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)

Sertoli cells play a major role in the regulation of spermatogenic cell energy metabolism and differentiation. This study demonstrates that Sertoli cells are essential for the maintenance of spermatogenic cell glutathione (GSH), an important intracellular reductant and detoxicant. Primary spermatocytes and round spermatids isolated from Xenopus laevis contained 1.5 +/- 0.1 mM GSH, but sperm lacked detectable GSH. During a 5-day culture period, isolated spermatocytes and spermatids lost 80% of the initial GSH (t 1/2 = 55 h). The levels of GSH were unaffected by L-buthionine-SR-sulfoximine (BSO), a selective inhibitor of GSH synthesis. Cultures of testicular lobules and spermatocysts (composed of germ cells and Sertoli cells) depleted of interstitial tissue lost only 30% of their initial GSH in 4.5 days; the GSH levels decreased during treatment with BSO. Spermatogenic cells in cultured testes maintained their GSH levels for 7 days by a BSO-sensitive mechanism. These results demonstrate that the intracellular GSH levels of spermatogenic cells are dependent upon germ cell-somatic cell interactions. Spermatogenic cells were shown to possess gamma-glutamyl transpeptidase, glutathione synthetase, 5-oxoprolinase, and gamma-glutamylcysteine synthetase activities. [35S] Cysteine incorporation and distribution as analyzed by high performance liquid chromatography (HPLC) showed that isolated spermatogenic cells are capable of GSH synthesis. The rate of GSH synthesis, however, was insufficient to compensate for GSH turnover. These results demonstrate that production of spermatogenic cell GSH is dependent upon Sertoli cells. To our knowledge, this is the first evidence that interactions between different cell types may be of significance in GSH metabolism.
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PMID:Spermatogenic cell-somatic cell interactions are required for maintenance of spermatogenic cell glutathione. 272 29

A procedure for synthesis of glutathione selectivity labeled with isotopes is described. A strain of Escherichia coli enriched in its content of gamma-glutamylcysteine synthetase and glutathione synthetase by recombinant DNA techniques is immobilized in a carrageenan matrix and treated with toluene to render the cells more permeable to the substrates. The immobilized cell matrix is incubated with a mixture containing the appropriately labeled amino acid, the other amino acid constituents of glutathione, ATP, and acetylphosphate. The radiolabeled product is isolated by column chromatography.
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PMID:Glutathione specifically labeled with isotopes. 286 14

The low activity level of lenticular gamma-glutamylcysteine synthetase appears to be an evolutionary phenomenon restricted to higher primates. Rapid reduction with age of the activity of both enzymes (gamma-glutamylcysteine synthetase and glutathione synthetase) required for glutathione synthesis in the human lens was demonstrated in an earlier study. The activities of gamma-glutamylcysteine synthetase and glutathione synthetase, the two enzymes responsible for glutathione synthesis, were determined in 39 lenses from the rhesus monkey (Macaca mulatta) as a function of age. The ages ranged from 137 day old fetuses to a 34 year old monkey. Glutathione synthetase activity decreased 8-fold (units/g lens), 7-fold (units/mg soluble protein) and 2-fold (units/lens) over the age span studied. gamma-Glutamylcysteine synthetase activity decreased 3-fold (units/g lens), 4-fold (units/mg soluble protein) and less than 2-fold (units/lens) over the same age span. A small increase in gamma-glutamylcysteine synthetase activity (units/lens) from embryonic lenses to birth and one year of age was followed in later years by a decrease in activity. In adults, the overall ratio of glutathione synthetase activity to gamma-glutamylcysteine synthetase activity was 42:1 as compared to 77:1 for the human and 2:1 to 4:1 for common domestic species. The aging study data indicate that the rhesus monkey lenticular glutathione synthesis system appears to be a good model for the human lens enzymic system.
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PMID:Activity of glutathione synthesis enzymes in the rhesus monkey lens related to age: a model for the human lens. 286 16

The activities of five enzymes of glutathione metabolism were determined in lenses from cataract-resistant and cataract-prone (Emory) mouse variants at three different ages (5 weeks, 10 weeks and 6 months). The enzymes included those required for glutathione synthesis, gamma-glutamylcysteine synthetase and glutathione synthetase, as well as glutathione-S-transferase, glutathione peroxidase and glutathione reductase. The differences in the activities of the five enzymes in the two mouse variants were not remarkable at any of the three ages. Activity of each enzyme was noted to be in excess of the preceding one in this integrated metabolic pathway, with the exception of glutathione reductase. gamma-Glutamylcysteine synthetase appears to be the pacesetting enzyme of this metabolic scheme in the mouse lens. The activity of each enzyme was compared with that earlier reported for human, rabbit and dog lenses.
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PMID:Glutathione metabolism in lenses of Emory and cataract-resistant mice: activity of five enzymes. 287 Aug 75

New methods for the estimation of red cell gamma-glutamylcysteine synthetase and glutathione synthetase have been developed. gamma-32P ATP is allowed to equilibrate until the gamma and beta phosphate groups are equally labelled. The amount of 32Pi released in the presence of glutamic acid and cysteine, the substrates for GC-S or in the presence of gamma-glutamylcysteine and glycine, the substrates of GSH-S, is measured. This is accomplished by extraction of the phosphomolybdate complex into isobutanol-benzene. The methods are linear with time and hemolysate concentration. Normal values are presented.
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PMID:Improved assay of the enzymes of glutathione synthesis: gamma-glutamylcysteine synthetase and glutathione synthetase. 287 3

