Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0272170 (SDS)
 50,377 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

An elastomeric polypeptide was produced, with the sequence G-(VPGVG)19-VPGV, as a fusion to glutathione S-transferase using the vector pGEX-3X. The fusion protein was expressed to high levels in Escherichia coli as indicated by SDS-PAGE analysis of induced cells. The fusion protein was affinity purified and cleaved with protease factor Xa, and the elastomeric polypeptide was recovered to a high degree of purity as indicated by SDS-PAGE followed by staining with CuCl2. The physical characterizations of carbon-13 and proton nuclear magnetic resonance and of the temperature profile for turbidity formation for the inverse temperature transition of hydrophobic folding and assembly attest to the successful microbial synthesis of the polypentapeptide of elastin. The results of these studies provide the initial progress toward achieving a more economical and practical means of producing material for elastic protein-based polymer research and applications.
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PMID:Production and purification of a recombinant elastomeric polypeptide, G-(VPGVG)19-VPGV, from Escherichia coli. 136 56

Leukotriene (LT) C4 synthase, the enzyme that catalyzes the conjugation of LTA4 with reduced glutathione to form LTC4, was purified to homogeneity from the KG-1 myeloid cell line after solubilization of the microsomes utilizing a combination of 0.4% sodium deoxycholate and 0.4% Triton X-102. The solubilized enzyme was then applied to an S-hexyl-glutathione-agarose column that was eluted by the use of 7.5 mM probenecid. After removal of the probenecid by sequential concentration and dilution in an Amicon concentrator, the enzyme was additionally purified and concentrated by binding to and elution from approximately 75 mg of S-hexyl-glutathione-agarose. The enzyme was further resolved by electrophoresis with a nondenaturing Tris-glycine gel, and the LTC4 synthase activity was localized to slices 3 and 4. When the remainder of the eluate from the nondenaturing gel was precipitated by acetone and analyzed by 14% SDS/PAGE with silver staining, a single protein band of 18 kDa was associated with LTC4 synthase activity and was not present in the eluates of slices lacking activity. The overall recovery was 12.5%. In a separate preliminary purification, in which the yield was only approximately 1%, the eluates of the nondenaturing gel had also revealed a single protein of 18 kDa by SDS/PAGE, which was present only in the eluates with LTC4 synthase activity. These data identify LTC4 synthase as a protein of 18 kDa, a size consistent with its membership in the microsomal glutathione S-transferase family.
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PMID:Purification of human leukotriene C4 synthase. 145 53

Fasciola hepatica glutathione S-transferase (FhGST) was isolated from adult worms by glutathione agarose affinity chromatography. SDS-PAGE shows three proteins of M(r) ranging from 29-27.8 kDa. Western immunoblot analyses using SDS-PAGE separated adult worm extracts and probed with a rabbit anti-FhGST antiserum reveal two bands in the same M(r) range. Mice and rabbits immunized with purified FhGST develop copious amounts of anti-FhGST antibodies. Moreover, antisera to F. hepatica adult worms and excretion-secretion products also react with FhGST. Cross-reactivity with schistosomes is evidenced in the reactivity with FhGST of anti-Schistosoma mansoni adult worm antisera and, to a lesser extent, antisera to S. mansoni-soluble egg antigens. The time of appearance of anti-FhGST antibodies in different species of animals infected with F. hepatica was determined. Sheep and a New Zealand white rabbit developed anti-FhGST antibodies detectable by ELISA as early as 2 weeks postexposure with F. hepatica. However, neither mice nor calves infected with F. hepatica developed antibodies to FhGST through the 5-10 weeks of infection tested. But mice infected with S. mansoni developed anti-FhGST cross-reacting antibodies by 6 weeks of infection. Calves immunized with a Fasciola/Schistosoma cross-reactive, cross-protective antigen complex in which a 12,000-kDa protein (Fh12) has been shown to contain immunoprophylactic activity, also developed antibodies to FhGST. Since FhGST is a novel potential vaccine, its protection-inducing capability in a multivalent vaccine combined with Fh12 clearly warrants study. In summary, it appears that hosts with fascioliasis are either responders to FhGST (rabbits, sheep) or nonresponders (mice, cattle), offering interesting models for studying the immune response.
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PMID:Fasciola hepatica: host responders and nonresponders to parasite glutathione S-transferase. 151 66

