UMLS:C1260386 - FACTA Search

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Query: UMLS:C1260386 (GSH)
38,102 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The activity of 3-hydroxy-3-methylglutaryl-coenzyme A reductase (hydroxymethylglutaryl-CoA reductase, EC 1.1.1.34) in preparations of thiol-deficient rat liver microsomes and microsomes containing thiols have been compared. Unlike microsomes containing thiols, which possess an active hydroxymethylglutaryl-CoA reductase (Ea), thiol-deficient microsomes contain an inactive, latent enzyme (E1) which can be activated by addition of thiols. Ea can be converted to E1 by dialysis. The maximal degree of activation of E1 depends on the activating thiol with the order of effectiveness: dithioerythritol = dithiothreitol greater than glutathione (GSH) greater than cysteine. Ea is inhibited by oxidized glutathione (GSSG). The degree of the inhibition of Ea by GSSG is proportional to the ratio GSSG/thiol in the reaction. E1 was solubilized from microsomes and purified. Its molecular weight is estimated to be 104 000 by gel filtration chromatography on Sepharose 6B. The reducing agents NaBH4, dithionite and ascorbate failed to activate E1. NaBH4 did not inhibit Ea whereas only partial inhibition was caused by ascorbate and dithionite. Soluble Ea binds to both blue dextran/Sepharose 4B and agarose/hexane-3-hydroxy-3-methylglutaryl Coenzyme A affinity resins at low-salt concentrations. By contrast, soluble E1 did not bind to agarose/hexane-hydroxymethylglutaryl-CoA whereas quantitative binding of E1 to blue dextran/Sepharose 4B was still observed at low salt concentrations. These results indicate that thiols are necessary cofactors for hydroxymethylglutaryl-CoA reductase reaction. Their effect on the activation of E1 is not caused by change in the state of aggregation of the enzyme. Rather, the reversible change of the enzyme from E1 to Ea is affected by increasing the affinity of the enzyme to the substrate hydroxymethylglutaryl-CoA.
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PMID:Thiol-disulfide-dependent interconversion of active and latent forms of rat hepatic 3-hydroxy-3-methylglutaryl-coenzyme A reductase. 715 Jun 22

Peroxynitrite (ONOO-) is widely recognized as mediator of NO toxicity, but recent studies have indicated that this compound may also have physiological activity and induce vascular relaxation as well as inhibition of platelet aggregation. We found that ONOO- induced a pronounced increase in endothelial cyclic GMP levels, and that this effect was significantly attenuated by pretreatment of the cells with GSH-depleting agents. In the presence of 2 mM GSH, ONOO- stimulated purified soluble guanylyl cyclase with a half-maximally effective concentration of about 20 microM. In contrast to the NO donor 2,2-Diethyl-1-nitroso-oxyhydrazine sodium salt (DEA/NO), ONOO- was completely inactive in the absence of GSH, indicating that thiol-mediated bioactivation of ONOO- is involved in enzyme stimulation. Studies on the reaction between ONOO- and GSH revealed that about 1% of ONOO- was non-enzymatically converted to S-nitrosoglutathione. The authentic nitrosothiol was found to be stable in solution, but slowly decomposed in the presence of GSH. GSH-induced decomposition of S-nitrosoglutathione was apparently catalyzed by trace metals and was accompanied by a sustained release of NO and a 40-100-fold increase in its potency to stimulate purified soluble guanylyl cyclase. Our data suggest that the biologic activity of ONOO- involves S-nitrosation of cellular thiols resulting in NO-mediated cyclic GMP accumulation.
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PMID:Peroxynitrite-induced accumulation of cyclic GMP in endothelial cells and stimulation of purified soluble guanylyl cyclase. Dependence on glutathione and possible role of S-nitrosation. 761 39

