)

    Fukao, T.; Mitchell, G. A.; Song, X. Q.; Nakamura, H.; Kassovska-Bratinova, S.; Orii, K. E.; Wraith, J. E.; Besley, G.; Wanders, R. J.; Niezen-Koning, K. E.; Berry, G. T.; Palmieri, M.; Kondo, N.

    2000-01-01

    The activity of succinyl-CoA:3-ketoacid CoA transferase (SCOT; locus symbol OXCT; EC 2.8.3.5) is the main determinant of the ketolytic capacity of tissues. Hereditary SCOT deficiency causes episodic ketoacidosis. Here we describe the human SCOT gene, which spans more than 100 kb and contains 17

  16. Mucin-type O-glycosylation is controlled by short- and long-range glycopeptide substrate recognition that varies among members of the polypeptide GalNAc transferase family

    DEFF Research Database (Denmark)

    Revoredo, Leslie; Wang, Shengjun; Bennett, Eric Paul

    2016-01-01

    A large family of UDP-GalNAc:polypeptide GalNAc transferases (ppGalNAc-Ts) initiates and defines sites of mucin-type Ser/Thr-O-GalNAc glycosylation. Family members have been classified into peptide- and glycopeptide-preferring subfamilies, although both families possess variable activities agains...

  17. Molecular Electronics

    DEFF Research Database (Denmark)

    Jennum, Karsten Stein

    This thesis includes the synthesis and characterisation of organic compounds designed for molecular electronics. The synthesised organic molecules are mainly based on two motifs, the obigo(phenyleneethynylenes) (OPE)s and tetrathiafulvalene (TTF) as shown below. These two scaffolds (OPE and TTF......) are chemically merged together to form cruciform-like structures that are an essential part of the thesis. The cruciform molecules were subjected to molecular conductance measurements to explore their capability towards single-crystal field-effect transistors (Part 1), molecular wires, and single electron......, however, was obtained by a study of a single molecular transistor. The investigated OPE5-TTF compound was captured in a three-terminal experiment, whereby manipulation of the molecule’s electronic spin was possible in different charge states. Thus, we demonstrated how the cruciform molecules could...

  18. Electron tube

    Science.gov (United States)

    Suyama, Motohiro [Hamamatsu, JP; Fukasawa, Atsuhito [Hamamatsu, JP; Arisaka, Katsushi [Los Angeles, CA; Wang, Hanguo [North Hills, CA

    2011-12-20

    An electron tube of the present invention includes: a vacuum vessel including a face plate portion made of synthetic silica and having a surface on which a photoelectric surface is provided, a stem portion arranged facing the photoelectric surface and made of synthetic silica, and a side tube portion having one end connected to the face plate portion and the other end connected to the stem portion and made of synthetic silica; a projection portion arranged in the vacuum vessel, extending from the stem portion toward the photoelectric surface, and made of synthetic silica; and an electron detector arranged on the projection portion, for detecting electrons from the photoelectric surface, and made of silicon.

  19. Spin electronics

    CERN Document Server

    Buhrman, Robert; Daughton, James; Molnár, Stephan; Roukes, Michael

    2004-01-01

    This report is a comparative review of spin electronics ("spintronics") research and development activities in the United States, Japan, and Western Europe conducted by a panel of leading U.S. experts in the field. It covers materials, fabrication and characterization of magnetic nanostructures, magnetism and spin control in magnetic nanostructures, magneto-optical properties of semiconductors, and magnetoelectronics and devices. The panel's conclusions are based on a literature review and a series of site visits to leading spin electronics research centers in Japan and Western Europe. The panel found that Japan is clearly the world leader in new material synthesis and characterization; it is also a leader in magneto-optical properties of semiconductor devices. Europe is strong in theory pertaining to spin electronics, including injection device structures such as tunneling devices, and band structure predictions of materials properties, and in development of magnetic semiconductors and semiconductor heterost...

  20. Electronic Commerce

    Energy Technology Data Exchange (ETDEWEB)

    Laird, N. [NRG Information Services Inc., Calgary, AB (Canada)

    1995-11-01

    The concept of electronic commerce in the gas industry was discussed. It was defined as the integration of communication technology, advanced information processing capability and business standards, to improve effectiveness of the business process. Examples of electronic data interchange from the automotive, airline, and banking industry were given. The objective of using this technology in the gas industry was described as the provision of one electronic facility to make seamless contractual and operational arrangements for moving natural gas across participating pipelines. The benefit of seamless integration - one readily available standard system used by several companies - was highlighted. A list of value-added services such as the free movement of bulletins, directories, nominations,and other documents was provided.

  1. Electron accelerator

    International Nuclear Information System (INIS)

    Abramyan.

    1981-01-01

    The USSR produces an electron accelerator family of a simple design powered straight from the mains. The specifications are given of accelerators ELITA-400, ELITA-3, ELT-2, TEUS-3 and RIUS-5 with maximum electron energies of 0.3 to 5 MeV, a mean power of 10 to 70 kW operating in both the pulsed and the continuous (TEUS-3) modes. Pulsed accelerators ELITA-400 and ELITA-3 and RIUS-5 in which TESLA resonance transformers are used are characterized by their compact size. (Ha)

  2. Electronic cigarette

    OpenAIRE

    Wang, Tao

    2016-01-01

    As we know E-cigarette is becoming increasingly popular all over the world. It is a new product that the most of smoking people would like to buy and use. However, we are not realizing advantages and disadvantages of e-cigarette clearly. My objective was to research the development of electronic cigarette whether it is under control or a good way of marketing. The thesis has two main parts. They include answers to questions what is electronic cigarette and how to manage the whole industry...

  3. Electronic School.

    Science.gov (United States)

    Executive Educator, 1994

    1994-01-01

    This issue of "The Electronic School" features a special forum on computer networking. Articles specifically focus on network operating systems, cabling requirements, and network architecture. Tom Wall argues that virtual reality is not yet ready for classroom use. B.J. Novitsky profiles two high schools experimenting with CD-ROM…

  4. Electronic Government

    DEFF Research Database (Denmark)

    Wimmer, Maria A.; Traunmüller, Roland; Grönlund, Åke

    This book constitutes the refereed proceedings of the 4th International Conference on Electronic Government, EGOV 2005, held in Copenhagen, Denmark, in August 2005. The 30 revised papers presented were carefully reviewed and selected from numerous submissions, and assess the state-of-the-art in e-government/e-governance...

  5. Electronics department

    International Nuclear Information System (INIS)

    1979-01-01

    This report summarizes the activities in 1978 of some of the groups within the Electronics Department. The work covered includes plant protection and operator studies, reliability techniques, application of nuclear techniques to mineral exploration, applied laser physics, computing and, lastly, research instrumentation. (author)

  6. Power electronics

    Indian Academy of Sciences (India)

    Kishore Chatterjee

    This special issue of Sadhana is a compilation of papers selected from those presented at the 7th National Power. Electronics Conference (NPEC), held at the Indian Institute of Technology, Bombay, on 21–23 December 2015. From among the papers presented in NPEC-2017, selected papers were peer-reviewed for ...

  7. Electron linacs

    Energy Technology Data Exchange (ETDEWEB)

    Loew, G A; Schriber, S O [ed.

    1976-11-01

    A study was made of the present status of the thousand or so electron linacs in the world, and future trends in the field. These machines were classified according to their use: medical, industrial, and nuclear physics. In the medical category, two types of electron linacs are discussed: the conventional ones which are used for x-ray and electron therapy, and those which may in the future be used for negative pion therapy. Industrial machines discussed include linacs for radiographic and other specialized applications. In the nuclear physics category, the status of conventional low- and medium-energy as well as high duty cycle linacs is reviewed. The question of how one might obtain a c-w, 1 GeV, 100..mu..A electron linac is raised, and various options using recirculation and stretchers are examined. In this connection, the status of rf superconductivity is summarized. A review is given of linacs for injectors into synchrotrons and e/sup +-/ storage rings, and recent work done to upgrade the only multi-GeV linac, namely SLAC, is described.

  8. Greening Electronics

    DEFF Research Database (Denmark)

    Pizzol, Massimo; Søes Kokborg, Morten; Thomsen, Marianne

    Based on a literature review with focus on hazardous substances in waste electric and electronic equipment (WEEE) and numbers from a Danish treatment facility a flow analysis for specific substances has been conducted. Further, the accessible knowledge on human and environmental effects due...

  9. Electronic seal

    International Nuclear Information System (INIS)

    Musyck, E.

    1981-01-01

    An electronic seal is presented for a volume such as container for fissile materials. The seal encloses a lock for barring the space as well as a device for the detection and the recording of the intervention of the lock. (AF)

  10. Nuclear electronics

    International Nuclear Information System (INIS)

    Friese, T.

    1981-09-01

    A short survey is given on nuclear radiation detectors and nuclear electronics. It is written for newcomers and those, who are not very familiar with this technique. Some additional information is given on typical failures in nuclear measurement systems. (orig.) [de

  11. Electron linacs

    International Nuclear Information System (INIS)

    Loew, G.A.

    1976-01-01

    To study the present status of the thousand or so electron linacs in the world, and future trends in the field, we have classified these machines according to their use: medical, industrial, and nuclear physics. In the medical category, two types of electron linacs are discussed: the conventional ones which are used for X-ray and electron therapy, and those which may in the future be used for negative pion therapy. The section on industrial machines includes linacs for radiographic and other specialized applications. In the nuclear physics category, the status of conventional low- and medium-energy as well as high duty cycle linacs is reviewed. The question of how one might obtain a C.W., 1 GeV, 100 μA electron linac is raised and various options using recirculation and stretchers are examined. In this connection, the status of RF superconductivity is summarized. Following, there is a review of linacs for injectors into synchrotrons and e +- storage rings. The paper ends with a description of recent work done to upgrade the only multi-GeV linac, namely SLAC. (author)

  12. Electronic Nose and Electronic Tongue

    Science.gov (United States)

    Bhattacharyya, Nabarun; Bandhopadhyay, Rajib

    Human beings have five senses, namely, vision, hearing, touch, smell and taste. The sensors for vision, hearing and touch have been developed for several years. The need for sensors capable of mimicking the senses of smell and taste have been felt only recently in food industry, environmental monitoring and several industrial applications. In the ever-widening horizon of frontier research in the field of electronics and advanced computing, emergence of electronic nose (E-Nose) and electronic tongue (E-Tongue) have been drawing attention of scientists and technologists for more than a decade. By intelligent integration of multitudes of technologies like chemometrics, microelectronics and advanced soft computing, human olfaction has been successfully mimicked by such new techniques called machine olfaction (Pearce et al. 2002). But the very essence of such research and development efforts has centered on development of customized electronic nose and electronic tongue solutions specific to individual applications. In fact, research trends as of date clearly points to the fact that a machine olfaction system as versatile, universal and broadband as human nose and human tongue may not be feasible in the decades to come. But application specific solutions may definitely be demonstrated and commercialized by modulation in sensor design and fine-tuning the soft computing solutions. This chapter deals with theory, developments of E-Nose and E-Tongue technology and their applications. Also a succinct account of future trends of R&D efforts in this field with an objective of establishing co-relation between machine olfaction and human perception has been included.

  13. Electronics Industry

    Science.gov (United States)

    2007-01-01

    countries in developing market nations in Asia (such as Korea, Taiwan, Singapore, Malaysia , China and Vietnam). The competition for the knowledge, economic...Intel, Infineon Technologies, STMicroelectronics, Samsung Electronics, Texas Instruments, AMD Spansion, Philips Semiconductor, Freescale... Samsung ($19.7B), #5 Toshiba ($9.8B), #6 TSMC ($9.7B), #7 Hynix ($8.0B) and #8 Renesas ($7.9B) (McGrath, 2007, p. 3). Samsung , headquartered in

  14. Electronic banking

    OpenAIRE

    Gradišnik, Monika

    2017-01-01

    The development of information and communication technology is one of the most important reasons for the incredibly fast changes in business. Electronic commerce is spreading unstoppably in the operations of companies. The creation of new models, such as online banking, online shopping and the like, has sped up the development of the World Wide Web. Owing to the rapid progress of the World Wide Web and technologies for secure business operations, we can barely imagine life today without e...

  15. Electronic Aggression

    Centers for Disease Control (CDC) Podcasts

    2007-11-20

    Aggression is no longer limited to the school yard. New forms of electronic media, such as blogs, instant messaging, chat rooms, email, text messaging, and the internet are providing new arenas for youth violence to occur.  Created: 11/20/2007 by National Center for Injury Prevention and Control, Division of Violence Prevention.   Date Released: 11/28/2007.

  16. ELECTRON GUN

    Science.gov (United States)

    Christofilos, N.C.; Ehlers, K.W.

    1960-04-01

    A pulsed electron gun capable of delivering pulses at voltages of the order of 1 mv and currents of the order of 100 amperes is described. The principal novelty resides in a transformer construction which is disposed in the same vacuum housing as the electron source and accelerating electrode structure of the gun to supply the accelerating potential thereto. The transformer is provided by a plurality of magnetic cores disposed in circumferentially spaced relation and having a plurality of primary windings each inductively coupled to a different one of the cores, and a helical secondary winding which is disposed coaxially of the cores and passes therethrough in circumferential succession. Additional novelty resides in the disposition of the electron source cathode filament input leads interiorly of the transformer secondary winding which is hollow, as well as in the employment of a half-wave filament supply which is synchronously operated with the transformer supply such that the transformer is pulsed during the zero current portions of the half-wave cycle.

  17. Electronic sputtering

    International Nuclear Information System (INIS)

    Johnson, R.E.

    1989-01-01

    Electronic sputtering covers a range of phenomena from electron and photon stimulated desorption from multilayers to fast heavy ion-induced desorption (sputtering) of biomolecules. In this talk the author attempted. Therefore, to connect the detailed studies of argon ejection from solid argon by MeV ions and keV electrons to the sputtering of low temperatures molecular ices by MeV ions then to biomolecule ejection from organic solids. These are related via changing (dE/dx) e , molecular size, and transport processes occurring in materials. In this regard three distinct regions of (dE/dx) e have been identified. Since the talk this picture has been made explicit using a simple spike model for individual impulsive events in which spike interactions are combined linearly. Since that time also the molecular dynamics programs (at Virginia and Uppsala) have quantified both single atom and dimer processes in solid Ar and the momentum transport in large biomolecule sputtering. 5 refs

  18. Glutathione S-transferase genes and the risk of type 2 diabetes mellitus: Role of sexual dimorphism, gene-gene and gene-smoking interactions in disease susceptibility.

    Science.gov (United States)

    Azarova, Iuliia; Bushueva, Olga; Konoplya, Alexander; Polonikov, Alexey

    2018-05-01

    Compromised defense against reactive oxygen species (ROS) is considered important in the pathogenesis of type 2 diabetes mellitus (T2DM); therefore, genes encoding antioxidant defense enzymes may contribute to disease susceptibility. This study investigated whether polymorphisms in genes encoding glutathione S-transferase M1 (GSTM1), T1 (GSTT1), and P1 (GSTP1) jointly contribute to the risk of T2DM. In all, 1120 unrelated Russian subjects (600 T2DM patients, 520 age- and sex-matched healthy subjects), were recruited to the study. Genotyping was performed by multiplex polymerase chain reaction (PCR; del/del polymorphisms of GSTM1 and GSTT1) and TaqMan-based PCR (polymorphisms I105V and A114V of GSTP1). Plasma ROS and glutathione levels in study subjects were analyzed by fluorometric and colorimetric assays, respectively. Genotype del/del GSTT1 was significantly associated with the risk of T2DM (odds ratio [OR] 1.60, 95% confidence interval [CI] 1.17-2.21, P = 0.003). Gender-stratified analysis showed that the deletion genotypes of GSTM1 (OR 1.99, 95% CI 1.30-3.05; P = 0.0002, Q = 0.016) and GSTT1 (OR 2.23, 95% CI 1.22-4.09; P = 0.008, Q = 0.0216), as well as genotype 114A/V of GSTP1 (OR 2.85, 95% CI 1.44-5.62; P = 0.005, Q = 0.02) were associated with an increased risk of T2DM exclusively in males. Three genotype combinations (i.e. GSTM1+ × GSTT1+, GSTM1+ × GSTP1 114A/A and GSTT1+ × GSTP1 114A/A) showed significant associations with a decreased risk of T2DM in males. This study demonstrates, for the first time, that genes encoding glutathione S-transferases jointly contribute to the risk of T2DM, and that their effects on disease susceptibility are gender specific. © 2017 Ruijin Hospital, Shanghai Jiaotong University School of Medicine and John Wiley & Sons Australia, Ltd.

  19. Detectors - Electronics

    International Nuclear Information System (INIS)

    Bregeault, J.; Gabriel, J.L.; Hierle, G.; Lebotlan, P.; Leconte, A.; Lelandais, J.; Mosrin, P.; Munsch, P.; Saur, H.; Tillier, J.

