Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:6.3.2.3 (glutathione synthetase)
 678 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The activities of gamma-glutamylcysteine synthetase (gamma-GCS) and glutathione synthetase, the two enzymes responsible for glutathione synthesis, were determined in adult lenses from representative species of eight mammalian orders. Lenses from Old World higher simians, including man, exhibited remarkably low gamma-GCS activity when compared to a prosimian and the other seven orders. In contrast, glutathione synthetase activity was comparable and relatively high in all orders. This, together with knowledge of its known lability and control mechanisms, suggests that gamma-GCS is a critical enzyme in the lens of the aging higher primate, whose very low and rate-limiting activity is a latent factor in the development of age-related cataract.
 ...
PMID:Glutathione synthesis in evolution: an Achilles' heel of human and other old world simian lenses. 287 29

Glutathione (GSH), an important physiological antioxidant, is synthesized de novo by the sequential reactions of gamma-glutamylcysteine synthetase (gamma GCS) and GSH synthetase. In the present studies, incubation with the quinones 2,3-dimethoxy-1,4-naphthoquinone (DMNQ) and menadione (MQ), which generate superoxide and hydrogen peroxide, was used to investigate GSH synthesis in bovine pulmonary artery endothelial cells under oxidative stress. MQ can also cause initial depletion of GSH through conjugation, whereas DMNQ cannot. during continuous exposure to DMNQ (5 or 10 microM), elevation of GSH by DMNQ started after 6 h, almost doubled after 24 h, and remained at this level to 48 h. The elevation of GSH by DMNQ was mostly in the reduced form, and the ratio of reduced to oxidized glutathione remained unchanged for the first 24 h. Treatment with MQ (25 or 50 microM) for 30 min caused a significant decrease in GSH and total glutathione. After changing the medium to remove any residual MQ, GSH content doubled during the next 12 h. The enzymatic activity of gamma GCS, the rate-limiting enzyme of GSH biosynthesis, increased twofold after 12 h of exposure of cells to either 5 microM DMNQ or 50 microM MQ. Both DMNQ and MQ treatment caused concentration- and time-dependent increases in gamma GCS-mRNA expression. The elevation of gamma GCS-mRNA content by DMNQ for 12 h was completely blocked by coincubation with 0.05 microgram/ml actinomycin D but not 0.5 microgram/ml cycloheximide, suggesting the elevation of gamma GCS-mRNA content occurred through increased transcription. Our results suggest that increased de novo GSH synthesis, mediated by an elevation in gamma GCS, constitutes an adaptive response to oxidative stress.
 ...
PMID:gamma-Glutamylcysteine synthetase and GSH increase in quinone-induced oxidative stress in BPAEC. 794 45

Tumor necrosis factor (TNF) is an inflammatory cytokine that causes cell injury by generation of oxidative stress. Since glutathione (GSH) is a key cellular antioxidant that detoxifies reactive oxygen species, the purpose of our work was to examine the regulation of cellular GSH, the expression of heavy subunit chain of gamma-glutamylcysteine synthetase (gamma-GCS-HS), and control of intracellular generation of reactive oxygen species in cultured rat hepatocytes treated with TNF. Exposure of cells to TNF (10,000 units/ml) resulted in depletion of cellular GSH levels (50-70%) and overproduction of hydrogen peroxide (2-3-fold) and lipid peroxidation. However, cells treated with lower doses of TNF (250-500 units/ml) exhibited increased levels of GSH (60-80% over control). TNF treatment increased (70-100%) the levels of gamma-GCS-HS mRNA, the catalytic subunit of the regulating enzyme in GSH biosynthesis. Furthermore, intact nuclei isolated from hepatocytes treated with TNF transcribed the gamma-GCS-HS gene to a greater extent than control cells, indicating that TNF regulates gamma-GCS-HS at the transcriptional level. The capacity to synthesize GSH de novo determined in cell-free extracts incubated with GSH precursors was greater (50-70%) in hepatocytes that were treated with TNF; however, the activity of GSH synthetase remained unaltered by TNF treatment indicating that TNF selectively increased the activity of gamma-GCS. Despite activation of nuclear factor-kappaB (NF-kappaB) by TNF, this transcription factor was not required for TNF-induced transcription of gamma-GCS-HS as revealed by deletion constructs of the gamma-GCS-HS promoter subcloned in a chloramphenicol acetyltransferase reporter vector and transfected into HepG2 cells. In contrast, a construct containing AP-1 like/metal response regulatory elements increased chloramphenicol acetyltransferase activity upon exposure to TNF. Thus, TNF increases hepatocellular GSH levels by transcriptional regulation of gamma-GCS-HS gene, probably through AP-1/metal response element-like binding site(s) in its promoter, which may constitute a protective mechanism in the control of oxidative stress induced by inflammatory cytokines.
 ...
PMID:Tumor necrosis factor increases hepatocellular glutathione by transcriptional regulation of the heavy subunit chain of gamma-glutamylcysteine synthetase. 937 27

