Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0027960 (mole)
 21,279 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Bovine serum albumin (BSA) was phosphorylated by the catalytic subunit of cAMP-dependent protein kinase under general protein phosphorylation conditions. The optimal pH for this phosphorylation was 9.0. The K0.5 (the concentration required for 50% of maximal phosphorylation) for BSA at pH 7.5 was 15 microM. One mole of phosphate was incorporated per mole of BSA, and only one phosphopeptide fragment was obtained after extensive proteolysis with trypsin. BSA phosphorylation required dithiothreitol or GSH, but GSH was only one-fiftieth as effective as dithiothreitol. GSSG counteracted the effect of dithiothreitol and GSH. Phosphorylation increased in a time-dependent and dithiothreitol concentration-dependent manner when BSA was preincubated with dithiothreitol. The increase in the incorporation of 32P correlated with the appearance of up to six free sulfhydryl groups. The effect of dithiothreitol on BSA appeared reversible, since reoxidation of reduced BSA decreased its susceptibility to phosphorylation. These experiments showed that this in vitro phosphorylation is dependent on the sulfhydryl-disulfide state of BSA. The possible implications of the sulfhydryl-disulfide state of proteins in the regulation of phosphorylation are discussed.
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PMID:Effect of sulfhydryl-disulfide state on protein phosphorylation: phosphorylation of bovine serum albumin. 298 43

Peroxidase catalysed the formation of active oxygen in the presence of NADH or GSH and traces of H2O2 and arylamine or phenolic substrates. Some oxygen activation occurred with some arylamines even in the absence of NADH or GSH. Oxygen consumption was proportional to the NADH oxidized or GSSG formed. Approximately 0.80 and 0.40 mol of oxygen were consumed per mole of NADH or GSH oxidized respectively. The requirement for trace amounts of hydrogen peroxide and arylamine or phenolic substrates suggest that redox cycling resulted in H2O2 formation. It is proposed that initially formed phenoxy radicals or arylamine cation radicals oxidize NADH or GSH to radicals which react with oxygen to form superoxide radicals and H2O2.
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PMID:Peroxidase catalysed oxygen activation by arylamine carcinogens and phenol. 300 Jun 37

Pure glutathione reductase from Saccharomyces cerevisiae catalyzed under anaerobic conditions the enzymatic reduction of GSSG using electrochemically reduced methyl viologen as electron donor. The new assay was completely dependent on the amount of active enzyme present, and involved the formation of 1 mol GSH per mole of reduced methyl viologen consumed. The enzyme followed a standard Michaelis-Menten kinetics; a Km = 230 microM for reduced methyl viologen and a turnover number of 969 mumol GSSG reduced per minute per micromole enzyme were determined. The enzymatic activity seemed to depend on the redox potential, showing half-maximal activity at -0.407 V. The enzyme was quite specific: the activity using reduced benzyl viologen as electron donor was just 1.5% of that obtained with reduced methyl viologen at the same concentration and potential. Glutathione reductase was totally inactivated after a brief anaerobic exposure with reduced methyl viologen in the absence of GSSG; a partial reactivation was observed following addition of glutathione disulfide. No inhibition of the methyl viologen-dependent activity was observed in the presence of 2',5'-ADP or 2'-P-5'-ADP-ribose, two NADP(H) analogs, at concentrations which drastically inhibited the NADPH-dependent activity, thus suggesting that the reduced viologen does not interact with the pyridine nucleotide-binding site.
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PMID:Electron transfer between reduced methyl viologen and oxidized glutathione: a new assay of Saccharomyces cerevisiae glutathione reductase. 353 78

Glutathione peroxidase (GSHPx), (glutathione:H2O2 oxidoreductase, EC 1.11.1.9) was purified to homogeneity from human plasma. This resulted in a 6800-fold purification of the enzyme with a 2.8% yield. The purification process involved ammonium sulfate fractionation, DEAE-cellulose batch and column chromatographies, hydroxyapatite, and Sephadex G-200 and DEAE-Sephadex A-25 chromatographies. The major peak on DEAE-Sephadex A-25 column chromatography was found to be homogeneous on polyacrylamide gel electrophoresis in the presence or absence of sodium dodecyl sulfate (SDS). Relative mobility in nondenaturing polyacrylamide gel electrophoresis at pH 8.2 was 0.5 for the purified enzyme as detected by both protein staining and enzyme activity compared with 0.38 for erythrocyte GSHPx. The molecular weight of the plasma enzyme as determined by gel filtration was found to be approximately 100,000. SDS-gel electrophoresis of the plasma enzyme gave a subunit molecular weight of approximately 23,000. This suggests that the plasma enzyme exists as a tetramer in its native state, similar to that seen for the erythrocyte enzyme, but with slightly different mobility on SDS-gel electrophoresis. Plasma GSHPx, like the erythrocyte enzyme, was found to contain approximately four atoms of selenium per mole of protein. Utilizing iodinated concanavalin A, it was found that plasma GSHPx, but not the erythrocyte GSPx, is a glycoprotein. Purified plasma enzyme catalyzes both the reduction of tertiary butyl hydroperoxide and hydrogen peroxide. The apparent Km of plasma GSHPx for GSH is 5.3 mM and for tertiary butyl hydroperoxide it is 0.57 mM. Copper, mercury, and zinc strongly inhibit the enzyme activity of plasma GSHPx. Rabbit antibodies directed against the human erythrocyte GSHPx do not precipitate the enzyme activity of the purified plasma enzyme. Radioimmunoassay utilizing erythrocyte GSHPx and anti-erythrocyte GSHPx antibodies showed that less than 0.13% of the antigenically detectable protein is found in the purified GSHPx from plasma.
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PMID:Purification and characterization of human plasma glutathione peroxidase: a selenoglycoprotein distinct from the known cellular enzyme. 361 51