The common substrate for glutathione S-transferases (GSTs), 1-chloro-2,4-dinitrobenzene (CDNB), is an inhibitor of Escherichia coli growth. This growth inhibition by CDNB is enhanced when E. coli expresses a functional GST. Cells under growth inhibition have reduced intracellular GSH levels and form filaments when they resume growth. Based on this differential growth inhibition by CDNB we have developed a simple procedure to select for null-mutants of a human GST in E. coli. Null mutations in the human GST gene from hydroxylamine mutagenesis or oligonucleotide-directed mutagenesis can be selected for on agar plates containing CDNB after transformation. The molecular nature of each mutation can be identified by DNA sequence analysis of the mutant GST gene. We have identified three essential amino acid residues in an alpha class human GST at Glu31, Glu96, and Gly97. Single substitution at each of these residues, E31K, E96K, G97D, resulted in mutant GST proteins with loss of CDNB conjugation activity and failure in binding to the S-hexyl GSH affinity matrix. In contrast, a mutant GST (Y8F) resulting from substitution of the conserved tyrosine near the N terminus has much reduced CDNB conjugation activity but was still capable of binding to the S-hexyl GSH-agarose. Additional mutant GSTs with substitutions at position 96 (E96F, E96Y) and 97 (G97P, G97T, G97S) resulted in changes in both Km and kcat to different extents. The in vitro CDNB conjugation activity of the purified mutant enzymes correlate negatively with the plating efficiencies of strains encoding them in the presence of CDNB. Based on the x-ray structure model of human GST 1-1, two of these residues are involved in salt bridges (Arg19-Glu31, Arg68-Glu96) and the third Gly97 is in the middle of the helix alpha 4. Our results provide evidence in vivo that Tyr8, Gly97, and the two salt bridges are important for GST structure-function. This molecular genetic approach for the identification of essential amino acids in GSTs should be applicable to any GSTs with CDNB conjugation activity. It should also complement the x-ray crystallographic approach in understanding the structure and function of GSTs.
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PMID:A molecular genetic approach for the identification of essential residues in human glutathione S-transferase function in Escherichia coli. 781 27

Male C57BL/6 mice were fed with normal diet (ND) or diets containing 0.3 or 0.5% ursodeoxycholic acid (UDCA) for 3 weeks. Glutathione S-transferase (GST) activities in the liver cytosolic fraction of these animals toward 1,2-dichloro,4-nitrobenzene (DCNB) as well as to 1-chloro,2,4-dinitrobenzene (CDNB) were significantly increased in a dose dependent manner in UDCA-treated groups compared with the control (ND-fed) animal group (one-way ANOVA). Reduced glutathione (GSH) levels tended to slightly decrease with UDCA diets but the difference did not attain a statistical significance (P > 0.05, one-way ANOVA). Twenty four hr survival rates after an oral challenge of 3.5 mg/kg of DCNB were significantly higher (P < 0.05, Chi-square test) in the two UDCA fed groups (10/10 for 0.5% group, 8/11 for 0.3% group) compared with the control group (3/11). Thus, UDCA appears to reduce the systemic toxicity of DCNB which is detoxified by the hepatic GST system. Although UDCA has been shown to exert hepatoprotective effects in experimental animals and humans in the past, to the best of our knowledge, the present study is the first report that UDCA reduces the systemic toxicity of a toxicant which is detoxified by the hepatic GST system. Although a direct proof is not available, it is most likely that the reduction of the systemic toxicity of DCNB was achieved by the increase in GST activity caused by UDCA feeding. This finding may open a new research field with regard to the unique biological properties of this bile salt in modulating hepatic detoxifying enzymes.
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PMID:Ursodeoxycholic acid reduces the systemic toxicity of 1,2-dichloro,4-nitrobenzene by stimulating hepatic glutathione S-transferase in mice. 813 88

Prostaglandin (PG) A2 has been shown to be actively incorporated into mammalian cells and transferred to the cell nucleus. To characterize the properties of the PGA2 transfer system, we examined the status of PGA2 in L-1210 cells with modified cellular glutathione (GSH) levels. PGA2 in the cytosol fraction of the cells existed in its free-form, the GSH conjugate-form and a macromolecule associate-form (protein bound-form). When the GSH level was lowered under culture conditions, the amount of free-form increased while that of the protein bound-form was unchanged. When PGA2-loaded cells were incubated in a salt solution, free- and conjugate-forms were emitted from the cells. A concomitant decrease and increase of protein bound PGA2 in cytosol and nuclei, respectively, were observed. Subsequent studies with isolated cellular fractions revealed that PGA2 bound to cytosolic protein was transported into the nuclear interior in a temperature-dependent manner. The binding of PGA2 to the protein and subsequent transport to the nuclei were inhibited by PGJ2 and 4-hydroxy-cyclopentenone, but not by PGB2, PGD2, PGE2, PGF2 alpha, arachidonic acid or oleic acid. N-Ethylmaleimide (NEM) and p-chloromercuribenzoate (PCMB) strongly interfered with these transfer processes, suggesting that sulfhydryl components are involved in the transport of PGA2. Subfractionation of cytosol by gel chromatography proved the presence of two proteins (100-150 kDa and 25-35 kDa) that are capable of transporting PGA2 to cell nuclei. Though 40-45 kDa proteins, which correspond to GSH S-transferases, bound to PGA2, they lacked the nuclear transport activities.
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PMID:Characterization of the transport system of prostaglandin A2 in L-1210 murine leukemia cells. 836 39

