Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:6.3.2.3 (glutathione synthetase)
678 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

In the roots of pea plants (Pisum sativum L.) cultivated with 20 [mu]M CdCl2 for 3 d, synthesis of phytochelatins [PCs or ([gamma]EC)nG, where [gamma]EC is [gamma]glutamylcysteine and G is glycine] and homophytochelatins [h-PCs, ([gamma]EC)n[beta]-alanine] is accompanied by a drastic decrease in glutathione (GSH) content, but an increase in homoglutathione (h-GSH) content. In contrast, the in vitro activity of GSH synthetase increases 5-fold, whereas h-GSH synthetase activity increases regardless of Cd exposure. The consititutive enzyme PC synthase, which catalyzes the transfer of the [gamma]-EC moiety of GSH to an acceptor GSH molecule thus producing ([gamma]EC)2G, is activated by heavy metals, with Cd and Cu being strong activators and Zn being a very poor activator. Using h-GSH or hm-GSH for substrate, the synthesis rate of([gamma]EC)2[beta]-alanine and [gamma]EC)2-serine is only 2.4 and 0.3%, respectively, of the sythesis rate of ([gamma]EC)2G with GSH as substrate. However, in the presence of a constant GSH level, increasing the concentration of h-GSH or hm-GSH results in increased synthesis of ([gamma]EC)2[beta]-alanine or ([gamma]EC)2-serine, respecively; simultaneously, the synthesis of ([gamma]EC)2G is inhibited. [gamma]EC is not a substrate of PC synthase. These results are best explained by assuming that PC synthase has a [gamma]EC donor binding site, which is very specific for GSH, and a [gamma]EC acceptor binding site, which is less specific and accepts several tripeptides, namely GSH, h-GSH, and hm-GSH.
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PMID:Synthesis of Phytochelatins and Homo-Phytochelatins in Pisum sativum L. 1222 79

Proteins in the ATP-grasp superfamily of amide bond-forming ligases have evolved to function in a number of unrelated biosynthetic pathways. Previously identified homologs encoding glutathione synthetase, d-alanine:d-alanine ligase and the bacterial ribosomal protein S6:glutamate ligase have been vertically inherited within certain organismal lineages. Although members of this specificity-diverse superfamily share a common reaction mechanism, the nonoverlapping set of amino acid and peptide substrates recognized by each family provided few clues as to their evolutionary history. Two members of this family have been identified in the hyperthermophilic marine archaeon Methanococcus jannaschii and shown to catalyze the final reactions in two coenzyme biosynthetic pathways. The MJ0620 (mptN) locus encodes a tetrahydromethanopterin:alpha-l-glutamate ligase that forms tetrahydrosarcinapterin, a single carbon-carrying coenzyme. The MJ1001 (cofF) locus encodes a gamma-F420-2:alpha-l-glutamate ligase, which caps the gamma-glutamyl tail of the hydride carrier coenzyme F420. These two genes share a common ancestor with the ribosomal protein S6:glutamate ligase and a putative alpha-aminoadipate ligase, defining the first group of ATP-grasp enzymes with a shared amino acid substrate specificity. As in glutathione biosynthesis, two unrelated amino acid ligases catalyze sequential reactions in coenzyme F420 polyglutamate formation: a gamma-glutamyl ligase adds 1-3 l-glutamate residues and the ATP-grasp-type ligase described here caps the chain with a single alpha-linked l-glutamate residue. The analogous pathways for glutathione, F420, folate, and murein peptide biosyntheses illustrate convergent evolution of nonribosomal peptide biosynthesis through the recruitment of single-step amino acid ligases.
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PMID:Glutathione synthetase homologs encode alpha-L-glutamate ligases for methanogenic coenzyme F420 and tetrahydrosarcinapterin biosyntheses. 1290 15