1. A study of the mode and mechanism of Cu(2+)-induced mitochondrial swelling was carried out. 2. Mitochondrial swelling curves (E(520) turbidity changes) were obtained as a function of [Cu(2+)], pH, temperature and mitochondrial protein concentration. ED(50) was approx. 70mmumoles of Cu(2+). Calculation of the activation energy from the Arrhenius equation gave a value of 22900cal./mole per degree with Q(10) 4.02. 3. No lipid peroxides were formed during swelling. 4. Changes in oxygen consumption (Clark-type electrode) were dependent on the substrate used, but revealed no increased uptake in presence of Cu(2+). 5. Cu(2+)-induced swelling was inhibited by EDTA, 8-hydroxyquinoline, cyanide, citrate, bovine serum albumin, ATP, glutamate, GSH, dithiothreitol and sucrose. Azide, Amytal, antimycin A and oligomycin had no significant effect. Potentiation of swelling was seen with ascorbate, 2,4-dinitrophenol and succinate. 6. The occurrence of different types of mitochondrial swelling and the suggestion that Cu(2+)-induced swelling is mediated through a stoicheiometric interaction with a thiol-containing membrane receptor are discussed.
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PMID:Studies of copper ion-induced mitochondrial swelling in vitro. 566 92

Glutathione (GSH) is a strong nucleophile which reacts well with soft electrophiles, but poorly with both weak and strong electrophiles. Weak electrophiles have low reactivity with all nucleophiles while strong electrophiles react well with weak nucleophiles including superabundant H(2)O. There are enzymes, the GSH transferases, which catalyze GSH conjugation with all the types of electrophiles described above. In order to deal with the wide variety of potential substrates, a multiplicity of GSH transferases exists-each tissue having its own collection and each enzyme having a different substrate specificity. These enzymes are often very abundant, e.g., in the rat liver cytosol, their concentration is 0.2 mM. THE FOLLOWING SUBSTRATES ARE CONSIDERED IN SOME DETAIL: 1-chloro-2,4-dinitrobenzene, the electrophile derived metabolically from paracetamol N-acetyliminoquinone?), benzo(a)pyrene-4-5-oxide, cholesterol-5alpha,6alpha-oxide, benzo(a)pyrene-7,8-diol-9,10-oxide and the electrophiles derived metabolically from aflatoxin B(1) (the 2,3-oxide?). According to the substrate, optimal enzyme rates vary over seven orders of magnitude from 10(-5) to 10(-12) mole/min/mg. Despite the wide embrace of the GSH transferases, not all metabolically produced electrophiles are substrates. We know of the following examples: N-methylol-4-aminoazobenzene and its 4'-hydroxy derivative (these are soft electrophiles and react well with GSH noncatalytically), N-sulfonyloxy-N-methyl-4-aminoazobenzene, N-sulfonyloxy-N-acetyl-2-aminofluorene (these are strong electrophiles which do not react selectively with GSH) and N-hydroxy-2-aminofluorene which appears to react only slowly with GSH. It is of interest in the present context that all these compounds are derived from either arylamine or arylamide carcinogens. Whether the reaction be enzymic or nonenzymic, conjugation with GSH is a very important means of detoxication accounting in some cases for up to 60% of the biliary metabolites. As seen in the example of aflatoxin B(1), very low enzymic rates observed in vitro are sufficient to account for apparently high rates of biliary excretion of GSH conjugates.GSH transferases have evolved other functions apart from the catalysis of GSH conjugation. GSH transferase B participates in the hepatic uptake of bilirubin and the intracellular distribution of the heme prosthetic group. It also has GSH peroxidase activity which suggests that it might participate in the detoxication of by-products of oxygen utilization including those produced by the action of cytochrome P-450. It is shown that GSH transferase B inhibits lipid peroxidation in vitro.
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PMID:The role of glutathione in detoxication. 633 28

