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PubMed1985

Isoenzymes of glutathione transferase in rat kidney cytosol.

Glutathione transferases from rat kidney cytosol were purified about 40-fold by chromatography on S-hexylglutathione linked to epoxy-activated Sepharose 6B. Further purification by fast protein liquid chromatography with chromatofocusing in the pH interval 10.6-7.6 resolved five major peaks of activity with 1-chloro-2,4-dinitrobenzene as the second substrate. Four of the peaks were identified with rat liver transferases 1-1, 1-2, 2-2 and 4-4 respectively. The criteria used for identification included physical properties, reactions with specific antibodies, substrate specificities and sensitivities to several inhibitors. The fourth major peak is a 'new' form of transferase, which has not been found in rat liver. This isoenzyme, glutathione transferase 7-7, has a lower apparent subunit Mr than any of the transferases isolated from rat liver cytosol, and does not react with antibodies raised against the liver enzymes. Glutathione transferases 3-3 and 3-4, which are abundant in liver, were only present in very small amounts. In a separate chromatofocusing separation in a lower pH interval, an additional peak was eluted at pH 6.3. This isoenzyme is characterized by its high activity with ethacrynic acid.

Animals↗
PubMed1986

Two distinct forms of glutathione transferase from human foetal liver. Purification and comparison with isoenzymes isolated from adult liver and placenta.

Isoelectric focusing of a cytosol fraction from human foetal liver revealed the existence of an acidic and a basic isoenzyme of GSH transferase. The acidic and basic forms of GSH transferase were purified in good yield by use of ion-exchange chromatography on DEAE-cellulose followed by affinity chromatography on S-hexyl-GSH coupled to epoxy-activated Sepharose 6B. The content of the acidic and the basic isoenzymes of GSH transferase together was calculated to constitute 1-2% of the soluble proteins in the hepatic cytoplasm. Physical, catalytic and immunological analyses of the acidic and the basic isoenzymes from foetal liver demonstrated unambiguously that the two forms are different structures with distinct properties. On the other hand, the results show clearly extensive similarities between the foetal acidic transferase and transferase pi from human placenta as well as between the foetal basic form and the basic isoenzymes isolated from adult liver. An exception is that both foetal enzymes seem to be considerably more efficient in catalysing the conjugation of GSH with styrene 7,8-epoxide than the corresponding adult forms of GSH transferase.

Female↗
PubMed1988

Isoenzymes of glutathione transferase in rat small intestine.

The major glutathione transferases in the rat small-intestine cytosol were isolated and characterized. The enzymes active with 1-chloro-2,4-dinitrobenzene as second substrate were almost quantitatively recovered after affinity chromatography on immobilized S-hexylglutathione. The different basic forms of glutathione transferase, which account for 90% of the activity, were resolved by chromatofocusing. Fractions containing enzymes with lower isoelectric points were not further resolved. The isolated fractions were characterized by their elution position in chromatofocusing, apparent subunit Mr, reactions with specific antibodies, substrate specificities and inhibition characteristics. The major basic forms identified were glutathione transferases 1-1, 4-4 and 7-7. In addition, evidence for the presence of a variant form of subunit 1, as well as trace amounts of subunits 2 and 3, was obtained. A significant amount of transferase 8-8 in the fraction of acidic enzyme forms was demonstrated by immunoblot and Ouchterlony double-diffusion analysis. In the comparison of the occurrence of the different forms of glutathione transferase in liver, lung, kidney and small intestine, it was found that the small intestine is the richest source of glutathione transferase 7-7.

Animals↗
PubMed1988

Positive correlation exists between glutathione S-transferase activity and aflatoxin formation in Aspergillus flavus.

