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)

Dichloromethane dehalogenase from Methylophilus sp. DM11 is a glutathione S-transferase homolog that is specifically active with dihalomethane substrates. This bacterial enzyme and rat liver glutathione S-transferases were purified to investigate their relative reactivity with CH2Cl2 and related substrates. Rat liver alpha class glutathione transferases were inactive and mu class enzymes showed low activity (7-23 nmol/min/mg of protein) with CH2Cl2. theta class glutathione transferase 5-5 from rat liver and Methylophilus sp. dichloromethane dehalogenase showed specific activities of > or = 1 mumol/min/mg of protein. Apparent Kcat/Km were determined to be 3.3 x 10(4) and 6.0 x 10(4) L M-1 S-1 for the two enzymes, respectively. Dideutero-dichloromethane was processed to dideutereo-formaldehyde, consistent with a nucleophilic halide displacement mechanism. The possibility of a GSCH2X reaction intermediate (GS, glutathione; X, halide) was probed using CH2ClF to generate a more stable halomethylglutathione species (GSCH2F). The reaction of CH2ClF with dichloromethane dehalogenase produced a kinetically identifiable intermediate that decomposed to formaldehyde at a similar rate to synthetic HOCH2CH2SCH2F. 19F-NMR revealed the transient formation of an intermediate identified as GSCH2F by its chemical shift, its triplet resonance, and H-F coupling constant consistent with a fluoromethylthioether. Its decomposition was matched by a stoichiometric formation of fluoride. These studies indicated that the bacterial dichloromethane dehalogenase directs a nucleophilic attack of glutathione on CH2Cl2 to produce a halomethylthioether intermediate. This focuses attention on the mechanism used by theta class glutathione transferases to generate a halomethylthioeter from relatively unreactive dihalomethanes.
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PMID:Reaction of rat liver glutathione S-transferases and bacterial dichloromethane dehalogenase with dihalomethanes. 813 17

Glutathione conjugation has been identified as an important detoxication reaction. However, in recent years several glutathione-dependent bioactivation reactions have been identified. Current knowledge on the mechanisms and the possible biological importance of these reactions are discussed. 1. Dichloromethane is metabolized by glutathione conjugation to formaldehyde via S-(chloromethyl)glutathione. Both compounds are reactive intermediates and may be responsible for the dichloromethane-induced tumorigenesis in sensitive species. 2. Vicinal dihaloalkanes are transformed by glutathione S-transferase-catalyzed reactions to mutagenic and nephrotoxic S-(2-haloethyl)glutathione S-conjugates. Electrophilic episulphonium ions are the ultimate reactive intermediates formed. 3. Several polychlorinated alkenes are bioactivated in a complex, glutathione-dependent pathway. The first step is hepatic glutathione S-conjugate formation followed by cleavage to the corresponding cysteine S-conjugates, and, after translocation to the kidney, metabolism by renal cysteine conjugate beta-lyase. Beta-Lyase-dependent metabolism of halovinyl cysteine S-conjugates yields electrophilic thioketenes, whose covalent binding to cellular macromolecules is responsible for the observed toxicity of the parent compounds. 4. Finally, hepatic glutathione conjugate formation with hydroquinones and aminophenols yields conjugates that are directed to gamma-glutamyltransferase-rich tissues, such as the kidney, where they undergo alkylation or redox cycling reactions, or both, that cause organ-selective damage.
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PMID:Glutathione-dependent bioactivation of xenobiotics. 828 43

Chlorinated methanes are important industrial chemicals and significant environmental pollutants. While the highly chlorinated methanes, trichloromethane and tetrachloromethane, are not productively metabolized by bacteria, chloromethane and dichloromethane are used by both aerobic and anaerobic methylotrophic bacteria as carbon and energy sources. Some of the dehalogenation reactions involved in the utilization of the latter two compounds have been elucidated. In a strictly anaerobic acetogenic bacterium growing with chloromethane, an inducible enzyme forming methyltetrahydrofolate and chloride from chloromethane and tetrahydrofolate catalyzes dehalogenation of the growth substrate. A different mechanism for the nucleophilic displacement of chloride is observed in aerobic methylotrophic bacteria utilizing dichloromethane as the sole carbon and energy source. These organisms possess the enzyme dichloromethane dehalogenase which, in a glutathione-dependent reaction, converts dichloromethane to inorganic chloride and formaldehyde, a central metabolite of methylotrophic growth. Sequence comparisons have shown that bacterial dichloromethane dehalogenases belong to the glutathione S-transferase enzyme family, and within this family to class Theta. The dehalogenation reactions underlying aerobic utilization of chloromethane by a pure culture and anaerobic growth with dichloromethane by an acetogenic mixed culture are not known. It appears that they are based on mechanisms other than nucleophilic attack by tetrahydrofolate or glutathione.
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PMID:Bacterial growth with chlorinated methanes. 856 6