Bovine lens aldose reductase (ALR2) is readily modified by glutathione disulphide (GSSG) to an enzyme form (GS-ALR2) exhibiting a reduced catalytic efficiency with all the substrates tested and a reduced susceptibility to inhibition. The modification, which is completely reversed by reduced glutathione (GSH) or dithiothreitol occurs by a pseudo-first-order process with respect to the enzyme and a second order rate constant of 30 +/- 0.1 mol-1 min-1 at 25 degrees C was determined. By measuring the residual activity of ALR2 incubated in different glutathione redox buffers at 25 degrees C, an apparent redox equilibrium constant of 1.4 +/- 0.1 was evaluated. Thus the rate and the maximal extent of ALR2 inactivation are proportional to the redox ratio of the thiol used as modifying agent (i.e. [GSH]/[GSSG]). The stoichiometric reversibility of the enzyme modification might be impaired by a reduced solubility of GS-ALR2 with respect to ALR2 and by an increased susceptibility of the modified enzyme to proteolysis. While the native enzyme form is rather insensitive to proteolytic breakdown. GS-ALR2 is easily degraded by chymotrypsin with the generation of a peptide of 26 kDa with an aminoacid sequence at the aminoterminal side compatible with proteolysis at level of Tyr 7 of aldose reductase. A reduced efficiency in the enzyme-cofactor binding following the GSSG dependent modification of ALR2, appears to be associated to the thiol accessibility of GS-ALR2 measured at different temperatures. GS-ALR2 is characterized by the presence of one glutathione residue, linked through a mixed disulphide bond. This is sustained by: (i) the isoelectric point for the modified enzyme of 4.75, which is 0.1 pH units lower than that observed for the native enzyme, which indicates the contribution of an acidic residue to the pI of GS-ALR2; (ii) the incorporation of radioactivity coming from [3H] labelled GSSG accounting for the presence of one equivalent of glutathione per mole of enzyme. Besides being a general feature of protein reactivity in oxidative conditions, the glutathione-mediated ALR2 modification might be part of a cell strategy to preserve reducing power in conditions of oxidative stress.
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PMID:Glutathione dependent modification of bovine lens aldose reductase. 792 85

Superoxide, generated by a xanthine oxidase/hypoxanthine system, reacts with reduced glutathione (GSH) to cause an increase in oxygen consumption and oxidized glutathione (GSSG) formation, both of which are fully inhibited by superoxide dismutase. In this study we have shown that little, if any, of the additional oxygen consumed is converted to hydrogen peroxide. We have confirmed that approximately 90% of the GSH is oxidized to GSSG, the remainder being converted to the sulfonic acid. Approximately 1.2 mol of GSSG was formed for each additional mole of oxygen consumed in the presence of GSH. The efficiency of the reaction increased with increasing GSH concentration (1-8 mM), pH, and pO2 and with decreasing superoxide generation rate. The results are consistent with a superoxide-dependent chain that does not produce hydrogen peroxide and that is terminated primarily by superoxide dismutation. We propose that this occurs via an initial reaction of superoxide with GSH to produce a sulfinyl radical rather than hydrogen transfer to give the thiyl radical. Our data suggest a rate constant for the superoxide/GSH reaction in the 10(2)-10(3) M-1s-1 range. GSH at the millimolar concentrations found intracellular should react with superoxide, but because superoxide is regenerated, it will not be an effective scavenger. Physiologically, superoxide dismutase is required to prevent chain oxidation of GSH.
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PMID:The reaction of superoxide with reduced glutathione. 797 67

Rates of basal and glucose-stimulated insulin and glutathione secretion were studied in experiments with isolated rat pancreas, as were prooxidant effects on these values. The rate of oxidized and recovered glutathione release was found increased at glucose concentration increase to 16.7 mmoles in perfusion solution. Addition of prooxidants (tert-butyl hydroperoxide and Fe2+) in concentrations 10(-4) mole did not change basal insulin secretion but resulted in reduction of glucose-stimulated hormone release. Under such conditions a reduction of the rate of oxidized and recovered glutathione release by the pancreas was observed which was adequate to changed GSH/GSSG ratio in isolated Langerhans' islets. It may be supposed that lipid peroxidation results in changed thiol-disulfide ratio in Langerhans' islets B cells and in reduction of their sensitivity to secretogen effect.
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PMID:[Insulin secretion by isolated rat pancreas as affected by prooxidants; relationship to glutathione release]. 807 2

The pH-Vmax/KmGSH plot of glutathione S-transferase P (GST-P) showed a bell-shaped profile, indicating bifunctional catalysis for glutathione (GSH) conjugation. The ionization constant (Ke) and the heat of ionization (delta He) of the essential ionizable group in the GSH binding site were measured and the value of pKe1 was 5.9 and that of pKe2, 8.4, while the delta He1 and delta He2 were -0.2 and 7.9 kcal/mole, respectively. In a solvent containing 25% ethanol, pKe1 and pKe2 shifted to the alkaline side by 0.47 and 0.2, respectively. These kinetic results indicated that carboxyl and phenolic groups were ionizable groups essential for the GSH conjugation. Chemical modifications using aminomethane sulfonic acid and N-acetylimidazole supported the results of the kinetic studies.
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PMID:Identification of ionizable groups essential for the enzyme catalysis on glutathione S-transferase P. 835 5


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