A chromatographic method is described to determine the distribution of selenium between selenoprotein P, glutathione peroxidase (GSH-Px), and albumin in plasma, using two small columns of heparin-Sepharose and reactive blue 2-Sepharose linked together in tandem. One milliliter of plasma was diluted to 12 ml with 0.02 M sodium phosphate buffer, pH 7.0 (the equilibration buffer), applied to the heparin-Sepharose column, and eluted at a flow rate of 30 ml per hour. GSH-Px was not retained by either of these columns but selenoprotein P was retained by heparin-Sepharose and albumin by reactive blue. After the two columns were separated, selenoprotein P was eluted with heparin from heparin-Sepharose and albumin eluted from reactive blue with high salt. Analytical work confirmed the presence of selenoprotein P, GSH-Px, and albumin in the respective fractions. When rats were injected with 75Se as either selenite or selenomethionine most of the radioactivity was incorporated into the selenoprotein P fraction, with the next greatest amount into GSH-Px, and the least amount into albumin. Slab gel electrophoresis was used to determine that most of the selenium in each of the three fractions was associated with each of these selenium containing proteins. This method indicated that the majority of the selenium in plasma is associated with selenoprotein P, and the only time this was found not to be true was with high levels of dietary selenomethionine.
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PMID:Determination of the distribution of selenium between glutathione peroxidase, selenoprotein P, and albumin in plasma. 843 86

This study was performed to explore the mechanisms by which sulphobromophthalein (BSP) reduces the secretion of biliary lipid using Sprague-Dawley rats (SDR) and mutant rats with congenital conjugated hyperbilirubinaemia bred from SDR (EHBR). We infused the bile-salt-pool-depleted rats with sodium taurocholate at a constant rate of 160 nmol/min per 100 g body wt. with BSP (12.5, 25 and 50 nmol/min per 100 g body wt.) or BSP-GSH (12.5, 25 and 50 nmol/min per 100 g body wt.). The biliary secretion of BSP and BSP-GSH was markedly impaired in EHBR as compared with that in SDR. BSP reduced the biliary secretion of cholesterol and phospholipids in a dose-dependent manner without affecting the secretion of bile salts and composition of fatty acids in phospholipids in SDR, but had no effect on lipid secretion in EHBR. In contrast, BSP-GSH had no such effect on biliary lipids, either in the SDR or EHBR. In addition, the amount of BSP in the liver of EHBR was in the same range as that of SDR. Therefore it is unlikely that an intracellular mechanism is involved in the phenomenon of uncoupling by BSP. We conclude that the uncoupling of biliary lipids from bile-salt secretion by BSP occurs at the level of the bile canaliculus following the secretion of unconjugated BSP.
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PMID:Partial characterization of mechanism(s) by which sulphobromophthalein reduces biliary lipid secretion. 847 Oct 36

Nacystelyn (NAL), a recently-developed lysine salt of N-acetylcysteine (NAC), and NAG, both known to have excellent mucolytic capabilities, were tested for their ability to enhance cellular antioxidant defence mechanisms. To accomplish this, both drugs were tested in vitro for their capacity: (1) to inhibit O2- and H2O2 in cell-free assay systems; (2) to reduce O2- and H2O2 released by polymorphonuclear leukocytes (PMN); and (3) for their cellular glutathione (GSH) precursor effect. In comparison with GSH, NAL and NAC inhibited H2O2, but not O2-, in cell-free, in vitro test systems in a similar manner. The anti-H2O2 effect of these drugs was as potent as that of GSH, an important antioxidant in mammalian cells. To enhance cellular GSH levels, increasing concentrations (0-2 x 10(-4) mol l-1) of both substances were added to a transformed alveolar cell line (A549 cells). After NAC administration (2 x 10(-4) mol l-1), total intracellular GSH (GSH + 2GSSG) levels reached 4.5 +/- 1.1 x 10(-6) mol per 10(6) cells, whereas NAL increased GSH to 8.3 +/- 1.6 x 10(-6) mol per 10(6) cells. NAC and NAL administration also induced extracellular GSH secretion; about two-fold (NAC), and 1.5-fold (NAL), respectively. The GSH precursor potency of cystine was about two-fold higher than that of NAL and NAC, indicating that the deacetylation process of NAL and NAC slows the ability of both drugs to induce cellular glut production and secretion. Buthionine-sulphoximine, which is an inhibitor of GSH synthetase, blocked the cellular GSH precursor effect of all substances. In addition, these data demonstrate that NAC and NAL reduce H2O2 released by freshly-isolated cultured blood PMN from smokers with chronic obstructive pulmonary disease (COPD) (n = 10) in a similar manner (about 45% reduction of H2O2 activity by NAC or NAL at 4 x 10(-6) mol l-1). In accordance with the results obtained from cell-free, in vitro assays, O2- released by PMN was not affected. Ambroxol (concentrations: 10(-9)-10(-3) mol l-1) did not reduce activity levels of H2O2 and O2- in vitro. Due to the basic effect of dissolved lysine, which separates easily in solution from NAL, the acidic function of the remaining NAC molecule is almost completely neutralized [at concentration 2 x 10(-4) M: pH 3.6 (NAC), pH 6.4 (NAL)]. Due to their function as H2O2 scavengers, and due to their ability to enhance cellular glutathione levels, NAL and NAC both have potent antioxidant capabilities in vitro. The advantage of NAL over NAC is two-fold; it enhances intracellular GSH levels twice as effectively, and it forms neutral pH solutions whereas NAC is acidic. Concluding from these in vitro results, NAL could be an interesting alternative to enhance the antioxidant capacity at the epithelial surface of the lung by aerosol administration.
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PMID:Nacystelyn, a novel lysine salt of N-acetylcysteine, to augment cellular antioxidant defence in vitro. 913 55

