Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

---

Query: EC:2.5.1.18 (glutathione S-transferase)
22,582 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

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.
...
PMID:Purification of human leukotriene C4 synthase. 145 53

Glutathione transferase (GST) was purified from the microsomes of rat liver by glutathione affinity chromatography. The interaction of 2,4-dichlorophenoxyacetic acid (2,4-D) and 1,4-benzoquinone with microsomal GST was investigated and compared with cytosolic GST. The kinetic inhibition pattern of 1,4-benzoquinone towards microsomal GST was found to be different from that towards cytosolic GST. Microsomal GST purified by affinity chromatography was inhibited by 2,4-D in a non dose-dependent manner, while the crude microsomal GST was inhibited in a dose-dependent manner. This difference was shown to be induced by a reaction on the affinity column, and not by Triton X-100 (also shown to be a GST inhibitor), glutathione, or the elution buffer 0.2% Triton X-100 and 5 mM glutathione in 50 mM Tris-HCl, pH 9.6. The binding of microsomal GST to the affinity matrix caused a partial inactivation of the active site for 2,4-D interaction. The results show that the properties of soluble GST enzymes may not be extrapolated to the microsomal ones.
...
PMID:Interaction of 1,4-benzoquinone and 2,4-dichlorophenoxyacetic acid with microsomal glutathione transferase from rat liver. 246 44

Radiation inactivation analysis was used to determine the target size of rat liver microsomal glutathione S-transferase both in situ and following purification. When Tris-HCl-washed microsomes were irradiated, there was a 1.5-2.0-fold increase in enzymatic activity over the first 3-6 megarads followed by a decrease in enzymatic activity. Above 48 megarads the radiation inactivation curve of the Tris-HCl-washed microsomes was described by a monoexponential function which gave a target size of 48 kDa. The enzymatic activity of the microsomal enzyme was selectively increased by treating the Tris-HCl-washed microsomes either with N-ethylmaleimide or washing the microsomes with small unilamellar vesicles made from phosphatidylcholine. The inactivation curves obtained with both types of treated microsomes were simple monoexponential decays in enzymatic activity with target sizes of 46 kDa (N-ethylmaleimide) and 44 kDa (unilamellar vesicles). The microsomal enzyme was detergent solubilized and purified. The Mr value of the purified protein was 15,500 (sodium dodecyl sulfate-polyacrylamide gel electrophoresis). These data suggest that the functional unit of the microsomal form of glutathione S-transferase in situ is a trimer. The target size of the purified enzyme solubilized in Triton X-100 was 85 kDa, and no increase in activity was observed at the lower radiation doses. The increase in the target size of the purified enzyme could not be ascribed solely to the presence of the detergent. This result suggests that the microsomal form of this enzyme can exist as catalytically active oligomers of different sizes depending on its environment.
...
PMID:Radiation inactivation of microsomal glutathione S-transferase. 378 49

Five cytosolic glutathione transferases were isolated from the liver of the male little skate, Raja erinacea, a marine elasmobranch. They were designated E-1 through E-5 in order of their elution from a DEAE-cellulose column with a 0 to 100 mM KCl gradient in 0.01 M Tris (pH 8.0). Each eluted peak of glutathione transferase activity, after concentration, was applied to an affinity column prepared by reaction of epoxy-activated Sepharose 6B with glutathione (GSH). Elution of the various glutathione transferases from this column with GSH resulted in the further purification of each enzyme; the major glutathione transferase, E-4 and E-1, were purified to apparent homogeneity by this procedure. Skate glutathione transferase E-4 is dimeric and the subunits are either very similar or identical in molecular weight (about 26 000 daltons). Enzymes E-2 through E-5 were acidic proteins (pI less than 7.0) and had high specific glutathione transferase activity (0.3--12 mumol/min/mg protein) with benzo[a]pyrene 4,5-oxide (BPO) as substrate, whereas the other enzyme (E-1) had low activity (0.01 mumol/min/mg) with BPO and a basic pI (greater than 9.5). Bilirubin and hematin, non-substrate ligands, bound tightly to homogeneous E-4, with dissociation constants in the micromolar range.
...
PMID:The hepatic glutathione transferases of the male little skate, Raja erinacea. 653 76