The tripeptide glutathione plays a pivotal role in the maintenance of the thiol redox state of the cell and for the detoxification of reactive oxygen species. Glutathione is synthesized in two consecutive reactions by y-glutamylcysteine synthetase (gamma-GCS) and glutathione synthetase, respectively. The former enzyme represents the rate limiting step of the synthetic pathway. We have cloned the cDNA and gene of a putative gamma-GCS from Plasmodium falciparum. The contiguous cDNA sequences obtained from various cDNA libraries of P. falciparum K1 and 3D7 encompass 4206 bp or 4038 bp and encode polypeptides of 1119 and 1063 amino acids, respectively. The deduced amino acid sequences show four regions of homology (identity: 31.3-43.9%) to human and Trypanosoma brucei gamma-GCS. These regions are interrupted by three large insertions between 94 and 239 amino acids. Within the first insert a variable repetitive motif was identified, which is responsible for the differing sizes of the sequences. We have analysed this phenomenon in five additional P. falciparum strains and found a high degree of variability in the number of the repeated octamer (Y/C)S(N/D)LQQ(Q/R). Therefore the predicted molecular mass of the proteins from different P. falciparum strains ranges from 124.4 to 133.2 kDa, which is almost twice that of the catalytic subunit of the human host enzyme. Isolation of three genomic clones revealed that the gene does not contain introns. P. falciparum gamma-GCS transcription peaks in trophozoites (24-30 h) suggesting that the antioxidant glutathione is predominantly produced at a time where hemoglobin degradation and the simultaneous formation of reactive oxygen species is maximal.
 ...
PMID:The putative gamma-glutamylcysteine synthetase from Plasmodium falciparum contains large insertions and a variable tandem repeat. 1002 15

Glutathione (L-gamma-glutamyl-L-cysteinylglycine, GSH) is synthesized from its constituent amino acids by the sequential action of gamma-glutamylcysteine synthetase (gamma-GCS) and GSH synthetase. The intracellular GSH concentration, typically 1-8 mM, reflects a dynamic balance between the rate of GSH synthesis and the combined rate of GSH consumption within the cell and loss through efflux. The gamma-GCS reaction is rate limiting for GSH synthesis, and regulation of gamma-GCS expression and activity is critical for GSH homeostasis. Transcription of the gamma-GCS subunit genes is controlled by a variety of factors through mechanisms that are not yet fully elucidated. Glutathione synthesis is also modulated by the availability of gamma-GCS substrates, primarily L-cysteine, by feedback inhibition of gamma-GCS by GSH, and by covalent inhibition of gamma-GCS by phosphorylation or nitrosation. Because GSH plays a critical role in cellular defenses against electrophiles, oxidative stress and nitrosating species, pharmacologic manipulation of GSH synthesis has received much attention. Administration of L-cysteine precursors and other strategies allow GSH levels to be maintained under conditions that would otherwise result in GSH depletion and cytotoxicity. Conversely, inhibitors of gamma-GCS have been used to deplete GSH as a strategy for increasing the sensitivity of tumors and parasites to certain therapeutic interventions.
 ...
PMID:Biologic and pharmacologic regulation of mammalian glutathione synthesis. 1056 25

To search for compounds that reverse the drug resistance induced by glutathione (GSH), an original screening system to detect intracellular GSH depleters was established. Among 8843 microbes derived from the soil samples tested, the extracts of two Streptomyces species named KS6701 and KS8846, lowered the intracellular GSH level of Saccharomyces cerevisiae 5 x 47. From both the microbes, 5-hydroxy-4-oxo-L-norvaline (HON) was isolated as the active compound. At a concentration of 50-100 micrograms/ml, HON also decreased the GSH/protein level of the human ovarian tumor cell line, 2008/C13*5.25 and reversed its resistance to cisplatin. We also investigated the mechanism of the depletion. HON had little effect on gamma-glutamylcysteine synthetase (gamma-GCS) or glutathione synthetase, but HON decreased the quantity of thiol substances when it was spontaneously reacted with them. This suggested that the GSH depletion by HON occurred through a mechanism different from that of buthionine sulfoximine, a selective gamma-GCS inhibitor.
 ...
PMID:5-Hydroxy-4-oxo-L-norvaline depletes intracellular glutathione: a new modulator of drug resistance. 1068 Nov 31

Glutathione (L-gamma-glutamyl-L-cysteinylglycine, GSH), is a vital intra- and extracellular protective antioxidant. Glutathione is synthesized from its constituent amino acids by the sequential action of gamma-glutamylcysteine synthetase (gamma-GCS) and GSH synthetase. The rate-limiting enzyme in GSH synthesis is gamma-GCS. Gamma-GCS expression is modulated by oxidants, phenolic antioxidants, and inflammatory and anti-inflammatory agents in various mammalian cells. The intracellular GSH redox homeostasis is strictly regulated to govern cell metabolism and protect cells against oxidative stress. Growing evidence has suggested that cellular oxidative processes have a fundamental role in inflammation through the activation of stress kinases (JNK, MAPK, p38) and redox-sensitive transcription factors such as NF-kappaB and AP-1, which differentially regulate the genes for proinflammatory mediators and protective antioxidant genes such as gamma-GCS, Mn-SOD, and heme oxygenase-1. The critical balance between the induction of proinflammatory mediators and antioxidant genes and the regulation of the levels of GSH in response to oxidative stress at the site of inflammation is not known. Knowledge of the mechanisms of redox GSH regulation and gene transcription in inflammation could lead to the development of novel therapies based on the pharmacological manipulation of the production of this important antioxidant in inflammation and injury. This FORUM article features the role of GSH levels in the regulation of transcription factors, whose activation and DNA binding leads to proinflammatory and antioxidant gene transcription. The potential role of thiol antioxidants as a therapeutic approach in inflammatory lung diseases is also discussed.
 ...
PMID:Regulation of redox glutathione levels and gene transcription in lung inflammation: therapeutic approaches. 1092 59

