Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.5.1.18 (glutathione S-transferase)
 22,582 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The peroxisome proliferators perfluorooctanoic acid (PFOA; 0.02% w/w), perfluorodecanoic acid (PFDA; 0.02%, w/w), nafenopin (0.125%, w/w), clofibrate (0.5%, w/w), and acetylsalicylic acid (ASA; 1%, w/w) were administered to male C57 BL/6 mice in their diet for two weeks. Parameters for Fe3+ ADP, NADPH or ascorbic acid-initiated lipid peroxidation in vitro were measured. Approximately a twofold increase in susceptibility to lipid peroxidation was obtained for all the peroxisome proliferators tested. Cotreatment of mice with the peroxisome proliferator ASA (1%, w/w) and a catalase inhibitor, 3-amino-1,2,4-triazole (AT; 0.4%, w/w) for 7 days resulted in little inhibition of peroxisome proliferation, an elevated level of H2O2 in vivo, and total inhibition of the increased susceptibility to lipid peroxidation in vitro. No increase in lipid peroxidation in vivo was observed. Certain antioxidant enzymes (DT-diaphorase, superoxide dismutase, glutathione transferase, glutathione peroxidase, and glutathione reductase) and components (ubiquinone and alpha-tocopherol) were also measured. The results showed that there was some induction of these antioxidant enzymes and components by ASA or aminotriazole, except for glutathione peroxidase and superoxide dismutase, which were inhibited. The possible involvement of oxidative stress in the carcinogenicity of peroxisome proliferators is discussed.
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PMID:Hepatic oxidative stress and related defenses during treatment of mice with acetylsalicylic acid and other peroxisome proliferators. 756 57

The cDNA encoding QPc-9.5 kDa (subunit VII) of bovine heart mitochondrial ubiquinol-cytochrome c reductase was cloned and sequenced. This cDNA is 665 base pairs long with an open reading frame of 246 base pairs that encodes an 81-amino acid mature QPc-9.5 kDa. The insert contains 395 base pairs of a 3'-noncoding sequence with a poly(A) tail. The amino acid sequence of QPc-9.5 kDa deduced from this nucleotide sequence is the same as that obtained by protein sequencing except that residue 61 is tryptophan instead of cysteine. The QPc-9.5 kDa was overexpressed in Escherichia coli JM109 cells as a glutathione S-transferase fusion protein (GST-QPc) using the expression vector, pGEX/QPc. The yield of soluble active recombinant GST-QPc fusion protein depends on the induction growth time, temperature, and medium. Maximum yield of recombinant fusion protein was obtained from cells harvested 3 h postinduction of growth at 27 degrees C on LB medium containing betaine and sorbitol. QPc-9.5 kDa was released from the fusion protein by proteolytic cleavage with thrombin. Isolated recombinant QPc-9.5 kDa showed one protein band in SDS-polyacrylamide gel electrophroesis corresponding to subunit VII of mitochondrial ubiquinol-cytochrome c reductase. Although the isolated recombinant QPc-9.5 kDa is soluble in aqueous solution, it is in a highly aggregated form, with an apparent molecular mass of over 1 million. Addition of detergent deaggreates the isolated protein to the monomeric state, suggesting that the recombinant protein exists as a hydrophobic aggregation in aqueous solution. The recombinant QPc-9.5 kDa binds ubiquinone and shows a spectral blue shift. Upon titration of the recombinant protein with ubiquinone, a saturation behavior is observed, suggesting that the binding is specific and that the recombinant protein may be in the functionally active state.
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PMID:Cloning, gene sequencing, and expression of the small molecular mass ubiquinone-binding protein of mitochondrial ubiquinol-cytochrome c reductase. 759 38