Monosodium methanearsonate (MSMA)-resistant and -susceptible common cocklebur (Xanthium strumarium L.) and cotton (Gossypium hirsutum L.) were treated with MSMA. Plant parameters analyzed were: glutathione synthetase activity, selected amino acid (arginine, glutamic acid, alanine, citrulline, glutamine, and glutathione) content and arsenic content (MSMA, total arsenic, and arsonate). No reduction of arsenic from the parent pentavalent form present in MSMA to the trivalent form was detected. Arginine, glutamic acid, and glutamine content increased in tissue three days after MSMA treatment. Glutathione content decreased during the first three days after treatment; however, five days after treatment the resistant biotype of cocklebur and cotton had elevated glutathione levels (8-20 times greater, respectively). Glutathione Synthetase activity was higher in cotton than in either of the cocklebur biotypes; MSMA did not affect its activity in cotton or either cocklebur biotype. Resistant biotypes have a slightly higher activity than the susceptible biotype. Tolerance of cotton to MSMA may be related to glutathione synthetase activity and possibly to the presence of phytochelatins. Also, increased glutathione levels in the resistant biotype may implicate phytochelatin involvement in the resistance mechanism.
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PMID:MSMA resistance studies. 1547 45

In most organisms, glutathione (GSH) is synthesized by the sequential action of distinct enzymes, gamma-glutamylcysteine synthetase (gamma-GCS) and GSH synthetase (GS). In Streptococcus agalactiae, GSH synthesis is catalyzed by a single enzyme, gamma-glutamylcysteine synthetase-glutathione synthetase (gamma-GCS-GS). The N-terminal sequence of gamma-GCS-GS is similar to Escherichia coli gamma-GCS, but the C-terminal sequence is an ATP-grasp domain more similar to d-Ala, d-Ala ligase than to any known GS. In the present studies, C-terminally and N-terminally truncated constructs were characterized in order to define the limits of the gamma-GCS and GS domains, respectively. Although WT gamma-GCS-GS is nearly uninhibited by GSH (K(i) approximately 140 mM), shorter gamma-GCS domain constructs were unexpectedly found to be strongly inhibited (K(i) approximately 15 mM), reproducing a physiologically important regulation seen in monofunctional gamma-GCS enzymes. Because studies with E. coli gamma-GCS implicate a flexible loop region in GSH binding, chimeras of S. agalactiae gamma-GCS-GS were made containing gamma-GCS domain flexible loop sequences from Enterococcus faecalis and Pasteurella multocida gamma-GCS-GS, isoforms that are inhibited by GSH. Inhibition remained S. agalactiae-like (i.e., very weak). C-Terminal constructs of gamma-GCS-GS have GS activity (0.01-0.04% of WT), but proper folding and significant GS activity required a covalently linked gamma-GCS domain. In addition, site-directed mutants in the middle region of the gamma-GCS-GS sequence established that GS activity depends on residues in a region that is also part of the gamma-GCS domain. Our results provide new insights into the structure of gamma-GCS-GS and suggest gamma-GCS-GS evolved from a monomeric gamma-GCS that became C-terminally fused to a multimeric ATP-grasp protein.
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PMID:Gamma-glutamylcysteine synthetase-glutathione synthetase: domain structure and identification of residues important in substrate and glutathione binding. 1693 98

In higher plants and some fungi, heavy metals induce the synthesis of chelating peptides known as phytochelatins (PCs). They are characterized by the general structure (gamma-Glu-Cys)n-Gly, but in some plant species, the C-terminal glycine can be replaced by serine, glutamine, glutamate or alanine, leading to iso-phytochelatins (iso-PCs). Although the distribution of iso-PCs is considered to differ from one species to another, we previously showed that Arabidopsis thaliana (A. thaliana) cells are able to synthesize most PC-related peptides (PCs and iso-PCs) described in the literature. We also observed an accumulation of the dipeptide gamma-glutamylcysteine (gamma-EC) when cadmium (Cd) (200 microM) was added to the culture medium, suggesting that either glutathione synthetase or glycine availability could be a limiting factor for the biosynthesis of PC-related peptides. In this context, the aim of the present work was to seek new insights into the regulation of PC synthesis by performing metabolic profiling using liquid chromatography-mass spectrometry. The levels of PC-related peptides and their precursors were measured in A. thaliana cells following Cd exposure. A range of doses (0, 50, 200 and 400 microM CdNO3) and kinetic studies (from 1 to 48 h) showed a dose threshold (50 microM CdNO3) and a lag time between the appearance of PCs and iso-PCs concomitant with the gamma-EC accumulation induced by Cd, occurring at cadmium concentrations above 50 microM. This accumulation was suppressed by supplementation of the culture medium with 25 mM glycine. Glycine supplementation had a limited impact on the concentrations of glutathione and PCs whereas the levels of most iso-PCs were significantly increased. Taken together, these results indicate that GSH is involved in the biosynthesis of the iso-PCs in vivo, and that the biosynthesis of PC-related peptides is limited by the availability of glycine in the presence of high cadmium concentrations.
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PMID:New insights into the regulation of phytochelatin biosynthesis in A. thaliana cells from metabolite profiling analyses. 1699 93