The presence of glutathione (GSH) S-transferase activity, using 1-chloro-2, 4-dinitrobenzene (CDNB) as a substrate, has been established in the cytosolic fraction of the toxigenic (aflatoxin producing) and nontoxigenic strains of Aspergillus flavus. Significant differences in the GSH S-transferase activity were observed between the toxigenic and non-toxigenic strains. A positive correlation has been demonstrated for the first time between aflatoxin formation and a biochemical parameter, namely GSH S-transferase activity. The evidence in support of A. flavus GSH S-transferase induction by endogenous aflatoxins is as follows: (i) the age-related production of aflatoxin follows the same pattern as the cytosolic GSH S-transferase activity profile; (ii) significantly higher enzyme activity was associated with mycelia of a toxigenic strain grown in medium supporting high aflatoxin production (sucrose-low-salts medium) while the enzyme activity was low in medium producing less aflatoxin (glucose-ammonium nitrate medium). The GSH S-transferase activity of the non-toxigenic strain was hardly affected by a change in the medium as it produces no aflatoxins; and (iii) the toxigenic strain demonstrated significantly higher apparent Vmax. with no change in Km as compared with the non-toxigenic strain. This indicates that the enzyme induction by endogenous aflatoxins is similar to the action of phenobarbitol and other inducing drugs (Kaplowitz et al., 1975).

Acetates↗
PubMed1990

Increase in the amount of glutathione transferase 4-4 in the rat adrenal gland after hypophysectomy and down-regulation by subsequent treatment with adrenocorticotrophic hormone.

The effect of hypophysectomy and subsequent treatment with adrenocorticotropic hormone (adrenocorticotropin, ACTH) on the isoenzymes of glutathione transferase in the rat adrenal gland was investigated. A large increase (approx. 11-fold) in the level of transferase subunit 4 was observed in hypophysectomized animals by immunoblotting. When the activity of glutathione transferase 4-4 was measured in adrenal cytosol using trans-stilbene oxide as a selective substrate, a 15-fold increase was noted. Lack of the pituitary hormone ACTH is apparently related to this increase, since treatment of hypophysectomized animals with ACTH for 2 weeks partially down-regulated subunit 4. Glutathione transferase subunits 3 and 8 in the adrenal were also increased in amount by hypophysectomy, but not at all to the same extent. The activity of glutathione transferase 4-4 was elevated also in the liver and ovary (5 and 1.5 times respectively) after hypophysectomy. These elevated enzyme levels were, however, not affected by ACTH treatment. This down-regulation of glutathione transferases in the rat adrenal by ACTH may be related to the fact that, under normal conditions, this organ is highly susceptible to the toxic effects of various polycyclic hydrocarbons, whereas under circumstances where there is no ACTH production, as in hypophysectomized rats, the adrenal is resistant to these same hydrocarbons.

Adrenal Glands↗
PubMed1991

The potency of inducers of NAD(P)H:(quinone-acceptor) oxidoreductase parallels their efficiency as substrates for glutathione transferases. Structural and electronic correlations.

Induction of glutathione transferases (EC. 2.5.1.18), NAD(P)H:(quinone-acceptor) oxidoreductase (EC 1.6.99.2; quinone reductase) and other detoxification enzymes is a major mechanism for protecting cells against the toxicities of electrophiles, including many carcinogens. Although inducers of these two enzymes belong to many different chemical classes, they nevertheless contain (or acquire by metabolism) electrophilic centres that appear to be essential for inclusive activity, and many inducers are Michael reaction acceptors [Talalay, De Long & Prochaska (1988) Proc. Natl. Acad. Sci. U.S.A., 85, 8261-8265]. The inducers therefore share structural and electronic features with glutathione transferase substrates. To define these features more precisely, we examined the inductive potencies (by measuring quinone reductase in murine hepatoma cells) of two types of glutathione transferase substrates: a series of 1-chloro-2-nitrobenzenes bearing para-oriented electron-donating or -withdrawing substituents and a wide variety of other commonly used and structurally unrelated glutathione transferase substrates. We conclude that virtually all glutathione transferase substrates are inducers, and their potencies in the nitrobenzene series correlate linearly with the Hammett sigma or sigma- values of the aromatic substituents, precisely as previously reported for their efficiencies as glutathione transferase substrates. More detailed information on the electronic requirements for inductive activity was obtained with a series of methyl trans-cinnamates bearing electron-withdrawing or -donating substituents on the aromatic ring, and in which the electronic densities at the olefinic and adjacent carbon atoms were measured by 13C n.m.r. Electron-withdrawing meta-substituents markedly enhance inductive potency in parallel with their increased non-enzymic reactivity with GSH. Thus, methyl 3-bromo-, 3-nitro- and 3-chloro-cinnamates are 21, 14 and 8 times more potent inducers than the parent methyl cinnamate. This finding permits the design of more potent inducers, which are important for elucidation of the molecular mechanisms of induction.