Increased glutathione (GSH) level occurs early during liver regeneration and in many drug and/or radiation-resistant tumors. Whether GSH level is elevated in liver cancer is unknown. GSH levels and expression of GSH synthetic enzymes were measured in hepatocellular carcinoma (HCC) and normal liver. GSH levels doubled in HCC. The mRNA levels of g-glutamylcysteine synthetase heavy subunit (GCS-HS) and GSH synthetase (GS) doubled, whereas the expression of GCS light subunit was unchanged. Nuclear run-on assay showed that the rate of gene transcription doubled for both GCS-HS and GS. In HCC, there is increased binding to anti-oxidant response, AP-1 and NF-kB, three cis-acting elements in the 5'-flanking region of the human GCS-HS important for its transcriptional regulation. The role of GSH in cell growth was examined by using HepG2 cells. Cell GSH level was varied by treating cells with cystine (0 to 0.2 mM) with or without GSH ester or buthionine sulfoximine. Cell GSH level correlated directly with growth rate. Finally, preventing the increase in GSH after two-thirds partial hepatectomy blunted liver regeneration. Thus, GSH level is increased during liver growth as a result of up-regulation of GCS-HS and GS. This increase, in turn, facilitates growth.
 ...
PMID:Mechanism and significance of increased glutathione level in human hepatocellular carcinoma and liver regeneration. 1109 88

GSH is the major low-molecular-mass thiol in most organisms. The tripeptide maintains a reduced intracellular environment and protects cellular components from damaging oxidation. GSH is synthesized by the action of two ATP-dependent enzymic steps, in which gamma-glutamylcysteine synthetase (gamma-GCS) catalyses the ligation of glutamate and cysteine and subsequently glutathione synthetase (GS) adds glycine to the dipeptide. Recently it was shown that the synthesis of gamma-glutamylcysteine is crucial for the survival of the erythrocytic stages of the malaria parasite Plasmodium falciparum by using the specific gamma-GCS inhibitor buthionine sulphoximine. In order to investigate further the synthetic pathway of the tripeptide in the parasite, GS was cloned and expressed recombinantly. The deduced amino acid sequence of P. falciparum GS shares only a moderate degree of identity with other known GSs, but the residues responsible for substrate and co-factor binding are almost all conserved, with the exception of the ones involved in gamma-glutamylcysteine binding. The protein is active as a dimer, with a subunit molecular mass of 77 kDa, and the addition of reducing reagents such as dithiothreitol is essential in maintaining enzymic activity, indicating that thiol groups are important for stability and enzymic activity. The K(app)(m) values for gamma-glutamyl-alpha-aminobutyrate, ATP and glycine were determined to be 107.1 microM, 59.1 microM and 5.04 mM, respectively, and the V(max) of 5.24 +/- 0.7 micromol.min(-1).mg(-1) was in the same range as that of the mammalian enzymes. However, the negative co-operativity observed for gamma-glutamylcysteine binding to the rat enzyme was not found for the parasite protein. This may be due to the alteration of several amino acids in the gamma-glutamylcysteine-binding site.
 ...
PMID:Glutathione synthetase from Plasmodium falciparum. 1196 86

An assay of gamma-glutamylcysteine synthetase (gamma-GCS) and glutathione synthetase (GS) in crude extracts of cultured cells and tissues is described. It represents a novel combination of known methods, and is based on the formation of glutathione (GSH) from cysteine, glutamate and glycine in the presence of rat kidney GS for the assay of gamma-GCS, or from gamma-glutamylcysteine and glycine for the assay of GS. GSH is then quantified by the Tietze recycling method. Assay mixtures contain the gamma-glutamyl transpeptidase (GGT) inhibitor acivicin in order to prevent the degradation of gamma-glutamylcysteine and of the accumulating GSH, and dithiothreitol in order to prevent the oxidation of cysteine and gamma-glutamylcysteine. gamma-GCS and GS levels determined by this method are comparable to those determined by others. The method is suitable for the rapid determination of gamma-GCS GS in GGT-containing tissues and for the studies of induction of gamma-GCS and GS in tissue cultures.
 ...
PMID:A spectrophotometric assay of gamma-glutamylcysteine synthetase and glutathione synthetase in crude extracts from tissues and cultured mammalian cells. 1204 60


1 2 3 Next >>