    1998-01-01

    The reports presents the main results obtained in the fields of radiation detectors and associated electronics. In the domain of X-ray gas detectors for the keV range efforts were undertaken to rise the detector efficiency. Multiple gap parallel plate chambers of different types as well as different types of X → e - converters were tested to improve the efficiency (values of 2.4% at 60 KeV were reached). In the field of scintillators a study of new crystals has been carried out (among which Lutetium orthosilicate). CdTe diode strips for obtaining X-ray imaging were studied. The complete study of a linear array of 8 CdTe pixels has been performed and certified. The results are encouraging and point to this method as a satisfying solution. Also, a large dimension programmable chamber was used to study the influence of temperature on the inorganic scintillators in an interval from -40 deg. C to +150 deg. C. Temperature effects on other detectors and electronic circuits were also investigated. In the report mentioned is also the work carried out for the realization of the DEMON neutron multidetector. For neutron halo experiments different large area Si detectors associated with solid and gas position detectors were realized. In the frame of a contract with COGEMA a systematic study of Li doped glasses was undertaken aiming at replacing with a neutron probe the 3 He counters presently utilized in pollution monitoring. An industrial prototype has been realised. Other studies were related to integrated analog chains, materials for Cherenkov detectors, scintillation probes for experiments on fundamental processes, gas position sensitive detectors, etc. In the field of associated electronics there are mentioned the works related to the multidetector INDRA, data acquisition, software gamma spectrometry, automatic gas pressure regulation in detectors, etc

  20. Electronic materials

    CERN Document Server

    Kwok, H L

    2010-01-01

    The electronic properties of solids have become of increasing importance in the age of information technology. The study of solids and materials, while having originated from the disciplines of physics and chemistry, has evolved independently over the past few decades. The classical treatment of solid-state physics, which emphasized classifications, theories and fundamental physical principles, is no longer able to bridge the gap between materials advances and applications. In particular, the more recent developments in device physics and technology have not necessarily been driven by new conc

  1. Electronic wastes

    Science.gov (United States)

    Regel-Rosocka, Magdalena

    2018-03-01

    E-waste amount is growing at about 4% annually, and has become the fastest growing waste stream in the industrialized world. Over 50 million tons of e-waste are produced globally each year, and some of them end up in landfills causing danger of toxic chemicals leakage over time. E-waste is also sent to developing countries where informal processing of waste electrical and electronic equipment (WEEE) causes serious health and pollution problems. A huge interest in recovery of valuable metals from WEEE is clearly visible in a great number of scientific, popular scientific publications or government and industrial reports.

  2. Electron gun

    International Nuclear Information System (INIS)

    Chen, H.-Y.; Hughes, R.H.

    1979-01-01

    The invention described relates to cathode ray tubes, and particularly to color picture tubes of the type useful in home television receivers and therefore to electron guns. The invention is especially applicable to self-converging tube-yoke combinations with shadow mask tubes of the type having plural-beam in-line guns disposed in a horizontal plane, an apertured mask with vertically oriented slit-shaped apertures, and a screen with vertically oriented phosphor stripes. The invention is not, however, limited to use in such tubes and may in fact be used, e.g., in dot-type shadow mask tubes and index-type tubes. (Auth.)

  3. Bolometer electronics

    International Nuclear Information System (INIS)

    Groenig, D.E.

    1981-01-01

    High quality is required to the electronic which works with bolometer made of metal for measuring the radiation power in plasmaphysical experiments. If the bandwidth is to be 1 kHz, and the time constant of the bolometer is about 160 ms by high overall gain the critical parameters are the noise of the amplifier, pick up to the system, stability and decoupling of common mode signals. The high overall gain is necessary to be able to measure lowest radiation power. The design made is a good approach to the desired property. (orig.) [de

  4. Basic electronics

    CERN Document Server

    Tayal, DC

    2010-01-01

    The second edition of this book incorporates the comments and suggestions of my friends and students who have critically studied the first edition. In this edition the changes and additions have been made and subject matter has been rearranged at some places. The purpose of this text is to provide a comprehensive and up-to-date study of the principles of operation of solid state devices, their basic circuits and application of these circuits to various electronic systems, so that it can serve as a standard text not only for universities and colleges but also for technical institutes. This book

  5. Nuclear electronics

    International Nuclear Information System (INIS)

    Lucero B, E.

    1989-01-01

    The rapid technical development of Colombia over the past years, resulted among others, a considerable increase in the number of measuring instrumentation and testing laboratories, scientific research and metrology centers, in industry, agriculture, public health, education on the nuclear field, etc. IAN is a well organized institution with qualified management, trained staff and reasonably equipped laboratories to carry out tasks as: Metrology, standardization, quality control and maintenance and repair of nuclear instruments. The government of Colombia has adopted a policy to establish and operate through the country maintenance and repair facilities for nuclear instrumentation. This policy is reflected in the organization of electronic laboratories in Bogota-IAN

  6. Isolation of the human anionic glutathione S-transferase cDNA and the relation of its gene expression to estrogen-receptor content in primary breast cancer

    International Nuclear Information System (INIS)

    Moscow, J.A.; Townsend, A.J.; Goldsmith, M.E.; Whang-Peng, J.; Vickers, P.J.; Poisson, R.; Legault-Poisson, S.; Myers, C.E.; Cowan, K.H.

    1988-01-01

    The development of multidrug resistance in MCF7 human breast cancer cells is associated with overexpression of P-glycoprotein, changes in activities of several detoxication enzymes, and loss of hormone sensitivity and estrogen receptors (ERs). The authors have cloned the cDNA for one of the drug-detoxifying enzymes overexpressed in multidrug-resistant MCF7 cells (Adr R MCF7), the anionic isozyme of glutathione S-transferase (GSTπ). Hybridization with this GSTπ cDNA, GSTπ-1, demonstrated that increased GSTπ activity in Adr R MCF7 cells is associated with overexpression but not with amplification of the gene. They mapped the GSTπ gene to human chromosome 11q13 by in situ hybridization. Since multidrug resistance and GSTπ overexpression are associated with the loss of ERs in Adr R MCF7 cells, they examined several other breast cancer cell lines that were not selected for drug resistance. In each of these cell lines they found an inverse association between GSTπ expression and ER content. They also examined RNA from 21 primary breast cancers and found a similar association between GSTπ expression and ER content in vivo. The finding of similar patterns of expression of a drug-detoxifying enzyme and of ERs in vitro as well as in vivo suggests that ER-negative breast cancer cells may have greater protection against antineoplastic agents conferred by GSTπ than ER-positive tumors

  7. Molecular cloning of a cDNA and chromosomal localization of a human theta-class glutathione S-transferase gene (GSTT2) to chromosome 22

    Energy Technology Data Exchange (ETDEWEB)

    Tan, K.L.; Baker, R.T.; Board, P.G. [Australian National Univ., Canberra (Australia)] [and others

    1995-01-20

    Until recently the Theta-class glutathione S-transferases (GSTs) were largely overlooked due to their low activity with the model substrate 1-chloro-2,4-dinitrobenzene (CDNB) and their failure to bind to immobilized glutathione affinity matrices. Little is known about the number of genes in this class. Recently, Pemble et al. reported the cDNA cloning of a human Theta-class GST, termed GSTT1. In this study, we describe the molecular cloning of a cDNA encoding a second human Theta-class GST (GSTT2) from a {lambda}gt11 human liver 5{prime}-stretch cDNA library. The encoded protein contains 244 amino acids and has 78.3% sequence identity with the rat subunit 12 and only 55.0% identity with human GSTT1. GSTT2 has been mapped to chromosome 22 by somatic cell hybrid analysis. The precise position of the gene was localized to subband 22q11.2 by in situ hybridization. The absence of other regions of hybridization suggests that there are no closely related sequences (e.g., reverse transcribed pseudogenes) scattered throughout the genome and that if there are closely related genes, they must be clustered near GSTT2. Southern blot analysis of human DNA digested with BamHI shows that the size of the GSTT2 gene is relatively small, as the coding sequence falls within a 3.6-kb BamHI fragment. 35 refs., 6 figs.

  8. Molecular evolution of Theta-class glutathione transferase for enhanced activity with the anticancer drug 1,3-bis-(2-chloroethyl)-1-nitrosourea and other alkylating agents.

    Science.gov (United States)

    Larsson, Anna-Karin; Shokeer, Abeer; Mannervik, Bengt

    2010-05-01

    Glutathione transferase (GST) displaying enhanced activity with the cytostatic drug 1,3-bis-(2-chloroethyl)-1-nitrosourea (BCNU) and structurally related alkylating agents was obtained by molecular evolution. Mutant libraries created by recursive recombination of cDNA coding for human and rodent Theta-class GSTs were heterologously expressed in Escherichia coli and screened with the surrogate substrate 4-nitrophenethyl bromide (NPB) for enhanced alkyltransferase activity. A mutant with a 70-fold increased catalytic efficiency with NPB, compared to human GST T1-1, was isolated. The efficiency in degrading BCNU had improved 170-fold, significantly more than with the model substrate NPB. The enhanced catalytic activity of the mutant GST was also 2-fold higher with BCNU than wild-type mouse GST T1-1, which is 80-fold more efficient than wild-type human GST T1-1. We propose that GSTs catalyzing inactivation of anticancer drugs may find clinical use in protecting sensitive normal tissues to toxic side-effects in treated patients, and as selectable markers in gene therapy. Copyright 2010 Elsevier Inc. All rights reserved.

  9. Induction of the pi class of glutathione S-transferase by carnosic acid in rat Clone 9 cells via the p38/Nrf2 pathway.

    Science.gov (United States)

    Lin, Chia-Yuan; Wu, Chi-Rei; Chang, Shu-Wei; Wang, Yu-Jung; Wu, Jia-Jiuan; Tsai, Chia-Wen

    2015-06-01

    Induction of phase II enzymes is important in cancer chemoprevention. We compared the effect of rosemary diterpenes on the expression of the pi class of glutathione S-transferase (GSTP) in rat liver Clone 9 cells and the signaling pathways involved. Culturing cells with 1, 5, 10, or 20 μM carnosic acid (CA) or carnosol (CS) for 24 h in a dose-dependent manner increased the GSTP expression. CA was more potent than CS. The RNA level and the enzyme activity of GSTP were also enhanced by CA treatment. Treatment with 10 μM CA highly induced the reporter activity of the enhancer element GPEI. Furthermore, CA markedly increased the translocation of nuclear factor erythroid-2 related factor 2 (Nrf2) from the cytosol to the nucleus after 30 to 60 min. CA the stimulated the protein induction of p38, nuclear Nrf2, and GSTP was diminished in the presence of SB203580 (a p38 inhibitor). In addition, SB203580 pretreatment or silencing of Nrf2 by siRNA suppressed the CA-induced GPEI-DNA binding activity and GSTP protein expression. Knockdown of p38 or Nrf2 by siRNA abolished the activation of p38 and Nrf2 as well as the protein induction and enzyme activity of GSTP by CA. These results suggest that CA up-regulates the expression and enzyme activity of GSTP via the p38/Nrf2/GPEI pathway.

  10. Glutathione S-Transferase as biomarker in Sciades herzbergii (Siluriformes: Ariidae for environmental monitoring: the case study of São Marcos Bay, Maranhão, Brazil

    Directory of Open Access Journals (Sweden)

    Raimunda N.F Carvalho-Neta

    2013-04-01

    Full Text Available The Glutathione S-Transferase (GST activity has been proposed as a biomarker of susceptibility to the presence of potentially damaging xenobiotics in aquatic organisms. The aim of this work was to measure GST activity in the liver of Sciades herzbergii (catfish in order to evaluate the biochemical effects of pollutants. The catfish samples were collected along known pollution gradients areas (A1 and from areas regarded as relatively free of anthropogenic input (A2, in São Marcos Bay, São Luis de Maranhão, Brazil. The variables analyzed in fish were: length, weight, gonadal stages, gonadosomatic index and GST activity. The databases from this analysis were compiled, and generalized linear models were used to analyze the dependence of enzyme activity on the areas of sampling and on selected biological parameters of fish. A significant difference was observed in GST activity in the liver of S. herzbergii in the comparison between fish from the contaminated site and those from the reference site (P < 0.05. Morphometric (length and weight parameters and gonadosomatic index of collected fish were significant in the linear model of GST activity only in the reference site. These results may be due to the activity pattern of the enzyme, which increases with the sexual maturity of the animals in healthy environments. In the contaminated area (A1 these correlations do not exist, probably as a result of the energy used in the biotransformation of the various contaminants.

  11. Variable Levels of Glutathione S-Transferases Are Responsible for the Differential Tolerance to Metolachlor between Maize (Zea mays) Shoots and Roots.

    Science.gov (United States)

    Li, Dongzhi; Xu, Li; Pang, Sen; Liu, Zhiqian; Wang, Kai; Wang, Chengju

    2017-01-11

    Glutathione S-transferases (GSTs) play important roles in herbicide tolerance. However, studies on GST function in herbicide tolerance among plant tissues are still lacking. To explore the mechanism of metolachlor tolerance difference between maize shoots and roots, the effects of metolachlor on growth, GST activity, and the expression of the entire GST gene family were investigated. It was found that this differential tolerance to metolachlor was correlated with contrasting GST activity between the two tissues and can be eliminated by a GST inhibitor. An in vitro metolachlor-glutathione conjugation assay confirmed that the transformation of metolachlor is 2-fold faster in roots than in shoots. The expression analysis of the GST gene family revealed that most GST genes are expressed much higher in roots than shoots, both in control and in metolachlor-treated plants. Taken together, higher level expression of most GST genes, leading to higher GST activity and faster herbicide transformation, appears to be responsible for the higher tolerance to metolachlor of maize roots than shoots.

  12. Glutathione S-transferase M1-null genotype as risk factor for SOS in oxaliplatin-treated patients with metastatic colorectal cancer.

    Science.gov (United States)

    Vreuls, C P H; Olde Damink, S W M; Koek, G H; Winstanley, A; Wisse, E; Cloots, R H E; van den Broek, M A J; Dejong, C H C; Bosman, F T; Driessen, A

    2013-02-19

    Oxaliplatin is used as a neo-adjuvant therapy in hepatic colorectal carcinoma metastasis. This treatment has significant side effects, as oxaliplatin is toxic to the sinusoidal endothelial cells and can induce sinusoidal obstruction syndrome (SOS), which is related to decreased overall survival. Glutathione has an important role in the defence system, catalysed by glutathione S-transferase (GST), including two non-enzyme producing polymorphisms (GSTM1-null and GSTT1-null). We hypothesise that patients with a non-enzyme producing polymorphism have a higher risk of developing toxic injury owing to oxaliplatin. In the nontumour-bearing liver, the presence of SOS was studied histopathologically. The genotype was determined by a semi-nested PCR. Thirty-two of the 55 (58%) patients showed SOS lesions, consisting of 27% mild, 22% moderate and 9% severe lesions. The GSTM1-null genotype was present in 25 of the 55 (46%). Multivariate analysis showed that the GSTM1-null genotype significantly correlated with the presence of (moderate-severe) SOS (P=0.026). The GSTM1-null genotype is an independent risk factor for SOS. This finding allows us, in association with other risk factors, to conceive a potential risk profile predicting whether the patient is at risk of developing SOS, before starting oxaliplatin, and subsequently might result in adjustment of treatment.

  13. Are serum gamma-glutamyl transferase, high-sensitivity C-reactive protein, and ischaemia-modified albumin useful in diagnosing PCOS?

    Science.gov (United States)

    Ozturk, Mustafa; Keskin, Ugur; Ozturk, Ozlem; Ulubay, Mustafa; Alanbay, İbrahim; Aydin, Aytekin; Yenen, Müfit Cemal

    2016-10-01

    We assessed the serum levels of gamma-glutamyl transferase (GGT), high-sensitivity C-reactive protein (hsCRP) and ischaemia-modified albumin (IMA) in patients with polycystic ovary syndrome (PCOS). Fifty-three patients with PCOS were included in our study along with 40 women with no PCOS as the control group. The patients were divided according to their body mass index (BMI). GGT levels were significantly higher in the women with PCOS than the women in the control group (p PCOS women who were normoweight and overweight than the normoweight and overweight women in the control group (p PCOS and the controls or between the normoweight and overweight subgroups. GGT may be associated with the diagnosis of PCOS when the threshold is set at >15.5 U/L. With the application of this threshold, raised GGT levels had 83% sensitivity (95% CI 0.70-0.90) and 67.5% specificity (95% CI 0.52-0.79), for the diagnosis of PCOS. In our study, GGT levels were elevated in the PCOS patients independent of BMI and could thus be an important marker of PCOS.

  14. Identification and characterization of an Apis cerana cerana Delta class glutathione S-transferase gene ( AccGSTD) in response to thermal stress

    Science.gov (United States)

    Yan, Huiru; Jia, Haihong; Wang, Xiuling; Gao, Hongru; Guo, Xingqi; Xu, Baohua

    2013-02-01

    Glutathione S-transferases (GSTs) are members of a multifunctional enzyme super family that plays a pivotal role in both insecticide resistance and protection against oxidative stress. In this study, we identified a single-copy gene, AccGSTD, as being a Delta class GST in the Chinese honey bee ( Apis cerana cerana). A predicted antioxidant response element, CREB, was found in the 1,492-bp 5'-flanking region, suggesting that AccGSTD may be involved in oxidative stress response pathways. Real-time PCR and immunolocalization studies demonstrated that AccGSTD exhibited both developmental- and tissue-specific expression patterns. During development, AccGSTD transcript was increased in adults. The AccGSTD expression level was the highest in the honey bee brain. Thermal stress experiments demonstrated that AccGSTD could be significantly upregulated by temperature changes in a time-dependent manner. It is hypothesized that high expression levels might be due to the increased levels of oxidative stress caused by the temperature challenges. Additionally, functional assays of the recombinant AccGSTD protein revealed that AccGSTD has the capability to protect DNA from oxidative damage. Taken together, these data suggest that AccGSTD may be responsible for antioxidant defense in adult honey bees.

