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

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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)

The 3-dimensional crystal structure of glutathione S-transferase (GST) of Schistosoma japonicum (Sj) fused with a conserved neutralizing epitope on gp41 (glycoprotein, 41 kDa) of human immunodeficiency virus type 1 (HIV-1) (Muster T et al., 1993, J Virol 67:6642-6647) was determined at 2.5 A resolution. The structure of the 3-3 isozyme rat GST of the mu gene class (Ji X, Zhang P, Armstrong RN, Gilliland GL, 1992, Biochemistry 31:10169-10184) was used as a molecular replacement model. The structure consists of a 4-stranded beta-sheet and 3 alpha-helices in domain 1 and 5 alpha-helices in domain 2. The space group of the Sj GST crystal is P4(3)2(1)2, with unit cell dimensions of a = b = 94.7 A, and c = 58.1 A. The crystal has 1 GST monomer per asymmetric unit, and 2 monomers that form an active dimer are related by crystallographic 2-fold symmetry. In the binding site, the ordered structure of reduced glutathione is observed. The gp41 peptide (Glu-Leu-Asp-Lys-Trp-Ala) fused to the C-terminus of Sj GST forms a loop stabilized by symmetry-related GSTs. The Sj GST structure is compared with previously determined GST structures of mammalian gene classes mu, alpha, and pi. Conserved amino acid residues among the 4 GSTs that are important for hydrophobic and hydrophilic interactions for dimer association and glutathione binding are discussed.
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PMID:Three-dimensional structure of Schistosoma japonicum glutathione S-transferase fused with a six-amino acid conserved neutralizing epitope of gp41 from HIV. 753 46

The niaD and niiA genes of Aspergillus nidulans, which code, respectively, for nitrate and nitrite reductases, are divergently transcribed, and their ATGs are separated by 1,200 bp. The genes are under the control of the positively acting NirA transcription factor, which mediates nitrate induction. The DNA binding domain of NirA was expressed as a fusion protein with the glutathione S-transferase of Schistosoma japonicum. Gel shift and footprint experiments have shown that in the intergenic region there are four binding sites for the NirA transcription factor. These sites can be represented by the nonpalindromic consensus 5'CTCCGHGG3'. Making use of a bidirectional expression vector, we have analyzed the role of each of the sites in niaD and niiA expression. The sites were numbered from the niiA side. It appeared that site 1 is necessary for the inducibility of niiA only, while sites 2, 3, and to a lesser extent 4 (which is nearer to and strongly affects niaD) act bidirectionally. The results also suggest that of the 10 binding sites for the AreA protein, which mediates nitrogen metabolite repression, those which are centrally located are physiologically important. The insertion of an unrelated upstream activating sequence into the intergenic region strongly affected the expression of both genes, irrespective of the orientation in which the element was inserted.
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PMID:The intergenic region between the divergently transcribed niiA and niaD genes of Aspergillus nidulans contains multiple NirA binding sites which act bidirectionally. 756 20

We used direct RNA sequencing to determine the genomic organization of the region downstream from the G gene of viral hemorrhagic septicemia virus (VHSV), a fish rhabdovirus. This region contains a gene coding for a protein, identified as nonvirion protein (NV), and the gene coding for the RNA polymerase (L). Thus, VHSV genome organization was confirmed to be 3'-N-P-M-G-NV-L-5'. In both a virulent European (07-71) and an avirulent North American (Makah) strain, the NV gene is transcribed into a small mRNA that codes for a protein of 122 amino acids. It has no significant sequence similarity with the infectious hematopoietic necrosis virus NV protein nor with any other known protein. We expressed the NV protein as a fusion protein with the glutathione S-transferase of Schistosoma japonicum and used the purified fusion protein to immunize rabbits. The rabbit antiserum precipitated from infected cell extracts--and not from noninfected cells or purified virions--a protein of 14 kDa, well in accordance with the expected NV gene product size. The prediction that the NV protein is a nonstructural protein is supported by its absence from mature virions although it is present in infected cells.
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PMID:Distant strains of the fish rhabdovirus VHSV maintain a sixth functional cistron which codes for a nonstructural protein of unknown function. 757 46

