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)

Drosophila ribosomal protein PO was overexpressed in Escherichia coli to allow for its purification, biochemical characterization and to generate polyclonal antibodies for Western analysis. Biochemical tests were originally performed to see if overexpressed PO contained DNase activity similar to that recently reported for the apurinic/apyrimidinic (AP) lyase activity associated with Drosophila ribosomal protein S3. The overexpressed ribosomal protein was subsequently found to act on AP DNA, producing scissions that were in this case 5' of a baseless site instead of 3', as has been observed for S3. As a means of confirming that the source of AP endonuclease activity was in fact due to PO, glutathione S-transferase (GST) fusions containing a Factor Xa cleavage site between GST and PO were constructed, overexpressed in an E.coli strain defective for the major 5'-acting AP endonucleases and the fusions purified using glutathione-agarose affinity column chromatography. Isolated fractions containing purified GST-PO fusion proteins were subsequently found to have authentic AP endonuclease activity. Moreover, glutathione-agarose was able to deplete AP endonuclease activity from GST-PO fusion protein preparations, whereas the resin was ineffective in lowering DNA repair activity for PO that had been liberated from the fusion construct by Factor Xa cleavage. These results suggested that PO was a multifunctional protein with possible roles in DNA repair beyond its known participation in protein translation. In support of this notion, tests were performed that show that GST-PO, but not GST, was able to rescue an E.coli mutant lacking the major 5'-acting AP endonucleases from sensitivity to an alkylating agent. We furthermore show that GST-PO can be located in both the nucleus and ribosomes. Its nuclear location can be further traced to the nuclear matrix, thus placing PO in a subcellular location where it could act as a DNA repair protein. Other roles beyond DNA repair seem possible, however, since GST-PO also exhibited significant nuclease activity for both single- and double-stranded DNA.
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PMID:Drosophila ribosomal protein PO contains apurinic/apyrimidinic endonuclease activity. 893 86

Rad23 is an evolutionarily conserved protein that is important for nucleotide excision repair. A regulatory role has been proposed for Rad23 because rad23 mutants are sensitive to ultraviolet light but are still capable of incising damaged DNA. Here we show that Rad23 interacts with the 26S proteasome through an amino-terminal ubiquitin-like domain (UbL[R23]). The carboxy terminus of Rad23 binds to the Rad4 DNA repair protein and creates a link between the DNA repair and proteasome pathways. The ultraviolet sensitivity caused by deletion of the UbL(R23) domain may therefore arise from its inability to interact with the proteasome. The fusion proteins glutathione S-transferase (GST)-Rad23 and Rad4-haemagglutinin (HA), and the proteasome subunits Cim3 and Cim5, cofractionate through consecutive chromatography steps. The ubiquitin-like domain of human Rad23 (UbL[HRB]) also interacts with the human proteasome. These results demonstrate that ubiquitin-like domains (UbLs) represent a new class of proteasome-interacting motifs.
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PMID:Rad23 links DNA repair to the ubiquitin/proteasome pathway. 949 Apr 18

The presence of the DNA repair protein O(6)-alkylguanine-DNA alkyltransferase (AGT) paradoxically increases the mutagenicity and cytotoxicity of 1,2-dibromoethane (DBE) in Escherichia coli. This enhancement of genotoxicity did not occur when the inactive C145A mutant of human AGT (hAGT) was used. Also, hAGT did not enhance the genotoxicity of S-(2-haloethyl)glutathiones that mimic the reactive product of the reaction of DBE with glutathione, which is catalyzed by glutathione S-transferase. These experiments support a mechanism by which hAGT activates DBE. Studies in vitro showed a direct reaction between purified recombinant hAGT and DBE resulting in a loss of AGT repair activity and a formation of an hAGT-DBE conjugate at Cys(145). A 2-hydroxyethyl adduct was found by mass spectrometry to be present in the Gly(136)-Arg(147) peptide from tryptic digests of AGT reacted with DBE. Incubation of AGT with DBE and oligodeoxyribonucleotides led to the formation of covalent AGT-oligonucleotide complexes. These results indicate that DBE reacts at the active site of AGT to generate an S-(2-bromoethyl) intermediate, which forms a highly reactive half-mustard at Cys(145). In the presence of DNA, the DNA-binding function of AGT facilitates formation of DNA adducts. In the absence of DNA, the intermediate undergoes hydrolytic decomposition to form AGT-Cys(145)-SCH(2)CH(2)OH.
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PMID:Paradoxical enhancement of the toxicity of 1,2-dibromoethane by O6-alkylguanine-DNA alkyltransferase. 1215 4

