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)

Highly larvicidal strains of Bacillus sphaericus produce a binary toxin composed of 51 and 42 kDa proteins which binds to sharply delineated regions of the gastric caecum and posterior midgut of susceptible larvae of the mosquito Culex quinquefasciatus. To investigate the role of the individual subunits and the organization of functional binding regions within the toxin, plasmids were constructed for the expression in Escherichia coli of the toxin proteins and their NH2- and COOH-terminal deletion derivatives as fusions with glutathione S-transferase (GST). Toxin proteins were purified by affinity chromatography followed by cleavage from the GST carrier with thrombin. The LC50 values for the purified toxin proteins and their deletion derivatives were determined. The binding patterns of fluorescently labelled toxin suggested that the 51 kDa protein is the primary binding component of the toxin and mediates the regional binding and internalization of the 42 kDa protein. Examination of the toxin deletion derivatives revealed that the NH2-terminal region of the 51 kDa protein was required for binding to the larval gut, whilst the COOH-terminal region was responsible for interacting with the 42 kDa protein. Toxicity was strongly correlated with the subsequent internalization of the toxin, probably by endocytosis.
...
PMID:Binding of purified Bacillus sphaericus binary toxin and its deletion derivatives to Culex quinquefasciatus gut: elucidation of functional binding domains. 151 80

Different glucokinase isoforms are produced by tissue-specific alternative RNA splicing in the liver and pancreatic islet, the only tissues in which glucokinase activity has been detected. To determine whether differences in protein structure brought about by alternative RNA splicing have an effect on glucose phosphorylating activity, we expressed cDNAs encoding four different hepatic and islet glucokinase isoforms and determined the Km and Vmax of each. When the glucokinase B1 and L1 isoforms were expressed in eukaryotic cells, both high Km glucose phosphorylating activity and immunoreactive protein were detected. However, when the glucokinase B2 and L2 isoforms were expressed, both of which differ by deletion of 17 amino acids in a region between the putative glucose and ATP-binding domains, no high Km glucose phosphorylating activity and much less immunoreactive protein were detected. When the glucokinase B1 and B2 isoforms were expressed in Escherichia coli as fusion proteins with glutathione S-transferase, affinity-purified B1 fusion protein was able to phosphorylate glucose whereas the B2 fusion protein was not, thus indicating that the lack of glucose phosphorylating activity from both the B2 and L2 isoforms is due to lack of intrinsic activity in addition to accumulation of less protein. The Km values of the B1 and L1 isoforms, which differ from each other by 15 amino acids at the NH2 terminus, were similar, but the Vmax of the B1 isoform was 2.8-fold higher than that of the L1 isoform. Mutagenesis of the first two potential initiation codons in the glucokinase B1 cDNA from ATG to GTC (methionine to valine) indicated that the first ATG was crucial for activity and is, therefore, the likely translation initiation codon. Messenger RNAs encoding both the B2 and L2 isoforms of glucokinase were detected in islet and liver by polymerase chain reaction amplification of total cDNA, indicating that mRNAs utilizing this weak alternate splice acceptor site in the fourth exon are normally present in both the liver and islet but as minor components. A regulatory role for weak alternate splice acceptor and donor sites in the glucokinase gene was suggested by examining the expression of the gene in the pituitary and in AtT-20 cells. Interestingly, although glucokinase mRNAs of appropriate sizes were detected in both the AtT-20 cells and rat pituitaries, neither exhibited any detectable high Km glucose phosphorylating activity.(ABSTRACT TRUNCATED AT 400 WORDS)
...
PMID:Effects of alternate RNA splicing on glucokinase isoform activities in the pancreatic islet, liver, and pituitary. 201 11