We have expressed human glutathione S-transferases GSTA1-1 and GSTP1-1 in Salmonella typhimurium TA100 in order to assess the ability of these enzymes to modulate the mutagenicity of 1,2-dibromo-3-chloropropane (DBCP) and tris(2,3-dibromopropyl)phosphate (Tris-BP). Both compounds were mutagenic when activated by Aroclor-induced rat liver microsomes. However, when Aroclor-induced rat liver microsomes were used together with the GST-expressing strains the mutagenicity of both DBCP and Tris-BP was markedly potentiated. Neither of the GST-expressing strains potentiated the mutagenicity in the absence of microsomes, indicating that cytochrome P450-mediated metabolism was a prerequisite for GST-mediated potentiation. With DBCP both isozymes had comparable effects on mutagenic frequency, although the highest dose of DBCP was toxic in strains expressing GSTP1-1. In the case of Tris-BP, GSTP1-1 was much more active in potentiating the mutagenicity. These results indicate that human GSTs can play an important role in the activation of compounds such as DBCP and Tris-BP to mutagenic metabolites.
...
PMID:Increased mutagenicity of 1,2-dibromo-3-chloropropane and tris(2,3-dibromopropyl)phosphate in Salmonella TA100 expressing human glutathione S-transferases. 824 59

Thiopropyl Sepharose 6B in the 2-thiopyridyl-activated form was used for the reversible immobilisation of reduced glutathione (GSH). The resulting affinity matrix was successfully tested as a sorbent for the partial purification of glutathione S-transferase (GST) from pig kidney. The specific elution of the enzyme was performed with 10 mM GSH in Tris-HCl buffer (pH 7.8), non-specific elution with 20 mM dithiotreitol (DTT) in the same buffer.
...
PMID:Use of thiopropyl sepharose for the synthesis of an adsorbent for the affinity chromatography of glutathione S-transferase. 906 61

Until quite recently, high-level expression of full-length cellular prion protein (Prp(c)) in bacterial cells was not possible. We describe here the effective purification of mature Syrian golden hamster PrPc (residues 23-231) as a C-terminal fusion to glutathione S-transferase (GST) from inclusion bodies expressed in Escherichia coli. Purification of the denatured fusion protein was simplified greatly by the introduction of a C-terminal histidine anchor, leading to 255 mg pure GST-PrPc-His6/l bacterial broth, which could be refolded easily by dilution in 20 mM Tris, 5 mM dithiothreitol, 1 mM EDTA, pH 9.0. Refolding was monitored by following GST activity. Mature Syrian hamster PrPc (residues 23-231) was released from the refolded fusion protein by thrombin digestion, yielding 73 mg homogeneous protein/l bacterial culture after purification. The recombinant protein was identified by monoclonal antibodies, Edman sequencing and matrix-assisted laser-desorption/ionization MS. Correct folding was confirmed by near-ultraviolet circular dichroism spectroscopy. Samples resulting from different purification steps were sensitive to proteinase K digestion and showed no signs of infectivity in animal experiments, demonstrating that the PrPc produced is identical with the cellular isoform. The presented purification procedure should prove useful for the production of other GST-fusion proteins.
...
PMID:Large-scale production, purification and refolding of the full-length cellular prion protein from Syrian golden hamster in Escherichia coli using the glutathione S-transferase-fusion system. 949 19

The flame retardant tris(2,3-dibromopropyl)phosphate (Tris-BP) and its metabolite 2-bromoacrolein (2BA) are very potent bacterial mutagens in Salmonella typhimurium (S. typhimurium) TA 100. In this study, we showed that 2BA and Tris-BP are also mutagenic in S. typhimurium TA 104, which detects mutations at AT base pairs, while TA 100 detects mutations at CG basepairs. We also studied the mutagenicity of 2BA in mammalian cells in vitro and in the rat in vivo. Firstly, 2BA was tested in the human lymphoblastoid cell line TK6. The results showed that there was no increase in mutation frequency at the hprt locus, whereas there was a large decrease in cell survival. Secondly, a shuttle vector system was used to study the induction of mutations by 2BA:DNA adducts. The vector was modified by insertion of a single-stranded oligonucleotide containing on average one 2BA:DNA adduct. No increase in mutation frequency above background was detected after replication of this vector in SV40 transformed normal human fibroblasts. Because the liver is a major site for bioactivation of Tris-BP to 2BA in vivo, we tested the initiating capacity of Tris-BP in the rat liver in a modified Solt & Farber initiation and promotion system. Administration of Tris-BP resulted in a small increase in the number of preneoplastic gamma-glutamyl-transpeptidase positive (GGT+) foci in the liver compared to control animals (only significant in the lowest size class). Modification of the experimental protocol by performing partial hepatectomy 24 h after the administration of Tris-BP, did not increase the number of GGT+ or glutathione S-transferase-P (GST-P+) positive foci above the control level. Taken together, these results indicate that, in spite of a high mutagenicity in S. typhimurium, 2BA and Tris-BP have low or negligible mutagenic effects in mammalian systems. The lack of mutagenic activity may explain why Tris-BP is not a carcinogen in the rat liver.
...
PMID:Lack of mammalian mutagenicity of the potent bacterial mutagen tris(2,3-dibromopropyl) phosphate and its metabolite 2-bromoacrolein. 971 6

