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

---

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

The zonal distribution of GSH metabolism was investigated by comparing hepatocytes obtained from the periportal (zone 1) or perivenous (zone 3) region by digitonin/collagenase perfusion. Freshly isolated periportal and perivenous cells had similar viability (dye exclusion, lactate dehydrogenase leakage and ATP content) and GSH content (2.4 and 2.7 mumol/g respectively). During incubation, periportal cells slowly accumulated GSH (0.35 mumol/h per g), whereas in perivenous cells a decrease occurred (-0.14 mumol/h per g). Also, in the presence of either L-methionine or L-cysteine (0.5 mM) periportal hepatocytes accumulated GSH much faster (3.5 mumol/h per g) than did perivenous cells (1.9 mumol/h per g). These periportal-perivenous differences were also found in cells from fasted rats. Efflux of GSH was faster from perivenous cells than from periportal cells, but this difference only explained 10-20% of the periportal-perivenous difference in accumulation. Furthermore, periportal cells accumulated GSH to a plateau 26-40% higher than in perivenous cells. There was no significant difference in gamma-glutamylcysteine synthetase or glutathione synthetase activity between the periportal and perivenous cell preparations. The periportal-perivenous difference in GSH accumulation was unaffected by inhibition of gamma-glutamyl transpeptidase or by 5 mM-glutamate or -glutamine, but was slightly diminished by 2 mM-L-methionine. This suggests differences between periportal and perivenous cells in their metabolism and/or transport of (sulphur) amino acids. Our results suggest that a lower GSH replenishment capacity of the hepatocytes from the perivenous region may contribute to the greater vulnerability of this region to xenobiotic damage.
...
PMID:Glutathione replenishment capacity is lower in isolated perivenous than in periportal hepatocytes. 290 50

Normal human platelets metabolise [3H]-LTA4 into [3H]-LTC4. Platelets from patients with glutathione synthetase deficiency possessing 10-30% of normal levels of cellular glutathione showed marked reduction in capacity to form [3H]-LTC4 (8-10% of normal) even though exogenous reduced glutathione was added to the incubation medium. To our knowledge this is the first demonstration of a genetic defect in LTC4 synthetase coupled to a defect in cellular glutathione levels.
...
PMID:Human genetic defect in leukotriene C4 synthesis. 302 37

Escherichia coli B glutathione synthetase is composed of four identical subunits; each subunit contains 4 cysteine residues (Cys-122, -195, -222, and -289). We constructed seven different mutant enzymes containing 3, 2, or no cysteine residues/subunit by replacement of cysteine codons with those of alanine in the gsh II gene using site-directed mutagenesis. Three mutant enzymes, Ala289, Ala222/289, Cys-free (Ala122/195/222/289), in which cysteine at residue 289 was replaced with alanine, were not inactivated by 5,5'-dithiobis(2-nitrobenzoate) (DTNB), while the other four mutants retaining Cys-289 were inactivated at the wild-type rate. From these selective inactivations of mutant enzymes by DTNB, the sulfhydryl group modified by DTNB was unambiguously identified as Cys-289. In this way, Cys-289 was found to be also a target of modification with 2-nitrothiocyanobenzoate and N-ethylmaleimide, while Cys-195 was of p-chloromercuribenzoate. These results suggest that both Cys-195 and Cys-289 were not essential for the activity of the glutathione synthetase, but chemical modification of either one of the two sulfhydryl groups resulted in complete loss of the activity. Replacement of Cys-122 to Ala-122 enhanced the reactivity of Cys-289 with sulfhydryl reagents.
...
PMID:Role of cysteine residues in glutathione synthetase from Escherichia coli B. Chemical modification and oligonucleotide site-directed mutagenesis. 304 75