  15. Defective Pollen Wall 2 ( DPW2 ) Encodes an Acyl Transferase Required for Rice Pollen Development

    Energy Technology Data Exchange (ETDEWEB)

    Xu, Dawei [Shanghai Jiao Tong Univ. (China). Joint International Research Lab. of Metabolic and Developmental Sciences; Shi, Jianxin [Shanghai Jiao Tong Univ. (China). Joint International Research Lab. of Metabolic and Developmental Sciences; Rautengarten, Carsten [Univ. of Melbourne (Australia). ARC Centre of Excellence in Plant Cell Walls; Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Joint BioEnergy Inst. and Biological Systems and Engineering Division; Yang, Li [Shanghai Jiao Tong Univ. (China). Joint International Research Lab. of Metabolic and Developmental Sciences; Qian, Xiaoling [Shanghai Jiao Tong Univ. (China). Joint International Research Lab. of Metabolic and Developmental Sciences; Uzair, Muhammad [Shanghai Jiao Tong Univ. (China). Joint International Research Lab. of Metabolic and Developmental Sciences; Zhu, Lu [Shanghai Jiao Tong Univ. (China). Joint International Research Lab. of Metabolic and Developmental Sciences; Luo, Qian [Shanghai Jiao Tong Univ. (China). Joint International Research Lab. of Metabolic and Developmental Sciences; An, Gynheung [Kyung Hee Univ., Yongin (Korea). Crop Biotech Inst.; Waßmann, Fritz [Univ. of Bonn (Germany). Inst. of Cellular and Molecular Botany; Schreiber, Lukas [Univ. of Bonn (Germany). Inst. of Cellular and Molecular Botany; Heazlewood, Joshua L. [Univ. of Melbourne (Australia). ARC Centre of Excellence in Plant Cell Walls; Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Joint BioEnergy Inst. and Biological Systems and Engineering Division; Scheller, Henrik Vibe [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Joint BioEnergy Inst. and Biological Systems and Engineering Division; Univ. of California, Berkeley, CA (United States). Dept. of Plant and Microbial Biology; Hu, Jianping [Michigan State Univ., East Lansing, MI (United States). Dept. of Energy Plant Research Lab.; Zhang, Dabing [Shanghai Jiao Tong Univ. (China). Joint International Research Lab. of Metabolic and Developmental Sciences; Univ. of Adelaide, SA (Australia). School of Agriculture, Food and Wine; Liang, Wanqi [Shanghai Jiao Tong Univ. (China). Joint International Research Lab. of Metabolic and Developmental Sciences

    2016-05-31

    Aliphatic and aromatic lipids are both essential structural components of the plant cuticle, an important interface between the plant and environment. Although cross links between aromatic and aliphatic or other moieties are known to be associated with the formation of leaf cutin and root and seed suberin, the contribution of aromatic lipids to the biosynthesis of anther cuticles and pollen walls remains elusive. In this study, we characterized the rice (Oryza sativa) male sterile mutant, defective pollen wall 2 (dpw2), which showed an abnormal anther cuticle, a defective pollen wall, and complete male sterility. Compared with the wild type, dpw2 anthers have increased amounts of cutin and waxes and decreased levels of lipidic and phenolic compounds. DPW2 encodes a cytoplasmically localized BAHD acyltransferase. In vitro assays demonstrated that recombinant DPW2 specifically transfers hydroxycinnamic acid moieties, using v-hydroxy fatty acids as acyl acceptors and hydroxycinnamoyl-CoAs as acyl donors. Thus, The cytoplasmic hydroxycinnamoyl-CoA:v-hydroxy fatty acid transferase DPW2 plays a fundamental role in male reproduction via the biosynthesis of key components of the anther cuticle and pollen wall.

  16. Glutathione S-transferase M1 and T1 gene polymorphisms with consumption of high fruit-juice and vegetable diet affect antioxidant capacity in healthy adults.

    Science.gov (United States)

    Yuan, Linhong; Zhang, Ling; Ma, Weiwei; Zhou, Xin; Ji, Jian; Li, Nan; Xiao, Rong

    2013-01-01

    To our knowledge, no data have yet shown the combined effects of GSTM1/GSTT1 gene polymorphisms with high consumption of a fruit and vegetable diet on the body's antioxidant capacity. A 2-wk dietary intervention in healthy participants was conducted to test the hypothesis that the antioxidant biomarkers in individuals with different glutathione-S-transferases (GST) genotypes will be different in response to a high fruit-juice and vegetable diet. In our study, 24 healthy volunteers with different GST genotypes (12 GSTM1+/GSTT1+ and 12 GSTM1-/GSTT1- participants) consumed a controlled diet high in fruit-juice and vegetables for 2 wk. Blood and first-void urine specimens were obtained at baseline, 1-wk, and 2-wk intervals. The antioxidant capacity-related biomarkers in blood and urine were observed and recorded at the scheduled times. Erythrocyte GST and glutathione reductase (GR) activities response to a high fruit-juice and vegetable diet are GST genotype-dependent. Two weeks on the high fruit-juice and vegetable diet increased GST and GR activities in the GSTM1+/GSTT1+ group (P juice and vegetable diet than GSTM1-/GSTT1- participants. The diet intervention was effective in enhancing glutathione peroxidase and catalase activities in all participants (P 0.05). The effects of a diet rich in fruit-juice and vegetables on antioxidant capacity were dependent on GSTM1/GSTT1 genotypes. Copyright © 2013 Elsevier Inc. All rights reserved.

  17. Oxidative Stress and Modulatory effects of the root extract of Phlogacanthus tubiflorus on the activity of Glutathione-S-Transferase in Hydrogen Peroxide treated Lymphocyte

    Directory of Open Access Journals (Sweden)

    Ramteke A

    2012-04-01

    Full Text Available Glutathione-S-transferase is one of the important enzyme systems that plays vital role in decomposition of lipid hydro-peroxides formed due to oxidative stress. In the present study GST activity increased in the lymphocytes treated with increasing concentration of H2O2, and decrease in the levels of GSH was observed. For similar treatment conditions LDH activity and MDA levels increased significantly leading to decrease in the cell viability. Treatment of lymphocytes with the root extract of Phlogacanthus tubiflorus (PTE resulted in dose dependent decline in the GST activity and rise in GSH levels. LDH activity and MDA levels also declined that led to the increase of cell viability. Lymphocytes pre-treated with the PTE followed by H2O2 (0.1 and 1% treatment, decline in the activity of GST and increase in GSH levels was observed. Also we have observed decline in the activity of LDH and MDA levels in the lymphocytes for both 0.1 and 1% of H2O2 though the magnitude of change was higher in the lymphocytes pre-treated with the PTE followed with 1% of H2O2 treatment. Significant increase in the cell viability for similar conditions was also observed. These findings suggest protective function of the root extracts might be through modulation of GST activity and levels of GSH and might find application in Chemomodulation in future.

  18. Nickel in Soil Modifies Sensitivity to Diazinon Measured by the Activity of Acetylcholinesterase, Catalase, and Glutathione S-Transferase in Earthworm Eisenia fetida

    Directory of Open Access Journals (Sweden)

    Agnieszka Zawisza-Raszka

    2013-01-01

    Full Text Available Nickel in typical soils is present in a very low concentration, but in the contaminated soils it occurs in locally elevated concentrations. The aim of this study was to examine the effect of nickel in the concentrations of 300 (very high, close to LOEC for reproduction and 900 (extremely high, close to LOEC for mortality mg/kg dry soil on the life history and acetylcholinesterase, catalase, and glutathione S-transferase activities in earthworm Eisenia fetida and to establish how nickel modifies the sensitivity to organophosphorous pesticide—diazinon. Cocoons production and juveniles’ number were significantly lower only in groups exposed to Ni in the concentration of 900 mg/kg dry soil for two months. Diazinon administration diminished the AChE activity in the GI tract and in the body wall. The interaction between diazinon and nickel was observed, and, in consequence, the AChE activity after the pesticide treatment was similar to controls in worms preexposed to nickel. Both pesticide administration and exposure to nickel caused an increase in the GST activity in examined organs and CAT activity in body wall. Both biometric and development data and simple enzymatic analysis, especially the AChE and GST, show a Ni pretreatment effect on the subsequent susceptibility to pesticide.

  19. Purification, molecular cloning and heterologous expression of a glutathione S-transferase involved in insecticide resistance from the rice brown planthopper, Nilaparvata lugens.

    Science.gov (United States)

    Vontas, John G; Small, Graham J; Nikou, Dimitra C; Ranson, Hilary; Hemingway, Janet

    2002-03-01

    A novel glutathione S-transferase (GST)-based pyrethroid resistance mechanism was recently identified in Nilaparvata lugens [Vontas, Small and Hemingway (2001) Biochem. J. 357, 65-72]. To determine the nature of GSTs involved in conferring this resistance, the GSTs from resistant and susceptible strains of N. lugens were partially purified by anion exchange and affinity chromatography. The majority of peroxidase activity, previously correlated with resistance, was confined to the fraction that bound to the affinity column, which was considerably elevated in the resistant insects. A cDNA clone encoding a GST (nlgst1-1) - the first reported GST sequence from Hemiptera with up to 54% deduced amino-acid identity with other insect class I GSTs - was isolated from a pyrethroid-resistant strain. Northern analysis showed that nlgst1-1 was overexpressed in resistant insects. nlgst1-1 was expressed in Escherichia coli, purified and characterized. The ability of the recombinant protein to bind to the S-hexylglutathione affinity matrix, its substrate specificities and its immunological properties confirmed that this GST was one from the elevated subset of N. lugens GSTs. Peroxidase activity of the recombinant nlgst1-1 indicated that it had a role in resistance, through detoxification of lipid peroxidation products induced by pyrethroids. Southern analysis of genomic DNA from the resistant and susceptible strains indicated that GST-based insecticide resistance may be associated with gene amplification in N. lugens.

  20. Human cytosolic glutathione-S-transferases: quantitative analysis of expression, comparative analysis of structures and inhibition strategies of isozymes involved in drug resistance.

    Science.gov (United States)

    Mohana, Krishnamoorthy; Achary, Anant

    2017-08-01

    Glutathione-S-transferase (GST) inhibition is a strategy to overcome drug resistance. Several isoforms of human GSTs are present and they are expressed in almost all the organs. Specific expression levels of GSTs in various organs are collected from the human transcriptome data and analysis of the organ-specific expression of GST isoforms is carried out. The variations in the level of expressions of GST isoforms are statistically significant. The GST expression differs in diseased conditions as reported by many investigators and some of the isoforms of GSTs are disease markers or drug targets. Structure analysis of various isoforms is carried out and literature mining has been performed to identify the differences in the active sites of the GSTs. The xenobiotic binding H site is classified into H1, H2, and H3 and the differences in the amino acid composition, the hydrophobicity and other structural features of H site of GSTs are discussed. The existing inhibition strategies are compared. The advent of rational drug design, mechanism-based inhibition strategies, availability of high-throughput screening, target specific, and selective inhibition of GST isoforms involved in drug resistance could be achieved for the reversal of drug resistance and aid in the treatment of diseases.

  1. Succinyl-CoA:3-ketoacid CoA transferase (SCOT) deficiency: two pathogenic mutations, V133E and C456F, in Japanese siblings.

    Science.gov (United States)

    Song, X Q; Fukao, T; Watanabe, H; Shintaku, H; Hirayama, K; Kassovska-Bratinova, S; Kondo, N; Mitchell, G A

    1998-01-01

    Succinyl-CoA:3-ketoacid CoA transferase (SCOT; EC 2.8.3.5; locus symbol OXCT) is the key enzyme of ketone body utilization. Hereditary SCOT deficiency (MIM 245050) causes episodes of severe ketoacidosis. We developed a transient expression system for mutant SCOT cDNAs, using immortalized SCOT-deficient fibroblasts. This paper describes and characterizes three missense mutations in two SCOT-deficient siblings from Japan. They are genetic compounds who inherited the mutation C456F (c1367 G-->T) from their mother. Their paternal allele contains two mutations in cis, T58M (c173 C-->T) and V133E (c398T-->A). Expression of SCOT cDNAs containing either V133E or C456F produces no detectable SCOT activity, whereas T58M is functionally neutral. T58M is a rare sequence variant not detected in 100 control Japanese alleles. In fibroblasts from the proband (GS02), in whom immunoblot demonstrated no detectable SCOT peptide, we measured an apparent residual SCOT activity of 20-35%. We hypothesize that the high residual SCOT activity in homogenates may be an artifact caused by use of the substrate, acetoacetyl-CoA by other enzymes. Expression of mutant SCOT cDNAs more accurately reflects the residual activity of SCOT than do currently available assays in cell or tissue homogenates.

  2. Regulation of epigenetic traits of the glutathione S-transferase P1 gene:From detoxification towards cancer prevention and diagnosis

    Directory of Open Access Journals (Sweden)

    Marc eDiederich

    2014-07-01

    Full Text Available Glutathione S-transferases (GSTs are phase II drug detoxifying enzymes that play an essential role in maintenance of cell integrity and protection against DNA damage by catalyzing the conjugation of glutathione to a wide variety of exo- and endogenous electrophilic substrates. GSTP1, the gene encoding the pi­class GST is frequently inactivated by acquired somatic CpG island promoter hypermethylation in multiple cancer subtypes including prostate, breast, liver and blood cancers. Epigenetically mediated GSTP1 silencing is associated with enhanced cancer susceptibility by decreasing its caretaker gene function, which tends to promote neoplastic transformation allowing the cell to acquire additional alterations. Thus, this epigenetic alteration is now considered as a cancer biomarker but could as well play a driving role in multistep cancer development especially well documented in prostate cancer development.The present review discusses application of epigenetic alterations affecting GSTP1 in cancer medicine used alone or in combination with other biomarkers for cancer detection and diagnosis as well as for future targeted preventive and therapeutic interventions including by dietary agents.

  3. Enhanced phytoremediation of mixed heavy metal (mercury)-organic pollutants (trichloroethylene) with transgenic alfalfa co-expressing glutathione S-transferase and human P450 2E1.

    Science.gov (United States)

    Zhang, Yuanyuan; Liu, Junhong; Zhou, Yuanming; Gong, Tingyun; Wang, Jing; Ge, Yinlin

    2013-09-15

    Soil contamination is a global environmental problem and many efforts have been made to find efficient remediation methods over the last decade. Moreover, remediation of mixed contaminated soils are more difficult. In the present study, transgenic alfalfa plants pKHCG co-expressing glutathione S-transferase (GST) and human P450 2E1 (CYP2E1) genes were used for phytoremediation of mixed mercury (Hg)-trichloroethylene (TCE) contaminants. Simultaneous expression of GST and CYP2E1 may produce a significant synergistic effect, and leads to improved resistance and accumulation to heavy metal-organic complex contaminants. Based on the tolerance and accumulation assays, pKHCG transgenic plants were more resistant to Hg/TCE complex pollutants and many folds higher in Hg/TCE-accumulation than the non-transgenic control plants in mixed contaminated soil. It is confirmed that GST and CYP2E1 co-expression may be a useful strategy to help achieve mixed heavy metal-organic pollutants phytoremediation. Copyright © 2013 Elsevier B.V. All rights reserved.

  4. Glutathione S-transferase Pi expression predicts response to adjuvant chemotherapy for stage C colon cancer: a matched historical control study

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    Jankova Lucy

    2012-05-01

    Full Text Available Abstract Background This study examined the association between overall survival and Glutathione S-transferase Pi (GST Pi expression and genetic polymorphism in stage C colon cancer patients after resection alone versus resection plus 5-fluourouracil-based adjuvant chemotherapy. Methods Patients were drawn from a hospital registry of colorectal cancer resections. Those receiving chemotherapy after it was introduced in 1992 were compared with an age and sex matched control group from the preceding period. GST Pi expression was assessed by immunohistochemistry. Overall survival was analysed by the Kaplan-Meier method and Cox regression. Results From an initial 104 patients treated with chemotherapy and 104 matched controls, 26 were excluded because of non-informative immunohistochemistry, leaving 95 in the treated group and 87 controls. Survival did not differ significantly among patients with low GST Pi who did or did not receive chemotherapy and those with high GST Pi who received chemotherapy (lowest pair-wise p = 0.11 whereas patients with high GST Pi who did not receive chemotherapy experienced markedly poorer survival than any of the other three groups (all pair-wise p Conclusion Stage C colon cancer patients with low GST Pi did not benefit from 5-fluourouracil-based adjuvant chemotherapy whereas those with high GST Pi did.

  5. Co-downregulation of the hydroxycinnamoyl-CoA:shikimate hydroxycinnamoyl transferase and coumarate 3-hydroxylase significantly increases cellulose content in transgenic alfalfa (Medicago sativa L.).

    Science.gov (United States)

    Tong, Zongyong; Li, Heng; Zhang, Rongxue; Ma, Lei; Dong, Jiangli; Wang, Tao

    2015-10-01

    Lignin is a component of the cell wall that is essential for growth, development, structure and pathogen resistance in plants, but high lignin is an obstacle to the conversion of cellulose to ethanol for biofuel. Genetically modifying lignin and cellulose contents can be a good approach to overcoming that obstacle. Alfalfa (Medicago sativa L.) is rich in lignocellulose biomass and used as a model plant for the genetic modification of lignin in this study. Two key enzymes in the lignin biosynthesis pathway-hydroxycinnamoyl -CoA:shikimate hydroxycinnamoyl transferase (HCT) and coumarate 3-hydroxylase (C3H)-were co-downregulated. Compared to wild-type plants, the lignin content in the modified strain was reduced by 38%, cellulose was increased by 86.1%, enzyme saccharification efficiency was increased by 10.9%, and cell wall digestibility was increased by 13.0%. The modified alfalfa exhibited a dwarf phenotype, but normal above ground biomass. This approach provides a new strategy for reducing lignin and increasing cellulose contents and creates a new genetically modified crop with enhanced value for biofuel. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  6. A phase II trial of R115777, an oral farnesyl transferase inhibitor, in      patients with advanced urothelial tract transitional cell carcinoma

    DEFF Research Database (Denmark)

    Rosenberg, Jonathan E.; Maase, Hans von der; Seigne, John D.