We have used three kinds of stresses, including the signaling compound jasmonic acid, an environmental stressor, UV irradiation, and a heavy metal salt copper chloride, to study changes in the protein patterns in rice (Oryza sativa L.) leaf tissues using two-dimensional polyacrylamide gel electrophoresis. However, instead of using lysis buffer containing urea (O'Farrell, J. Biol. Chem. 1975, 250, 4007-4021) for extraction of proteins from rice seedling tissues, we used Tris-HCl buffer (commonly used for extraction of proteins for separation by sodium dodecyl sulfate-polyacrylamide gel electrophoresis) for extraction of proteins and resolved these extracted proteins by the usual method of O'Farrell. Furthermore, the induction of a large number of proteins was clearly observed over controls. No spots corresponding to these induced proteins were found in the control experiment, indicating qualitative changes in protein patterns after various stress treatments. A total of 12 out of 13 proteins could be N-terminally sequenced from jasmonic acid-treated rice leaf tissues, and one protein was sequenced from UV-irradiated leaf tissues. These proteins showed high homology to pathogenesis-related (thaumatin-like protein, a PR5 class protein; a beta-1,3-glucanase precursor; an intracellular PR protein encoded by PBZ1 gene, and an antifungal protein) and cellular protectant (glutathione transferase, EC 2.5.1.18; and ascorbate peroxidase) proteins, from plants, including rice. Results presented here suggest a role for jasmonic acid in the self-defense mechanisms of rice plants.
...
PMID:Separation of proteins from stressed rice (Oryza sativa L.) leaf tissues by two-dimensional polyacrylamide gel electrophoresis: induction of pathogenesis-related and cellular protectant proteins by jasmonic acid, UV irradiation and copper chloride. 1060 17

S-Adenosyl-l-methionine-dependent protein arginine N-methyltransferases (PRMTs) catalyze the methylation of arginine residues within a variety of proteins. At least four distinct mammalian family members have now been described, including PRMT1, PRMT3, CARM1/PRMT4, and JBP1/PRMT5. To more fully define the physiological role of PRMT3, we characterized its unique putative zinc-finger domain and how it can affect its enzymatic activity. Here we show that PRMT3 does contain a single zinc-finger domain in its amino terminus. Although the zinc-liganded form of this domain is not required for methylation of an artificial substrate such as the glutathione S-transferase-fibrillarin amino-terminal fusion protein (GST-GAR), it is required for the enzyme to recognize RNA-associated substrates in RAT1 cell extracts. The recombinant form of PRMT3 is inhibited by high concentrations of ZnCl(2) as well as N-ethylmaleimide, reagents that can modify cysteine sulfhydryl groups. We found that we could distinguish PRMT family members by their sensitivity to these reagents; JBP1/PRMT5 and Hsl7 methyltransferases were inhibited in a similar manner as PRMT3, whereas Rmt1, PRMT1, and CARM1/PRMT4 were not affected. We were also able to define differences in these enzymes by their sensitivity to inhibition by Tris and free arginine. Finally, we found that the treatment of RAT1 cell extracts with N-ethylmaleimide leads to a loss of the major PRMT1-associated activity that was immune to inhibition under the same conditions as a GST fusion protein. These results suggest that native forms of PRMTs can have different properties than their GST-catalytic chain fusion protein counterparts, which may lack associated noncatalytic subunits.
...
PMID:PRMT3 is a distinct member of the protein arginine N-methyltransferase family. Conferral of substrate specificity by a zinc-finger domain. 1093 50


1 2 Next >>

---