Two modes of killing of Escherichia coli by hydrogen peroxide can be distinguished. Mode-one killing is maximal at 1-2 mM; at higher concentrations the killing rate is approximately half-maximal and is independent of H2O2 concentration but first order with respect to exposure time. Mutagenesis and induction of a phage lambda lysogen are similarly affected by H2O2 concentration, with reduced levels of response above 1-2 mM-H2O2. Mutagenesis is not affected by inactivation of umuC. Mode-one killing requires active metabolism during the H2O2 challenge and it results in sfiA-independent filamentation of both cells that survive and those that are killed by the challenge. This mode of killing is enhanced in xth, polA, recA and recB strains; however, it is unaffected by mutations in the nth, uvrA, uvrB, uvrC, uvrD, rep, gyrA, htpR and rel loci. Mode-one killing is normal in strains totally lacking catalase activity (katE, katG), glutathione reductase (gor) or glutathione synthetase (gshB), but enhanced in a strain lacking NADH dehydrogenase (ndh). Mode-one killing is accelerated by the presence of CN- or by an unidentified function that is induced by anoxic growth and is under the control of the fnr locus. A strain carrying both xth and recA mutations and certain polA mutants appear to undergo spontaneous mode-one killing only under aerobic conditions. Taken together, these observations imply that mode-one killing results from DNA damage that normally occurs at a low, non-lethal level during aerobic growth. Models for the resistance to mode-one killing at dose above 1-2 mM-H2O2 will be discussed. Mode-two killing occurs at high concentrations of H2O2 and longer times. It does not require active metabolism, and cells that are killed do not filament, although survivors demonstrate a dose-dependent growth lag followed by a period of filamentation. Mode-two killing is accompanied by enhanced mutagenesis, but strains with DNA repair defects were not observed to be especially sensitive to this mode of killing.
...
PMID:Toxicity, mutagenesis and stress responses induced in Escherichia coli by hydrogen peroxide. 330 21

Blood samples from 722 unrelated patients with anemia and/or reticulocytosis were submitted to our laboratory for red cell enzyme assay during the past 7 years. Among these 722 cases, we found 82 cases of 7 different red cell enzyme deficiencies and 2 of unstable hemoglobin. Abnormalities of pyruvate kinase (PK) were found to cause hemolysis in 55 patients. Although their average PK activity was about 35% of the normal level, 5 showed normal and 2 demonstrated high PK activity. Among 17 patients in whom pyruvate kinase assays or screening tests had been carried out in routine laboratories, the correct diagnoses had been made in only 4. Glucose-6-phosphate dehydrogenase (G6PD) deficiency was found in 15 patients, pyrimidine 5'-nucleotidase deficiency in 5, glucose phosphate isomerase deficiency in 3, adenylate kinase deficiency in 2, phosphoglycerate kinase deficiency in 1, and glutathione synthetase deficiency in 1 patient. Even after we performed a panel of over 20 different red cell enzyme assays, 519 patients still remained undiagnosed.
...
PMID:Enzymatic diagnosis in non-spherocytic hemolytic anemia. 335 12

Evaluation of idiosyncratic drug reactions in predisposed individuals is limited by ethical concerns arising from rechallenge with the suspected offending agent. A previously developed in vitro method using human lymphocytes and a murine microsomal drug metabolizing system has been used to examine toxicity due to acetaminophen (APAP), sulfonamide antibiotics and aromatic anticonvulsants. An improved method is described in which toxic APAP metabolites are generated by a purified and reconstituted cytochrome P-450 system, minimizing the amount of exogenous detoxification enzymes in the assay. Toxicity is assessed by an objective, automated method based on the reduction of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide to an insoluble purple formazan by the mitochondria of viable cells and correlates with that based on trypan blue exclusion. Toxicity required cytochrome P-450 and NADPH, and was inhibited by SKF 525A. Exogenous glutathione also decreased toxicity in a concentration-dependent manner. Lymphocytes from a glutathione synthetase-deficient patient exhibited markedly enhanced toxicity to APAP exceeding the 95% CL of 10 control subjects over a concentration range of 10 to 1000 micrograms/ml. The data are consistent with the generation of cytochrome P-450-dependent reactive metabolites which subsequently can be detoxified by glutathione. This method allows one to address specifically individual differences in detoxification pathways. The use of an automated assessment of cell viability may prove useful in preclinical screening of new compounds for their propensity to cause "idiosyncratic" drug reactions in a predisposed population.
...
PMID:Drug metabolite toxicity assessed in human lymphocytes with a purified, reconstituted cytochrome P-450 system. 338 48