A novel class alpha glutathione S-transferase (GST) isozyme is expressed in the hepatic cytosol of rabbits treated with 4-picoline. SDS-PAGE analysis revealed the presence of a new 28-kDa band which cross-reacted with class alpha GST-specific IgG. This new GST isozyme was isolated from the hepatic cytosol of 4-picoline-treated rabbits and purified to homogeneity using S-hexylglutathione-agarose, CM-Sepharose, and PBE118 chromatofocusing chromatography. The isozyme was determined by SDS-PAGE and gel filtration analyses to be a homodimer of approximately 28 kDa with blocked N-terminus. A heterodimer consisting of 25 and 28 kDa subunits with activity toward the substrate 1-chloro-2,4-dinitrobenzene was also purified. Immunoblot analysis revealed that the 25, 26.5, and 28 kDa bands cross-reacted with class alpha GST-specific IgG and failed to react with either class mu or class pi GST-specific antibodies. The 28 kDa enzyme had a pI of 8.2 as determined by nonequilibrium pH gel electrophoresis. The purified 28 kDa enzyme exhibited activity toward 1-chloro-2,4-dinitrobenzene (Km = 1.60 mM and Vmax = 73.5 mumol/min/mg) and cumene hydroperoxide (Km = 1.02 mM and Vmax = 6.92 mumol/min/mg). Amino acid sequence analysis of several fragments resulting from cyanogen bromide cleavage of the 28 kDa GST isozyme revealed a class alpha GST consensus sequence. In addition, proteolytic digestion with alpha-chymotrypsin yielded peptide maps which showed distinct differences between the purified 28 kDa GST and another purified class alpha GST isozyme present in rabbit liver. These results provide evidence that class alpha GST isozymes containing a novel 28 kDa subunit are expressed following treatment with 4-picoline.
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PMID:Enhanced expression, purification, and characterization of a novel class alpha glutathione S-transferase isozyme appearing in rabbit hepatic cytosol following treatment with 4-picoline. 153 65

Glutathione S-transferase (GST) expression was examined in hepatic cytosol from rats and rabbits treated with 4-picoline, pyrrole, pyridine, pyrazine, imidazole, or piperidine using enzymatic activity, SDS-PAGE, and immunoblot analyses and the results were compared to those obtained with phenobarbital and 3-methylcholanthrene. SDS-PAGE and immunoblot analyses of hepatic cytosol prepared from rats treated with pyrazine revealed the induction of class alpha (Ya and Yc) and mu (Yb) bands with a corresponding 2.4-fold increase in metabolic activity using 1-chloro-2,4-dinitrobenzene as substrate. A new class alpha band migrating in the region of the Yc band was observed in the SDS-PAGE and detected in the immunoblot of cytosol from pyrrole-treated rats, whereas treatment with 4-picoline, imidazole, or piperidine failed to alter the expression of the major classes of GST isozymes in this species. SDS-PAGE and immunoblot analyses of rabbit hepatic cytosol revealed a unique species-dependent difference in the expression of GSTs. While phenobarbital and 3-methylcholanthrene induce class alpha and mu GST expression in rat hepatic cytosol, one of the most interesting observations was that neither of these agents stimulated GST expression in the rabbit. Immunoblot analysis of cytosol isolated from 4-picoline-treated rabbits using GST class alpha-specific IgG showed the appearance of a novel class alpha 28-kDa GST band and the concomitant disappearance of a class alpha 29-kDa GST band. In addition, SDS-PAGE and immunoblot analyses showed that treatment of rabbits with pyrrole, pyrazine, imidazole, or piperidine resulted in the disappearance of this class alpha 29-kDa GST band with no detectable expression of the class alpha 28-kDa GST band; the level of the class alpha 29-kDa band was unaffected by pyridine treatment. In contrast, immunoblot analyses of hepatic cytosol revealed that a 25.5-kDa class mu GST band disappeared following treatment with pyridine, but was unaffected by treatment with other nitrogen heterocycles. The Vmax of glutathione conjugation to the substrate 1-chloro-2,4-dinitrobenzene decreased by 52, 36, 59, 41, 37, and 32% in hepatic cytosol isolated from 4-picoline-, pyrrole-, pyridine-, pyrazine-, imidazole-, and piperidine-treated rabbits, respectively. The results suggest that nitrogen heterocycles differ in their ability to modulate glutathione S-transferase isozyme expression in rat and rabbit hepatic tissue and that rabbit hepatic GSTs are refractory to induction by agents such as pyrazine, phenobarbital, or 3-methylcholanthrene and hence these xenobiotics do not appear to be bifunctional inducers in this species.
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PMID:Differences between rats and rabbits in hepatic cytosolic glutathione S-transferase expression in response to nitrogen heterocycle and other inducers. 155 56

Bovine adrenal cortex tissue expresses high levels of glutathione S-transferase (GST) from each of the alpha, mu and pi gene families. We describe the purification and characterization of an abundant alpha-class GST from this tissue that has not been identified previously because of its failure to bind to S-hexylglutathione-Sepharose 6B (S-hexG-Ag). This enzyme has been affinity purified on glutathione-Sepharose 6B (GSH-Ag) and was obtained in a highly purified form by employing S-hexG-Ag to remove the bulk of GST before chromatography on GSH-Ag. The purified GST eluted from GSH-Ag was found to exhibit marked peroxidase and delta 5-ketosteroid isomerase activities (19.2 and 1.67 U/mg respectively). The bovine enzyme also showed high GST activity towards 4-hydroxynonenal (5.09 U/mg). Sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis revealed that the bovine GST contains two distinct polypeptides, one with an Mr of 25,900 and the other with an Mr of 26,500. An abundant alpha-class GST was also purified from human adrenal cortex that possessed properties which were similar to the bovine alpha-class GST described above; however, unlike the bovine enzyme, the corresponding human alpha-class GST bound to S-hexG-Ag. As with the bovine enzyme, the purified human GST displayed marked peroxidase and isomerase activities (27 and 4.02 U/mg respectively). Further analysis on SDS-PAGE (Mr 25,800) and reverse-phase high-performance liquid chromatography established that this abundant alpha-class GST in human adrenal cortex is equivalent to the human liver GST B1B1 enzyme. As both human and bovine adrenal cortex contain high levels of alpha-class GST with similar catalytic properties, we discuss the possible functions of these enzymes in this tissue.
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PMID:Expression of an abundant alpha-class glutathione S-transferase in bovine and human adrenal cortex tissues. 173 63