Alkylating agents, which are metabolized by glutathione S-transferase (GST), have an important role in the etiology of cancer by forming mutagenic DNA adducts. Previous studies have shown that DNA repair protein, O6-methylguanine DNA methyltransferase, repairs these mutagenic DNA adducts and its activity is correlated with the resistance of human tumors to alkylating agent-based anti-cancer drugs. However, little is known about GST and O6-methylguanine DNA methyltransferase activities in patients with thyroid cancer in vivo. We measured the activities of GST and O6-methylguanine DNA methyltransferase in the leukocytes from patients with papillary thyroid carcinoma and healthy controls. The GST activity was significantly higher in men than in women, and it was negative correlated with age in men whereas it was unchanged in women in the control group. Both GST and O6-methylguanine DNA methyltransferase activities were significantly increased in the patient group. There were no age and sex-related changes in the O6-methylguanine DNA methyltransferase activity in both the control and patient groups. These results suggest that leukocyte GST and O6-methylguanine DNA methyltransferase activities were increased with thyroid cancer. This may be related to the resistance to chemotherapy exhibited by patients with thyroid cancer.
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PMID:Glutathione S-transferase and O6-methylguanine DNA methyl transferase activities in patients with thyroid papillary carcinoma. 1244 29

A lower rate of colon cancer was observed in consumers of coffee with a high content of the diterpenes Kahweol and Cafestol (K/C). In animal models, K/C have been found to protect against the mutagenic/carcinogenic effects of compounds such as 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP), aflatoxin B1, and 7,12-dimethylbenz[a]anthracene. Thus far, such chemoprotection by K/C has been attributed to modifications of xenobiotic metabolism, e.g. enhanced detoxification by UDP-glucuronosyltransferase (UDPGT) and/or glutathione transferase (GST). In the present study, we investigated the potential of several coffee-related treatments (K/C [1:1], Cafestol-alone, Turkish coffee) to modify the expression level of the DNA repair protein O(6)-methylguanine-DNA methyltransferase (MGMT) which is involved in the reversal of the precarcinogenic DNA damage O(6)-alkylguanine induced by alkylating agents. The results show that, in the male F344 rat, K/C and Cafestol increase hepatic MGMT in a dose-dependent manner up to a maximum of 2.6-fold at 0.122% K/C in the feed. Turkish coffee led to enhancements of up to 16%, the more moderate increase being associated with the lower estimated K/C intake through the beverage. In the livers of the rats receiving Turkish coffee, we also found 10-30% increases in several GST-related parameters (overall GST, GST-pi, glutathione, gamma-glutamylcysteine-synthetase) and a two-fold increase in UDPGT activity. Dose-response studies with K/C revealed that MGMT increased in parallel with three of the four GST-related parameters whereas the dose-response curves of UDPGT and of GST-pi activity displayed a steeper slope. Increased expression level of MGMT may extend the antimutagenic/anticarcinogenic potential of coffee components to protection against DNA alkylating agents.
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PMID:Coffee and its chemopreventive components Kahweol and Cafestol increase the activity of O6-methylguanine-DNA methyltransferase in rat liver--comparison with phase II xenobiotic metabolism. 1251 12

O6-methylguanine-DNA methyltransferase (MGMT) is a DNA repair protein which protects the cellular genome and critical oncogenic genes from the mutagenic action of endogenous and exogenous alkylating agents. An expedited elimination of O6-alkylguanines by increasing MGMT activity levels is likely to be a successful chemoprevention strategy. Here, we report for the first time that cysteine/glutathione enhancing drugs and certain plant antioxidants possess the ability to increase human MGMT expression beyond its steady-state levels that may afford protection. The non-toxic cysteine prodrugs, 2-oxothiazolidine-4-carboxylic acid (OTC) and N-acetyl-L-cysteine (NAC), metabolized, respectively by 5-oxoprolinase and acylases, increased the MGMT protein and its repair activity levels in a dose- and time-dependent manner in several cancer cell lines and peripheral blood lymphocytes with a maximum of 3-fold increase by 72 h. The natural antioxidants, namely, curcumin, silymarin, sulforaphane and resveratrol were also effective in raising the MGMT levels to different extents. Among the synthetic agents, oltipraz and N-(4-hydroxyphenyl) retinamide (4-HPR) also increased MGMT expression, albeit to a lesser extent. Augmented mRNA levels accounted at least, in part, for the increased activity of MGMT in this setting. However, evidence from cysteine/methionine deprivation, acivicin treatment, and protein synthesis measurements in OTC-treated cells suggested that an increased cysteine flux also contributed significantly to enhanced MGMT expression. Many of these treatments increased the glutathione S-transferase-P1 (GSTP1) levels as well. These findings raise the possibility of MGMT-targeted chemoprevention strategies through dietary supplementation of OTC and herbal antioxidants. Further, the studies reveal the antioxidant responsiveness of the human MGMT gene.
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PMID:Increased expression of the MGMT repair protein mediated by cysteine prodrugs and chemopreventative natural products in human lymphocytes and tumor cell lines. 1695 Jul 96