Animals↗
PubMed2000

Low levels of colonic glutathione S-transferase in patients with X-linked agammaglobulinaemia.

BACKGROUND: Patients with X-linked agammaglobulinaemia, a primary immunodeficiency disorder, suffer from recurrent infections of the respiratory and intestinal tract. Rapidly progressive colorectal cancer was diagnosed in three unrelated young adults with X-linked agammaglobulinaemia. This finding implies a 30-fold increase of risk for this cancer in this patient group. Glutathione S-transferases are a family of biotransformation enzymes involved in the detoxification of cytotoxic and carcinogenic compounds, that may function in the prevention of carcinogenesis. We investigated the possible role of the glutathione S-transferase enzyme system in the apparently increased colorectal cancer risk in X-linked agammaglobulinaemia patients. MATERIALS AND METHODS: We analysed the glutathione levels and the glutathione S-transferase enzyme activity and iso-enzyme composition in normal colonic biopsies of eight X-linked agammaglobulinaemia patients, 25 patients with a recent history of colonic adenomas and 10 healthy volunteers. RESULTS: X-linked agammaglobulinaemia patients had significantly lower glutathione S-transferase enzyme activities at all sites in the normal colonic mucosa as compared to adenoma patients. In X-linked agammaglobulinaemia patients the rectal glutathione S-transferase enzyme activity was lower than in the proximal colon and significantly lower as compared to controls. CONCLUSION: This lower glutathione S-transferase enzyme activity might play a role in the apparently increased colorectal cancer risk in X-linked agammaglobulinaemia patients, assuming that detoxification of carcinogenic compounds plays a role in the aetiology of colon cancer of these patients.

Adenoma↗
PubMed1998

Cloning and characterization of 3-deoxy-D-manno-oct-2-ulosonic acid (Kdo) transferase genes (kdtA) from Acinetobacter baumannii and Acinetobacter haemolyticus.

3-Deoxy-D-manno-oct-2-ulosonic acid (Kdo) transferases (KdtA) are multifunctional glycosyltransferases with primary structures of low similarity. Totally degenerated primers were deduced from two stretches of identical amino acids between known KdtA sequences and used to amplify by PCR a kdtA-specific fragment from Acinetobacter baumannii ATCC 15308 DNA which was then applied as a probe for the cloning and sequencing of the complete Kdo transferase gene. With conserved PCR primers for this structural gene from A. baumannii ATCC 15308, also kdtA genes of A. baumannii ATCC 19606 and A. haemolyticus ATCC 17906 were obtained, cloned from the chromosome and sequenced. The genes coded for proteins with similarities to known Kdo transferases. Within the genus Acinetobacter, the identity and similarity of the deduced amino acid sequences were 71% and 84.5%, respectively. The kdtA sequences of both A. baumannii strains were identical and possessed a TTG start codon, whereas ATG was found in the case of A. haemolyticus. The genes from Acinetobacter and kdtA from Escherichia coli K-12 were expressed in the Gram-positive bacterium Corynebacterium glutamicum. In vitro tests confirmed the function of the gene products as Kdo transferases, which transferred mainly two Kdo residues to a synthetic lipid A precursor of E. coli. Also, no differences between the cloned kdtA genes from A. baumanniii, A. haemnolyticus and E. coli were observed when tetraacyl or hexaacyl lipid A were tested, since all transferases acted more efficiently on the former. With limiting amounts of acceptor, all Kdo transferases were able to transfer a third Kdo residue with varying efficiency.