A full length cDNA clone, pGTB38 (C. B. Pickett et al. (1984) J. Biol. Chem. 259, 5182-5188), complementary to a rat liver glutathione S-transferase Ya mRNA has been expressed in Escherichia coli. The cDNA insert was isolated from pGTB38 using MaeI endonuclease digestion and was inserted into the expression vector pKK2.7 under the control of the tac promoter. Upon transformation of the expression vector into E. coli, two protein bands with molecular weights lower than the full-length Ya subunit were detected by Western blot analysis in the cell lysate of E. coli. These lower-molecular-weight proteins most likely result from incorrect initiation of translation at internal AUG codons instead of the first AUG codon of the mRNA. In order to eliminate the problem of incorrect initiation, the glutathione S-transferase Ya cDNA was isolated from the expression vector and digested with Bal31 to remove extra nucleotides from the 5' noncoding region. The protein expressed by this expression plasmid, pKK-GTB34, comigrated with the Ya subunit on sodium dodecyl sulfate polyacrylamide gels and was recognized by antibodies against the YaYc heterodimer. The expressed Ya homodimer was purified by S-hexylglutathione affinity and ion-exchange chromatographies. Approximately 50 mg pure protein was obtained from 9 liters of E. coli culture. The expressed Ya homodimer displayed glutathione-conjugating, peroxidase, and isomerase activities, which are identical to those of the native enzyme purified from rat liver cytosol. Protein sequencing indicates that the expressed protein has a serine as the NH2 terminus whereas the NH2 terminus of the glutathione S-transferase Ya homodimer purified from rat liver cytosol is apparently blocked.
...
PMID:Expression of a cDNA encoding a rat liver glutathione S-transferase Ya subunit in Escherichia coli. 264 28

A series of GSH analogues with modifications at the gamma-glutamyl moiety was synthesized and purified by following peptide chemistry methodology. Benzyl, benzyloxycarbonyl and t-butyloxycarbonyl protective groups were used to protect individual amino acid functional groups. The formation of peptide bonds was accomplished through coupling of free amino groups with active esters, generated by reaction of the carboxylate functions with dicyclohexylcarbodi-imide and 1-hydroxybenzotriazole. The protecting groups in the tripeptides were removed in a single step by using Na in liquid NH3. Precautions were taken in order to prevent oxidation of the thiol function in the cysteine residue. Thus GSH analogues containing both L- and D-glutamic acid and L- and D-aspartic acid, coupled to cysteinylglycine through both the alpha- and the omega-carboxylate group, were synthesized. Also, decarboxy-GSH and deamino-GSH, lacking one functional group in the glutamate moiety, were prepared. The spontaneous non-enzyme-catalysed nucleophilic reaction of these GSH analogues with the electrophilic model substrate 1-chloro-2,4-dinitrobenzene showed appreciable rate differences, indicating the importance of intramolecular interactions in determining the nucleophilic reactivity of the thiol function in the cysteine residue. In particular, the free amino group in the gamma-L-glutamic acid residue appears to play a crucial role in activating the thiol group in GSH. In an adjacent paper [Adang, Brussee, Meyer, Coles, Ketterer, van der Gen & Mulder (1988) Biochem. J. 255, 721-724] these results are compared with those obtained in a study on the ability of these GSH analogues to act as a co-substrate in the glutathione S-transferase-catalysed conjugation reaction with 1-chloro-2,4-dinitrobenzene.
...
PMID:Synthesis and nucleophilic reactivity of a series of glutathione analogues, modified at the gamma-glutamyl moiety. 290 8

Using polysomal immunoselected rat liver glutathione S-transferase mRNAs, we have constructed cDNA clones using DNA polymerase I, RNase H, and Escherichia coli ligase (NAD+)-mediated second strand cDNA synthesis as described by Gubler and Hoffman (Gubler, U., and Hoffman, B. S. (1983) Gene 25, 263-269). Recombinant clone, pGTB42, contained a cDNA insert of 900 base pairs whose 3' end showed specificity for the Yc mRNA in hybrid-select translation experiments. The nucleotide sequence of pGTB42 has been determined, and the complete amino acid sequence of a Yc subunit has been deduced. The cDNA clone contains an open reading frame of 663 nucleotides encoding a polypeptide comprising 221 amino acids with a molecular weight of 25,322. The NH2-terminal sequence deduced from pGTB42 is in agreement with the first 39 amino acids determined for a Ya-Yc heterodimer by conventional protein-sequencing techniques. A comparison of the nucleotide sequence of pGTB42 with the sequence of a Ya clone, pGTB38, described previously by our laboratory (Pickett, C. B., Telakowski-Hopkins, C. A., Ding, G. J.-F., Argenbright, L., and Lu, A.Y.H. (1984) J. Biol. Chem. 259, 5182-5188) reveals a sequence homology of 66% over the same regions of both clones; however, the 5'- and 3'-untranslated regions of the Ya and Yc mRNAs are totally divergent in their sequences. The overall amino acid sequence homology between the Ya and Yc subunits is 68%, however, the NH2-terminal domain is more highly conserved than the middle or carboxyl-terminal domains. Our data suggest that the Ya and Yc subunits of the rat liver glutathione S-transferases are products of two different mRNAs which are derived from two related yet different genes.
...
PMID:Rat liver glutathione S-transferases. Construction of a cDNA clone complementary to a Yc mRNA and prediction of the complete amino acid sequence of a Yc subunit. 298 14

