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

HA-1 hamster fibroblasts receiving fresh media every 24 h were continuously passaged in progressively increasing O2 concentrations for 18 mo (designated O2R95). These cells were significantly more resistant than parental HA-1 to clonogenic inactivation mediated by 95% O2 without media replacement. The O2R95 cell line exhibited increases in the activities of catalase (CAT), Mn superoxide dismutase (MnSOD), Cu,Zn superoxide dismutase (Cu,Zn SOD), and glutathione peroxidase (GPx). O2R95 cells demonstrated uniformly distributed increased staining for CAT, MnSOD, Cu,Zn SOD, and GPx proteins, as determined by immunohistochemistry. Cellular resistance to and metabolism of 4-hydroxy-2-nonenal (4HNE), a toxic byproduct of lipid peroxidation implicated in mechanisms of O2 toxicity, was examined in HA-1 and O2R95 cell lines. O2R95 cells were significantly more resistant to 4HNE cytotoxicity, which was accompanied by a significant increase in 4HNE metabolism. O2R95 cells also demonstrated an increase in total glutathione (GSH) and glutathione S-transferase (GST) activity, an enzymatic system believed to be involved with 4HNE metabolism. Furthermore, homogenates from O2R95 cells consumed greater quantities of 4HNE in the presence of NADPH (but not NADH, NAD+, or NADP+), suggesting that an enzyme(s) utilizing NADPH contributes to 4HNE metabolism, resistance to 95% O2 and 4HNE as well as increased total GSH, antioxidant enzyme activities, and NADPH-dependent metabolism of 4HNE, persisted in O2R95 cells for 75 days of growth in 21% O2. These findings are compatible with the hypothesis that aldehydic byproducts of lipid peroxidation contribute to mechanisms of O2 toxicity and the selective pressure exerted by exposure of cells to hyperoxia.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:A stable O2-resistant cell line: role of lipid peroxidation byproducts in O2-mediated injury. 161 58

We found that Adriamycin increased the pentose phosphate shunt activity in both Adriamycin-sensitive (WT) and Adriamycin-resistant (ADRR) human breast cancer MCF-7 cells. In contrast, hydrogen peroxide and cumene hydroperoxide markedly stimulated pentose-shunt activity in ADRR but only moderately increased the activity in WT cells. Furthermore, the altered oxidation-reduction regulation is associated with changes intrinsic to the key enzymes of the pentose-shunt pathway, glucose-6-phosphate dehydrogenase, and 6-phosphogluconate dehydrogenase and with glutathione peroxidase. We found the Vmax values for glucose-6-phosphate dehydrogenase and 6-phosphogluconate dehydrogenase were 50- and 4-fold lower, respectively, in ADRR than WT cells and the Kms of NADP+ were 10-fold lower in ADRR than WT. The activity of glutathione reductase in ADRR is 42% of that in WT. In spite of these changes, the response of the cells to both hydrogen peroxide and organic peroxide is not limited by either the capacity of the pentose shunt or glutathione reductase, but is determined by the activity of glutathione peroxidase and a glutathione transferase which possess peroxidase activity. The kinetic properties of the glucose-6-phosphate dehydrogenase in ADRR may, however, seriously limit the activity of cytochrome P-450 reductase, a major enzyme of Adriamycin conversion to a free radical.
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PMID:Adriamycin resistance in human tumor cells associated with marked alteration in the regulation of the hexose monophosphate shunt and its response to oxidant stress. 366 3

The biotransformation of allyl alcohol and acrolein by rat lung and liver preparations was investigated by measuring acrolein, acrylic acid, glycidol, and glycidaldehyde. Acrolein was detected by high-pressure liquid chromatography from incubation mixtures containing allyl alcohol, NAD+, and liver 9000g supernatant fraction or cytosol. Acrolein was not formed when lung fractions were treated similarly. Addition of pyrazole in the incubation mixture inhibited the reaction. The metabolism of acrolein to acrylic acid by liver 9000g supernatant fraction, cytosol, and microsomes has been demonstrated; acrylic acid formation was greater with NAD+ than with NADP+ in all three fractions. Acrylic acid was also formed from allyl alcohol. Disulfiram inhibited the NAD+- and NADP+-dependent reactions. Acrylic acid was not formed when lung preparations were used. Lung and liver microsomal epoxidation products of allyl alcohol and acrolein have been identified. Conversion of glycidol to glycerol and glycidaldehyde to glyceraldehyde by liver epoxide hydrase has been demonstrated. Epoxides, glycidol, and glycidaldehyde were also found to be substrates for lung and liver cytosolic glutathione S-transferase.
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PMID:The biotransformation of allyl alcohol and acrolein in rat liver and lung preparations. 610 26