Dietary L-arginine (Arg) supplementation reduces white-fat gain in diet-induced obese rats but the underlying mechanisms are unknown. This study tested the hypothesis that Arg treatment affects expression of genes related to lipid metabolism in adipose tissue. Four-week-old male Sprague-Dawley rats were fed a low-fat (LF) or high-fat (HF) diet for 15 weeks. Thereafter, lean or obese rats continued to be fed their same respective diets and received drinking water containing 1.51% Arg-HCl or 2.55% L: -alanine (isonitrogenous control). After 12 weeks of Arg supplementation, rats were euthanized to obtain retroperitoneal adipose tissue for analyzing global changes in gene expression by microarray. The results were confirmed by RT-PCR analysis. HF feeding decreased mRNA levels for lipogenic enzymes, AMP-activated protein kinase, glucose transporters, heme oxygenase 3, glutathione synthetase, superoxide dismutase 3, peroxiredoxin 5, glutathione peroxidase 3, and stress-induced protein, while increasing expression of carboxypeptidase-A, peroxisome proliferator activated receptor (PPAR)-alpha, caspase 2, caveolin 3, and diacylglycerol kinase. In contrast, Arg supplementation reduced mRNA levels for fatty acid binding protein 1, glycogenin, protein phosphates 1B, caspases 1 and 2, and hepatic lipase, but increased expression of PPARgamma, heme oxygenase 3, glutathione synthetase, insulin-like growth factor II, sphingosine-1-phosphate receptor, and stress-induced protein. Biochemical analysis revealed oxidative stress in white adipose tissue of HF-fed rats, which was prevented by Arg supplementation. Collectively, these results indicate that HF diet and Arg supplementation differentially regulate gene expression to affect energy-substrate oxidation, redox state, fat accretion, and adipocyte differentiation in adipose tissue. Our findings provide a molecular mechanism to explain a beneficial effect of Arg on ameliorating diet-induced obesity in mammals.
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PMID:High fat feeding and dietary L-arginine supplementation differentially regulate gene expression in rat white adipose tissue. 1921 6

T-2 toxin is one of the most potent trichothecenes, and on exposure causes severe human and animal diseases. We investigated the dose- and time-dependent effect of T-2 toxin on certain biochemical variables, oxidative damage in terms of antioxidant enzyme activity, and gene expression profile in mice. Mice treated intraperitoneally with either 1 LD50 or 2 LD50 dose (5.61 and 11.22 mg/kg body weight, respectively) of T-2 toxin showed significant alterations in hepatic alanine amino transferase, aspartate amino transferase, and lactate dehydrogenase. Significant changes in hepatic lipid peroxidation, depletion of glutathione (GSH), and expression of heat shock protein-70 indicated oxidative damage. We also evaluated the activity of antioxidant enzymes and compared the gene expression profile by quantitative real-time reverse transcriptase-polymerase chain reaction. Except for glutathione reductase (GR), there was a significant increase in activity of glutathione-S-transferase (GST), glutathione peroxidase (GPx), superoxide dismutase (SOD), and catalase at 1 LD50 dose. At 2 LD50 dose, SOD showed decrease in activity, whereas GST, GPx, and catalase showed significant increase. In contrast, gene expression profile showed downregulation in GR, GPx, GST, and catalase at 1 LD50 dose. At 2 LD50 dose except GSH synthetase, all other genes were downregulated. The results clearly show oxidative stress as one of the mechanisms of T-2 toxin-mediated toxicity.
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PMID:Oxidative damage and gene expression profile of antioxidant enzymes after T-2 toxin exposure in mice. 1952 62