Acinetobacter↗
PubMed1984

Regulation of glutathione transferase and DT-diaphorase mRNAs in persistent hepatocyte nodules during chemical hepatocarcinogenesis.

We have utilized cDNA probes and in vitro translation analysis to quantitate the levels of rat liver glutathione transferase (glutathione S-aralkyltransferase; RX:glutathione R-transferase, EC 2.5.1.18) and DT-diaphorase [NAD-(P)H:quinone-acceptor oxidoreductase, EC 1.6.99.2] mRNAs in persistent hepatocyte nodules induced by chemical carcinogens. Our results indicate that within the nodules, glutathione transferase mRNAs specific for the Ya/Yc and Yb subunits are increased 3-fold and 5-fold, respectively, over the levels observed in normal liver or in the liver tissue surrounding the nodules. Similarly, the level of DT-diaphorase mRNA is increased 5- to 7-fold within the nodules as compared to surrounding liver tissue or normal liver. When animals were administered 3-methylcholanthrene, a typical inducer of these mRNAs in normal animals, a further increase in the glutathione transferase Yb mRNA(s) and DT-diaphorase mRNA was observed in the nodules; however, the Ya/Yc mRNA levels remained unaffected. Our data indicate that during chemically induced neoplastic transformation, the mRNA levels for the Yb subunit of glutathione transferase and DT-diaphorase are increased in the nodules but still retain the capacity to be regulated by 3-methylcholanthrene. Although the glutathione transferase Ya/Yc mRNAs are also increased in the nodules, they lost their ability to be regulated by 3-methylcholanthrene. These latter data suggest that within the nodules there is a specific defect in the regulatory mechanism(s) that leads to an induction of the Ya/Yc mRNAs in normal tissue by xenobiotics.

2-Acetylaminofluorene↗
PubMed1997

Structural and functional aspects of rat microsomal glutathione transferase. The roles of cysteine 49, arginine 107, lysine 67, histidine, and tyrosine residues.

Rat liver microsomal glutathione transferase is rapidly inactivated upon treatment with the arginine-selective reagent phenylglyoxal or the lysine-selective 1,3,5-trinitrobenzenesulfonate. Glutathione sulfonate, an inhibitor of the enzyme, gives nearly complete protection against inactivation and prevents modification, indicating that these residues form part of or reside close to the active site. Sequence analysis of peptides from peptic and tryptic digests of [7-14C]phenylglyoxal- and 1,3,5-trinitrobenzenesulfonate-treated microsomal glutathione transferase indicated arginine 107 and lysine 67 as the sites of modification. A set of mutant forms of microsomal glutathione transferase was constructed by site-directed mutagenesis and heterologously expressed in Escherichia coli BL21(DE3). Arginine 107 was exchanged for alanine and lysine residues. The alanine mutant (R107A) exhibited an activity and inhibition profile similar to that of the wild type enzyme but displayed a decreased thermostability. Thus, arginine 107 does not appear to participate in catalysis or substrate binding; instead, an important structural role is suggested for this residue. Lysine 67 was mutated to alanine and arginine with no effect on activity. All three histidines were replaced by glutamine, and the resulting mutant proteins had activities comparable with that of the wild type. It can thus be concluded that the chemical modification experiments indicating that arginine 107, lysine 67, and one of the histidines partake in catalysis can be disproved. However, protection from modification by a competitive inhibitor indicates that these residues could be close to the glutathione binding site. All tyrosine to phenylalanine substitutions resulted in mutants with activities similar to that of the wild type. Interestingly, the exchange of tyrosine 137 appears to result in activation of the enzyme. Thus, the microsomal glutathione transferase must display an alternate stabilization of the thiolate anion of glutathione other than through interaction with the phenolic hydroxyl group of a tyrosine residue. Substitution of cysteine 49 with alanine resulted in a semiactivated mutant enzyme with enzymatic properties partly resembling the activated form of microsomal glutathione transferase. The function of this mutant was not altered upon reaction with N-ethylmaleimide, and cysteine 49 is thus demonstrated as the site of modification that results in activation of microsomal glutathione transferase.