    2005-01-01

    BACKGROUND: R115777 is a potent farnesyl transferase inhibitor and has       significant antitumor effects in vitro and in vivo. METHODS: The objective       of the current study was to determine the objective response proportion in       patients with metastatic transitional cell carcinoma (TCC......) of the       urothelial tract who received treatment with R115777 at a dose of 300 mg       orally given twice daily for 21 days followed by 7 days of rest for every       4-week cycle. Thirty-four patients with TCC were enrolled in this Phase II       study. Patients were allowed to have received a maximum of one prior......       observed. CONCLUSIONS: The objective response rate of R115777 was not       sufficient to warrant future investigation in TCC as a single agent.       Preliminary evidence of the activity of R115777 in 2 chemotherapy-naive       patients may warrant further investigation in combination with first...

  7. Loss of Function Mutation in the Palmitoyl-Transferase HHAT Leads to Syndromic 46,XY Disorder of Sex Development by Impeding Hedgehog Protein Palmitoylation and Signaling

    Science.gov (United States)

    Makrythanasis, Periklis; Bernard, Pascal; Kurosaka, Hiroshi; Vannier, Anne; Thauvin-Robinet, Christel; Borel, Christelle; Mazaud-Guittot, Séverine; Rolland, Antoine; Desdoits-Lethimonier, Christèle; Guipponi, Michel; Zimmermann, Céline; Stévant, Isabelle; Kuhne, Françoise; Conne, Béatrice; Santoni, Federico; Lambert, Sandy; Huet, Frederic; Mugneret, Francine; Jaruzelska, Jadwiga; Faivre, Laurence; Wilhelm, Dagmar; Jégou, Bernard; Trainor, Paul A.; Resh, Marilyn D.; Antonarakis, Stylianos E.; Nef, Serge

    2014-01-01

    The Hedgehog (Hh) family of secreted proteins act as morphogens to control embryonic patterning and development in a variety of organ systems. Post-translational covalent attachment of cholesterol and palmitate to Hh proteins are critical for multimerization and long range signaling potency. However, the biological impact of lipid modifications on Hh ligand distribution and signal reception in humans remains unclear. In the present study, we report a unique case of autosomal recessive syndromic 46,XY Disorder of Sex Development (DSD) with testicular dysgenesis and chondrodysplasia resulting from a homozygous G287V missense mutation in the hedgehog acyl-transferase (HHAT) gene. This mutation occurred in the conserved membrane bound O-acyltransferase (MBOAT) domain and experimentally disrupted the ability of HHAT to palmitoylate Hh proteins such as DHH and SHH. Consistent with the patient phenotype, HHAT was found to be expressed in the somatic cells of both XX and XY gonads at the time of sex determination, and Hhat loss of function in mice recapitulates most of the testicular, skeletal, neuronal and growth defects observed in humans. In the developing testis, HHAT is not required for Sertoli cell commitment but plays a role in proper testis cord formation and the differentiation of fetal Leydig cells. Altogether, these results shed new light on the mechanisms of action of Hh proteins. Furthermore, they provide the first clinical evidence of the essential role played by lipid modification of Hh proteins in human testicular organogenesis and embryonic development. PMID:24784881

  8. Cholesterol esterification by mouse liver homogenate. Contribution to the study of ACYL-CoA: Cholesterol ACYL transferase in mammalian liver

    International Nuclear Information System (INIS)

    Soares, M.G.C.B.

    1976-01-01

    A cholesterol- esterifying enzyme from mouse liver has been partially characterized. The enzyme which showed optimum activity at pH 7,1 and required ATP and CoA, was identified as an acyl CoA: cholesterol acyl transferase (E.C.2.3.1.26). As a fuction of time the percentage of esterified cholesterol increased linearly during the first hour of incubation and continued to increase but not linearly with 4 hours, after which time no further net esterefication was observed. The relative concentration of esterified cholesterol remained constant between the fourth and twelveth hours of incubation but afterwards decreased when the incubation continued until 24 hours. The cholesterol- esterifying activity was 24,0+- 2,9 nmoles cholesterol esterified per gram tissue wet weight per minute. The mean percentages of free cholesterol esterified in and 24 hours respectively were 14,8+- 1,6 e 21,9+- 4,5. The subfractionation of labelled cholesteryl esters after one hour incubation of liver homogenate with 4-C 14 -Cholesterol showed the order of preference for the formation of the different ester classes to be monounsatured > diunsatured ≥ saturated >> polyunsaturated. The properties of the enzyme frommouse liver do not markedly differ from those of the previously recorded ACAT activity of rat liver. (Author) [pt

  9. Activation versus inhibition of microsomal glutathione S-transferase activity by acrolein. Dependence on the concentration and time of acrolein exposure.

    Science.gov (United States)

    Sthijns, Mireille M J P E; den Hartog, Gertjan J M; Scasso, Caterina; Haenen, Jan P; Bast, Aalt; Haenen, Guido R M M

    2017-09-25

    The toxicity of acrolein, an α,β-unsaturated aldehyde, is due to its soft electrophilic nature and primarily involves the adduction of protein thiols. The thiol glutathione (GSH) forms the first line of defense against acrolein. The present study confirms that acrolein added to isolated rat liver microsomes can increase microsomal GSH transferase (MGST) activity 2-3 fold, which can be seen as a direct adaptive increase in the protection against acrolein. At a relatively high exposure level, acrolein appeared to inhibit MGST. The activation is due to adduction of thiol groups, and the inactivation probably involves adduction of amino groups in the enzyme by acrolein. The preference of acrolein to react with thiol groups over amino groups can explain why the enzyme is activated at a low exposure level and inhibited at a high exposure level of acrolein. These opposite forms of direct adaptation on the level of enzyme activity further narrow the thin line between survival and promotion of cell death, governed by the level of exposure. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

  10. The link between antioxidant enzymes catalase and glutathione S-transferase and physiological condition of a control population of terrestrial isopod (Porcellio scaber).

    Science.gov (United States)

    Jemec, Anita; Lešer, Vladka; Drobne, Damjana

    2012-05-01

    The aim of this work was to investigate if the activities of catalase and glutathione S-transferase in a control population of terrestrial isopods (Porcellio scaber) are correlated with the physiological condition of the isopods. For this purpose, the activities of these enzymes were analysed in isopods from a stock population and in parallel, the physiological condition of the same specimens was assessed using a histological approach based on epithelial thickness and lipid droplets. We found a correlation between antioxidant enzymes and the physiological condition of the isopods. This implies that these enzymes could be used as predictive indicators of the physiological condition in a stock population before comprehensive toxicological studies are conducted and also in control group after the experiment. When a control group is found to be very heterogeneous in terms of physiological condition, the experiment should be repeated with a larger number of experimental animals. The findings of this study will contribute to more accurate experimental design of toxicity tests when using biomarkers. This should encourage other researchers to increase their effort to know the physiological state of their test organisms. Copyright © 2011 Elsevier Inc. All rights reserved.

  11. Expression of P-glycoprotein, multidrug resistance-associated protein, glutathione-S-transferase pi and p53 in canine transmissible venereal tumor

    Directory of Open Access Journals (Sweden)

    Daniel G. Gerardi

    2014-01-01

    Full Text Available The overexpression of proteins P-glycoprotein (P-gp, multidrug resistance-associated protein (MRP1, mutant p53, and the enzyme glutathione-S-transferase (GSTpi are related to resistance to chemotherapy in neoplasms. This study evaluated the expression of these markers by immunohistochemistry in two groups of canine TVT, without history of prior chemotherapy (TVT1, n=9 and in TVTs presented unsatisfactory clinical response to vincristine sulfate (TVT2, n=5. The percentage of specimens positively stained for P-gp, MRP1, GSTpi and p53 were, respectively 88.8%, 0%, 44.5% and 22.2% in TVT1 and 80%, 0%, 80% and 0% in TVT2. In TVT1, one specimen presented positive expression for three markers and four specimens for two markers. In TVT2, three specimens expressed P-gp and GSTpi. In conclusion, the canine TVTs studied expressed the four markers evaluated, but just P-gp and GSTpi were significantly expressed, mainly at cytoplasm and cytoplasm and nuclei, respectively, either before chemotherapy as after vincristine sulfate exposure. Future studies are needed to demonstrate the function of these two markers in conferring multidrug resistance (MDR or predict the response to chemotherapy in canine TVT.

  12. A study of the association of glutathione S-transferase M1/T1 polymorphisms with susceptibility to vitiligo in Egyptian patients.

    Science.gov (United States)

    Aly, Dalia Gamal; Salem, Samar Abdallah; Amr, Khalda Sayed; El-Hamid, Mahmoud Fawzy Abd

    2018-01-01

    The association of glutathione S-transferases M1/T1 (GSTM1/T1) null polymorphisms with vitiligo was proposed in several studies including two Egyptian studies with contradictory results. The aim here was to assess the association between GSTM1/T1 null polymorphisms and the susceptibility to vitiligo in a larger sample of Egyptian patients with generalized vitiligo. This study included 122 vitiligo patients and 200 healthy controls that were age, and gender matched. Assessment of GSTM1/T1 gene polymorphisms was done using a multiplex polymerase chain reaction (PCR). Increased odds of generalized vitiligo was observed with the null genotypes of GSTM1 and GSTT1 polymorphisms (Pvitiligo (OR=2.97, 95%CI=1.1-7.7) (P=0.02) compared with patients. Small sample size of patients. This study showed a significant trend towards an association with the combination of the GSTM1/GSTT1 double null polymorphism and generalized vitiligo. Individuals with GSTM1 null/GSTT1+ heterozygosis have a 2.97 odds protection from having generalized vitiligo compared with patients. It was is the first time, to our knowledge, that such an association has been reported.

  13. Comprehensive genome-wide analysis of Glutathione S-transferase gene family in potato (Solanum tuberosum L.) and their expression profiling in various anatomical tissues and perturbation conditions.

    Science.gov (United States)

    Islam, Md Shiful; Choudhury, Mouraj; Majlish, Al-Nahian Khan; Islam, Tahmina; Ghosh, Ajit

    2018-01-10

    Glutathione S-transferases (GSTs) are ubiquitous enzymes which play versatile functions including cellular detoxification and stress tolerance. In this study, a comprehensive genome-wide identification of GST gene family was carried out in potato (Solanum tuberosum L.). The result demonstrated the presence of at least 90 GST genes in potato which is greater than any other reported species. According to the phylogenetic analyses of Arabidopsis, rice and potato GST members, GSTs could be subdivided into ten different classes and each class is found to be highly conserved. The largest class of potato GST family is tau with 66 members, followed by phi and lambda. The chromosomal localization analysis revealed the highly uneven distribution of StGST genes across the potato genome. Transcript profiling of 55 StGST genes showed the tissue-specific expression for most of the members. Moreover, expression of StGST genes were mainly repressed in response to abiotic stresses, while largely induced in response to biotic and hormonal elicitations. Further analysis of StGST gene's promoter identified the presence of various stress responsive cis-regulatory elements. Moreover, one of the highly stress responsive StGST members, StGSTU46, showed strong affinity towards flurazole with lowest binding energy of -7.6kcal/mol that could be used as antidote to protect crop against herbicides. These findings will facilitate the further functional and evolutionary characterization of GST genes in potato. Copyright © 2017 Elsevier B.V. All rights reserved.

  14. T Cell Reactivity against Mycolyl Transferase Antigen 85 of M. tuberculosis in HIV-TB Coinfected Subjects and in AIDS Patients Suffering from Tuberculosis and Nontuberculous Mycobacterial Infections

    Directory of Open Access Journals (Sweden)

    Pascal Launois

    2011-01-01

    Full Text Available The mycolyl transferase antigen 85 complex is a major secreted protein family from mycobacterial culture filtrate, demonstrating powerful T cell stimulatory properties in most HIV-negative, tuberculin-positive volunteers with latent M.tuberculosis infection and only weak responses in HIV-negative tuberculosis patients. Here, we have analyzed T cell reactivity against PPD and Ag85 in HIV-infected individuals, without or with clinical symptoms of tuberculosis, and in AIDS patients with disease caused by nontuberculous mycobacteria. Whereas responses to PPD were not significantly different in HIV-negative and HIV-positive tuberculin-positive volunteers, responses to Ag85 were significantly decreased in the HIV-positive (CDC-A and CDC-B group. Tuberculosis patients demonstrated low T cell reactivity against Ag85, irrespective of HIV infection, and finally AIDS patients suffering from NTM infections were completely nonreactive to Ag85. A one-year follow-up of twelve HIV-positive tuberculin-positive individuals indicated a decreased reactivity against Ag85 in patients developing clinical tuberculosis, highlighting the protective potential of this antigen.

  15. Analyses of Genetic Variations of Glutathione S-Transferase Mu1 and Theta1 Genes in Bangladeshi Tannery Workers and Healthy Controls

    Directory of Open Access Journals (Sweden)

    Jobaida Akther

    2016-01-01

    Full Text Available Glutathione S-transferases (GSTs belong to a group of multigene detoxification enzymes, which defend cells against oxidative stress. Tannery workers are at risk of oxidative damage that is usually detoxified by GSTs. This study investigated the genotypic frequencies of GST Mu1 (GSTM1 and GST Theta1 (GSTT1 in Bangladeshi tannery workers and healthy controls followed by their status of oxidative stress and total GST activity. Of the 188 individuals, 50.0% had both GSTM1 and GSTT1 (+/+, 12.2% had GSTM1 (+/−, 31.4% had GSTT1 (−/+ alleles, and 6.4% had null genotypes (−/− with respect to both GSTM1 and GSTT1 alleles. Among 109 healthy controls, 54.1% were double positive, 9.2% had GSTM1 allele, 32.1% had GSTT1 allele, and 4.6% had null genotypes. Out of 79 tannery workers, 44.3% were +/+, 16.8% were +/−, 30.5% were −/+, and 8.4% were −/−. Though the polymorphic genotypes or allelic variants of GSTM1 and GSTT1 were distributed among the study subjects with different frequencies, the differences between the study groups were not statistically significant. GST activity did not vary significantly between the two groups and also among different genotypes while level of lipid peroxidation was significantly higher in tannery workers compared to controls irrespective of their GST genotypes.

  16. Analyses of Genetic Variations of Glutathione S-Transferase Mu1 and Theta1 Genes in Bangladeshi Tannery Workers and Healthy Controls

    Science.gov (United States)

    Akther, Jobaida; Ebihara, Akio; Nakagawa, Tsutomu; Islam, Laila N.; Suzuki, Fumiaki; Hosen, Md. Ismail; Hossain, Mahmud; Nabi, A. H. M. Nurun

    2016-01-01

    Glutathione S-transferases (GSTs) belong to a group of multigene detoxification enzymes, which defend cells against oxidative stress. Tannery workers are at risk of oxidative damage that is usually detoxified by GSTs. This study investigated the genotypic frequencies of GST Mu1 (GSTM1) and GST Theta1 (GSTT1) in Bangladeshi tannery workers and healthy controls followed by their status of oxidative stress and total GST activity. Of the 188 individuals, 50.0% had both GSTM1 and GSTT1 (+/+), 12.2% had GSTM1 (+/−), 31.4% had GSTT1 (−/+) alleles, and 6.4% had null genotypes (−/−) with respect to both GSTM1 and GSTT1 alleles. Among 109 healthy controls, 54.1% were double positive, 9.2% had GSTM1 allele, 32.1% had GSTT1 allele, and 4.6% had null genotypes. Out of 79 tannery workers, 44.3% were +/+, 16.8% were +/−, 30.5% were −/+, and 8.4% were −/−. Though the polymorphic genotypes or allelic variants of GSTM1 and GSTT1 were distributed among the study subjects with different frequencies, the differences between the study groups were not statistically significant. GST activity did not vary significantly between the two groups and also among different genotypes while level of lipid peroxidation was significantly higher in tannery workers compared to controls irrespective of their GST genotypes. PMID:27294127

  17. A SAM-dependent methyltransferase cotranscribed with arsenate reductase alters resistance to peptidyl transferase center-binding antibiotics in Azospirillum brasilense Sp7.

    Science.gov (United States)

    Singh, Sudhir; Singh, Chhaya; Tripathi, Anil Kumar

    2014-05-01

    The genome of Azospirillum brasilense harbors a gene encoding S-adenosylmethionine-dependent methyltransferase, which is located downstream of an arsenate reductase gene. Both genes are cotranscribed and translationally coupled. When they were cloned and expressed individually in an arsenate-sensitive strain of Escherichia coli, arsenate reductase conferred tolerance to arsenate; however, methyltransferase failed to do so. Sequence analysis revealed that methyltransferase was more closely related to a PrmB-type N5-glutamine methyltransferase than to the arsenate detoxifying methyltransferase ArsM. Insertional inactivation of prmB gene in A. brasilense resulted in an increased sensitivity to chloramphenicol and resistance to tiamulin and clindamycin, which are known to bind at the peptidyl transferase center (PTC) in the ribosome. These observations suggested that the inability of prmB:km mutant to methylate L3 protein might alter hydrophobicity in the antibiotic-binding pocket of the PTC, which might affect the binding of chloramphenicol, clindamycin, and tiamulin differentially. This is the first report showing the role of PrmB-type N5-glutamine methyltransferases in conferring resistance to tiamulin and clindamycin in any bacterium.

  18. Glutathione S-transferase T1, O1 and O2 polymorphisms are associated with survival in muscle invasive bladder cancer patients.