We report a new convenient assay of the activity of gamma-glutamylcysteine synthetase (EC 6.3.2.2) and glutathione synthetase (EC 6.3.2.3) in crude microbial extracts as well as in purified enzyme preparations. The assay is based on the quantitative analysis of the reaction products by high-performance liquid chromatography after derivatization of the thiol group with 5,5'-dithiobis-(2-nitrobenzoic acid) as described by J. Reeve, J. Kuhlenkamp, and N. Kaplowitz [(1980) J. Chromatogr. 194, 424-428]. In addition, the procedure yields information on basal levels of gamma-glutamylcysteine and glutathione in crude microbial extracts. The two enzymes responsible for glutathione biosynthesis can be determined in parallel under the same chromatographic conditions. No prior separation from substrates and by-products is necessary. Product formation is linear with time for at least 30 min between 0.03 and 12 mU for both enzymes. Even in crude extracts 0.2-0.5 nmol of products formed can be detected with certainty. The method was found to be sensitive and highly reproducible.
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PMID:Assay of the glutathione-synthesizing enzymes by high-performance liquid chromatography. 287 17

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.
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PMID:Glutathione synthesis in evolution: an Achilles' heel of human and other old world simian lenses. 287 29

The oxidized form of glutathione transport was studied in human erythrocytes in pyrimidine 5'-nucleotidase (P5N) deficiency, a disorder in which the amounts of CTP and UTP in the erythrocytes are elevated. The inhibition of ATP-requiring oxidized glutathione (GSSG) transport by CTP and UTP is believed to play a role in elevating the levels of the reduced form of glutathione (GSH) in the erythrocytes of patients with P5N deficiency. The current investigation was undertaken to determine if GSSG transport actually decreases in the erythrocytes of such patients. Erythrocytes from a 17-year-old patient and a 13-year-old patient with P5N deficiency hemolytic anemia and from ten normal subjects were used as materials for the experiment. Erythrocytes, which had been previously incubated with [3H]glycine, were incubated at 37 degrees C, and the rate of [3H]GSSG transported by the cells was estimated. The velocity of GSSG transport out of the erythrocytes was quite low in the patients, 3.17-3.65 nmol GSSG/ml erythrocytes/hr at 37 degrees C in one case, and 3.30 nmol GSSG/ml erythrocytes/hr in the other case, vs that in the normal controls (6.00 +/- 0.80 nmol GSSG/ml erythrocytes/hr; mean +/- SD). The activity of gamma-glutamylcysteine synthetase and glutathione synthetase did not decrease in the patients. Decreased transport activity of GSSG in addition to a normal synthesis rate for GSH may explain the increased concentration of erythrocyte GSH in P5N deficiency.
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PMID:Erythrocyte-oxidized glutathione transport in pyrimidine 5'-nucleotidase deficiency. 288 6

Intracellular concentrations of glutathione and activities of the enzymes gamma-glutamylcysteine synthetase, glutathione synthetase, and gamma-glutamyl transpeptidase were measured in confluent cultured human fibroblasts cell lines from 14 normal cell lines and four cystinotic cell lines. gamma-Glutamyl transpeptidase had a wide range of variability while the glutathione synthetic enzymes, gamma-glutamylcysteine synthetase and glutathione synthetase, had narrower variations and also exhibited no apparent relationship to glutathione content. No differences in the activities of these enzymes were found between normal and cystinotic cells in confluent cell cultures. The activities of the above enzymes and the cell number and content of glutathione, cystine, DNA, and total protein in two normal and two cystinotic fibroblast cell lines were measured during growth. The following growth-dependency patterns were observed: (1) gamma-glutamylcysteine synthetase activity increased markedly in lag and early log phases in both normal and cystinotic cells and decreased rapidly to low confluent levels thereafter. (2) gamma-Glutamyl transpeptidase showed the same wide range of activity noted at confluency but activities decreased in the log phase of growth, a pattern also seen in cystinotic cells. (3) Glutathione synthetase activity remained relatively constant during growth of normal cells but exhibited a peak of activity during lag and early growth of cystinotic cells. (4) Comparative glutathione levels of normal and cystinotic cells were not significantly different and exhibited similar fluctuations with time. (5) The cystine content of normal and cystinotic cells unexpectedly rose to high levels in the lag phase, then decreased to 0.1 nmol 1/2 cystine/mg protein in normal cells and to 0.3 to 1.2 nmol 1/2 cystine/mg protein in cystinotic cells during the log phase. As confluency was approached, normal cell cystine remained at low levels while cystinotic cell cystine rose to characteristically high levels of 50- to 100-fold greater than normal cells at late confluency. These studies extend our understanding of the regulation of glutathione and cystine content in cultured fibroblasts and suggest that glutathione content is closely controlled throughout the cell cycle in the face of varying activities of its anabolic and catabolic enzymes.
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PMID:Glutathione metabolism in normal and cystinotic fibroblasts. 288 73

Schizosaccharomyces pombe synthesize small cadmium-binding peptides cadystin, structure of which is (gamma-Glu-Cys)n-Gly, in response to cadmium. Mutants unable to synthesize cadystin were found in the mutants hypersensitive to cadmium. Some of them lack activity of either gamma-glutamylcysteine synthetase (EC 6.3.2.2) or glutathione synthetase (EC 6.3.2.3), enzyme involved in glutathione biosynthesis. Some mutants have the same activity levels of these enzymes as wild type has. These results indicate that some steps of cadystin biosynthesis are catalyzed by the enzymes catalyzing glutathione biosynthesis.
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PMID:Isolation of mutants of Schizosaccharomyces pombe unable to synthesize cadystin, small cadmium-binding peptides. 289 29


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