Bile acid inhibition of basic and neutral glutathione S-transferases in rat liver.
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ABSTRACT:
SUBMITTER: Hayes JD
PROVIDER: S-EPMC1152439 | biostudies-other | 1983 Dec
REPOSITORIES: biostudies-other
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Project description:Cholic acid-binding activity in cytosol from rat livers appears to be mainly associated with enzymes having glutathione S-transferase activity; at least four of the enzymes in this group can bind the bile acid. Examination of the subunit compositions of different glutathione S-transferases indicated that cholic acid binding and the ability to conjugate reduced glutathione with 1,2-dichloro-4-nitrobenzene may be ascribed to different subunits.
| S-EPMC1161272 | biostudies-other
Project description:A purification scheme was devised that resulted in the resolution of a number of basic glutathione S-transferases from rat liver, three of which contained two subunits of molecular mass 23500 Da (i.e. Yb monomers). These were identified as transferases D, C and A by their elution positions from CM-cellulose and their specific activities towards a variety of substrates. Hybridization, immunotitration and peptide 'mapping' experiments demonstrated that transferases D, C and A comprise Yb2Yb2, Yb1Yb2 and Yb1Yb1 subunits.
| S-EPMC1152177 | biostudies-other
Project description:The olfactory epithelium is continuously exposed to exogenous chemicals, including odorants. During the past decade, the enzymes surrounding the olfactory receptors have been shown to make an important contribution to the process of olfaction. Mammalian xenobiotic metabolizing enzymes, such as cytochrome P450, esterases and glutathione transferases (GSTs), have been shown to participate in odorant clearance from the olfactory receptor environment, consequently contributing to the maintenance of sensitivity toward odorants. GSTs have previously been shown to be involved in numerous physiological processes, including detoxification, steroid hormone biosynthesis, and amino acid catabolism. These enzymes ensure either the capture or the glutathione conjugation of a large number of ligands. Using a multi-technique approach (proteomic, immunocytochemistry and activity assays), our results indicate that GSTs play an important role in the rat olfactory process. First, proteomic analysis demonstrated the presence of different putative odorant metabolizing enzymes, including different GSTs, in the rat nasal mucus. Second, GST expression was investigated in situ in rat olfactory tissues using immunohistochemical methods. Third, the activity of the main GST (GSTM2) odorant was studied with in vitro experiments. Recombinant GSTM2 was used to screen a set of odorants and characterize the nature of its interaction with the odorants. Our results support a significant role of GSTs in the modulation of odorant availability for receptors in the peripheral olfactory process.
| S-EPMC6656353 | biostudies-literature
Project description:The 55 Arabidopsis glutathione transferases (GSTs) are, with one microsomal exception, a monophyletic group of soluble enzymes that can be divided into phi, tau, theta, zeta, lambda, dehydroascorbate reductase (DHAR) and TCHQD classes. The populous phi and tau classes are often highly stress inducible and regularly crop up in proteomic and transcriptomic studies. Despite much study on their xenobiotic-detoxifying activities their natural roles are unclear, although roles in defence-related secondary metabolism are likely. The smaller DHAR and lambda classes are likely glutathione-dependent reductases, the zeta class functions in tyrosine catabolism and the theta class has a putative role in detoxifying oxidised lipids. This review describes the evidence for the functional roles of GSTs and the potential for these enzymes to perform diverse functions that in many cases are not "glutathione transferase" activities. As well as biochemical data, expression data from proteomic and transcriptomic studies are included, along with subcellular localisation experiments and the results of functional genomic studies.
