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)

The activities of gamma-glutamylcysteine synthetase and glutathione synthetase, the two enzymes required for glutathione synthesis, were determined as a function of age in lenses of three species of Old World higher primates: orangutan, pigtail monkey and olive baboon. These were compared to enzyme activities in lenses of two prosimians: mouse lemur and galago. gamma-Glutamylcysteine synthetase activity decreased as a function of age in all three Old World simians. The rate of decrease was greatest in the juvenile lenses. In contrast, the enzyme activity increased continuously with age in the galago lens. In the mouse lemur the enzyme activity increased per lens, but was constant when expressed as specific activity or as units per gram of lens. The loss of enzyme activity with age was limited to Old World higher primates apparently representing genetic change. Glutathione synthetase activity decreased logarithmically with age in the lenses of all five species.
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PMID:The effects of age on glutathione synthesis enzymes in lenses of Old World simians and prosimians. 135 6

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

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

Glutathione synthetase from Escherichia coli B showed amino acid sequence homology with mammalian and bacterial dihydrofolate reductases over 40 residues, although these two enzymes are different in their reaction mechanisms and ligand requirements. The effects of ligands of dihydrofolate reductase on the reaction of E. coli B glutathione synthetase were examined to find resemblances in catalytic function to dihydrofolate reductase. The E. coli B enzyme was potently inhibited by 7,8-dihydrofolate, methotrexate, and trimethoprim. Methotrexate was studied in detail and proved to bind to an ATP binding site of the E. coli B enzyme with K1 value of 0.1 mM. The homologous portion of the amino acid sequence in dihydrofolate reductases, which corresponds to the portion coded by exon 3 of mammalian dihydrofolate reductase genes, provided a binding site of the adenosine diphosphate moiety of NADPH in the crystal structure of dihydrofolate reductase. These analyses would indicate that the homologous portion of the amino acid sequence of the E. coli B enzyme provides the ATP binding site. This report gives experimental evidence that amino acid sequences related by sequence homology conserve functional similarity even in enzymes which differ in their catalytic mechanisms.
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PMID:Homology of Escherichia coli B glutathione synthetase with dihydrofolate reductase in amino acid sequence and substrate binding site. 355 73

The two enzymes required to synthesize glutathione de novo have been purified from human erythrocytes. Glutamylcysteine synthetase was purified 4300-fold and was approximately 80% pure based on polyacrylamide gel electrophoresis. The purified enzyme catalyzes the formation of 30.5 mumoles of gamma-glutamyl-cysteine per mg of protein per hr and is inhibited by sulfhydryl inhibitors. Glutathione synthetase was purified 6000-fold from erythrocytes to homogeneity as determined by polyacrylamide gel electrophoresis. The erythrocyte enzyme has a molecular weight of 150,000 and catalyzes the formation of 35.9 mumoles of glutathione per mg of protein per hr. Comparison of the amino acid composition and some kinetic parameters of yeast glutathione synthetase and the erythrocyte enzyme demonstrate similarities between these enzymes.
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PMID:Glutathione synthesis in human erythrocytes. II. Purification and properties of the enzymes of glutathione biosynthesis. 509 71

The two enzymes required for de novo glutathione synthesis, glutamyl cysteine synthetase and glutathione synthetase, have been demonstrated in hemolysates of human erythrocytes. Glutamyl cysteine synthetase requires glutamic acid, cysteine, adenosine triphosphate (ATP), and magnesium ions to form gamma-glutamyl cysteine. The activity of this enzyme in hemolysates from 25 normal subjects was 0.43+/-0.04 mumole glutamyl cysteine formed per g hemoglobin per min. Glutathione synthetase requires gamma-glutamyl cysteine, glycine, ATP, and magnesium ions to form glutathione. The activity of this enzyme in hemolysates from 25 normal subjects was 0.19+/-0.03 mumole glutathione formed per g hemoglobin per min. Glutathione synthetase also catalyzes an exchange reaction between glycine and glutathione, but this reaction is not significant under the conditions used for assay of hemolysates. The capacity for erythrocytes to synthesize glutathione exceeds the rate of glutathione turnover by 150-fold, indicating that there is considerable reserve capacity for glutathione synthesis. A patient with erythrocyte glutathione synthetase deficiency has been described. The inability of patients' extracts to synthesize glutathione is corrected by the addition of pure glutathione synthetase, indicating that there is no inhibitor in the patients' erythrocytes.
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PMID:Glutathione biosynthesis in human erythrocytes. I. Identification of the enzymes of glutathione synthesis in hemolysates. 554 17