Mutations in the human glucokinase (GK) gene are thought to cause maturity-onset diabetes of youth (MODY) by leading to the production of enzymes with reduced catalytic activities and increased glucose Km values. However, in some cases the diabetic phenotype is more severe than might be predicted from these apparent kinetic effects alone. To determine whether these mutations might also effect other characteristics of the enzyme, nine MODY-associated mutants were expressed as fusion proteins with Schistosoma japonicum glutathione S-transferase (GST) and compared with three wild-type human GK isoforms that were also expressed in the same manner. Three GST-GK isoforms (liver 1, liver 2 and islet) were kinetically indistinguishable from each other and from purified rat liver GK. Noteworthy is a glucose-induced fit effect for the interaction of trinitrophenyl (TNP)-ATP with GST-GK, whereby glucose significantly increased the affinity of TNP-ATP binding to GST-GK without changing the stoichiometry of binding. The nine MODY-associated mutations studied either showed diminished catalytic activity, substrate affinities, allosteric regulation, or stability of the fusion enzyme. We conclude that: (1) Gly261 and Lys414 are important for ATP binding; (2) Val203 may be essential for a glucose-induced fit effect; and (3) the stability of fusion protein may be significantly reduced when Glu300 is replaced by Lys. These results suggest that, in addition to effects on the Km and Vmax. of GK, a decrease in the ATP-binding affinity or stability of the mutated enzyme may also contribute to a reduction of GK activity in individuals with GK-MODY. In the B-cell this would have the effect of blunting glucose-stimulated insulin release, thereby contributing to the diabetic phenotype.
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PMID:Variable effects of maturity-onset-diabetes-of-youth (MODY)-associated glucokinase mutations on substrate interactions and stability of the enzyme. 761 52

Nucleic acid vaccination by intramuscular or intradermal delivery of DNA plasmids encoding antigenic proteins has been shown to confer protection in experimental animals against viruses and unicellular protozoan parasites. However, this revolutionary approach has not been tested for induction of immunity to multicellular parasites, such as trematode worms. We report here, for the first time, that murine antibodies can be induced by intramuscular injection with plasmid DNA encoding fragments of Schistosoma japonicum paramyosin (Sj97), a 97 kDa molecule and a promising vaccine candidate in schistosomiasis. An additional construct containing the gene encoding full-length glutathione S-transferase (Sj26), another recognised anti-schistosome vaccine target, failed to raise detectable levels of specific antibody.
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PMID:Antibodies to Schistosoma japonicum (Asian bloodfluke) paramyosin induced by nucleic acid vaccination. 762 89

We have developed a rapid and simple procedure for the production and the purification of Escherichia coli thioredoxins containing additional amino acid residues at the N-terminus. By the polymerase chain reaction, the complete gene encoding for E. coli thioredoxin was modified and amplified with the addition at its 5' end of a BamHI cloning site and a triplet coding for an arginine residue instead of the initiator methionine codon, whereas at the 3' end the stop codon was followed by an EcoRI cloning site. The synthetic DNA was ligated into the BamHI/EcoRI site of the vector plasmid pGEX-2T, and the novel plasmid [pFTG] was used for the transformation of E. coli cells. Following induction and cell disruption, a protein composed of Schistosoma japonicum glutathione S-transferase and E. coli thioredoxin was obtained in soluble form and purified by affinity chromatography on agarose columns bearing immobilized glutathione. This procedure yielded 50 mg of homogeneous fusion protein per liter of culture media. Digestion of the chimeric thioredoxin with bovine plasma thrombin followed by an additional chromatography on glutathione-agarose gave a protein that contained the entire sequence of E. coli thioredoxin and three additional amino acid residues [G-S-R-] at the N-terminal side. The structural characteristics and the protein disulfide oxidoreductase activity of this recombinant protein, in terms of variations of emission fluorescence and reduction of insulin disulfide bonds, respectively, were essentially identical to those of its counterpart obtained from wild-type cells by conventional techniques of proteins purification.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:A procedure for the generation and the purification of Escherichia coli thioredoxins with variable N-terminal sequences. 766 53

Diethylpyrocarbonate (DEP) inhibits the catalytic activity of a cloned glutathione S-transferase from Schistosoma japonicum (Sj26GST) with a second-order rate constant of 474 M-1 min-1 at pH 7.0 and 25 degrees C. There is an accompanying increase in absorbance at 242 nm due to the formation of N-carbethoxyhistidyl derivatives. There was no evidence that tyrosine or cysteine residues were modified by DEP treatment nor did the enzyme undergo any major conformational change. Activity can be restored by treating the DEP-modified enzyme with hydroxylamine and the pH curve for inactivation indicates involvement of a residue with a pKa of 7.3. Complete inactivation of Sj26GST requires the modification of six histidine residues per subunit. Statistical analysis of residual enzyme activity versus number of groups modified showed that of the six modifiable groups, only one is critical for activity. Substrate protection suggests that this essential histidine residue is at or near the active site.
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PMID:Chemical modification of a cloned glutathione S-transferase from Schistosoma japonicum: evidence for an essential histidine residue. 775 42