| S-EPMC3244946 | biostudies-literature
Project description:SUMMARY:The soluble glutathione transferases (GSTs, EC 2.5.1.18) are encoded by a large and diverse gene family in plants, which can be divided on the basis of sequence identity into the phi, tau, theta, zeta and lambda classes. The theta and zeta GSTs have counterparts in animals but the other classes are plant-specific and form the focus of this article. The genome of Arabidopsis thaliana contains 48 GST genes, with the tau and phi classes being the most numerous. The GST proteins have evolved by gene duplication to perform a range of functional roles using the tripeptide glutathione (GSH) as a cosubstrate or coenzyme. GSTs are predominantly expressed in the cytosol, where their GSH-dependent catalytic functions include the conjugation and resulting detoxification of herbicides, the reduction of organic hydroperoxides formed during oxidative stress and the isomerization of maleylacetoacetate to fumarylacetoacetate, a key step in the catabolism of tyrosine. GSTs also have non-catalytic roles, binding flavonoid natural products in the cytosol prior to their deposition in the vacuole. Recent studies have also implicated GSTs as components of ultraviolet-inducible cell signaling pathways and as potential regulators of apoptosis. Although sequence diversification has produced GSTs with multiple functions, the structure of these proteins has been highly conserved. The GSTs thus represent an excellent example of how protein families can diversify to fulfill multiple functions while conserving form and structure.
| S-EPMC139027 | biostudies-literature
Project description:Glutathione S-transferases are a complex family of dimeric proteins that play a dual role in cellular detoxification; they catalyse the first step in the synthesis of mercapturic acids, and they bind potentially harmful non-substrate ligands. Bile acids are quantitatively the major group of ligands encountered by the glutathione S-transferases. The enzymes from rat liver comprise Yk (Mr 25 000), Ya (Mr 25 500), Yn (Mr 26 500), Yb1, Yb2 (both Mr 27 000) and Yc (Mr 28 500) monomers. Although bile acids inhibited the catalytic activity of all transferases studied, the concentration of a particular bile acid required to produce 50% inhibition (I50) varies considerably. A comparison of the I50 values obtained with lithocholate (monohydroxylated), chenodeoxycholate (dihydroxylated) and cholate (trihydroxylated) showed that, in contrast with all other transferase monomers, the Ya subunit possesses a relatively hydrophobic bile-acid-binding site. The I50 values obtained with lithocholate and lithocholate 3-sulphate showed that only the Ya subunit is inhibited more effectively by lithocholate than by its sulphate ester. Other subunits (Yk, Yn, Yb1 and Yb2) were inhibited more by lithocholate 3-sulphate than by lithocholate, indicating the existence of a significant ionic interaction, in the bile-acid-binding domain, between (an) amino acid residue(s) and the steroid ring A. By contrast, increasing the assay pH from 6.0 to 7.5 decreased the inhibitory effect of all bile acids studied, suggesting that there is little significant ionic interaction between transferase subunits and the carboxy group of bile acids. Under alkaline conditions, low concentrations (sub-micellar) of nonsulphated bile acids activated Yb1, Yb2 and Yc subunits but not Yk, Ya and Yn subunits. The diverse effects of the various bile acids studied on transferase activity enables these ligands to be used to help establish the quaternary structure of individual enzymes. Since these inhibitors can discriminate between transferases that appear to be immunochemically identical (e.g. transferases F and L), bile acids can provide information about the subunit composition of forms that cannot otherwise be distinguished.
| S-EPMC1153041 | biostudies-other
Project description:The present study characterized conserved residues in a GST (glutathione transferase) in the active-site region that interacts with glutathione. This region of the active site is near the glycine moiety of glutathione and consists of a hydrogen bond network. In the GSTD (Delta class GST) studied, adGSTD4-4, the network consisted of His(38), Met(39), Asn(47), Gln(49), His(50) and Cys(51). In addition to contributing to glutathione binding, this region also had major effects on enzyme catalysis, as shown by changes in kinetic parameters and substrate-specific activity. The results also suggest that the electron distribution of this network plays a role in stabilization of the ionized thiol of glutathione as well as impacting on the catalytic rate-limiting step. This area constitutes a second glutathione active-site network involved in glutathione ionization distinct from a network previously observed interacting with the glutamyl end of glutathione. This second network also appears to be functionally conserved in GSTs. In the present study, His(50) is the key basic residue stabilized by this network, as shown by up to a 300-fold decrease in k(cat) and 5200-fold decrease in k(cat)/K(m) for glutathione. Although these network residues have a minor role in structural integrity, the replaced residues induced changes in active-site topography as well as generating positive co-operativity towards glutathione. Moreover, this network at the glycine moiety of GSH (glutathione) also contributed to the 'base-assisted deprotonation model' for GSH ionization. Taken together, the results indicate a critical role for the functionally conserved basic residue His(50) and this hydrogen bond network in the active site.