Amino Acid Sequence↗
PubMed2000

Farnesyl transferase inhibitors block the farnesylation of CENP-E and CENP-F and alter the association of CENP-E with the microtubules.

Human tumor cell lines that are sensitive to the effects of farnesyl transferase inhibitors accumulate in G(2) --> M (except for cells with an activated Ha-ras that accumulate in G(1)). A search for CAAX box proteins from Swiss-Prot revealed more than 300 peptides. Of these, the centromeric proteins CENP-E and CENP-F are preferentially expressed during mitosis and are implicated as mediators of the G(2) --> M checkpoint. Experiments performed here show that peptides from the COOH-terminal CAAX box of CENP-E and CENP-F are substrates for farnesyl transferase but not geranylgeranyl transferase-I. Although both proteins are prenylated in the human tumor cell line DLD-1, their prenylation is completely inhibited by the farnesyl transferase inhibitor, SCH 66336. Immunohistochemical data with the lung carcinoma cell line, A549, showed that preventing the farnesylation of CENP-E and CENP-F by treatment with the farnesyl transferase inhibitor SCH 66336 does not affect their localization to the kinetochores. However, the presence of farnesyl transferase inhibitors alters the association between CENP-E and the microtubules. Our results imply that the inhibition of CENP-E farnesylation results in the alteration of the microtubule-centromere interaction during mitosis and results in the accumulation of cells prior to metaphase.

Alkyl and Aryl Transferases↗
PubMed2003

Identification of regulatory domains in ADP-ribosyltransferase-1 that determine transferase and NAD glycohydrolase activities.

Mono-ADP-ribosyltransferases (ART1-7) transfer ADP-ribose from NAD+ to proteins (transferase activity) or water (NAD glycohydrolase activity). The mature proteins contain two domains, an alpha-helical amino terminus and a beta-sheet-rich carboxyl terminus. A basic region in the carboxyl termini is encoded in a separate exon in ART1 and ART5. Structural motifs are conserved among ART molecules. Successive amino- or carboxyl-terminal truncations of ART1, an arginine-specific transferase, identified regions that regulated transferase and NAD glycohydrolase activities. In mouse ART1, amino acids 24-38 (ART-specific extension) were needed to inhibit both activities; amino acids 39-45 (common ART coil) were required for both. Successive truncations of the alpha-helical region reduced transferase and NAD glycohydrolase activities; however, truncation to residue 106 enhanced both. Removal of the carboxyl-terminal basic domain decreased transferase, but enhanced NAD glycohydrolase, activity. Thus, amino- and carboxyl-terminal regions of ART1 are required for transferase activity. The enhanced glycohydrolase activity of the shorter mutants indicates that sequences, which are not part of the NAD binding, core catalytic site, exert structural constraints, modulating substrate specificity and catalytic activity. These functional domains, defined by discrete exons or structural motifs, are found in ART1 and other ARTs, consistent with conservation of structure and function across the ART family.

ADP Ribose Transferases↗
PubMed2004

Trypanothione S-transferase activity in a trypanosomatid ribosomal elongation factor 1B.