We found that NADPH-dependent ubiquinone reductase (NADPH-UQ reductase) in rat liver cytosol reduces ubiquinone (UQ) to ubiquinol (UQH2) in lipid membranes and consequently inhibits lipid peroxidation [Takahashi T., et al., Biochem. J., 309, 883-890 (1995)]. Here we examined whether or not this UQH2-regenerating system functions as a cellular antioxidant defense in animals. Rats were given UQ-10 for 2 weeks, and were then exposed to carbon tetrachloride (CCl4). The UQ-10 supplement increased only in the NADPH-UQ reductase and the UQH2-10 pool of rat liver without any appreciable change in the levels of other antioxidant factors. On the other hand, CCl4 markedly increased plasma aspartate aminotransferase and alanine aminotransferase, liver weight and thiobarbituric acid reacting substances formation, which are indicators of CCl4-hepatitis, and it decreased the liver levels of L-ascorbic acid, reduced form of glutathione (GSH), alpha-tocopherol, NADPH-UQ reductase and glutathione S-transferase. However, all the above indicators of CCl4-induced hepatitis were significantly improved in rats given UQ-10. Furthermore, alpha-tocopherol, but neither L-ascorbic acid nor GSH, was significantly saved. UQ-10 supplement also was recovered glutathione S-transferase and NADPH-UQ reductase activities slightly. These results indicated that UQ-10 given to rats increased the cellular UQH2-10 pool and cytosolic NADPH-UQ reductase activity in their livers, resulting in the inhibition of lipid peroxidation in the biomembranes, and consequently protected the rats from the CCl4-hepatotoxicity.
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PMID:Cellular antioxidant defense by a ubiquinol-regenerating system coupled with cytosolic NADPH-dependent ubiquinone reductase: protective effect against carbon tetrachloride-induced hepatotoxicity in the rat. 887 5

Recent studies have revealed binding of mitochondrial enoyl-CoA isomerase (ECI) to S-hexylglutathione-Sepharose, an affinity matrix used for purification of glutathione transferases (GSTs), and the enzyme has been suggested to be identical with the Alpha class form of GST with a subunit molecular mass of about 30 kDa. In the present study, S-hexylglutathione-binding proteins of human hepatocellular carcinomas were characterized to examine their identity. Supernatant fractions of carcinoma and surrounding tissues were applied to an affinity column, and bound fractions were resolved into three proteins with subunit molecular masses/pI values of 33 kDa/7.0, 30 kDa/5.8 and 29 kDa/5.8 in addition to the well-characterized four GST subunits, A1, A2, M1 and P1, by two-dimensional gel electrophoresis. The proteins were further purified by chromatofocusing at pH 7.4-4.0. The 30 and 29 kDa proteins were eluted at pH 4.9 and by 1 M NaCl respectively, and could be clearly separated from each other. The 29 kDa protein exhibited a low but significant activity towards 1-chloro-2,4-dinitrobenzene (4.25 micromol/min per mg of protein) and reacted with anti-(GST A1-2) antibody, suggesting that it is a member of the GST Alpha class. The 30 kDa protein did not react with anti-GST antibodies and was identified as ECI by immunoblotting and N-terminal-amino-acid-sequencing analyses. The results thus indicated that the Alpha class GST form composed of the 29 kDa subunits and ECI are two different proteins. The 33 kDa protein was eluted from the chromatofocusing column at pH 7.0 and did not react with either anti-GST antibodies or antibodies against mitochondrial enzymes involved in the beta-oxidation of fatty acids. However, it exhibited a carbonyl reductase activity with menadione and ubiquinone, and amino acid sequences of its peptides cleaved by Staphylococcus aureus V8 proteinase were consistent with those reported for the enzyme. Thus this protein binding to S-hexylglutathione-Sepharose was identified as carbonyl reductase.
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PMID:Characterization of S-hexylglutathione-binding proteins of human hepatocellular carcinoma: separation of enoyl-CoA isomerase from an Alpha class glutathione transferase form. 937 3