    Directory of Open Access Journals (Sweden)

    Tatjana I Djukic

    Full Text Available OBJECTIVE: To examine the association of six glutathione transferase (GST gene polymorphisms (GSTT1, GSTP1/rs1695, GSTO1/rs4925, GSTO2/rs156697, GSTM1, GSTA1/rs3957357 with the survival of patients with muscle invasive bladder cancer and the genotype modifying effect on chemotherapy. PATIENTS AND METHODS: A total of 105 patients with muscle invasive bladder cancer were included in the study. The follow-up lasted 5 years. The effect of GSTs polymorphisms on predicting mortality was analyzed by the Cox proportional hazard models, while Kaplan-Meier analysis was performed to assess differences in survival. RESULTS: GSTT1 active, GSTO1 Asp140Asp or GSTO2 Asp142Asp genotypes were independent predictors of a higher risk of death among bladder cancer patients (HR = 2.5, P = 0.028; HR = 2.9, P = 0.022; HR = 3.9, P = 0.001; respectively and significantly influenced the overall survival. There was no association between GSTP1, GSTM1 and GSTA1 gene variants with overall mortality. Only GSTO2 polymorphism showed a significant effect on the survival in the subgroup of patients who received chemotherapy (P = 0.006. CONCLUSION: GSTT1 active genotype and GSTO1 Asp140Asp and GSTO2 Asp142Asp genotypes may have a prognostic/pharmacogenomic role in patients with muscle invasive bladder cancer.

  19. O-linked N-acetylglucosamine transferase enhances secretory clusterin expression via liver X receptors and sterol response element binding protein regulation in cervical cancer.

    Science.gov (United States)

    Kim, Min Jun; Choi, Mee Young; Lee, Dong Hoon; Roh, Gu Seob; Kim, Hyun Joon; Kang, Sang Soo; Cho, Gyeong Jae; Kim, Yoon Sook; Choi, Wan Sung

    2018-01-12

    O-linked N-acetylglucosamine transferase (OGT) expression is increased in various cancer types, indicating the potential importance of O-GlcNAcylation in tumorigenesis. Secretory clusterin (sCLU) is involved in cancer cell proliferation and drug resistance, and recently, liver X receptors (LXRs) and sterol response element binding protein-1 (SREBP-1) were reported to regulate sCLU transcription. Here, we found that sCLU is significantly increased in cervical cancer cell lines, which have higher expression levels of O-GlcNAc and OGT than keratinocytes. OGT knockdown decreased expression of LXRs, SREBP-1 and sCLU through hypo-O-GlcNAcylation of LXRs. Additionally, treatment with Thiamet G, O-GlcNAcase OGA inhibitor, increased expression of O-GlcNAcylation and sCLU, and high glucose increased levels of LXRs, SREBP-1 and sCLU in HeLa cells. Moreover, OGT knockdown induced G 0 /G 1 phase cell cycle arrest and late apoptosis in cisplatin-treated HeLa cells, and decreased viability compared to OGT intact HeLa cells. Taken together, these findings suggest that OGT, O-GlcNAcylated LXRs, and SREBP-1 increase sCLU expression in cervical cancer cells, which contributes to drug resistance.

  20. Checkpoint kinase 1-induced phosphorylation of O-linked β-N-acetylglucosamine transferase regulates the intermediate filament network during cytokinesis.

    Science.gov (United States)

    Li, Zhe; Li, Xueyan; Nai, Shanshan; Geng, Qizhi; Liao, Ji; Xu, Xingzhi; Li, Jing

    2017-12-01

    Checkpoint kinase 1 (Chk1) is a kinase instrumental for orchestrating DNA replication, DNA damage checkpoints, the spindle assembly checkpoint, and cytokinesis. Despite Chk1's pivotal role in multiple cellular processes, many of its substrates remain elusive. Here, we identified O- linked β- N -acetylglucosamine ( O -GlcNAc)-transferase (OGT) as one of Chk1's substrates. We found that Chk1 interacts with and phosphorylates OGT at Ser-20, which not only stabilizes OGT, but also is required for cytokinesis. Phospho-specific antibodies of OGT-pSer-20 exhibited specific signals at the midbody of the cell, consistent with midbody localization of OGT as reported previously. Moreover, phospho-deficient OGT (S20A) cells attenuated cellular O -GlcNAcylation levels and also reduced phosphorylation of Ser-71 in the cytoskeletal protein vimentin, a modification critical for severing vimentin filament during cytokinesis. Consequently, elongated vimentin bridges were observed in cells depleted of OGT via an si OGT- based approach. Lastly, expression of plasmids resistant to si OGT efficiently rescued the vimentin bridge phenotype, but the OGT-S20A rescue plasmids did not. Our results suggest a Chk1-OGT-vimentin pathway that regulates the intermediate filament network during cytokinesis. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  1. Clinical implications of thymidylate synthetase, dihydropyrimidine dehydrogenase and orotate phosphoribosyl transferase activity levels in colorectal carcinoma following radical resection and administration of adjuvant 5-FU chemotherapy

    International Nuclear Information System (INIS)

    Ishikawa, Masashi; Miyauchi, Takayuki; Kashiwagi, Yutaka

    2008-01-01

    A number of studies have investigated whether the activity levels of enzymes involved in 5-fluorouracil (5-FU) metabolism are prognostic factors for survival in patients with colorectal carcinoma. Most reports have examined thymidylate synthetase (TS) and dihydropyrimidine dehydrogenase (DPD) in unresectable or metastatic cases, therefore it is unclear whether the activity of these enzymes is of prognostic value in colorectal cancer patients treated with radical resection and adjuvant chemotherapy with 5-FU. This study examined fresh frozen specimens of colorectal carcinoma from 40 patients who had undergone curative operation and were orally administered adjuvant tegafur/uracil (UFT) chemotherapy. TS, DPD and orotate phosphoribosyl transferase (OPRT) activities were assayed in cancer tissue and adjacent normal tissue and their association with clinicopathological variables was investigated. In addition, the relationships between TS, DPD and OPRT activities and patient survival were examined to determine whether any of these enzymes could be useful prognostic factors. While there was no clear relationship between pathological findings and TS or DPD activity, OPRT activity was significantly lower in tumors with lymph node metastasis than in tumors lacking lymph node metastasis. Postoperative survival was significantly better in the groups with low TS activity and/or high OPRT activity. TS and OPRT activity levels in tumor tissue may be important prognostic factors for survival in Dukes' B and C colorectal carcinoma with radical resection and adjuvant chemotherapy with UFT

  2. Evaluation of glutathione S-transferase T1 deletion polymorphism on type 2 diabetes mellitus risk in Zoroastrian females in Yazd, Iran

    Science.gov (United States)

    Afrand, Mohammadhosain; Khalilzadeh, Saeedhossein; Bashardoost, Nasrollah; Sheikhha, Mohammad Hasan

    2015-01-01

    Background: There has been much interest in the role of free radicals and oxidative stress in the pathogenesis of diabetes mellitus (DM). The aim of this study was to assess the possible association between genetic polymorphisms of the glutathione S-transferase-Theta (GSTT1) and the risk of the development of DM in Zoroastrian females in Yazd, Iran. Materials and Methods: This was a case-control study in which GSTT1 polymorphism was genotyped in 51 randomly selected DM patients and 50 randomly selected healthy controls among Zoroastrian females whose ages ranged from 40 to 70. Results: The frequencies of GSTT1 null genotype and GSTT1 present were 72% and 28%, respectively, in control samples, while in patients with type 2 diabetes (T2DM), the frequencies of GSTT1 null genotype and GSTT1 present were 27.5% and 72.5%, respectively. There were higher levels of triglyceride (TG), fasting blood sugar (FBS), total cholesterol (TC), low-density lipoprotein (LDL), Urea, and high-density lipoprotein (HDL) in cases of GSTT1 null genotype compared to the GSTT1 present genotype in controls. Conclusions: Our results indicated that healthy subjects had a higher frequency of the GSTT1 null genotype than patients with T2DM. However, we observed no significant association between the GSTT1 null genotype and T2DM in the current study. PMID:25593839

  3. Localization of three human polypeptide GalNAc-transferases in HeLa cells suggests initiation of O-linked glycosylation throughout the Golgi apparatus

    DEFF Research Database (Denmark)

    Röttger, S; White, J; Wandall, H H

    1998-01-01

    O-glycosylation of proteins is initiated by a family of UDP-N-acetylgalactosamine:polypeptide N-acetylgalactos-aminyltransferases (GalNAc-T). In this study, we have localized endogenous and epitope-tagged human GalNAc-T1, -T2 and -T3 to the Golgi apparatus in HeLa cells by subcellular fractionation......, immunofluorescence and immunoelectron microscopy. We show that all three GalNAc-transferases are concentrated about tenfold in Golgi stacks over Golgi associated tubular-vesicular membrane structures. Surprisingly, we find that GalNAc-T1, -T2 and -T3 are present throughout the Golgi stack suggesting that initiation...... of O-glycosylation may not be restricted to the cis Golgi, but occur at multiple sites within the Golgi apparatus. GalNAc-T1 distributes evenly across the Golgi stack whereas GalNAc-T2 and -T3 reside preferentially on the trans side and in the medial part of the Golgi stack, respectively. Moreover, we...

  4. Determination of serum neuron specific enolase and glutathion S transferases levels in patients with acute cerebral infarction and its clinical significance

    International Nuclear Information System (INIS)

    Guo Jianyi; Lu Tianhe; Bao Yanmei

    2002-01-01

    Objective: To evaluate the variation of serum neuron specific enolase (NSE) and glutathion S transferases (GST) levels in patients with cerebral infarction and its clinical significance. Methods: The serum levels of NSE in cerebral infarction patients were determined with immunoradiometric assay (IRMA), and the serum level of GST were determined by enzyme immuno sandwich assay (ELISA). Results: Serum NSE levels linked in patients were significantly higher (p<0.01) and GST serum levels were significantly lower (p < 0.01) within 3 days after onset of disease than those at two weeks and those in the controls. There was a positive correlation between serum NSE levels and neurological deficit scores (p < 0.001) and a negative correlation with serum GST levels (p < 0.05). There was also a close relationship between the serum NSE levels and the volume of infarction (p < 0.001). Conclusion: There was a close relationship between the Serum levels of NSE, GST and clinical features of Patients in the early stage of cerebral infarction

  5. Practical electronics handbook

    CERN Document Server

    Sinclair, Ian R

    2013-01-01

    Practical Electronics Handbook, Third Edition provides the frequently used and highly applicable principles of electronics and electronic circuits.The book contains relevant information in electronics. The topics discussed in the text include passive and active discrete components; linear and digital I.C.s; microprocessors and microprocessor systems; digital-analogue conversions; computer aids in electronics design; and electronic hardware components.Electronic circuit constructors, service engineers, electronic design engineers, and anyone with an interest in electronics will find the book ve

  6. Atividade de glutationa S-transferase na metabolização de acetochlor, atrazine e oxyfluorfen em milho (Zea mays L., sorgo (Sorghum bicolor L. e trigo (Triticum aestivum L. (Poaceae Glutathione S-transferase activity in acetochlor, atrazine and oxyfluorfen metabolization in maize (Zea mays L., sorghum (Sorghum bicolor L. and wheat (Triticum aestivumL. (Poaceae

    Directory of Open Access Journals (Sweden)

    Ethel Lourenzi Barbosa Novelli

    2002-05-01

    Full Text Available Este experimento foi conduzido para avaliar a seletividade em plantas dos herbicidas acetochlor, atrazine e oxyfluorfen em relação à atividade da glutationa S-transferase (GST em plantas de milho (Zea mays L., sorgo (Sorghum bicolor L. e trigo (Triticum aestivum L. (Poaceae. A atividade da GST foi detectada às 24, 48 e 72 horas após as aplicaç��es dos tratamentos. Os tratamentos do experimento consistiram de aplicação com água (controle, acetochlor (3 L.ha-1, atrazine (4 L.ha-1 e oxyfluorfen (1 L.ha-1. As maiores atividades de GST foram observadas na presença de acetochlor, principalmente às 48 horas após o tratamento. Esses aumentos foram 105, 148 e 118% em relação ao controle para milho, sorgo e trigo, respectivamente. É sugerido que a GST pode ter papel na degradação de acetochlor e pode ser uma das razões para a seletividade desse herbicida para essas culturas.This experiment was conducted to evaluate the acetochlor, atrazine and oxyfluorfen herbicides plant selectivity, in relation to glutathione S-transferase activity (GST in maize (Zea mays L., sorghum (Sorghum bicolor L. and wheat (Triticum aestivum L (Poaceae plants. GST activity was detected 24, 48 and 72 hours after treatment applications. The experiment's treatments consisted of spraying plants with water (control, acetochlor (3 L.ha-1`, atrazine (4 L.ha-1 and oxyfluorfen (1 L.ha-1. The highest GST activities were observed in presence of acetochlor, mainly at 48 hours after treatment. These increments were 105, 148 and 118% when compared to maize, sorghum and wheat control groups, respectively. It is suggested that the GST may have a role in acetochlor degradation and it may be a reason for this herbicide's selectivity in these crops.

  7. Methylated Glutathione S-transferase 1 (mGSTP1) is a potential plasma free DNA epigenetic marker of prognosis and response to chemotherapy in castrate-resistant prostate cancer

    OpenAIRE

    Mahon, K L; Qu, W; Devaney, J; Paul, C; Castillo, L; Wykes, R J; Chatfield, M D; Boyer, M J; Stockler, M R; Marx, G; Gurney, H; Mallesara, G; Molloy, P L; Horvath, L G; Clark, S J

    2014-01-01

    Background: Glutathione S-transferase 1 (GSTP1) inactivation is associated with CpG island promoter hypermethylation in the majority of prostate cancers (PCs). This study assessed whether the level of circulating methylated GSTP1 (mGSTP1) in plasma DNA is associated with chemotherapy response and overall survival (OS). Methods: Plasma samples were collected prospectively from a Phase I exploratory cohort of 75 men with castrate-resistant PC (CRPC) and a Phase II independent validation cohort ...

  8. Electron foreshock

    International Nuclear Information System (INIS)

    Klimas, A.J.

    1985-01-01

    ISEE particle and wave data are noted to furnish substantial support for the basic features of the velocity dispersed model at the foreshock boundary that was proposed by Filbert and Kellogg (1979). Among many remaining discrepancies between this model and observation, it is noted that unstable reduced velocity distributions have been discovered behind the thin boundary proposed by the model, and that these are at suprathermal energies lying far below those explainable in terms of an oscillating, two-stream instability. Although the long-theorized unstable beam of electrons has been found in the foreshock, there is still no ready explanation of the means by which it could have gotten there. 16 references

  9. Oxidative stress markers and genetic polymorphisms of glutathione S-transferase T1, M1, and P1 in a subset of children with autism spectrum disorder in Lagos, Nigeria.

    Science.gov (United States)

    Oshodi, Y; Ojewunmi, O; Oshodi, T A; Ijarogbe, G T; Ogun, O C; Aina, O F; Lesi, Fea

    2017-09-01

    The role of oxidative stress has been identified in the development of autism spectrum disorder (ASD), and polymorphisms of glutathione S-transferase have been associated with some diseases linked to oxidative stress. Hence, we evaluated the serum levels of oxidative stress markers and investigated genetic polymorphisms of glutathione S-transferase associated with autism. Forty-two children clinically diagnosed with ASD using the Diagnostic and Statistical Manual for Mental Disorders (DSM-5) criteria and a clinical interview were included in the study. Twenty-three age-matched controls without any known genetic/developmental disorder were also recruited. Oxidative stress markers along with the genetic polymorphisms of glutathione S-transferase were determined. Reduced glutathione in ASD patients was significantly lower than the control (P = 0.008), whereas other oxidative stress markers measured were not significantly different in both the control and case populations. The frequencies of GSTT1 and GSTM1 null genotypes were lower among the controls compared with the cases, however, no association risk was observed. The observed risk of carrying Val/Val genotype among the cases was approximately six times that of the controls. Individuals with ASD showed a significant diminished level of reduced glutathione, however, the distribution of GSTT1, GSTM1, and GSTP1 polymorphisms was not found to be associated with autism in this study population.

  10. Sustainable Management of Electronics

    Science.gov (United States)

    To provide information on EPAs strategy for electronics stewardship, certified electronics recyclers and the Challenge; as well as where to donate unwanted electronics, how to calculate benefits, and what's going on with electronics mgmt in their states.

  11. Advanced electron beam techniques

    International Nuclear Information System (INIS)

    Hirotsu, Yoshihiko; Yoshida, Yoichi

    2007-01-01

    After 100 years from the time of discovery of electron, we now have many applications of electron beam in science and technology. In this report, we review two important applications of electron beam: electron microscopy and pulsed-electron beam. Advanced electron microscopy techniques to investigate atomic and electronic structures, and pulsed-electron beam for investigating time-resolved structural change are described. (author)

  12. EDITORIAL: Synaptic electronics Synaptic electronics

    Science.gov (United States)

    Demming, Anna; Gimzewski, James K.; Vuillaume, Dominique

    2013-09-01

    Conventional computers excel in logic and accurate scientific calculations but make hard work of open ended problems that human brains handle easily. Even von Neumann—the mathematician and polymath who first developed the programming architecture that forms the basis of today's computers—was already looking to the brain for future developments before his death in 1957 [1]. Neuromorphic computing uses approaches that better mimic the working of the human brain. Recent developments in nanotechnology are now providing structures with very accommodating properties for neuromorphic approaches. This special issue, with guest editors James K Gimzewski and Dominique Vuillaume, is devoted to research at the serendipitous interface between the two disciplines. 'Synaptic electronics', looks at artificial devices with connections that demonstrate behaviour similar to synapses in the nervous system allowing a new and more powerful approach to computing. Synapses and connecting neurons respond differently to incident signals depending on the history of signals previously experienced, ultimately leading to short term and long term memory behaviour. The basic characteristics of a synapse can be replicated with around ten simple transistors. However with the human brain having around 1011 neurons and 1015 synapses, artificial neurons and synapses from basic transistors are unlikely to accommodate the scalability required. The discovery of nanoscale elements that function as 'memristors' has provided a key tool for the implementation of synaptic connections [2]. Leon Chua first developed the concept of the 'The memristor—the missing circuit element' in 1971 [3]. In this special issue he presents a tutorial describing how memristor research has fed into our understanding of synaptic behaviour and how they can be applied in information processing [4]. He also describes, 'The new principle of local activity, which uncovers a minuscule life-enabling "Goldilocks zone", dubbed the

  13. A Putative O-Linked β-N-Acetylglucosamine Transferase Is Essential for Hormogonium Development and Motility in the Filamentous Cyanobacterium Nostoc punctiforme.