The NH2-terminal amino acid sequence of the Mr 26 000 glutathione S-transferase (EC 2.5.1.18) of Schistosoma japonicum (Sj26) has been deduced by RNA and protein sequence analysis. Using this information, a bacterial plasmid has been constructed that directs the synthesis of the entire Sj26 molecule in Escherichia coli. Recombinant Sj26 exhibits glutathione S-transferase activity and can be readily purified from bacteria in a one-step procedure under non-denaturing conditions. The availability of recombinant Sj26 in essentially unlimited quantities will aid its assessment as a candidate vaccine molecule in schistosomiasis and could eventually lead to the rational design of a drug targetted on schistosome glutathione S-transferases.
...
PMID:Expression of an enzymatically active parasite molecule in Escherichia coli: Schistosoma japonicum glutathione S-transferase. 327 28

We have constructed a nearly full length cDNA clone, pGTA/C44, complementary to the rat liver glutathione S-transferase Yb1 mRNA. The nucleotide sequence of pGTA/C44 has been determined, and the complete amino acid sequence of the Yb1 subunit has been deduced. The cDNA clone contains an open reading frame of 654 nucleotides encoding a polypeptide comprising 218 amino acids with Mr = 25,919. The NH2-terminal sequence deduced from DNA sequence analysis of pGTA/C44 is in agreement with the first 19 amino acids determined for purified glutathione S-transferase A, a Yb1 homodimer, by Frey et al. (Frey, A. B., Friedberg, T., Oesch, F., and Kreibich, G. (1983) J. Biol. Chem. 258, 11321-11325). The DNA sequence of pGTA/C44 shares significant sequence homology with a cDNA clone, pGT55, which is complementary to a mouse liver glutathione S-transferase (Pearson, W. R., Windle, J. J., Morrow, J. F., Benson, A. M., and Talalay, P. (1983) J. Biol. Chem. 258, 2052-2062). We have also determined 37 nucleotides of the 5'-untranslated region and 348 nucleotides of the 3'-untranslated region of the Yb1 mRNA. The Yb1 mRNA and subunit do not share any sequence homology with the rat liver glutathione S-transferase Ya or Yc mRNAs or their corresponding subunits. These data provide the first direct evidence that the Yb1 subunit is derived from a gene or gene family which is distinct from the Ya-Yc gene family.
...
PMID:Rat liver glutathione S-transferases. Nucleotide sequence analysis of a Yb1 cDNA clone and prediction of the complete amino acid sequence of the Yb1 subunit. 384 Apr 77