The smallest membrane-anchoring subunit (QPs3) of bovine heart succinate:ubiquinone reductase was overexpressed in Escherichia coli JM109 as a glutathione S-transferase fusion protein using the expression vector pGEX2T/QPs3. The yield of soluble active recombinant glutathione S-transferase-QPs3 fusion protein was isopropyl-1-thio-beta-D-galactopyranoside concentration-, induction growth time-, temperature-, and medium-dependent. Maximum yield of soluble recombinant fusion protein was obtained from cells harvested 3.5 h post-isopropyl-1-thio-beta-D-galactopyranoside (0.4 mM)-induction growth at 25 degrees C in 2.0% tryptone, 0.5% yeast extract, 10 mM NaCl, 2.5 mM KCl, 10 mM MgCl2, 20 mM glucose (SOC medium) containing 440 mM sorbitol and 2.5 mM betaine. QPs3 was released from the fusion protein by proteolytic cleavage with thrombin. Isolated recombinant QPs3 shows one protein band in sodium dodecyl sulfate-polyacrylamide gel electrophoresis that corresponds to subunit V of mitochondrial succinate:ubiquinone reductase. Although purified recombinant QPs3 is dispersed in 0.01% dodecylmaltoside, it is in a highly aggregated form, with an apparent molecular mass of more than 1 million. The recombinant QPs3 binds ubiquinone, causing a spectral blue shift. Upon titration of the recombinant protein with ubiquinone, a saturation behavior is observed, suggesting that the binding is specific and that recombinant QPs3 may be in the functionally active state. Two amino acid residues, serine 33 and tyrosine 37, in the putative ubiquinone binding domain of QPs3 are involved in ubiquinone binding because the S33A- or Y37A-substituted recombinant QPs3s do not cause the spectral blue shift of ubiquinone. Although recombinant QPs3 contains little cytochrome b560 heme, the spectral characteristics of cytochrome b560 are reconstituted upon addition of hemin chloride. Reconstituted cytochrome b560 in recombinant QPs3 shows a EPR signal at g = 2.92. Histidine residues at positions 46 and 60 are responsible for heme ligation because the H46N- or H60N-substituted QPs3 fail to restore cytochrome b560 upon addition of hemin chloride.
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PMID:Identification of quinone-binding and heme-ligating residues of the smallest membrane-anchoring subunit (QPs3) of bovine heart mitochondrial succinate:ubiquinone reductase. 1008 11

Protective effect of the cellular ubiquinone (UQ) reducing system linked to cytosolic NADPH-dependent ubiquinone reductase (NADPH-UQ reductase) against hydrogen peroxide (H2O2)-induced lipid peroxidation was investigated using UQ and control hepatocytes freshly isolated from rats injected with UQ-10 and the vehicles 14 d in advance, respectively. The UQ hepatocytes had higher levels of ubiquinol (UQH2)-10 content and NADPH-UQ reductase activity than the control hepatocytes but did not differ in other antioxidant factors from the latter cells. The UQ hepatocytes exhibited higher cell viability and lower release of lactate dehydrogenase than the control hepatocytes when they were exposed to H2O2 of up to 100 mM for 1 h at 37 degrees C. Furthermore, the formation of thiobarbituric acid reactive substances (TBARS) by H2O2 was almost completely inhibited in the UQ hepatocytes. Decreases in UQH2 and alpha-tocopherol contents and NADPH-UQ reductase activity by H2O2 exposure were observed in both types of the hepatocytes, but those levels in the UQ hepatocytes after the exposure were still higher than in the control hepatocytes. The decreases in ascorbic acid, reduced glutathione and protein thiol contents and DT-diaphorase activity by H2O2 were not different between in the two types of hepatocytes. Antioxidant enzyme activities of catalase, superoxide dismutase, glutathione peroxidase, glutathione S-transferase and glutathione reductase in the hepatocytes were not inhibited by H2O2. From these results, it was concluded that the cellular UQ reducing system linked to cytosolic NADPH-UQ reductase functions mainly as an antioxidant defense for cellular membranes.
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PMID:Antioxidant roles of cellular ubiquinone and related redox cycles: potentiated resistance of rat hepatocytes having stimulated NADPH-dependent ubiquinone reductase against hydrogen peroxide toxicity. 1059 33