The redox active peptide glutathione is ubiquitous in nature, but some plants also synthesize glutathione analogs in response to environmental stresses. To understand the evolution of chemical diversity in the closely related enzymes homoglutathione synthetase (hGS) and glutathione synthetase (GS), we determined the structures of soybean (Glycine max) hGS in three states: apoenzyme, bound to gamma-glutamylcysteine (gammaEC), and with hGSH, ADP, and a sulfate ion bound in the active site. Domain movements and rearrangement of active site loops change the structure from an open active site form (apoenzyme and gammaEC complex) to a closed active site form (hGSH*ADP*SO(4)(2-) complex). The structure of hGS shows that two amino acid differences in an active site loop provide extra space to accommodate the longer beta-Ala moiety of hGSH in comparison to the glycinyl group of glutathione. Mutation of either Leu-487 or Pro-488 to an Ala improves catalytic efficiency using Gly, but a double mutation (L487A/P488A) is required to convert the substrate preference of hGS from beta-Ala to Gly. These structures, combined with site-directed mutagenesis, reveal the molecular changes that define the substrate preference of hGS, explain the product diversity within evolutionarily related GS-like enzymes, and reinforce the critical role of active site loops in the adaptation and diversification of enzyme function.
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PMID:Structural basis for evolution of product diversity in soybean glutathione biosynthesis. 1994 90

Glutathione (GSH) is an abundant metabolite and a major antioxidant in plant cells. However, in the Leguminosae, homoglutathione (hGSH) may replace glutathione (GSH) partially or completely. To date, cowpea (Vigna unguiculata) has been considered a non-hGSH-producing species, and no hGSHS cDNA has been isolated. Here we report on the cloning of a full-length cDNA coding for a hGSHS (EC 6.3.2.23) and the cloning of a partial cDNA coding for a putative glutathione synthetase (GSHS; EC 6.3.2.3) in cowpea leaf extracts. These cDNAs possess, respectively, the leucine/proline hGSHS signature and the alanine/alanine GSHS signature at the 3' end. Expression analysis showed a significant up-regulation of hGSHS during progressive drought stress that could be directly related to the drought tolerance of the cowpea cultivar used, while GSHS was mainly constitutively expressed. Nevertheless, quantification of low-molecular-weight thiols confirmed the previous findings that cowpea is essentially a GSH producing plant, as no hGSH was detected in the leaves. These findings raise new questions regarding the function, activity and substrate specificity of the cloned hGSHS cDNA. These questions are discussed.
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PMID:Homoglutathione synthetase and glutathione synthetase in drought-stressed cowpea leaves: expression patterns and accumulation of low-molecular-weight thiols. 2003 31

Alkanolamines are surface-active chemicals used in a wide range of industrial, agricultural and pharmaceutical applications and products. Of particular interest is the use of alkanolamines such as diethanolamine (DEA) in the removal of CO(2) from natural gas and for CO(2) capture following fossil fuel combustion. Despite this widespread use, relatively little is known about the ecotoxicological impacts of these compounds. In an attempt to assess the potential effects of alkanolamines in the marine environment, a key species in the North Atlantic, the planktonic copepod Calanus finmarchicus, was studied for molecular effects following sublethal exposure to DEA. DEA-induced alterations in transcriptome and metabolome profiling were assessed using a suppression subtractive hybridization (SSH) gene library method and high resolution magic angle spinning nuclear magnetic resonance (HR-MAS NMR), respectively. Effects were observed on transcription of genes reportedly involved in lipid metabolism, antioxidant systems, metal binding, and amino acid and protein catabolism. These effects were accompanied by altered expression of fatty acid derivates, amino acids (threonine, methionine, glutamine, arginine, alanine and leucine) and cholines (choline, phosphocholine and glycerophosphocholine). Together, SSH and HR-MAS NMR offer complementary screening tools for the assessment of molecular responses of C. finmarchicus to DEA and can be used in the study of other chemicals and organisms. Concentration-response and time-response relationships between DEA exposure and single gene transcription were investigated using quantitative PCR. Specific relationships were found between DEA exposure and the transcription of genes involved in protein catabolism (ubiquitin-specific protease-7), metal ion homeostasis (ferritin) and defence against oxidative stress (gamma-glutamylcysteine synthase, glutathione synthase and Cu/Zn-superoxide dismutase). At the lowest alkanolamine concentration used in these experiments, which corresponded to 0.5% of the LC(50) concentration, no transcriptional effects were observed, giving information regarding the lower molecular effect level. Finally, similar transcription patterns were observed for a number of different genes following exposure to DEA, which indicates analogous mechanisms of toxicity and response.
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PMID:Molecular effects of diethanolamine exposure on Calanus finmarchicus (Crustacea: Copepoda). 2053 12


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