GDP-D-mannose dehydratase (GMD) catalyzes the first step of the pathway that converts GDP-D-mannose to GDP-L-fucose in bacteria, plants and mammals. Recently, the gene coding for GMD has been identified and sequenced in E. coli. Based on this sequence, we have expressed and purified GMD in E. coli as a glutathione transferase (GST) fusion protein. The fused GST-GMD protein and the thrombin-cleaved GMD were then characterized. The catalytically active form of both enzyme species seems to be a hexamer of 410 and 250 kDa, respectively. The GST-GMD fusion protein has a Km of 0.22 +/- 0.04 mM and a specific activity of 2.3 +/- 0.2 micromol/h/mg. Ca2+ and Mg2+ activate GMD, while GDP-L-beta-fucose, the end-product of the pathway, inhibits it specifically. The GST-GMD fusion protein contains one mole of tightly bound NADP+ per mole of hexamer. Apparently, this NADP+ is involved in the catalytic mechanism of GMD.
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PMID:Expression, purification and characterization of GDP-D-mannose 4,6-dehydratase from Escherichia coli. 925 4

The GDP-4-keto-6-deoxy-D-mannose epimerase/reductase (GM_ER) isolated from E. coli has been overexpressed as a GST-fusion protein and purified to homogeneity. The enzyme, an NADP+(H)-binding homodimer of 70 kDa, is responsible for the production of GDP-L-fucose. GM_ER shows significant structural homology to the human erythrocyte protein FX, which is involved in blood-group glycoconjugate biosynthesis, displaying 3,5 epimerase/reductase activity on GDP-4-keto-6-deoxy-D-mannose. GM_ER has been crystallized in a trigonal crystalline form, containing one molecule per asymmetric unit, suitable for high-resolution crystallographic investigations.
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PMID:Preliminary crystallographic investigations of recombinant GDP-4-keto-6-deoxy-D-mannose epimerase/reductase from E. coli. 976 75

One of the major soluble antigenic proteins of Entamoeba histolytica was purified to homogeneity and identified on a molecular basis. Its recombinant protein was expressed in Escherichia coli as a fusion protein with Shistosoma japonicum glutathione S-transferase. Apparent molecular weight of the purified antigenic protein was estimated to be 40-kDa and molecular-based analysis indicated that the purified protein was NADP+-dependent alcohol dehydrogenase (EhADH1). The application of the purified protein for the serodiagnosis of amebiasis was evaluated using an enzyme-linked immunosorbent assay applied to sera obtained from patients with amebiasis and healthy human controls. The purified protein was well recognized by the sera from asymptomatic amebiasis humans (22/22, 100%), whereas, it was less recognized by the sera from symptomatic amebiasis patients (5/16, 31%) with amebic colitis or liver abscess. To confirm the antigenicity of EhADH1, the recombinant glutathione S-transferase-EhADH1 fusion protein was also evaluated by the enzyme-linked immunosorbent assay using the same sera. The recombinant protein was also recognized by the sera from asymptomatic amebiasis humans (14/22, 64%) and less recognized by the sera from symptomatic amebiasis patients (2/16, 13%). These results suggest that the purified protein is applicable antigen for serodiagnostic screening of asymptomatic amebiasis humans.
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PMID:Purification and identification of major soluble 40-kDa antigenic protein from Entamoeba histolytica: its application for serodiagnosis of asymptomatic amebiasis. 1143 29

We have cloned cDNA for leukotriene B4 12-hydroxydehydrogenase (LTB4 12-HD)/15-ketoprostaglandin 13-reductase (PGR) from guinea-pig liver. LTB4 12-HD catalyzes the conversion of LTB4 into 12-keto-LTB4 in the presence of NADP+, and plays an important role in inactivating LTB4. The cDNA contained an ORF of 987 bp that encodes a protein of 329 amino-acid residues with a 78% identity with porcine LTB4 12-HD. The amino acids in the putative NAD+/NADP+ binding domain are well conserved among the pig, guinea-pig, human, rat, and rabbit enzymes. The guinea-pig LTB4 12-HD (gpLTB4 12-HD) was expressed as a glutathione S-transferase (GST) fusion protein in Escherichia coli, which exhibited similar enzyme activities to porcine LTB4 12-HD. We examined the 15-ketoprostaglandin 13-reductase (PGR) activity of recombinant gpLTB4 12-HD, and confirmed that the Kcat of the PGR activity is higher than that of LTB4 12-HD activity by 200-fold. Northern and Western blot analyses revealed that gpLTB4 12-HD/PGR is widely expressed in guinea-pig tissues such as liver, kidney, small intestine, spleen, and stomach. We carried out immunohistochemical analyses of this enzyme in various guinea-pig tissues. Epithelial cells of calyx and collecting tubules in kidney, epithelial cells of airway, alveoli, epithelial cells in small intestine and stomach, and hepatocytes were found to express the enzyme. These findings will lead to the identification of the unrevealed roles of PGs and LTs in these tissues.
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PMID:Immunohistochemical localization of guinea-pig leukotriene B4 12-hydroxydehydrogenase/15-ketoprostaglandin 13-reductase. 1173 4