Synthesis in E. coli of native coat protein of Johnsongrass mosaic virus, and hybrid protein molecules containing foreign antigens, resulted in the intracellular formation of potyvirus-like particles (PVLPs). The foreign antigens used were an octapeptide epitope from Plasmodium falciparum and a decapeptide hormone (luteinizing hormone releasing hormone) at the N- or at both N- and C-terminal regions of the coat protein molecule, and a full length protein antigen (Sj26-glutathione S-transferase of 26 kD from Schistosoma japonicum) replacing the N-terminal 62 amino acids of the coat protein. Electron microscopy of ultrathin sections of E. coli revealed that PVLPs resulting from coat protein molecules containing peptide fusions appeared in vast arrays of parallel strands within the cytoplasm sometimes extending the length of the cell and at times the cells were strung together, with "threads" of PVLPs appearing to connect individual bacterial cells. PVLPs resulting from the fusion of the 26 kD antigen Sj26 to coat protein were shorter and wider. The physical form of the high molecular weight PVLPs enabled purification by simple size exclusion column chromatography. The Sj26-PVLPs administered to mice without adjuvant elicited antibody responses comparable to monomeric Sj26 administered with Freund's Complete Adjuvant.
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PMID:High level production of hybrid potyvirus-like particles carrying repetitive copies of foreign antigens in Escherichia coli. 776 98

The nef gene of an infectious molecular clone of SIVSMM isolate PBj14 was fused to the glutathione S-transferase gene of Schistosoma japonicum to generate plasmid pEMC100. The recombinant plasmid was placed in an aroA live vaccine Salmonella dublin strain, and the production of GST-Nef protein was induced by exposure to IPTG. The fusion protein was purified and administered as vaccine to BALB/c mice by i.p. injection. Several doses of the purified fusion protein produced an earlier anti-GST-Nef response, without an anti-GST response, than did IPTG-induced Salmonella live vaccine containing an equal amount (0.1 microgram) of fusion protein, apparently because of the transient immunosuppressive effect of live vaccine given by injection. The highest anti-GST-Nef titers were obtained by a third immunization schedule in which mice were treated with a priming inoculum of induced live vaccine followed, after the predicted immunosuppressed interval, by two i.p. doses of 1 microgram of purified GST-Nef protein with Ribi adjuvant. The data presented here demonstrate that SL5928 aroA, an attenuated S. dublin strain, can be used as a live vaccine carrier to express Nef protein of SIVSMM-PBj14, one of the most acutely pathogenic primate lentiviruses so far described.
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PMID:Immunogenicity of Nef protein of SIVSMM-PBj14 expressed in a live vaccine strain of Salmonella species. 781 32

Schistosomiasis research within the framework of the Commission of the European Communities 'Science and Technology for Development' (CEC/STD) Programme is targeted at three specific problems: diagnosis of infection and disease; the dynamics of transmission, immunity, and morbidity; and the need for improved tools and strategies for control. Several important advances have been made over the past decade. Improved methods of diagnosis by detection of circulating antigens are in an advanced stage of development and have already undergone field trials in several epidemiological settings. Treatment and reinfection studies combined with immunological observations have allowed the elucidation of possible mechanisms leading to acquired resistance, and have shown that repeated chemotherapy with praziquantel can substantially reduce morbidity. Other projects have studied the epidemiological and ecological determinants of transmission, infection and disease in various endemic situations and also in newly established, epidemic foci where remarkable observations on chemotherapeutic responses were made. Important advances have been made towards the development of a vaccine. The glutathione-S-transferases of the major species of schistosomes have been cloned, sequenced and expressed, and their biological function studied. In a variety of vaccine formulations and animal systems GST has been able to confer protection against infection and to reduce worm fecundity. GST and a series of other crude and defined antigens have been evaluated with varying results in Schistosoma japonicum and S. bovis in cattle. Much work has yet been done, however. Recommendations as to possible future directions for research are provided.
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PMID:Schistosomiasis research and the European Community. 782 31


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