Although methylene chloride (MC) is readily detectable as a bacterial mutagen, published studies in mammalian cells have been inconclusive. We have previously shown (Graves et al., 1995) that glutathione S-transferase (GST)-mediated metabolism of MC by mouse liver cytosol (S100 fraction) causes DNA single-strand (ss) breaks in CHO cells. In this study, MC GST metabolites were shown to cause mutations at the HPRT locus of CHO cells. The mutagenicity of MC was enhanced by exposing the cells in suspension rather than as attached cultures. The MC GST metabolite formaldehyde was mutagenic in independent experiments, although the number of mutants induced was lower than with the MC. CHO HPRT mutations were also induced by the reference genotoxin 1,2-dibromoethane (1,2-DBE), which is activated to a mutagen by GST-mediated metabolism. Assay of DNA ss breaks and DNA-protein cross-links at mutagenic concentrations of MC, formaldehyde or 1,2-DBE, showed that all three compounds induced DNA ss breaks, but only formaldehyde induced significant DNA-protein cross-linking. These results suggest that whilst formaldehyde may play a role in MC mutagenesis, its weak mutagenicity and the absence of significant DNA-protein cross-linking after MC exposure, leads to the conclusion that the MC DNA damage and resulting mutations are induced by the glutathione conjugate of MC, S-chloromethylglutathione.
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PMID:Mouse liver glutathione S-transferase mediated metabolism of methylene chloride to a mutagen in the CHO/HPRT assay. 860 Mar 70

Formaldehyde, acetaldehyde, and acrolein are well-known upper respiratory tract irritants and occur simultaneously as pollutants in many indoor and outdoor environments. The upper respiratory tract, and especially the nose, is the prime target for inhaled aldehydes. To study possible additive or interactive effects on the nasal epithelium we carried out 1- and 3-day inhalation studies (6 hr/day) with formaldehyde (1.0, 3.2, and 6.4 ppm), acetaldehyde (750 and 1500 ppm), acrolein (0.25, 0.67, and 1.40 ppm), or mixtures of these aldehydes, using male Wistar rats and exposure concentrations varying from clearly nontoxic to toxic. The (mixtures of) aldehydes were studied for histopathological and biochemical changes in the respiratory and olfactory epithelium of the nose. In addition, cell proliferation was determined by incorporation of bromodeoxyuridine and proliferating cell nuclear antigen expression. Effects were primarily observed after 3 days of exposure. Histopathological changes and cell proliferation of the nasal epithelium induced by mixtures of the three aldehydes appeared to be more severe and more extensive in both the respiratory and the olfactory part of the nose than those observed after exposure to the individual aldehydes at comparable exposure levels. As far as nasal histopathological changes and cell proliferation are concerned neither dose addition nor potentiating interactions occurred at no-toxic-effect levels, except for a possible potentiating effect of acetaldehyde at noneffect levels. The results did not indicate a major role for aldehyde dehydrogenases in the biotransformation of the aldehydes studied. Activities of glutathione S-transferase and glutathione reductase after 3 days of exposure to acrolein, alone or in combination with formaldehyde and acetaldehyde, were depressed whereas the glutathione peroxidase activity was elevated. No decrease of nonprotein sulphydryl levels were observed. These findings suggest that, for no-toxic-effect levels, combined exposure to these aldehydes with the same target organ (nose) and exerting the same type of adverse effect (nasal cytotoxicity), but partly with different target sites (different regions of the nasal mucosa), is not associated with a greater hazard than that associated with exposure to the individual chemicals.
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PMID:Changes in the nasal epithelium of rats exposed by inhalation to mixtures of formaldehyde, acetaldehyde, and acrolein. 874 18