NAD+-dependent 15-hydroxyprostaglandin dehydrogenase (15-PGDH), a member of the short chain dehydrogenase/reductase (SDR) family, is responsible for the biological inactivation of prostaglandins. Sequence alignment within SDR coupled with molecular modeling analysis has suggested that Gln-15, Asp-36, and Trp-37 of 15-PGDH may determine the coenzyme specificity of this enzyme. Site-directed mutagenesis was used to examine the important roles of these residues. Several single mutants (Q15K, Q15R, W37K, and W37R), double mutants (Q15K-W37K, Q15K-W37R, Q15R-W37K, and Q15R-W37R), and triple mutants (Q15K-D36A-W37R and Q15K-D36S-W37R) were prepared and expressed as glutathione S-transferase (GST) fusion proteins in Escherichia coli and purified by GSH-agarose affinity chromatography. Mutants Q15K, Q15R, W37K, W37R, Q15K-W37K, and Q15R-W37K were found to be inactive or almost inactive with NADP+ but still retained substantial activity with NAD+. Mutant Q15K-W37R and mutant Q15R-W37R showed comparable activity for NAD+ and NADP+ with an increase in activity nearly 3-fold over that of the wild type. However, approximately 30-fold higher in K(m) for NADP+ than that of the wild type enzyme for NAD+ was found for mutants Q15K-W37R and Q15R-W37R. Similarly, the K(m) values for PGE(2) of mutants were also shown to increase over that of the wild type. Further mutation of Asp-36 to either an alanine or a serine of the double mutant Q15K-W37R (i.e., triple mutants Q15K-D36A-W37R and Q15K-D36S-W37R) rendered the mutants exhibiting exclusive activity with NADP+ but not with NAD+. The triple mutants showed a decrease in K(m) for NADP+ but an increase in K(m) for PGE(2). Further mutation at Ala-14 to a serine of a triple mutant (Q15K-D36S-W37R) decreased the K(m) values for both NADP+ and PGE(2) to levels comparable to those of the wild type. These results indicate that the coenzyme specificity of 15-PGDH can be altered from NAD+ to NADP+ by changing a few critical residues near the N-terminal end.
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PMID:Critical residues for the coenzyme specificity of NAD+-dependent 15-hydroxyprostaglandin dehydrogenase. 1459 57

Ribitol dehydrogenase (RDH) catalyzes the conversion of ribitol to D-ribulose. A novel RDH gene was cloned from Zymomonas mobilis subsp. mobilis ZM4 and overexpressed in Escherichia coli BL21(DE3). DNA sequence analysis revealed an open reading frame of 795 bp, capable of encoding a polypeptide of 266 amino acid residues with a calculated molecular mass of 28,426 Da. The gene was overexpressed in E. coli BL21(DE3) and the protein was purified as an active soluble form using glutathione S-transferase affinity chromatography. The molecular mass of the purified enzyme was estimated to be approximately 28 kDa by sodium dodecyl sulfate-polyacrylamide gel and approximately 58 KDa with gel filtration chromatography, suggesting that the enzyme is a homodimer. The enzyme had an optimal pH and temperature of 9.5 and 65 degrees C, respectively. Unlike previously characterized RDHs, Z. mobilis RDH (ZmRDH) showed an unusual dual coenzyme specificity, with a k(cat) of 4.83 s(-1) for NADH (k(cat)/K(m) = 27.3 s(-1) mM(-1)) and k(cat) of 2.79 s(-1) for NADPH (k(cat)/K(m) = 10.8 s(-1) mM(-1)). Homology modeling and docking studies of NAD+ and NADP+ into the active site of ZmRDH shed light on the dual coenzyme specificity of ZmRDH.
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PMID:Cloning and characterization of a ribitol dehydrogenase from Zymomonas mobilis. 2012 34

The physiological and biochemical mechanisms on boron (B)-induced alleviation of aluminum (B)-toxicity in plants have been examined in some details, but our understanding of the molecular mechanisms underlying these processes is very limited. In this study, we first used the cDNA-AFLP to investigate the gene expression patterns in Citrus grandis roots responsive to B and Al interactions, and isolated 100 differentially expressed genes. Results showed that genes related to detoxification of reactive oxygen species (ROS) and aldehydes (i.e., glutathione S-transferase zeta class-like isoform X1, thioredoxin M-type 4, and 2-alkenal reductase (NADP+-dependent)-like), metabolism (i.e., carboxylesterases and lecithin-cholesterol acyltransferase-like 4-like, nicotianamine aminotransferase A-like isoform X3, thiosulfate sulfurtransferase 18-like isoform X1, and FNR, root isozyme 2), cell transport (i.e., non-specific lipid-transfer protein-like protein At2g13820-like and major facilitator superfamily protein), Ca signal and hormone (i.e., calcium-binding protein CML19-like and IAA-amino acid hydrolase ILR1-like 4-like), gene regulation (i.e., Gag-pol polyprotein) and cell wall modification (i.e., glycosyl hydrolase family 10 protein) might play a role in B-induced alleviation of Al-toxicity. Our results are useful not only for our understanding of molecular processes associated with B-induced alleviation of Al-toxicity, but also for obtaining key molecular genes to enhance Al-tolerance of plants in the future.
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PMID:Mechanisms on boron-induced alleviation of aluminum-toxicity in Citrus grandis seedlings at a transcriptional level revealed by cDNA-AFLP analysis. 2574 50


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