Glutathione S-transferase activities in mouse hepatic cytosols are elevated as much as 11-fold following the administration of BHA (2(3)-tert-butyl-4-hydroxyanisole), a widely used antioxidant food additive. Ethoxyquin (1,2-dihydro-6-ethoxy-2,2,4-trimethylquinoline) and disulfiram [bis(diethyldithiocarbamyl)disulfide] also enhance the activities of glutathione S-transferases and certain other enzymes. Each of these compounds protects rodents against mutagenic and carcinogenic metabolites. A major (pI 8.7) and a minor (pI 9.3) component of the family of mouse hepatic glutathione S-transferases have been purified to homogeneity. These transferases are immunologically cross-reactive, and have a high degree of NH2-terminal sequence homology (but are not identical). The enzymes differ in a number of molecular and catalytic properties. The transferases are 12-fold elevated by dietary BHA as demonstrated by immunotitration. The mRNA for the major glutathione S-transferase is increased more than 20-fold in the liver RNA of BHA-fed mice, as determined by translation of total liver mRNA and characterization of the products by immunoprecipitation and sodium dodecyl sulfate-gel electrophoresis or by two-dimensional gel electrophoresis. A cDNA plasmid complementary to glutathione S-transferase mRNA was constructed. Translation of liver mRNA selected by hybridization with this plasmid gave products similar to or identical with glutathione S-transferase polypeptides. The cDNA insert has been partially sequenced and its orientation has been determined. Its sequence corresponds to the NH2-terminal region (beginning at residue 9) of the amino acid sequence of the glutathione S-transferase with pI 9.3. Hybridization of the 32P-labeled cDNA plasmid with total liver RNA indicates a 26-fold increase in homologous mRNA in response to the feeding of BHA.
...
PMID:Increased synthesis of glutathione S-transferases in response to anticarcinogenic antioxidants. Cloning and measurement of messenger RNA. 682 48

Native glutathione S-transferases are composed of subunits with apparent molecular weights of 25,000, 23,500, or 22,000 which form either homo- or heterodimers. Glutathione S-transferases A, C, and X which contain two subunits with molecular weights of 23,500 yielded similar but nonidentical proteolytic fragmentation patterns. Fragments unique to the subunits of the homodimers A and X were present in decreased intensities in the patterns of form C. Two-dimensional electrophoresis under denaturing conditions showed single nonoverlapping spots for transferases A and X, while form C yielded two spots corresponding in position to those obtained from forms A and X. Renaturation of dissociated glutathione S-transferase C yielded enzymatically active transferases A, C, and X. These results indicate that form C is a heterodimer composed of one subunit from the homodimeric transferases A and X. This was substantiated by NH2-terminal sequence analysis showing extensive NH2-terminal homology amongst all three forms. However, in the positions where forms A and X yielded different residues, both amino acids were detected in the sequence of form C, indicating that the two subunits of Mr = 23,500 are the products of two different genes. NH2-terminal sequence analysis of the heterodimeric glutathione S-transferase B which is composed of subunits with molecular weights of 22,000 and 25,000 revealed a single unique sequence which bore no resemblance to the sequences of either forms A or X. Despite the identical NH2-terminal sequences, proteolytic fragmentation of the separated subunits showed markedly different fragmentation patterns. This indicates that two different mRNAs code for these two subunits.
...
PMID:Studies on the subunit composition of rat liver glutathione S-transferases. 688 19

Four cDNA fragments encoding different portions of the alpha-subunit of human H,K-adenosine triphosphatase (ATPase) were amplified by means of the polymerase chain reaction technique, ligated into the plasmid pGEX-2T, and expressed as glutathione S-transferase fusion proteins in Escherichia coli. The fragments A (residues 163-313), Ba (residues 360-797), Bb (residues 526-797), and C (residues 822-1031) together encompass 77% of the alpha-subunit and cover most of its cytosolic part. The reactivities of autoantibodies in the sera from patients with pernicious anaemia with the recombinant fusion proteins were analysed by immunoblotting. One autoantigenic epitope was found in the NH2-terminal part of the Ba fragment--that is, between residues 360 and 525. No epitope was detected in the other fragments. The Ba fragment was cleaved off from the glutathione S-transferase fusion protein by the action of thrombin and was then further purified. By means of enzyme-linked immunosorbent assay, 28 of 42 sera (67%) from patients with pernicious anaemia were positive against the purified Ba fragment. The present results provide a final proof that the human H,K-ATPase alpha-subunit is a major autoantigen in the parietal cell and that the major epitope is located between residues 360 to 525 on the cytosolic side of the secretory membrane.
...
PMID:Localization of a pernicious anaemia autoantibody epitope on the alpha-subunit of human H,K-adenosine triphosphatase. 751 38


1 2 3 4 5 6 7 8 9 10 Next >>

---