1. The hepatic glutathione S-transferase (GST) isoenzymes were isolated and characterized from salmon, sea trout and rainbow trout. 2. In all three species the predominant GST expressed comprised subunits of Mr 24,800. These subunits each co-migrated with the rat pi-class Yf polypeptide during SDS/polyacrylamide gel electrophoresis. 3. Western blotting experiments demonstrated immunochemical cross-reactivity between the major salmonid and the rat pi-class GSTs. 4. The salmon GST of subunit Mr 24,800 was digested with cyanogen bromide and the peptides, once purified by reverse-phase HPLC, were subjected to automated amino acid sequencing. 5. Over the region sequenced, the salmon GST possessed about 65% homology with the rat and human pi-class GST.
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PMID:The major glutathione S-transferase in salmonid fish livers is homologous to the mammalian pi-class GST. 175 23

An acidic form of glutathione S-transferase (GST) was purified from human fetal livers by means of affinity chromatography and chromatofocusing. The major peak of the acidic form of GST was focused between pH 4.8 and 4.9. Judging by SDS-PAGE, the purified acidic GST was apparently homogeneous; the subunit molecular weight was estimated to be 23,000. The acidic GST catalyzed the conjugations of glutathione (GSH) with 1-chloro-2,4-dinitrobenzene (CDNB) and ethacrynic acid (EA). The immunochemical properties of the purified acidic GST were indistinguishable from those of human placental GST-pi. The N-terminal amino acid sequence of the acidic GST was identical with that of GST-pi from human placenta. The level of expression of the acidic form of GST was clearly different between human adult and fetal livers as examined on the levels of mRNA and protein.
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PMID:Purification and characterization of acidic form of glutathione S-transferase in human fetal livers: high similarity to placental form. 178 5

The basal level of benzo(a)pyrene monooxygenase, epoxide hydrolase and glutathione S-transferase activity as well as the content of cytochrome P-450 were found the same in both compared benzo(a)pyrene (BP) sensitive D. simulans strain 364yv and BP-resistant wild one (Turku). Phenobarbital pretreatment resulted in the same increase level of these enzyme activities in both strains. BP-pretreatment of 364yv flies decreased the amount of the cytochrome P-450 but raised up the turnover of BP per molecule of cytochrome P-450. SDS-polyacrylamide gel electrophoresis of the microsomal proteins from BP-pretreated 364yv flies (but not from Turku) showed an increased hemoprotein content in the 56000 band. The relationship between BP-sensitivity of the strain 364yv and BP-induced aberrant isoform of the cytochrome P-450 has been discussed.
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PMID:[Benzo(a)pyrene pretreatment of Drosophila simulans mutant strain results in the induction of aberrant isoform of cytochrome P-450 with increased capacity to metabolize benzo(a)pyrene]. 178 90

The activity of glutathione S-transferase (GST) decreased progressively in Schistosoma mansoni from mice treated with oltipraz (OPZ). However, the peroxidase activity of GST (selenium-independent) and selenium-dependent glutathione peroxidase was not affected by OPZ treatment. Purification and quantification of GST from worms after OPZ treatment indicated that the decrease in enzyme activity was greater than could be accounted for by the decrease in GST protein content. SDS-polyacrylamide gel electrophoresis followed by Western blot analysis with GST isoenzyme specific antisera revealed a slight decrease in the quantity of both 26 and 28 kDa GSTs. Fractionation of cytosolic GSTs from male S. mansoni by chromatofocusing resolved three major isoenzymes (SmI, II and III) and a minor form which eluted first from the column. SmI, II and III all had a molecular weight of about 28 kDa on SDS-polyacrylamide gel electrophoresis. However, on electrophoresis in the absence of SDS, the three GST forms exhibited different mobilities. The pattern of SmI, II and III was similar in untreated and OPZ-treated worms, but the activities of the isoenzymes from treated worms were lower. The results suggest that OPZ interacts with the GST isoenzymes SmI, II and III in a similar manner; thus, the effects are not isoenzyme specific. Taken together, these results suggest that OPZ and/or its metabolites interact directly with GST resulting in inhibition of activity and reduction in total enzyme protein. This mechanism may be important in the antischistosomal action of OPZ.
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PMID:Oltipraz-induced decrease in the activity of cytosolic glutathione S-transferase in Schistosoma mansoni. 178 33


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