The high levels of both enzymes of glutathione synthesis found in the infant human lens rapidly reached lower levels by age 10, and thereafter the rate of decrease diminished. Glutathione synthetase activity in the 6 month old lens was six-fold (units/g lens), four-fold (units/mg soluble protein) and two-fold (units/lens) higher than that in the 83 year old, clear human lens. gamma-Glutamylcysteine synthetase activity in the 6 month old lens was sixteen-fold (units/g lens), ten-fold (units/mg soluble protein) and six-fold (units/lens) higher than that in the 83 year old, clear human lens. When lenses from the young adult beagle, rabbit, bovine, and humans are compared, glutathione synthetase activity (units/g lens) varies by about two-fold. gamma-Glutamylcysteine synthetase activity (units/g lens) is quite similar in the first three species, whereas the enzyme activity is more than a magnitude less in young adult human lenses, and becomes much less with increasing age and in a high proportion of life-support system organ donors. The enzyme activity was undetectable in a few of the latter lenses. Loss of activity was not due to increased susceptibility to heat denaturation. The low levels of the enzyme, and total loss in some situations, suggest that gamma-glutamylcysteine synthetase may be an Achilles' Heel of human lens metabolism.
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PMID:Activity of glutathione synthesis enzymes in human lens related to age. 613 16

Glutathione synthetase was purified about 60-fold with 8.5% of activity yield from the cell extracts of Escherichia coli C600 cells transformed with a recombinant plasmid for the glutathione synthetase gene of E. coli B. The purified enzyme had a Mr of 152,000 and was composed of four identical subunits each with a Mr of 38,000. The Km values of the enzyme for gamma-glutamylcysteine, glycine, and ATP were 2.6, 2.0, and 1.8 mM, respectively. The enzyme was most active at pH 8.5 and at 45 degrees C and required divalent cations such as Mg2+, Mn2+, and Co2+ for activity. The activity was inhibited by oxidized glutathione (Ki = 4.4 mM). Reduced glutathione showed no effect on glutathione synthetase activity.
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PMID:Purification and characterization of glutathione synthetase from Escherichia coli B. 638 79

Glutathione synthetase deficiency is a rare inborn error of metabolism. Low levels of and at times unstable molecules of glutathione synthetase leads to glutathione deficiency affecting various systems of the body. The inheritance is thought to be of autosomal recessive variety. We diagnosed the condition in a neonate and proceeded to investigate the family. The results are discussed below.
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PMID:Glutathione synthetase deficiency: a family report. 815 1

Glutathione synthetase (gamma-L-glutamyl-L-cysteine: glycine ligase (ADP-forming) EC 6.3.2.3: GSHase) catalyzes the synthesis of glutathione from gamma-L-glutamyl-L-cysteine and Gly in the presence of ATP. The enzyme from Escherichia coli is a tetramer with four identical subunits of 316 amino acid residues. The crystal structure of the enzyme has been determined by isomorphous replacement and refined to a 2.0 A resolution. Two regions, Gly164 to Gly167 and Ile226 to Arg241, are invisible on the electron density map. The refined model of the subunit includes 296 amino acid residues and 107 solvent molecules. The crystallographic R-factor is 18.6% for 17.914 reflections with F > 3 sigma between 6.0 A and 2.0 A. The structure consists of three domains: the N-terminal, central, and C-terminal domains. In the tetrameric molecule, two subunits that are in close contact form a tight dimer, two tight dimers forming a tetramer with two solvent regions. The ATP molecule is located in the cleft between the central and C-terminal domains. The ATP binding site is surrounded by two sets of the structural motif that belong to those respective domains. Each motif consists of an anti-parallel beta-sheet and a glycine-rich loop.
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PMID:Three-dimensional structure of the glutathione synthetase from Escherichia coli B at 2.0 A resolution. 844 37


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