A new methoxymorpholinyl derivative of Adriamycin (ADR), FCE 23762 (MRD), has recently been selected for phase I clinical trials for its reduced cardiotoxicity and for its cytotoxic activity against a broad spectrum of solid tumors and leukemias that are sensitive or resistant to ADR. The purpose of the present study was to compare the in vitro antitumor activity of MRD and ADR on human melanoma lines with different chemosensitivity to triazene compounds, among which dacarbazine remains a reference drug in the treatment of melanoma. Both MRD and ADR were tested in vitro on three melanoma lines, MI13443-MEL, SK-MEL-28, and M14, previously screened for their chemosensitivity to the triazene compound p-(3-methyl-1-triazeno) benzoic acid, potassium salt (MTBA). The three lines were also analyzed for P-170 expression, total glutathione (GSH) content, and GSH-related enzyme activity. All melanomas, whether sensitive or resistant to MTBA, were susceptible to anthracycline treatment. The cytotoxic activity of MRD was comparable with that of ADR, and no substantial difference was found in cell growth inhibition between the two drugs. When the relative chemosensitivity of the three lines was considered, SK-MEL-28 was found to be slightly less sensitive to MRD treatment than the other tumors. This finding seems to correlate with the higher GSH-peroxidase activity of this melanoma relative to that of the MI13443 and M14 lines. These results show a homogeneous response of melanoma lines to MRD treatment in vitro, suggesting that phase I clinical trials concerning this drug, which in vivo appears to be activated to a more cytotoxic metabolite, could be extended to metastatic melanomas, including those completely resistant to triazene compounds.
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PMID:In vitro antitumor activity of 3'-desamino-3'(2-methoxy-4-morpholinyl) doxorubicin on human melanoma cells sensitive or resistant to triazene compounds. 918 41

Ca2+-activated K+ channels (K(Ca2+)) constitute key regulators of the endothelial cell electrophysiological response to InsP3-mobilizing agonists. Inside-out and outside-out patch clamp experiments were thus undertaken to determine if the gating properties of a voltage-insensitive K(Ca2+) channel of intermediate conductance present in bovine aortic endothelial (BAE) cells could be modified by specific sulfhydryl (SH) oxidative and/or reducing reagents. The results obtained first indicate that cytosolic application of hydrophilic oxidative reagents such as 5,5'-dithio-bis(2-nitrobenzoic acid) (DTNB) (0.2 to 5 mM) or [(O-carboxyphenyl)thio]ethyl mercury sodium salt (thimerosal) (0.5 to 5 mM) reduces gradually the K(Ca2+) channel activity with no modification of the channel unitary conductance. The inhibitory action of DTNB (1 to 5 mM) or thimerosal (1 to 5 mM) was not reserved following withdrawal of the oxidative agents, but channel activity could partly be restored by the addition of the SH group reducing agents dithiothreitol (DTT) (5 mM) or reduced glutathione (GSH) (5 mM) in 53% and 50% of the inside-out experiments performed with DTNB and thimerosal respectively. Similar results were obtained using H2O2 at concentrations ranging from 500 microM to 10 mm as oxidative reagent. In contrast, the lipid soluble oxidative agent 4,4'-dithiodipyridine (4-PDS) (1 mM) appeared in inside-out experiments less potent than DTNB and thimerosal at inhibiting the K(Ca2+) channel activity, suggesting that the critical SH groups involved in channel gating are localized at the inner face of the cell membrane. This conclusion was further substantiated by a series of outside-out patch clamp experiments which showed that DTNB (5 mM) and thimerosal (5 mM) were unable to inhibit the K(Ca2+) channel activity when applied to the external surface of the excised membrane. Finally, no significant changes of the gating properties of the K(Ca2+) channel were observed in inside-out experiments where the SH group reducing agents DTT and GSH were applied immediately following membrane excision. However, the application of either GSH or DTT was found to partly restore channel activity in experiments where the K(Ca2+) channels showed significant rundown.
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PMID:Effects of thiol-modifying agents on a K(Ca2+) channel of intermediate conductance in bovine aortic endothelial cells. 923 92

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