Male and female C57B1/6 mice were rendered vitamin A-deficient, and the effects of this deficiency on certain xenobiotic-metabolizing enzymes and defenses against oxidative stress were examined. Vitamin A deficiency significantly increased the levels of DT-diaphorase, glutathione transferase, and catalase in the hepatic cytosolic fraction from male mice (5.2-, 1.6-, and 3.5-fold, respectively), as well as from female mice (4.8-, 3.3-, and 2.4-fold, respectively). In the hepatic mitochondrial fraction (containing peroxisomes) from male animals, the activities of urate oxidase and catalase were increased 3.4- and 1.7-fold, respectively. The activity of catalase in the mitochondrial fraction from female mice was not affected by vitamin A deficiency, whereas the activity of peroxisomal urate oxidase was increased 2.9-fold. The hepatic level of ubiquinone was increased somewhat. The significance of the increases observed here is presently unclear, but it may be speculated that vitamin A and/or its metabolites are somehow involved in the down-regulation of these proteins. Another possibility is that these enzymes are increased as a result of hepatic oxidative stress caused by vitamin A deficiency. However, vitamin A deficiency had no effect on the activity of superoxide dismutase in this study, whereas the activity of glutathione peroxidase was slightly decreased (27%) in the hepatic cytosolic fraction from male mice. In addition, the hepatic level of alpha-tocopherol was decreased dramatically in the vitamin A-deficient animals.
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PMID:Effects of vitamin A deficiency on selected xenobiotic-metabolizing enzymes and defenses against oxidative stress in mouse liver. 1064 45

Polychlorinated biphenyls (PCBs) have been shown to be embryotoxic. The mechanism(s) of action is not clearly understood. The toxic effects could be either direct or indirect. Furthermore, PCB congeners vary in their toxic potential. They can be classified in coplanar PCBs binding to the transcription factor aryl hydrocarbon receptor (AhR), which induce subsequent changes in gene expression, and noncoplanar PCBs exhibiting AhR-independent effects. In order to investigate possible mechanisms, 5 and 6 days old preimplantation rabbit embryos were exposed in vitro to low levels of coplanar (PCB 77, 126, and 169) or noncoplanar PCBs (PCB 28, 52, 101, 118, 138, 153, and 180). The PCB effects were studied by semiquantitative RT-PCR analysis of AhR target genes (cytochrome P450 (CYP) 1A1, 1A2, UDP-glucuronosyl transferase 1, glutathione S-transferase pi1 and aldehyde dehydrogenase) and dioxin-responsive genes (IL 1beta, PAI 2, Cox 2, TGFalpha, EGF, erbB 1-4, c-fos, c-jun, HSP 90, cyclophilin 40), and by differential display (DD) RT-PCR. CYP 1B1 mRNA and AhR protein were localized by in situ hybridization and immunohistochemistry, respectively. From the AhR target genes studied only CYP 1B1, and cyclooxygenase 2 showed an increase in mRNA levels after coplanar and noncoplanar PCB. Interleukin 1beta and plasminogen activator inhibitor 2 were downregulated. CYP 1B1 mRNA showed a stage specific inducibility at day 6, but not at day 5. By DD RT-PCR we identified six new genes previously not reported to be regulated by PCBs. The mRNAs encoding the subunits 1, 2, 4, and 5 of the NADH ubiquinone oxidoreductase and beta-globin showed a decrease, whereas trichohyalin mRNA was increased after PCB exposure. Coplanar and noncoplanar PCB congeners elicited similar responses on the mRNA levels of the studied genes. Exposure to coplanar PCBs did not result in the AhR being translocated to the nucleus. Our results show that (i). PCBs induce changes in gene expression in rabbit day 5 and 6 preimplantation embryos and imply (ii). that the transcriptional changes observed were not mediated by the nuclear AhR.
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PMID:Polychlorinated biphenyls affect gene expression in the rabbit preimplantation embryo. 1254 57