Trypanothione is a thiol unique to the Kinetoplastida and has been shown to be a vital component of their antioxidant defenses. However, little is known as to the role of trypanothione in xenobiotic metabolism. A trypanothione S-transferase activity was detected in extracts of Leishmania major, L. infantum, L. tarentolae, Trypanosoma brucei, and Crithidia fasciculata, but not Trypanosoma cruzi. No glutathione S-transferase activity was detected in any of these parasites. Trypanothione S-transferase was purified from C. fasciculata and shown to be a hexadecameric complex of three subunits with a relative molecular weight of 650,000. This enzyme complex was specific for the thiols trypanothione and glutathionylspermidine and only used 1-chloro-2,4-dinitrobenzene from a range of glutathione S-transferase substrates. Peptide sequencing revealed that the three components were the alpha, beta, and gamma subunits of ribosomal eukaryotic elongation factor 1B (eEF1B). Partial dissociation of the complex suggested that the S-transferase activity was associated with the gamma subunit. Moreover, Cibacron blue was found to be a tight binding inhibitor and reactive blue 4 an irreversible time-dependent inhibitor that covalently modified only the gamma subunit. The rate of inactivation by reactive blue 4 was increased more than 600-fold in the presence of trypanothione, and Cibacron blue protected the enzyme from inactivation by 1-chloro-2,4-dinitrobenzene, confirming that these dyes interact with the active site region. Two eEF1Bgamma genes were cloned from C. fasciculata, but recombinant C. fasciculata eEF1Bgamma had no S-transferase activity, suggesting that eEF1Bgamma is unstable in the absence of the other subunits.

Amino Acid Sequence↗
PubMed2004

Kinetic and mechanistic characterization of the formyl-CoA transferase from Oxalobacter formigenes.

Oxalobacter formigenes is an obligate anaerobe that colonizes the human gastrointestinal tract and employs oxalate breakdown to generate ATP in a novel process involving the interplay of two coupled enzymes and a membrane-bound oxalate:formate antiporter. Formyl-CoA transferase is a critical enzyme in oxalate-dependent ATP synthesis and is the first Class III CoA-transferase for which a high resolution, three-dimensional structure has been determined (Ricagno, S., Jonsson, S., Richards, N., and Lindqvist, Y. (2003) EMBO J. 22, 3210-3219). We now report the first detailed kinetic characterizations of recombinant, wild type formyl-CoA transferase and a number of site-specific mutants, which suggest that catalysis proceeds via a series of anhydride intermediates. Further evidence for this mechanistic proposal is provided by the x-ray crystallographic observation of an acylenzyme intermediate that is formed when formyl-CoA transferase is incubated with oxalyl-CoA. The catalytic mechanism of formyl-CoA transferase is therefore established and is almost certainly employed by all other members of the Class III CoA-transferase family.

Bacterial Proteins↗
PubMed2005

Crystallographic trapping of the glutamyl-CoA thioester intermediate of family I CoA transferases.

Coenzyme A transferases are involved in a broad range of biochemical processes in both prokaryotes and eukaryotes, and exhibit a diverse range of substrate specificities. The YdiF protein from Escherichia coli O157:H7 is an acyl-CoA transferase of unknown physiological function, and belongs to a large sequence family of CoA transferases, present in bacteria to humans, which utilize oxoacids as acceptors. In vitro measurements showed that YdiF displays enzymatic activity with short-chain acyl-CoAs. The crystal structures of YdiF and its complex with CoA, the first co-crystal structure for any Family I CoA transferase, have been determined and refined at 1.9 and 2.0 A resolution, respectively. YdiF is organized into tetramers, with each monomer having an open alpha/beta structure characteristic of Family I CoA transferases. Co-crystallization of YdiF with a variety of CoA thioesters in the absence of acceptor carboxylic acid resulted in trapping a covalent gamma-glutamyl-CoA thioester intermediate. The CoA binds within a well defined pocket at the N- and C-terminal domain interface, but makes contact only with the C-terminal domain. The structure of the YdiF complex provides a basis for understanding the different catalytic steps in the reaction of Family I CoA transferases.

Binding Sites↗
PubMed1995

Human leukocyte glutathione S-transferase isozyme (class mu) and susceptibility to smoking-related cancers.

Glutathione S-transferase isozyme class mu from human leukocytes has been shown to be dominantly inherited and can be determined by activity measurement directed toward the substrate trans-stilbene oxide. The activity distribution of leukocyte glutathione S-transferase class mu was determined from control healthy nonsmokers, smokers, and smoking-related cancer patients. In a control healthy nonsmoker population, 54% (n = 50) of the subjects showed high levels of glutathione S-transferase class mu activity. In patients with cancers known to be related to smoking, 46% (n = 50) showed higher levels of glutathione S-transferase class mu. Noncancer smokers matched for age and smoking history with cancer patients showed an increased likelihood of having glutathione S-transferase (GST) class mu activity (76%). These results suggest that GST mu may be a cancer susceptibility marker in the case of smokers. In rats, benzo[a]pyrene (1 mg/kg, ip) administration daily for 3 d produced a significant increase in liver glutathione S-transferase class mu. Although these induction studies in experimental animals may not be relevant to humans, there is a possibility that, as in rats, this enzyme may be inducible in humans by polycyclic aromatic hydrocarbons.