Dichloromethane (DCM) (methylene chloride; CH2Cl2) is metabolized via a glutathione S-transferase-mediated pathway to formaldehyde (HCHO), a mutagenic compound that could play a role in the carcinogenic effects of DCM observed in the liver and lungs of B6C3F1 mice at 2000 and 4000 ppm. Mice but not hamsters formed DNA-protein cross-links (DPX) in the liver at DCM concentrations ranging from approximately 500 to 4000 ppm. The formation of DPX was a nonlinear function of the airborne concentration of DCM. In addition, mice exposed to DCM (6 hr/day, 3 days) at concentrations ranging from approximately 1500 to 4000 ppm showed an increased rate of DNA synthesis in the lung indicating cell proliferation, but increased cell turnover was not detected in mouse lung at exposure concentrations of 150 or 500 ppm. Hamsters showed no evidence of cell proliferation in the lung at any concentration, and cell proliferation was not apparent in the livers of either mice or hamsters. An extended physiologically based pharmacokinetic (PBPK) model for DPX formation in mouse liver was developed, based on a published PBPK model for DCM (Andersen, M.E., Clewell, H.J., III, Gargas, M.L., Smith, F.A., and Reitz, R.H. (1987). Toxicol. Appl. Pharmacol. 87, 185-205). The extended PBPK model was fitted to the DPX data using the PBPK model-estimated area under the curve for DCM in mouse liver as the independent variable. Parameter estimates for HCHO disposition in the livers of mice exposed to dichloromethane were similar to previously published estimates for HCHO disposition in the nasal mucosa of rats exposed to formaldehyde. Using the extended PBPK model, estimates were made of the yields of DPX presumably formed in mouse liver at the DCM concentrations used in a bioassay (Mennear, J.H., McConnell, E.E., Huff, J.E., Renne, R.A., and Giddens, E. (1988). Ann. NY Acad. Sci. 534, 343-351). The tumor incidence data in mice were fitted to the DPX yields and to the airborne concentration of DCM as alternative measures of exposure using the linearized multistage (LMS) model. The two dose measures yielded similar maximum likelihood estimates for the cancer risk at concentrations from 10 to 100 ppm, but the upper 95% confidence limit on the risk was reduced by two orders of magnitude when DPX rather than the airborne concentration was used as the measure of exposure. The results demonstrate that an internal dosimeter such as DPX can markedly improve the precision of low-dose risk estimates, while having only a minor effect on the maximum likelihood estimates calculated with the LMS model.
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PMID:DNA-protein cross-links (DPX) and cell proliferation in B6C3F1 mice but not Syrian golden hamsters exposed to dichloromethane: pharmacokinetics and risk assessment with DPX as dosimeter. 899 46

Metabolism of dichloromethane (DCM) to formaldehyde (HCHO) via a glutathione S-transferase (GST) pathway is thought to be required for its carcinogenic effects in B6C3F1 mice. In humans, this reaction is catalyzed primarily by the protein product of the gene GSTT1, a member of the Theta class of GST, and perhaps to a small extent by the protein product of the gene GSTM1. Humans are polymorphic with respect to both genes. Since HCHO may bind to both DNA and RNA forming DNA-protein crosslinks (DPX) and RNA-formaldehyde adducts (RFA), respectively, these products were determined in isolated hepatocytes from B6C3F1 mice, F344 rats, Syrian golden hamsters, and humans to compare species with respect to the production of HCHO from DCM and its reaction with nucleic acids. Only mouse hepatocytes formed detectable amounts of DPX, the quantities of which corresponded well with quantities of DPX formed in the livers of mice exposed to DCM in vivo [Casanova, M., Conolly, R.B., and Heck, H. d'A. (1996). Fundam. Appl. Toxicol. 31, 103-116]. Hepatocytes from all rodent species and from humans with functional GSTT1 and GSTM1 genes formed RFA. No RFA were detected in human cells lacking these genes. Yields of RFA in hepatocytes of mice were 4-fold higher than in those of rats, 7-fold higher than in those of humans, and 14-fold higher than in those of hamsters. The RFA:DPX ratio in mouse hepatocytes incubated with DCM was approximately 9.0 +/- 1.4, but it was 1.1 +/- 0.3 when HCHO was added directly to the medium, indicating that HCHO generated internally from DCM is not equivalent to that added externally to cells and that it may occupy separate pools. DPX were not detected in human hepatocytes even at concentrations equivalent to an in vivo exposure of 10,000 ppm; however, the possibility that very small amounts of DPX were produced from DCM cannot be excluded, since HCHO was formed in human cells. Maximal amounts of DPXliver that might be formed in humans were predicted from the amounts in mice and the relative amounts of RFA in hepatocytes of both species. With predicted DPXliver as the dosimeter, the unit risk, the upper 95% confidence limit on the cancer risk, and the margin of exposure were calculated at several concentrations using the linearized multistage and benchmark dose methods. Since the actual delivered dose is smaller than that predicted, the results suggest that DCM poses at most a very low risk of liver cancer to humans.
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PMID:Dichloromethane metabolism to formaldehyde and reaction of formaldehyde with nucleic acids in hepatocytes of rodents and humans with and without glutathione S-transferase T1 and M1 genes. 924 90