The environmental contaminant 1,2-dibromoethane and diepoxybutane, an oxidation product of the important industrial chemical butadiene, are bis-functional electrophiles and are known to be mutagenic and carcinogenic. One mechanism by which bis-electrophiles can exert their toxic effects is through the induction of genotoxic and mutagenic DNA-peptide cross-links. This mechanism has been shown in systems overexpressing the DNA repair protein O6 -alkylguanine DNA-alkyltransferase (AGT) or glutathione S-transferase and involves reactions with nucleophilic cysteine residues. The hypothesis that DNA-protein cross-link formation is a more general mechanism for genotoxicity by bis-electrophiles was investigated by screening nuclear proteins for reactivity with model monofunctional electrophiles. Glyceraldehyde 3-phosphate dehydrogenase (GAPDH) was identified as a candidate because of the nucleophilicity of two cysteine residues (Cys152 and Cys246) in reaction screens with model electrophiles (Dennehy, M. K. et al. (2006) Chem. Res. Toxicol. 19, 20-29). Incubation of GAPDH with bis-electrophiles resulted in inhibition of its catalytic activity, but only at high concentrations of diepoxybutane. In vitro assays indicated DNA-GAPDH cross-link formation in the presence of diepoxybutane, and bis-electrophile reactivity at Cys246 was confirmed using mass spectral analysis. In contrast to AGT, overexpression of human GAPDH in Escherichia coli did not enhance mutagenesis by diepoxybutane. We propose that the lack of mutational enhancement is in part due to the inherently lower reactivity of GAPDH toward bis-electrophiles as well as the reduced DNA binding ability relative to AGT, preventing the in vivo formation of DNA-protein cross-links and enhanced mutagenesis.
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PMID:Reactions of glyceraldehyde 3-phosphate dehydrogenase sulfhydryl groups with bis-electrophiles produce DNA-protein cross-links but not mutations. 1816 42

The Ku heterodimer, a DNA repair protein complex consisting of 70- and 80-kDa subunits, is involved in the non-homologous end-joining (NHEJ) pathway. Plants are thought to use the NHEJ pathway primarily for the repair of DNA double-strand breaks (DSBs). The Ku70/80 protein has been identified in many plants and been shown to possess several similar functions to its counter protein complex in mammals. In the present study, ovate family protein 1 (AtOFP1) was demonstrated to be a plant Ku-interacting protein by yeast two-hybrid screening and the GST pull-down assay. Truncation analysis revealed that the C-terminal domain of AtKu70 contains interacting sites for AtOFP1. The electrophoretic mobility shift assay (EMSA) indicated that AtOFP1 is also a DNA binding protein with its binding domain at the N-terminus. In 3-week-old seedlings, expression of the AtOFP1 gene increased after exposure to DNA-damaging agents (such as methyl methanesulfonate (MMS) and menadione) in a time dependent manner. Seedlings lacking the AtOFP1 protein were more sensitive to MMS and menadione as compared with wild-type. Furthermore, similar to AtKu70(-/-) and AtKu80(-/-), the AtOFP1(-/-) mutant showed relatively lower NHEJ activity in vivo. Taken together, these results suggest that AtOFP1 may play a role in DNA repair through the NHEJ pathway accompanying with the AtKu protein.
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PMID:Ovate family protein 1 as a plant Ku70 interacting protein involving in DNA double-strand break repair. 2084 35

Cytoplasmic male sterility (CMS), widely used in the production of hybrid seeds, is a maternally inherited trait resulting in a failure to produce functional pollen. In order to identify some specific proteins associated with CMS in pepper, two-dimensional gel electrophoresis (2-DE) was applied to proteomic analysis of anthers/buds between a CMS line (designated NA3) and its maintainer (designated NB3) in Capsicum annuum L. Thirty-three spots showed more than 1.5-fold in either CMS or its maintainer. Based on mass spectrometry, 27 spots representing 23 distinct proteins in these 33 spots were identified. Proteins down-regulated in CMS anthers/buds includes ATP synthase D chain, formate dehydrogenase, alpha-mannosidas, RuBisCO large subunit-binding protein subunit beta, chloroplast manganese stabilizing protein-II, glutathione S-transferase, adenosine kinase isoform 1T-like protein, putative DNA repair protein RAD23-4, putative caffeoyl-CoA 3-O-methyltransferase, glutamine synthetase (GS), annexin Cap32, glutelin, allene oxide cyclase, etc. In CMS anthers/buds, polyphenol oxidase, ATP synthase subunit beta, and actin are up-regulated. It was predicted that male sterility in NA3 might be related to energy metabolism turbulence, excessive ethylene synthesis, and suffocation of starch synthesis. The present study lays a foundation for future investigations of gene functions associated with pollen development and cytoplasmic male sterility, and explores the molecular mechanism of CMS in pepper.
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PMID:Differential proteomic analysis of anthers between cytoplasmic male sterile and maintainer lines in Capsicum annuum L. 2426 42