A brief period of ischemia followed by timely reperfusion may lead to prolonged, yet reversible, contractile dysfunction (myocardial stunning). Damage to the myocardium occurs not only during ischemia, but also during reperfusion, where a massive release of oxygen-free radicals (OFR) occurs. We have previously utilized 2-DE and MS to define 57 protein spot changes during brief ischemia/reperfusion (15 min ischemia, 60 min reperfusion; 15I/60R) injury in a rabbit model (White, M. Y., Cordwell, S. J., McCarron, H. C. K., Prasan, A. M. et al., Proteomics 2005, 5, 1395-1410) and shown that the majority of these occur because of physical and/or chemical PTMs. In this study, we subjected rabbit myocardium to 15I/60R in the presence of the OFR scavenger N-(2-mercaptopropionyl) glycine (MPG). Thirty-seven of 57 protein spots altered during 15I/60R remained at control levels in the presence of MPG (15I/60R + MPG). Changes to contractile proteins, including myosin light chain 2 (MLC-2) and troponin C (TnC), were prevented by the addition of MPG. To further investigate the individual effects of ischemia and reperfusion, we generated 2-DE gels from rabbit myocardium subjected to brief ischemia alone (15I/0R), and observed alterations of 33 protein spots, including 18/20 seen in both 15I/60R-treated and 15I/60R + MPG-treated tissue. The tissue was also subjected to ischemia in the presence of MPG (15I/0R + MPG), and 21 spot changes, representing 14 protein variants, remained altered despite the presence of the OFR scavenger. These ischemia-specific proteins comprised those involved in energy metabolism (lactate dehydrogenase and ATP synthase alpha), redox regulation (NADH ubiquinone oxidoreductase 51 kDa and GST Mu), and stress response (Hsp27 and 70, and deamidated alpha B-crystallin). We conclude that contractile dysfunction associated with myocardial stunning is predominantly caused by OFR damage at the onset of reperfusion, but that OFR-independent damage also occurs during ischemia. These ischemia-specific protein modifications may be indicative of early myocardial injury.
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PMID:Proteomics of ischemia and reperfusion injuries in rabbit myocardium with and without intervention by an oxygen-free radical scavenger. 1713 70

The mitochondrial permeability transition pore (MPTP) plays a key role in cell death, yet its molecular identity remains uncertain. Although knock-out studies have confirmed critical roles for both cyclophilin-D (CyP-D) and the adenine nucleotide translocase (ANT), given a strong enough stimulus MPTP opening can occur in the absence of either. Here we provide evidence that the mitochondrial phosphate carrier (PiC) may also be a critical component of the MPTP. Phenylarsine oxide (PAO) was found to activate MPTP opening in the presence of carboxyatractyloside (CAT) that prevents ANT binding to immobilized PAO. Only four proteins from solubilized CAT-treated beef heart inner mitochondrial membranes bound to immobilized PAO, one of which was the PiC. GST-CyP-D pull-down and co-immunoprecipitation studies revealed CsA-sensitive binding of PiC to CyP-D; this increased following diamide treatment. Co-immunoprecipitation of the ANT with the PiC was also observed but was insensitive to CsA treatment. N-ethylmaleimide and ubiquinone analogues (UQ(0) and Ro 68-3400) inhibited phosphate transport into rat liver mitochondria with the same concentration dependence as their inhibition of MPTP opening. UQ(0) and Ro 68-3400 also induced the "m" conformation of the ANT, as does NEM, and reduced the binding of both the PiC and ANT to the PAO column. We propose a model for the MPTP in which a calcium-triggered conformational change of the PiC, facilitated by CyP-D, induces pore opening. An interaction of the PiC with the ANT may enable agents that bind to either transporter to modulate pore opening.
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PMID:The mitochondrial phosphate carrier interacts with cyclophilin D and may play a key role in the permeability transition. 1866 15


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