The monohalomethane methyl iodide (MeI) is a site specific toxin within the nasal cavity of the rat, selectively damaging the olfactory epithelium (OE) whilst respiratory epithelium (RE) is spared. The aim of this study was to investigate the rates and routes of metabolism of MeI within the nasal cavity, in order to understand the reasons for the observed site-selectivity. Cytosolic glutathione S-transferases (GSTs) of both OE and RE catalysed the conjugation of MeI with glutathione (GSH), but rates were 4-fold higher in OE than RE. The product of this reaction was confirmed as S-methyl GSH. In both OE and liver the GST catalysing the conjugation of MeI was shown to belong to the theta class. No cytochrome P450-dependent oxidation of MeI to formaldehyde could be detected in incubations containing hepatic or olfactory microsomes. Intact nasal turbinates were incubated with [14C]-MeI, and a dose- and time-dependent covalent binding of MeI to olfactory protein was demonstrated. The rates of protein methylation were found to be similar in OE and RE. Thus the only parameter that correlates with the site-selectivity of the observed lesion is the rate of conjugation of MeI with GSH. Whether toxicity is due to production of a reactive metabolite or GSH depletion per se, remains to be elucidated.
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PMID:Investigations of the pathways of toxicity of methyl iodide in the rat nasal cavity. 977 95

Blood samples from 140 healthy German volunteers were used to further characterize the genetic polymorphism of the human theta class glutathione S-transferase 1 (GSTT1). For measurements of GSTT1 activity, hemolysates were incubated in vitro with different concentrations of dichloromethane. The resulting enzymatically mediated production of formaldehyde was determined colorimetrically by the Nash reaction. GSTT1 genotyping was performed by polymerase chain reaction (PCR) methods using genomic DNA from total white blood cells. The prevalence of homozygous deletion of the GSTT1 gene was 19.3% (95% confidence limits: 12.2-27.7%). There was a high agreement between genotyping and phenotyping data. The individuals with the null genotype had a rate of formaldehyde production below the limit of quantification. In addition, in the group of GSTT1-positive individuals, we could differentiate highly active people (35.7%) from individuals with an intermediate enzyme activity (45.0%). It can be concluded that the PCR method is suitable to quickly genotype large populations, whereas the phenotyping assay at present offers the advantage of differentiating heterozygously from homozygously active subjects. Our results confirm the ethnic differences in the prevalence of the homozygous deleted genotype which were previously observed and seem to exist even between closely related ethnic groups such as German and Swedish populations.
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PMID:Concordance between enzyme activity and genotype of glutathione S-transferase theta (GSTT1). 980 30

The kinetic properties of bacterial and rat liver glutathione S-transferases (GST) active with dichloromethane (DCM) were compared. The theta class glutathione S-transferase (rGSTTI-1) from rat liver had an affinity for dihalomethanes lower by three orders of magnitude (K(app) > 50 mM) than the bacterial DCM dehalogenase/GST from Methylophilus sp. DM11. Unlike the bacterial DCM dehalogenase, the rat enzyme was unable to support growth of the dehalogenase minus Methylobacterium sp. DM4-2cr mutant with DCM. Moreover, the presence of DCM inhibited growth with methanol of the DM4-2cr transconjugant expressing the rat liver GSTT1-1. In Salmonella typhimurium TA1535, expression of rat and bacterial DCM-active GST from a plasmid in the presence of DCM yielded up to 5.3 times more reversions to histidine prototrophy in the transconjugant expressing the rat enzyme. Under the same conditions, however, GST-mediated conversion of DCM to formaldehyde was lower in cell-free extracts of the transconjugant expressing the rat GSTT1 than in the corresponding strain expressing the bacterial DCM dehalogenase. This provided new evidence that formaldehyde was not the main toxicant associated with GST-mediated DCM conversion, and indicated that an intermediate in the transformation of DCM by GST, presumably S-chloromethylglutathione, was responsible for the observed effects. The marked differences in substrate affinity of rat and bacterial DCM-active GST, as well as in the toxicity and genotoxicity associated with expression of these enzymes in bacteria, suggest that bacterial DCM dehalogenases/GST have evolved to minimise the toxic effects associated with glutathione-mediated catalysis of DCM conversion.
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PMID:Enzyme-mediated dichloromethane toxicity and mutagenicity of bacterial and mammalian dichloromethane-active glutathione S-transferases. 1035 Jan 86


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