Adult↗
PubMed1990

Formation of the DNA adduct S-[2-(N7-guanyl)ethyl]glutathione from ethylene dibromide: effects of modulation of glutathione and glutathione S-transferase levels and lack of a role for sulfation.

Hepatic S-[2-(N7-guanyl)ethyl]glutathione DNA adducts were determined in several strains of rats and mice after i.p. injection of a dose of 37 mg ethylene dibromide/kg body wt. More adducts were formed in rats than in mice, while no difference was noted among strains within each species. Removal of adducts in liver DNA was relatively slow in all animals tested. On the contrary, in vitro incubation of calf thymus DNA with ethylene dibromide and either rat cytosol or mouse cytosol gave rise to similar amounts of adduct, yet mouse cytosol showed much higher glutathione (GSH) S-transferase activity toward 1-chloro-2,4-dinitrobenzene. Human cytosol also activated ethylene dibromide, with the extent of conjugation being approximately half that of rat cytosol. Pretreatment of rats with phenobarbital or beta-naphthoflavone induced GSH S-transferases but did not increase the in vivo formation of DNA adducts, suggesting that concomitant induction of cytochrome P450 might abolish the effect of induction of GSH S-transferase by increasing the oxidation of ethylene dibromide. Butylated hydroxytoluene induced GSH S-transferase and also markedly increased DNA adduct levels. Disulfiram, a known cytochrome P450 inhibitor, significantly increased the formation of DNA adducts whereas it did not affect GSH S-transferase activity. Depletion of GSH by pretreatment of rats with diethylmaleate or buthionine sulfoximine resulted in decreased in vivo DNA adduct levels and the degree of reduction was well correlated with the extent of GSH depletion. In vitro incubation of tritiated S-(2-hydroxyethyl)GSH with calf thymus DNA in the presence of 3'-phosphoadenosine-5'-phosphosulfate and rat liver cytosol did not result in significant binding to DNA, suggesting that sulfation of the alcohol does not readily occur to add a leaving group and regenerate an episulfonium ion. These results suggest that induction of the Phase II enzyme GSH S-transferase can be detrimental in the case of ethylene dibromide and that decreases in GSH levels reduce DNA alkylation in rats.

Animals↗
PubMed1996

Expression of cytochrome P450s and glutathione S-transferases in human esophagus with squamous-cell carcinomas.

In order to clarify the expression of cytochrome P450 and glutathione S-transferase in human esophagus, 41 samples of human esophagus with squamous-cell carcinoma were investigated by immunoblot analysis and enzyme assays. Cytochrome P450 1A2/1 was clearly expressed in microsomes, and the amount in samples with tumorous tissue was significantly greater than that in samples without tumourous tissues or in liver; cytochrome P450 2B6 and 3A4/3 were expressed polymorphically. Aryl hydrocarbon hydroxylase activity was detected in microsomes and was greater in samples from smokers than non-smokers. Patients who both smoked and drank alcohol, however, had activity similar to that of patients without these habits. Glutathione S-transferase M1 and A1/2 protein existed polymorphically in cytosol, and glutathione S-transferase P1-1 was detected in all samples. The frequency of expression of the glutathione S-transferase A1/2 protein was greater in patients with M1 protein than in those without; no difference in the expression was seen for glutathione S-transferase P1-1. Neither smoking nor drinking influenced the expression or activity of glutathione S-transferase. Our data support the idea that some carcinogens can be directly activated or inactivated in human esophageal epithelium.

Alcohol Drinking↗