    Science.gov (United States)

    Khayatan, Behzad; Bains, Divleen K; Cheng, Monica H; Cho, Ye Won; Huynh, Jessica; Kim, Rachelle; Omoruyi, Osagie H; Pantoja, Adriana P; Park, Jun Sang; Peng, Julia K; Splitt, Samantha D; Tian, Mason Y; Risser, Douglas D

    2017-05-01

    Most species of filamentous cyanobacteria are capable of gliding motility, likely via a conserved type IV pilus-like system that may also secrete a motility-associated polysaccharide. In a subset of these organisms, motility is achieved only after the transient differentiation of hormogonia, which are specialized filaments that enter a nongrowth state dedicated to motility. Despite the fundamental importance of hormogonia to the life cycles of many filamentous cyanobacteria, the molecular regulation of hormogonium development is largely undefined. To systematically identify genes essential for hormogonium development and motility in the model heterocyst-forming filamentous cyanobacterium Nostoc punctiforme , a forward genetic screen was employed. The first gene identified using this screen, designated ogtA , encodes a putative O-linked β- N -acetylglucosamine transferase (OGT). The deletion of ogtA abolished motility, while ectopic expression of ogtA induced hormogonium development even under hormogonium-repressing conditions. Transcription of ogtA is rapidly upregulated (1 h) following hormogonium induction, and an OgtA-GFPuv fusion protein localized to the cytoplasm. In developing hormogonia, accumulation of PilA but not HmpD is dependent on ogtA Reverse transcription-quantitative PCR (RT-qPCR) analysis indicated equivalent levels of pilA transcript in the wild-type and Δ ogtA mutant strains, while a reporter construct consisting of the intergenic region in the 5' direction of pilA fused to gfp produced lower levels of fluorescence in the Δ ogtA mutant strain than in the wild type. The production of hormogonium polysaccharide in the Δ ogtA mutant strain is reduced compared to that in the wild type but comparable to that in a pilA deletion strain. Collectively, these results imply that O -GlcNAc protein modification regulates the accumulation of PilA via a posttranscriptional mechanism in developing hormogonia. IMPORTANCE Filamentous cyanobacteria are among

  14. Influenza A Virus-Induced Expression of a GalNAc Transferase, GALNT3, via MicroRNAs Is Required for Enhanced Viral Replication.

    Science.gov (United States)

    Nakamura, Shoko; Horie, Masayuki; Daidoji, Tomo; Honda, Tomoyuki; Yasugi, Mayo; Kuno, Atsushi; Komori, Toshihisa; Okuzaki, Daisuke; Narimatsu, Hisashi; Nakaya, Takaaki; Tomonaga, Keizo

    2016-02-15

    Influenza A virus (IAV) affects the upper and lower respiratory tracts and rapidly induces the expression of mucins, which are common O-glycosylated proteins, on the epithelial surfaces of the respiratory tract. Although mucin production is associated with the inhibition of virus transmission as well as characteristic clinical symptoms, little is known regarding how mucins are produced on the surfaces of respiratory epithelial cells and how they affect IAV replication. In this study, we found that two microRNAs (miRNAs), miR-17-3p and miR-221, which target GalNAc transferase 3 (GALNT3) mRNA, are rapidly downregulated in human alveolar basal epithelial cells during the early stage of IAV infection. We demonstrated that the expression of GALNT3 mRNA is upregulated in an IAV replication-dependent fashion and leads to mucin production in bronchial epithelial cells. A lectin microarray analysis revealed that the stable expression of GALNT3 by human alveolar basal epithelial cells induces mucin-type O-glycosylation modifications similar to those present in IAV-infected cells, suggesting that GALNT3 promotes mucin-type O-linked glycosylation in IAV-infected cells. Notably, analyses using short interfering RNAs and miRNA mimics showed that GALNT3 knockdown significantly reduces IAV replication. Furthermore, IAV replication was markedly decreased in embryonic fibroblast cells obtained from galnt3-knockout mice. Interestingly, IAV-infected galnt3-knockout mice exhibited high mortality and severe pathological alterations in the lungs compared to those of wild-type mice. Our results demonstrate not only the molecular mechanism underlying rapid mucin production during IAV infection but also the contribution of O-linked glycosylation to the replication and propagation of IAV in lung cells. Viral infections that affect the upper or lower respiratory tracts, such as IAV, rapidly induce mucin production on the epithelial surfaces of respiratory cells. However, the details of how

  15. Analysis of selected glutathione S-transferase gene polymorphisms in Malaysian type 2 diabetes mellitus patients with and without cardiovascular disease.

    Science.gov (United States)

    Etemad, A; Vasudevan, R; Aziz, A F A; Yusof, A K M; Khazaei, S; Fawzi, N; Jamalpour, S; Arkani, M; Mohammad, N A; Ismail, P

    2016-04-07

    Type 2 diabetes mellitus (T2DM) is believed to be associated with excessive production of reactive oxygen species. Glutathione S-transferase (GST) polymorphisms result in decreased or absent enzyme activity and altered oxidative stress, and have been associated with cardiovascular disease (CVD). The present study assessed the effect of GST polymorphisms on the risk of developing T2DM in individuals of Malaysian Malay ethnicity. A total of 287 subjects, consisting of 87 T2DM and 64 CVD/T2DM patients, as well as 136 healthy gender- and age-matched controls were genotyped for selected polymorphisms to evaluate associations with T2DM susceptibility. Genomic DNA was extracted using commercially available kits, and GSTM1, GSTT1, and α-globin sequences were amplified by multiplex polymerase chain reaction. Biochemical parameters were measured with a Hitachi autoanalyzer. The Fisher exact test, the chi-square statistic, and means ± standard deviations were calculated using the SPSS software. Overall, we observed no significant differences regarding genotype and allele frequencies between each group (P = 0.224 and 0.199, respectively). However, in the combined analysis of genotypes and blood measurements, fasting plasma glucose, HbA1c, and triglyceride levels, followed by age, body mass index, waist-hip ratio, systolic blood pressure, and history of T2DM significantly differed according to GST polymorphism (P ˂ 0.05). Genetically induced absence of the GSTT1 enzyme is an independent and powerful predictor of premature vascular morbidity and death in individuals with T2DM, and might be triggered by cigarette smoking's oxidative effects. These polymorphisms could be screened in other ethnicities within Malaysia to determine further possible risk factors.

  16. Females with paired occurrence of cancers in the UADT and genital region have a higher frequency of either Glutathione S-transferase M1/T1 null genotype

    Directory of Open Access Journals (Sweden)

    Jhavar Sameer G

    2005-03-01

    Full Text Available Abstract Upper Aero digestive Tract (UADT is the commonest site for the development of second cancer in females after primary cervical cancer. Glutathione S-transferase (GSTM1 and / or T1 null genotype modulates the risk of developing UADT cancer (primary as well as second cancer. The aim of this study was to evaluate the difference in GST null genotype frequencies in females with paired cancers in the UADT and genital region as compared to females with paired cancers in the UADT and non-genital region. Forty-nine females with a cancer in the UADT and another cancer (at all sites-genital and non-genital were identified from a database of patients with multiple primary neoplasms and were analyzed for the GSTM1 and T1 genotype in addition to known factors such as age, tobacco habits, alcohol habits and family history of cancer. Frequencies of GSTM1 null, GSTT1 null, and either GSTM1/T1 null were higher in females with paired occurrence of cancer in the UADT and genital site (54%, 33% and 75% respectively in comparison to females with paired occurrence of cancer in the UADT and non-genital sites (22%, 6% and 24% respectively. The significantly higher inherited frequency of either GSTM1/T1 null genotype in females with a paired occurrence of cancers in UADT and genital region (p = 0.01, suggests that these females are more susceptible to damage by carcinogens as compared to females who have UADT cancers in association with cancers at non-genital sites.

  17. Genome-Wide Identification, Characterization, and Expression Profiling of Glutathione S-Transferase (GST) Family in Pumpkin Reveals Likely Role in Cold-Stress Tolerance

    Science.gov (United States)

    Abdul Kayum, Md.; Nath, Ujjal Kumar; Park, Jong-In; Choi, Eung Kyoo; Song, Jae-Young; Kim, Hoy-Taek; Nou, Ill-Sup

    2018-01-01

    Plant growth and development can be adversely affected by cold stress, limiting productivity. The glutathione S-transferase (GST) family comprises important detoxifying enzymes, which play major roles in biotic and abiotic stress responses by reducing the oxidative damage caused by reactive oxygen species. Pumpkins (Cucurbita maxima) are widely grown, economically important, and nutritious; however, their yield can be severely affected by cold stress. The identification of putative candidate genes responsible for cold-stress tolerance, including the GST family genes, is therefore vital. For the first time, we identified 32 C. maxima GST (CmaGST) genes using a combination of bioinformatics approaches and characterized them by expression profiling. These CmaGST genes represent seven of the 14 known classes of plant GSTs, with 18 CmaGSTs categorized into the tau class. The CmaGSTs were distributed across 13 of pumpkin’s 20 chromosomes, with the highest numbers found on chromosomes 4 and 6. The large number of CmaGST genes resulted from gene duplication; 11 and 5 pairs of CmaGST genes were segmental- and tandem-duplicated, respectively. In addition, all CmaGST genes showed organ-specific expression. The expression of the putative GST genes in pumpkin was examined under cold stress in two lines with contrasting cold tolerance: cold-tolerant CP-1 (C. maxima) and cold-susceptible EP-1 (Cucurbita moschata). Seven genes (CmaGSTU3, CmaGSTU7, CmaGSTU8, CmaGSTU9, CmaGSTU11, CmaGSTU12, and CmaGSTU14) were highly expressed in the cold-tolerant line and are putative candidates for use in breeding cold-tolerant crop varieties. These results increase our understanding of the cold-stress-related functions of the GST family, as well as potentially enhancing pumpkin breeding programs. PMID:29439434

  18. Uridine diphosphate glucuronide transferase 1A1FNx0128 gene polymorphism and the toxicity of irinotecan in recurrent and refractory small cell lung cancer

    Directory of Open Access Journals (Sweden)

    Fan Yun

    2014-01-01

    Full Text Available Objective: The aim was to investigate the association between uridine diphosphate glucuronide transferase 1A1 (UGT1A1 gene promoter region polymorphism and irinotecan-related adverse effects and efficacy on recurrent and refractory small cell lung cancer (SCLC. Materials and Methods: A total of 31 patients with recurrent and refractory SCLC were enrolled in this study from June 2012 to August 2013 and received at least two cycles of single-agent irinotecan chemotherapy. The efficacy and adverse effects of irinotecan were evaluated. DNA was extracted from peripheral blood and direct sequencing method was employed to test UGT1A1FNx0128 polymorphism, thus analyzing the correlation between UGT1A1FNx0128 polymorphism and irinotecan-related side-effects and efficacy. Results: A total of 25 cases (80.6% were UGT1A1FNx0128 wild-type (TA 6 /(TA 6 ; 6 cases (19.4% were heterozygous mutant (TA 6 /(TA 7 , no homozygous mutant genotype (TA 7 /(TA 7 was found. The incidences of grade 3/4 neutropenia, diarrhea and thrombocytopenia were 35.5%, 25.8% and 22.6% in all the patients, respectively. The incidence of 3/4 adverse effects in patients with genotype (TA 6 /(TA 6 and heterozygous (TA 6 /(TA 7 had no statistical difference (P > 0.05 for all. The overall response rate (ORR was 32.3%. Median progression free survival (PFS and overall survival (OS were 4 months and 7.5 months in all patients, respectively. There was no statistical difference in ORR, PFS and OS between genotype (TA 6 /(TA 6 patients and heterozygous (TA 6 /(TA 7 patients. Conclusion: Irinotecan showed efficacy in patients with recurrent and refractory SCLC; UGT1A1 FNx01 28 polymorphism failed to predict the incidence of serious adverse effects and efficacy of irinotecan.

  19. Effects of deletion of the Streptococcus pneumoniae lipoprotein diacylglyceryl transferase gene lgt on ABC transporter function and on growth in vivo.

    Directory of Open Access Journals (Sweden)

    Suneeta Chimalapati

    Full Text Available Lipoproteins are an important class of surface associated proteins that have diverse roles and frequently are involved in the virulence of bacterial pathogens. As prolipoproteins are attached to the cell membrane by a single enzyme, prolipoprotein diacylglyceryl transferase (Lgt, deletion of the corresponding gene potentially allows the characterisation of the overall importance of lipoproteins for specific bacterial functions. We have used a Δlgt mutant strain of Streptococcus pneumoniae to investigate the effects of loss of lipoprotein attachment on cation acquisition, growth in media containing specific carbon sources, and virulence in different infection models. Immunoblots of triton X-114 extracts, flow cytometry and immuno-fluorescence microscopy confirmed the Δlgt mutant had markedly reduced lipoprotein expression on the cell surface. The Δlgt mutant had reduced growth in cation depleted medium, increased sensitivity to oxidative stress, reduced zinc uptake, and reduced intracellular levels of several cations. Doubling time of the Δlgt mutant was also increased slightly when grown in medium with glucose, raffinose and maltotriose as sole carbon sources. These multiple defects in cation and sugar ABC transporter function for the Δlgt mutant were associated with only slightly delayed growth in complete medium. However the Δlgt mutant had significantly reduced growth in blood or bronchoalveolar lavage fluid and a marked impairment in virulence in mouse models of nasopharyngeal colonisation, sepsis and pneumonia. These data suggest that for S. pneumoniae loss of surface localisation of lipoproteins has widespread effects on ABC transporter functions that collectively prevent the Δlgt mutant from establishing invasive infection.

  20. The Potato Aphid Salivary Effector Me47 Is a Glutathione-S-Transferase Involved in Modifying Plant Responses to Aphid Infestation.

    Science.gov (United States)

    Kettles, Graeme J; Kaloshian, Isgouhi

    2016-01-01

    Polyphagous aphid pests cause considerable economic damage to crop plants, primarily through the depletion of photoassimilates and transfer of viruses. The potato aphid (Macrosiphum euphorbiae) is a notable pest of solanaceous crops, however, the molecular mechanisms that underpin the ability to colonize these hosts are unknown. It has recently been demonstrated that like other aphid species, M. euphorbiae injects a battery of salivary proteins into host plants during feeding. It is speculated that these proteins function in a manner analagous to secreted effectors from phytopathogenic bacteria, fungi and oomycetes. Here, we describe a novel aphid effector (Me47) which was identified from the potato aphid salivary secretome as a putative glutathione-S-transferase (GST). Expression of Me47 in Nicotiana benthamiana enhanced reproductive performance of green peach aphid (Myzus persicae). Similarly, delivery of Me47 into leaves of tomato (Solanum lycopersicum) by Pseudomonas spp. enhanced potato aphid fecundity. In contrast, delivery of Me47 into Arabidopsis thaliana reduced GPA reproductive performance, indicating that Me47 impacts the outcome of plant-aphid interactions differently depending on the host species. Delivery of Me47 by the non-pathogenic Pseudomonas fluorescens revealed that Me47 protein or activity triggers defense gene transcriptional upregulation in tomato but not Arabidopsis. Recombinant Me47 was purified and demonstrated to have GST activity against two specific isothiocyanates (ITCs), compounds implicated in herbivore defense. Whilst GSTs have previously been associated with development of aphid resistance to synthetic insecticides, the findings described here highlight a novel function as both an elicitor and suppressor of plant defense when delivered into host tissues.

  1. The potato aphid salivary effector Me47 is a glutathione-S-transferase involved in modifying plant responses to aphid infestation

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    Graeme James Kettles

    2016-08-01

    Full Text Available Polyphagous aphid pests cause considerable economic damage to crop plants, primarily through the depletion of photoassimilates and transfer of viruses. The potato aphid (Macrosiphum euphorbiae is a notable pest of solanaceous crops, however the molecular mechanisms that underpin the ability to colonize these hosts are unknown. It has recently been demonstrated that like other aphid species, M. euphorbiae injects a battery of salivary proteins into host plants during feeding. It is speculated that these proteins function in a manner analagous to secreted effectors from phytopathogenic bacteria, fungi and oomycetes. Here we describe a novel aphid effector (Me47 which was identified from the potato aphid salivary secretome as a putative glutathione-S-transferase (GST. Expression of Me47 in Nicotiana benthamiana enhanced reproductive performance of green peach aphid (Myzus persicae. Similarly, delivery of Me47 into leaves of tomato (Solanum lycopersicum by Pseudomonas spp. enhanced potato aphid fecundity. In contrast, delivery of Me47 into Arabidopsis thaliana reduced GPA reproductive performance, indicating that Me47 impacts the outcome of plant-aphid interactions differently depending on the host species. Delivery of Me47 by the non-pathogenic Pseudomonas fluorescens revealed that Me47 protein or activity triggers defense gene transcriptional upregulation in tomato but not Arabidopsis. Recombinant Me47 was purified and demonstrated to have GST activity against two specific isothiocyanates (ITCs, compounds implicated in herbivore defense. Whilst GSTs have previously been associated with development of aphid resistance to synthetic insecticides, the findings described here highlight a novel function as both an elicitor and suppressor of plant defense when delivered into host tissues.