Perchloric acid extracts of LLC-PK1/Cl4 cells, a renal epithelial cell line, incubated with either [2-13C]glycine L-[3-13C]alanine, or D,L-[3-13C]aspartic acid were investigated by 13C-NMR spectroscopy. All amino acids, except labelled glycine, gave rise to glycolytic products and tricarboxylic acid cycle (TCA) intermediates. For the first time we also observed activity of gamma-glutamyltransferase activity and glutathione synthetase activity in LLC-PK1 cells, as is evident from enrichment of reduced glutathione. Time courses showed that only 6% of the labelled glycine was utilized in 30 min, whereas 31% of L-alanine and 60% of L-aspartic acid was utilized during the same period. 13C-NMR was also shown to be a useful tool for the determination of amino acid uptake in LLC-PK1 cells. These uptake experiments indicated that glycine, alanine and aspartic acid are transported into Cl4 cells via a sodium-dependent process. From the relative enrichment of the glutamate carbons, we calculated the activity of pyruvate dehydrogenase to be about 61% when labelled L-alanine was the only carbon source for LLC-PK1/Cl4 cells. Experiments with labelled D,L-aspartic, however, showed that about 40% of C-3-enriched oxaloacetate (arising from a de-amination of aspartic acid) reached the pyruvate pool.
...
PMID:A 13C-NMR study on the influxes into the tricarboxylic acid cycle of a renal epithelial cell line, LLC-PK1/Cl4: the metabolism of [2-13C]glycine, L-[3-13C]alanine and L-[3-13C]aspartic acid in renal epithelial cells. 340 8

The activity and the kinetic properties of glutathione synthetase and the concentrations of non-protein bound thiols of the gamma-glutamyl cycle were measured in 11 human fibroblast cell strains. Six of these strains were derived from patients suffering from 5-oxoprolinuria, a recessive genetic disease characterized by a deficiency in glutathione synthetase; the other cell strains were derived from healthy heterozygous or homozygous relatives of the patients. The glutathione synthetase activities of homozygous deficient strains were 1/3 of control values while those of heterozygous strains were 2/3 of control values. The total thiol concentration was lower in only 3 of the 6 deficient homozygotes and that of glutathione (GSH) was lower in only 4 of the 6 deficient homozygotes. This lower GSH level was at least partly offset by an accumulation of gamma-glutamylcysteine, a precursor of GSH, which is almost completely absent from control cells. The total quantities of thiols and GSH in plateau phase cells were about 50% and 30% respectively of the levels in growth phase cells. Approximately 80% of the GSH was in the reduced form in both quiescent and growing cells.
...
PMID:Low molecular weight thiol content in glutathione synthetase-deficient human fibroblasts. 343 51

The radioprotective role of endogenous and exogenous thiols was investigated, with survival as the end-point, after radiation exposure of cells under oxic and hypoxic conditions. Human cell strains originating from a 5-oxoprolinuria patient and from a related control were used. Due to a genetic deficiency in glutathione synthetase, the level of free SH groups, and in particular that of glutathione, is decreased in 5-oxoprolinuria cells. The glutathione synthetase deficient cells have a reduced oxygen enhancement ratio (1.5) compared to control cells (2.7). The radiosensitivity was assessed for both cell strains in the presence of different concentrations of an exogenous radioprotector:cysteamine. At concentrations varying between 0.1 and 20 mM, cysteamine protected the two cell strains to the same extent when irradiated under oxic and hypoxic conditions. The protective effect of cysteamine was lower under hypoxia than under oxic conditions for both cell strains. Consequently, the oxygen enhancement ratio decreased for both cell strains when cysteamine concentration increased. These results suggest that cysteamine cannot replace endogenous thiols as far as they are implicated in the radiobiological oxygen effect.
...
PMID:Radioprotective effect of cysteamine in glutathione synthetase-deficient cells. 348 71

Using a human fibroblast strain deficient in glutathione synthetase and a related proficient control strain, the role of glutathione (GSH) in repair of potentially lethal damage (PLD) has been investigated in determining survival by plating cells immediately or 24 h after irradiation. After oxic or hypoxic irradiation, both cell strains repair radiation-induced damage. However, under hypoxic conditions, the proficient cells repair PLD as well as under oxic conditions while the deficient cells repair less PLD after irradiation under hypoxic than under oxic conditions. Therefore, the oxygen enhancement ratio (o.e.r.) for proficient cells is similar whether the cells are plated immediately or 24 h later (2.0 and 2.13, respectively). In contrast, the o.e.r. for deficient cells is lower when the cells are plated 24 h after irradiation than when they are plated immediately thereafter (1.16 as compared to 1.55). The results indicate that GSH is involved in PLD repair and, in particular, in the repair of damage induced by radiation delivered under hypoxic conditions.
...
PMID:Reduced repair of potentially lethal radiation damage in glutathione synthetase-deficient human fibroblasts after X-irradiation. 348 89


<< Previous 1 2 3 4 5 6 7 8 9 10 Next >>

---