  2. Genetic polymorphisms of glutathione-s-transferase M1 and T1 genes with risk of diabetic retinopathy in Iranian population

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    Elham Moasser

    2014-05-01

    Full Text Available Objective(s:To the best of our knowledge, this is the first report on the contributions of GST genetic variants to the risk of diabetic retinopathy in an Iranian population. Therefore, the objective of this study was to determine whether sequence variation in glutathione S-transferase gene (GSTM1 and GSTT1 is associated with development of diabetic retinopathy in type 2 diabetes mellitus (T2DM Iranian patients. Materials and Methods: A total of 605 subjects were investigated in this case-control study; Study groups consisted of 201 patients with diabetic retinopathy (DR, 203 subjects with no clinically significant signs of DR and a group of 201 cases of healthy volunteers with no clinical evidence of diabetes mellitus or any other diseases. The GSTM1 and GSTT1 were genotyped by multiplex-polymerase chain reaction (multiplex-PCR analysis in all 404 T2DM patients and 201 healthy individuals served as control. Results: Increased odds ratio showed that GSTM1-null genotype had a moderately higher occurrence in T2DM patients (OR=1.43, 95% CI=1.01–2.04; P=0.03 than in healthy individuals. However, the frequency of GSTT1 genotype (OR=1.41; 95% CI=0.92-2.18; P=0.09 was not significantly different comparing both groups. Although, regression analysis in T2DM patients showed that GSTM1 and GSTT1 genotypes are not associated with T2DM retinopathy development. Conclusion: Our findings suggest that GSTM1 and GSTT1 genotypes might not be involved in the pathogenesis of type 2 diabetes mellitus retinopathy in the Southern Iranian population. However, further investigations are needed to confirm these results in other larger populations.

  3. Genetic polymorphism of glutathion S-transferase P1 (GSTP1 Ile105Val and susceptibility to atherogenesis in patients with type 2 diabetes mellitus

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    Grubiša Ivana

    2013-01-01

    Full Text Available One of the characteristics of type 2 diabetes mellitus (T2DM is the state of persistent oxidative stress (OS that has been implicated in the pathogenesis of diseases such is atherosclerosis mainly through chronic hyperglycemia that stimulates production of reactive oxygen species (ROS and increases OS. Glutathione S-transferase P1 (GSTP1 is a member of the cytosolic GST superfamily. It plays an important role in neutralizing OS as an enzyme. Also, it participates in regulation of stress signaling and protects cells against apoptosis via its noncatalytic ligand-binding activity. GSTP1 Ile105Val functional polymorphism influences protein catalytic activity and stability and the aim of this study was to determine whether this gene variation influences susceptibility to atherogenesis in T2DM patients. A total of 240 individuals (140 patients with T2DM, accompanied with clinical manifestations of atherosclerosis, and 100 healthy controls were included in this study. Genomic DNA was isolated from peripheral blood cells and genotyping was performed using polymerase chain reaction followed by restriction fragment length polymorphism (PCR-RFLP analysis. We obtained no statistically significant differences in the distribution of alleles and genotypes between cases and controls (P>0.05 but association between Ile/Val (OR=0.6, 95%CI=0.35-1.05, P=0.08 and Val/Val (OR=0.45, 95%CI=0.18-1.11, P=0.08 genotypes and disease approached significance (P=0.08. Our results indicated that a larger study group is needed to establish the true relationship between potentialiy protective allele Val and the disease, and to determine the influence of other GSTP1 polymorphisms on atherogenesis in T2DM patients. [Projekat Ministarstva nauke Republike Srbije, br. 175075

  4. The mitochondrial O-linked N-acetylglucosamine transferase (mOGT) in the diabetic patient could be the initial trigger to develop Alzheimer disease.

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    Lozano, Liliana; Lara-Lemus, Roberto; Zenteno, Edgar; Alvarado-Vásquez, Noé

    2014-10-01

    Diabetes mellitus (DM) is considered a risk factor for the development of Alzheimer disease (AD); however, how DM favors evolution of AD is still insufficiently understood. Hyperglycemia in DM is associated to an increase in mitochondrial reactive oxygen species (ROS) generation, as well as damage of hippocampal cells, reflected by changes in morphological and mitochondrial functionality. Similar mitochondrial damage has been observed when amyloid beta (Aβ) accumulates in the brain of AD patients. In DM, the excess of glucose in the brain induces higher activity of the hexosamine biosynthesis pathway (HBP), it synthesizes UDP-N-acetylglucosamine (UDP-GlcNAc), which is used by O-linked N-acetylglucosamine transferase (OGT) to catalyze O-GlcNAcylation of numerous proteins. Although O-GlcNAcylation plays an important role in maintaining structure and cellular functionality, chronic activity of this pathway has been associated with insulin resistance and hyperglycemia-induced glucose toxicity. Three different forms of OGT are known: nucleocytoplasmic (ncOGT), short (sOGT), and mitochondrial (mOGT). Previous reports showed that overexpression of ncOGT is not toxic to the cell; in contrast, overexpression of mOGT is associated with cellular apoptosis. In this work, we suggest that hyperglycemia in the diabetic patient could induce greater expression and activity of mOGT, modifying the structure and functionality of mitochondria in hippocampal cells, accelerating neuronal damage, and favoring the start of AD. In consequence, mOGT activity could be a key point for AD development in patients with DM. Copyright © 2014 Elsevier Inc. All rights reserved.

  5. Sulforaphane and alpha-lipoic acid upregulate the expression of the pi class of glutathione S-transferase through c-jun and Nrf2 activation.

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    Lii, Chong-Kuei; Liu, Kai-Li; Cheng, Yi-Ping; Lin, Ai-Hsuan; Chen, Haw-Wen; Tsai, Chia-Wen

    2010-05-01

    The anticarcinogenic effect of dietary organosulfur compounds has been partly attributed to their modulation of the activity and expression of phase II detoxification enzymes. Our previous studies indicated that garlic allyl sulfides upregulate the expression of the pi class of glutathione S-transferase (GSTP) through the activator protein-1 pathway. Here, we examined the modulatory effect of sulforaphane (SFN) and alpha-lipoic acid (LA) or dihydrolipoic acid (DHLA) on GSTP expression in rat Clone 9 liver cells. Cells were treated with LA or DHLA (50-600 micromol/L) or SFN (0.2-5 micromol/L) for 24 h. Immunoblots and real-time PCR showed that SFN, LA, and DHLA dose dependently induced GSTP protein and mRNA expression. Compared with the induction by the garlic organosulfur compound diallyl trisulfide (DATS), the effectiveness was in the order of SFN > DATS > LA = DHLA. The increase in GSTP enzyme activity in cells treated with 5 micromol/L SFN, 50 micromol/L DATS, and 600 micromol/L LA and DHLA was 172, 75, 122, and 117%, respectively (P GPEI) was required for GSTP induction by the organosulfur compounds. Electromobility gel shift assays showed that the DNA binding of GPEI to nuclear proteins reached a maximum at 0.5-1 h after SFN, LA, and DHLA treatment. Super-shift assay revealed that the transcription factors c-jun and nuclear factor erythroid-2 related factor 2 (Nrf2) were bound to GPEI. These results suggest that SFN and LA in either its oxidized or reduced form upregulate the transcription of the GSTP gene by activating c-jun and Nrf2 binding to the enhancer element GPEI.

  6. Activation of Nrf2 is required for up-regulation of the π class of glutathione S-transferase in rat primary hepatocytes with L-methionine starvation.

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    Lin, Ai-Hsuan; Chen, Haw-Wen; Liu, Cheng-Tze; Tsai, Chia-Wen; Lii, Chong-Kuei

    2012-07-04

    Numerous genes expression is regulated in response to amino acid shortage, which helps organisms adapt to amino acid limitation. The expression of the π class of glutathione (GSH) S-transferase (GSTP), a highly inducible phase II detoxification enzyme, is regulated mainly by activates activating protein 1 (AP-1) binding to the enhancer I of GSTP (GPEI). Here we show the critical role of nuclear factor erythroid-2-related factor 2 (Nrf2) in up-regulating GSTP gene transcription. Primary rat hepatocytes were cultured in a methionine-restricted medium, and immunoblotting and RT-PCR analyses showed that methionine restriction time-dependently increased GSTP protein and mRNA expression over a 48 h period. Nrf2 translocation to the nucleus, nuclear proteins binding to GPEI, and antioxidant response element (ARE) luciferase reporter activity were increased by methionine restriction as well as by l-buthionine sulfoximine (BSO), a GSH synthesis inhibitor. Transfection with Nrf2 siRNA knocked down Nrf2 expression and reversed the methionine-induced GSTP expression and GPEI binding activity. Chromatin immunoprecipitation assay confirmed the binding of Nrf2 to the GPEI. Phosphorylation of extracellular signal-regulated kinase 2 (ERK2) was increased in methionine-restricted and BSO-treated cells. ERK2 siRNA abolished methionine restriction-induced Nrf2 nuclear translocation, GPEI binding activity, ARE-luciferase reporter activity, and GSTP expression. Our results suggest that the up-regulation of GSTP gene transcription in response to methionine restriction likely occurs via the ERK-Nrf2-GPEI signaling pathway.

  7. Genomic insights into the glutathione S-transferase gene family of two rice planthoppers, Nilaparvata lugens (Stal and Sogatella furcifera (Horvath (Hemiptera: Delphacidae.

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    Wen-Wu Zhou

    Full Text Available BACKGROUND: Glutathione S-transferase (GST genes control crucial traits for the metabolism of various toxins encountered by insects in host plants and the wider environment, including insecticides. The planthoppers Nilaparvata lugens and Sogatella furcifera are serious specialist pests of rice throughout eastern Asia. Their capacity to rapidly adapt to resistant rice varieties and to develop resistance to various insecticides has led to severe outbreaks over the last decade. METHODOLOGY/PRINCIPAL FINDINGS: Using the genome sequence of N. lugens, we identified for the first time the complete GST gene family of a delphacid insect whilst nine GST gene orthologs were identified from the closely related species S. furcifera. Nilaparvata lugens has 11 GST genes belonging to six cytosolic subclasses and a microsomal class, many fewer than seen in other insects with known genomes. Sigma is the largest GST subclass, and the intron-exon pattern deviates significantly from that of other species. Higher GST gene expression in the N. lugens adult migratory form reflects the higher risk of this life stage in encountering the toxins of non-host plants. After exposure to a sub-lethal dose of four insecticides, chlorpyrifos, imidacloprid, buprofezin or beta-cypermethrin, more GST genes were upregulated in S. furcifera than in N. lugens. RNA interference targeting two N. lugens GST genes, NlGSTe1 and NlGSTm2, significantly increased the sensitivity of fourth instar nymphs to chlorpyrifos but not to beta-cypermethrin. CONCLUSIONS/SIGNIFICANCE: This study provides the first elucidation of the nature of the GST gene family in a delphacid species, offering new insights into the evolution of metabolic enzyme genes in insects. Further, the use of RNA interference to identify the GST genes induced by insecticides illustrates likely mechanisms for the tolerance of these insects.

  8. Superoxide dismutase, catalase, glutathione peroxidase and gluthatione S-transferases M1 and T1 gene polymorphisms in three Brazilian population groups.

    Science.gov (United States)

    de Oliveira Hiragi, Cássia; Miranda-Vilela, Ana Luisa; Rocha, Dulce Maria Sucena; de Oliveira, Silviene Fabiana; Hatagima, Ana; de Nazaré Klautau-Guimarães, Maria

    2011-01-01

    Antioxidants such as superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPX1) reduce the oxidation rates in the organism. Gluthatione S-transferases (GSTs) play a vital role in phase 2 of biotransformation of many substances. Variation in the expression of these enzymes suggests individual differences for the degree of antioxidant protection and geographical differences in the distribution of these variants. We described the distribution frequency of CAT (21A/T), SOD2 (Ala9Val), GPX1 (Pro198Leu), GSTM1 and GSTT1 polymorphisms in three Brazilian population groups: Kayabi Amerindians (n = 60), Kalunga Afro-descendants (n = 72), and an urban mixed population from Federal District (n = 162). Frequencies of the variants observed in Kalunga (18% to 58%) and Federal District (33% to 63%) were similar to those observed in Euro and Afro-descendants, while in Kayabi (3% to 68%), depending on the marker, frequencies were similar to the ones found in different ethnic groups. Except for SOD2 in all population groups studied here, and for GPX1 in Kalunga, the genotypic distributions were in accordance with Hardy-Weinberg Equilibrium. These data can clarify the contribution of different ethnicities in the formation of mixed populations, such as that of Brazil. Moreover, outcomes will be valuable resources for future functional studies and for genetic studies in specific populations. If these studies are designed to comprehensively explore the role of these genetic polymorphisms in the etiology of human diseases they may help to prevent inconsistent genotype-phenotype associations in pharmacogenetic studies.

  9. Polymorphisms of glutathione-S-transferase M1, T1, P1 and the risk of prostate cancer: a case-control study

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    Račay Peter

    2009-03-01

    Full Text Available Abstract Background It has been suggested that polymorphisms in glutathione-S-transferases (GST could predispose to prostate cancer through a heritable deficiency in detoxification pathways for environmental carcinogens. Yet, studies linking GST polymorphism and prostate cancer have so far failed to unambiguously establish this relation in patients. A retrospective study on healthy, unrelated subjects was conducted in order to estimate the population GST genotype frequencies in the Slovak population of men and compare our results with already published data (GSEC project-Genetic Susceptibility to Environmental Carcinogens. A further aim of the study was to evaluate polymorphisms in GST also in patients with prostate cancer in order to compare the evaluated proportions with those found in the control subjects. Methods We determined the GST genotypes in 228 healthy, unrelated subjects who attended regular prostate cancer screening between May 2005 and June 2007 and in 129 histologically verified prostate cancer patients. Analysis for the GST gene polymorphisms was performed by PCR and PCR-RFLP. Results We found that the GST frequencies are not significantly different from those estimated in a European multicentre study or from the results published by another group in Slovakia. Our results suggest that Val/Val genotype of GSTP1 gene could modulate the risk of prostate cancer, even if this association did not reach statistical significance. We did not observe significantly different crude rates of the GSTM1 and GSTT1 null genotypes in the men diagnosed with prostate cancer and those in the control group. Conclusion Understanding the contribution of GST gene polymorphisms and their interactions with other relevant factors may improve screening diagnostic assays for prostate cancer. We therefore discuss issues of study feasibility, study design, and statistical power, which should be taken into account in planning further trials.

  10. Association Study of Glutathione S-transferases Gene Polymorphisms (GSTM1 and GSTT1 with Ulcerative Colitis and Crohn's Disease in the South of Iran

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    Maryam Moini

    2017-01-01

    Full Text Available Background: Inflammatory bowel diseases (IBDs, including ulcerative colitis (UC and Crohn's disease (CD, are chronic inflammatory disorders of the gastrointestinal tract. A combination of environmental factors and interactions with a genetic predisposition are suggested to play an important role in the etiology and pathogenesis of the IBD. Glutathione S-transferases (GSTs are multifunctional enzymes involved in the cellular oxidative stress handling. Possible associations between GSTs gene polymorphisms and susceptibility to UC and CD have been reported in different population. The relationship between GSTM1 and GSTT1 deletion polymorphisms and susceptibility to UC and CD were investigated in the Iranian population. Materials and Methods: The study was performed in 106 IBD patients and 243 age- and sex-matched healthy Iranian controls consulting the IBD registry center of the Motahari Clinic, Shiraz University of Medical Sciences, Shiraz, Iran, between 2011 and 2013. GSTM1 and GSTT1 genotyping were performed using multiplex polymerase chain reaction and differences in the distribution of gene polymorphisms were analyzed statistically between the studied groups. Results: Statistically significant higher frequency of GSTM1 null genotype was observed in IBD patients (P = 0.01 and in the subgroup of patients with UC (P = 0.04 compared to healthy controls, whereas this was not true for CD patients. No significant association was found between GSTT1 gene polymorphism and UC or CD. Conclusions: Absence of GSTT1 functional gene does not play an important role in the pathophysiology and development of IBD, UC, and CD in Iranian population whereas GSTM1 null genotype could be considered as a possible genetic predisposing factor for more susceptibility to IBD and UC.

  11. Reverted glutathione S-transferase-like genes that influence flower color intensity of carnation (Dianthus caryophyllus L.) originated from excision of a transposable element.

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    Momose, Masaki; Itoh, Yoshio; Umemoto, Naoyuki; Nakayama, Masayoshi; Ozeki, Yoshihiro

    2013-12-01

    A glutathione S-transferase-like gene, DcGSTF2, is responsible for carnation (Dianthus caryophyllus L.) flower color intensity. Two defective genes, DcGSTF2mu with a nonsense mutation and DcGSTF2-dTac1 containing a transposable element dTac1, have been characterized in detail in this report. dTac1 is an active element that produces reverted functional genes by excision of the element. A pale-pink cultivar 'Daisy' carries both defective genes, whereas a spontaneous deep-colored mutant 'Daisy-VPR' lost the element from DcGSTF2-dTac1. This finding confirmed that dTac1 is active and that the resulting reverted gene, DcGSTF2rev1, missing the element is responsible for this color change. Crosses between the pale-colored cultivar '06-LA' and a deep-colored cultivar 'Spectrum' produced segregating progeny. Only the deep-colored progeny had DcGSTF2rev2 derived from the 'Spectrum' parent, whereas progeny with pale-colored flowers had defective forms from both parents, DcGSTF2mu and DcGSTF2-dTac1. Thus, DcGSTF2rev2 had functional activity and likely originated from excision of dTac1 since there was a footprint sequence at the vacated site of the dTac1 insertion. Characterizing the DcGSTF2 genes in several cultivars revealed that the two functional genes, DcGSTF2rev1 and DcGSTF2rev2, have been used for some time in carnation breeding with the latter in use for more than half a century.

  12. Modulation of xenobiotic metabolising enzymes by anticarcinogens-focus on glutathione S-transferases and their role as targets of dietary chemoprevention in colorectal carcinogenesis

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    Pool-Zobel, Beatrice [Department of Nutritional Toxicology, Institute for Nutrition, Friedrich Schiller University Jena, 07743 Jena (Germany)]. E-mail: b8pobe@uni-jena.de; Veeriah, Selvaraju [Department of Nutritional Toxicology, Institute for Nutrition, Friedrich Schiller University Jena, 07743 Jena (Germany); Boehmer, Frank-D. [Institute of Molecular Cell Biology, University Hospital, Friedrich Schiller University Jena, 07743 Jena (Germany)

    2005-12-11

    There is evidence that consumption of certain dietary ingredients may favourably modulate biotransformation of carcinogens. Associated with this is the hypothesis that the risk for developing colorectal cancer could be reduced, since its incidence is related to diet. Two main groups of biotransformation enzymes metabolize carcinogens, namely Phase I enzymes, which convert hydrophobic compounds to more water-soluble moieties, and Phase II enzymes (e.g. glutathione S-transferases [GST]), which primarily catalyze conjugation reactions. The conjugation of electrophilic Phase I intermediates with glutathione, for instance, frequently results in detoxification. Several possible colon carcinogens may serve as substrates for GST isoenzymes that can have marked substrate specificity. The conjugated products could be less toxic/genotoxic if GSTs are induced, thereby reducing exposure. Thus, numerous studies have shown that the induction of GSTs by antioxidants enables experimental animals to tolerate exposure to carcinogens. One important mechanism of GST induction involves an antioxidant-responsive response element (ARE) and the transcription factor nuclear factor E2-related factor 2 (Nrf2), which is bound to the Kelch-like ECH associated protein 1 (Keap1) in the cytoplasm. Antioxidants may disrupt the Keap-Nrf2 complex, allowing Nrf2 to translocate to the nucleus and mediate expression of Phase II genes via interaction with the ARE. GSTs are also induced by butyrate, a product of gut flora-derived fermentation of plant foods, which may act via different mechanisms, e.g. by increasing histone acetylation. GSTs are expressed with high inter-individual variability in human colonocytes, which points to large differences in cellular susceptibility to xenobiotics. Enhancing expression of GSTs in human colon tissue could therefore contribute to reducing cancer risks. However, it has not been demonstrated in humans that this mechanism is associated with cancer prevention. In the

  13. Modulation of xenobiotic metabolising enzymes by anticarcinogens-focus on glutathione S-transferases and their role as targets of dietary chemoprevention in colorectal carcinogenesis

    International Nuclear Information System (INIS)

    Pool-Zobel, Beatrice; Veeriah, Selvaraju; Boehmer, Frank-D.

    2005-01-01

    There is evidence that consumption of certain dietary ingredients may favourably modulate biotransformation of carcinogens. Associated with this is the hypothesis that the risk for developing colorectal cancer could be reduced, since its incidence is related to diet. Two main groups of biotransformation enzymes metabolize carcinogens, namely Phase I enzymes, which convert hydrophobic compounds to more water-soluble moieties, and Phase II enzymes (e.g. glutathione S-transferases [GST]), which primarily catalyze conjugation reactions. The conjugation of electrophilic Phase I intermediates with glutathione, for instance, frequently results in detoxification. Several possible colon carcinogens may serve as substrates for GST isoenzymes that can have marked substrate specificity. The conjugated products could be less toxic/genotoxic if GSTs are induced, thereby reducing exposure. Thus, numerous studies have shown that the induction of GSTs by antioxidants enables experimental animals to tolerate exposure to carcinogens. One important mechanism of GST induction involves an antioxidant-responsive response element (ARE) and the transcription factor nuclear factor E2-related factor 2 (Nrf2), which is bound to the Kelch-like ECH associated protein 1 (Keap1) in the cytoplasm. Antioxidants may disrupt the Keap-Nrf2 complex, allowing Nrf2 to translocate to the nucleus and mediate expression of Phase II genes via interaction with the ARE. GSTs are also induced by butyrate, a product of gut flora-derived fermentation of plant foods, which may act via different mechanisms, e.g. by increasing histone acetylation. GSTs are expressed with high inter-individual variability in human colonocytes, which points to large differences in cellular susceptibility to xenobiotics. Enhancing expression of GSTs in human colon tissue could therefore contribute to reducing cancer risks. However, it has not been demonstrated in humans that this mechanism is associated with cancer prevention. In the

  14. Isoenzyme-specific up-regulation of glutathione transferase and aldo-keto reductase mRNA expression by dietary quercetin in rat liver.

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    Odbayar, Tseye-Oidov; Kimura, Toshinori; Tsushida, Tojiro; Ide, Takashi

    2009-05-01

    The impact of quercetin on the mRNA expression of hepatic enzymes involved in drug metabolism was evaluated with a DNA microarray and real-time PCR. Male Sprague-Dawley rats were fed an experimental diet containing either 0, 2.5, 5, 10, or 20 g/kg of quercetin for 15 days. The DNA microarray analysis of the gene expression profile in pooled RNA samples from rats fed diets containing 0, 5, and 20 g/kg of quercetin revealed genes of some isoenzymes of glutathione transferase (Gst) and aldo-keto reductase (Akr) to be activated by this flavonoid. Real-time PCR conducted with RNA samples from individual rats fed varying amounts of quercetin together with the microarray analysis showed that quercetin caused marked dose-dependent increases in the mRNA expression of Gsta3, Gstp1, and Gstt3. Some moderate increases were also noted in the mRNA expression of isoenzymes belonging to the Gstm class. Quercetin also dose-dependently increased the mRNA expression of Akr1b8 and Akr7a3. However, it did not affect the parameters of the other Gst and Akr isoenzymes. It is apparent that quercetin increases the mRNA expression of Gst and Akr involved in drug metabolism in an isoenzyme-specific manner. Inasmuch as Gst and Akr isoenzymes up-regulated in their gene expression are involved in the prevention and attenuation of cancer development, this consequence may account for the chemopreventive propensity of quercetin.

  15. Germline glutathione S-transferase variants in breast cancer: Relation to diagnosis and cutaneous long-term adverse effects after two fractionation patterns of radiotherapy

    International Nuclear Information System (INIS)

    Edvardsen, Hege; Kristensen, Vessela N.; Grenaker Alnaes, Grethe Irene B.Sc.; Bohn, Mona; Erikstein, Bjorn; Helland, Aslaug; Borresen-Dale, Anne-Lise; Fossa, Sophie Dorothea

    2007-01-01

    Purpose: To explore whether certain glutathione S-transferase (GST) polymorphisms are associated with an increased risk of breast cancer or the level of radiation-induced adverse effects after two fractionation patterns of adjuvant radiotherapy. Methods and Materials: The prevalence of germline polymorphic variants in GSTM1, GSTP1, and GSTT1 was determined in 272 breast cancer patients and compared with that in a control group of 270 women from the general population with no known history of breast cancer. The genetic variants were determined using multiplex polymerase chain reaction followed by restriction enzyme fragment analysis. In 253 of the patients surveyed for radiotherapy-induced side effects after a median observation time of 13.7 years (range, 7-22.8 years), the genotypes were related to the long-term effects observed after two fractionation patterns (treatment A, 4.3 Gy in 10 fractions for 156 patients; and treatment B, 2.5 Gy in 20 fractions for 97; both administered within a 5-week period). Results: None of the GST polymorphisms conferred an increased risk of breast cancer, either alone or in combination. Compared with treatment B, treatment A was followed by an increased level of moderate to severe radiation-induced side effects for all the endpoints studied (i.e., degree of telangiectasia, subcutaneous fibrosis and atrophy, lung fibrosis, costal fractures, and pleural thickening; p <0.001 for all endpoints). A significant association was found between the level of pleural thickening and the GSTP1 Ile105Val variant. Conclusion: The results of this study have illustrated the impact of hypofractionation on the level of adverse effects and indicated that the specific alleles of GSTP1, M1, and T1 studied here may be significant in determining the level of adverse effects after radiotherapy

  16. Glutathione transferase-M2-2 secreted from glioblastoma cell protects SH-SY5Y cells from aminochrome neurotoxicity.

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    Cuevas, Carlos; Huenchuguala, Sandro; Muñoz, Patricia; Villa, Monica; Paris, Irmgard; Mannervik, Bengt; Segura-Aguilar, Juan

    2015-04-01

    U373MG cells are able to take up aminochrome that induces glutathione transferase M2-2 (GSTM2) expression in a concentration-dependent manner where 100 µM aminochrome increases GSTM2 expression by 2.1-fold (P protects SH-SY5Y cells incubated with 10 µM aminochrome. The significant protection provided by U373MG-conditioned medium in SH-SY5Y cells incubated with aminochrome was dependent on GSTM2 internalization into SH-SY5Y cells as evidenced by (i) uptake of (14)C-GSTM2 released from U373MG cells into SH-SY5Y cells, a process inhibited by anti-GSTM2 antiserum; (ii) lack of protection of U373MG-conditioned medium in the presence of anti-GSTM2 antiserum on SH-SY5Y cells treated with aminochrome; and (iii) lack of protection of conditioned medium from U373MGsiGST6 that expresses an siRNA directed against GSTM2 on SH-SY5Y cells treated with aminochrome. In conclusion, our results demonstrated that U373MG cells protect SH-SY5Y cells against aminochrome neurotoxicity by releasing GSTM2 into the conditioned medium and subsequent internalization of GSTM2 into SH-SY5Y cells. These results suggest a new mechanism of protection of dopaminergic neurons mediated by astrocytes by releasing GSTM2 into the intersynaptic space and subsequent internalization into dopaminergic neuron in order to protect these cells against aminochrome neurotoxicity.

  17. Doping test results dependent on genotype of uridine diphospho-glucuronosyl transferase 2B17, the major enzyme for testosterone glucuronidation.

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    Schulze, Jenny Jakobsson; Lundmark, Jonas; Garle, Mats; Skilving, Ilona; Ekström, Lena; Rane, Anders

    2008-07-01

    Testosterone abuse is conventionally assessed by the urinary testosterone/epitestosterone (T/E) ratio, levels above 4.0 being considered suspicious. The large variation in testosterone glucuronide (TG) excretion and its strong association with a deletion polymorphism in the uridine diphospho-glucuronosyl transferase (UGT) 2B17 gene challenge the accuracy of the T/E ratio test. Our objective was to investigate whether genotype-based cutoff values will improve the sensitivity and specificity of the test. This was an open three-armed comparative study. A total of 55 healthy male volunteers with either two, one, or no allele [insertion/insertion, insertion/deletion, or deletion/deletion (del/del)] of the UGT2B17 gene was included in the study. A single im dose of 500 mg testosterone enanthate was administered. Urinary excretion of TG after dose and the T/E ratio during 15 d were calculated. The degree and rate of increase in the TG excretion rate were highly dependent on the UGT2B17 genotype with a 20-fold higher average maximum increase in the insertion/insertion group compared with the del/del group. Of the del/del subjects, 40% never reached the T/E ratio of 4.0 on any of the 15 d after the dose. When differentiated cutoff levels for the del/del (1.0) and the other genotypes (6.0) were applied, the sensitivity increased substantially for the del/del group, and false positives in the other genotypes were eliminated. Consideration of the genetic variation in disposition of androgens will improve the sensitivity and specificity of the testosterone doping test. This is of interest not only for combating androgen doping in sports, but also for detecting and preventing androgen abuse in society.

  18. The radioprotector WR-2721 reduces neutron-induced mutations at the hypoxanthine-guanine phosphoribosyl transferase locus in mouse splenocytes when administered prior to or following irradiation

    International Nuclear Information System (INIS)

    Grdina, D.J.; Basic, I.

    1992-01-01

    An in vitro T-lymphocyte cloning technique has been applied to study the effects of JANUS fission-spectrum neutron irradiation and the radioprotector S-2-(3-aminopropylamino) ethylphosphorothioic acid (WR-2721) on the subsequent development of somatic mutations at the hypoxanthine-guanine phosphoribosyl transferase (hprt) locus in hybrid B6CF1 male mice. In control studies performed to establish an in vitro cloning technique, the mutant frequencies of splenic T-lymphocytes, as a result of exposure to a 100 cGy dose of neutrons, increased with time from a control level of 9 x 10 -7 to a maximum value of 1.7 x 10 -5 at 56 days following irradiation. Between 56 and 150 days after irradiation, mutant frequencies were observed to plateau and remain stable. All subsequent determinations were performed at 56 days following the experimental treatment of animals. WR-2721 at a dose of 400 mg/kg was effective in protecting against the induction of hprt mutants (i.e. a mutant frequency reduction factor, MFRF) following the largest dose of neutrons used (i.e. 150 cGy). The antimutagenic effectiveness of WR-2721 administered 30 min prior to irradiation was unaffected, even when the dose was reduced to 200 mg/kg. These findings confirm our earlier report using the radioprotector N-(2-mercaptoethyl)-1,2-diaminopropane (WR-1065) under in vitro conditions, and demonstrate that these agents can be used as effective antimutagens even when they are administered up to 3 h following radiation exposure. (Author)

  19. Molecular cloning, biochemical characterization, and expression analysis of two glutathione S-transferase paralogs from the big-belly seahorse (Hippocampus abdominalis).

    Science.gov (United States)

    Tharuka, M D Neranjan; Bathige, S D N K; Lee, Jehee

    2017-12-01

    Glutathione S-transferases (GSTs, EC 2.5.1.18) are important Phase II detoxifying enzymes that catalyze hydrophobic, electrophilic xenobiotic substance with the conjugation of reduced glutathione (GSH). In this study, GSTμ and GSTρ paralogs of GST in the big belly seahorse (Hippocampus abdominalis; HaGSTρ, HaGSTμ) were biochemically, molecularly, functionally characterized to determine their detoxification range and protective capacities upon different pathogenic stresses. HaGSTρ and HaGSTμ are composed of coding sequences of 681bp and 654bp, which encode proteins 225 and 217 amino acids, with predicted molecular masses of 26.06kDa and 25.74kDa respectively. Sequence analysis revealed that both HaGSTs comprise the characteristic GSH-binding site in the thioredoxin-like N-terminal domain and substrate binding site in the C-terminal domain. The recombinant HaGSTρ and HaGSTμ proteins catalyzed the model GST substrate 1-chloro-2, 4-dinitrobenzene (CDNB). Enzyme kinetic analysis revealed different K m and V max values for each rHaGST, suggesting that they have different conjugation rates. The optimum conditions (pH, temperature) and inhibitory assays of each protein demonstrated different optimal ranges. However, HaGSTμ was highly expressed in the ovary and gill, whereas HaGSTρ was highly expressed in the gill and pouch. mRNA expression of HaGSTρ and HaGSTμ was significantly elevated upon lipopolysaccharide, Poly (I:C), and Edwardsiella tarda challenges in liver and in blood cells as well as with Streptococcus iniae challenge in blood cells. From these collective experimental results, we propose that HaGSTρ and HaGSTμ are effective in detoxifying xenobiotic toxic agents, and importantly, their mRNA expression could be stimulated by immunological stress signals in the aquatic environment. Copyright © 2017 Elsevier Inc. All rights reserved.

  20. Phosphorylation of TET proteins is regulated via O-GlcNAcylation by the O-linked N-acetylglucosamine transferase (OGT).

    Science.gov (United States)

    Bauer, Christina; Göbel, Klaus; Nagaraj, Nagarjuna; Colantuoni, Christian; Wang, Mengxi; Müller, Udo; Kremmer, Elisabeth; Rottach, Andrea; Leonhardt, Heinrich

    2015-02-20

    TET proteins oxidize 5-methylcytosine to 5-hydroxymethylcytosine, 5-formylcytosine, and 5-carboxylcytosine and thus provide a possible means for active DNA demethylation in mammals. Although their catalytic mechanism is well characterized and the catalytic dioxygenase domain is highly conserved, the function of the regulatory regions (the N terminus and the low-complexity insert between the two parts of the dioxygenase domains) is only poorly understood. Here, we demonstrate that TET proteins are subject to a variety of post-translational modifications that mostly occur at these regulatory regions. We mapped TET modification sites at amino acid resolution and show for the first time that TET1, TET2, and TET3 are highly phosphorylated. The O-linked GlcNAc transferase, which we identified as a strong interactor with all three TET proteins, catalyzes the addition of a GlcNAc group to serine and threonine residues of TET proteins and thereby decreases both the number of phosphorylation sites and site occupancy. Interestingly, the different TET proteins display unique post-translational modification patterns, and some modifications occur in distinct combinations. In summary, our results provide a novel potential mechanism for TET protein regulation based on a dynamic interplay of phosphorylation and O-GlcNAcylation at the N terminus and the low-complexity insert region. Our data suggest strong cross-talk between the modification sites that could allow rapid adaption of TET protein localization, activity, or targeting due to changing environmental conditions as well as in response to external stimuli. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.