| S-EPMC1948969 | biostudies-literature
Project description:The glutathione S-transferases that were purified to homogeneity from liver cytosol have overlapping but distinct substrate specificities and different isoelectric points. This report explores the possibility of using preparative electrofocusing to compare the composition of the transferases in liver and kidney cytosol. Hepatic cytosol from adult male Sprague-Dawley rats was resolved by isoelectric focusing on Sephadex columns into five peaks of transferase activity, each with characteristic substrate specificity. The first four peaks of transferase activity (in order of decreasing basicity) are identified as transferases AA, B, A and C respectively, on the basis of substrate specificity, but the fifth peak (pI6.6) does not correspond to a previously described transferase. Isoelectric focusing of renal cytosol resolves only three major peaks of transferase activity, each with narrow substrate specificity. In the kidney, peak 1 (pI9.0) has most of the activity toward 1-chloro-2,4-dinitrobenzene, peak 2 (pI8.5) toward p-nitrobenzyl chloride, and peak 3 (pI7.0) toward trans-4-phenylbut-3-en-2-one. Renal transferase peak 1 (pI9.0) appears to correspond to transferase B on the basis of pI, substrate specificity and antigenicity. Kidney transferase peaks 2 (pI8.5) and 3 (pI7.0) do not correspond to previously described glutathione S-transferases, although kidney transferase peak 3 is similar to the transferase peak 5 from focused hepatic cytosol. Transferases A and C were not found in kidney cytosol, and transferase AA was detected in only one out of six replicates. Thus it is important to recognize the contribution of individual transferases to total transferase activity in that each transferase may be regulated independently.
| S-EPMC1186156 | biostudies-other
Project description:DNA peroxidized by exposure to ionizing radiation in the presence of oxygen is a substrate for the Se-independent GSH peroxidase activity of several GSH transferases, GSH transferases 5-5, 3-3 and 4-4 being the most active in the rat liver soluble supernatant fraction (500, 35 and 20 nmol/min per mg of protein respectively) and GSH transferases mu and pi the most active, so far found, in the human liver soluble supernatant fraction (80 and 10 nmol/min per mg respectively). Although the GSH transferase content of the rat nucleus was found to be much lower than that of the soluble supernatant, nuclear GSH transferases are likely to be more important in the detoxification of DNA hydroperoxide produced in vivo. Two nuclear fractions were studied, one extracted with 0.075 M-saline/0.025 M-EDTA, pH 8.0, and the other extracted from the residue with 8.5 M-urea. The saline/EDTA fraction contained subunits 1, 2, 3, 4 and a novel subunit, similar but not identical to 5, provisionally referred to as 5*, in the proportions 40:25:5:5:25 respectively. The 8.5 M-urea-extracted fraction contained principally subunit 5* together with a small amount of subunit 6 in the proportion 95:5 respectively. GSH transferase 5*-5* purified from the 8.5 M-urea extract has the highest activity towards DNA hydroperoxide of any GSH transferase so far studied (1.5 mumol/min per mg). A Se-dependent GSH peroxidase fraction from rat liver was also active towards DNA hydroperoxide; however, since this enzyme accounts for only 14% of the GSH peroxidase activity detectable in the nucleus, GSH transferases may be the more important source of this activity. The possible role of GSH transferases, in particular GSH transferase 5*-5*, in DNA repair is discussed.
| S-EPMC1135159 | biostudies-other
Project description:The human bile acid pool composition is composed of both primary bile acids (cholic acid and chenodeoxycholic acid) and secondary bile acids (deoxycholic acid and lithocholic acid). Secondary bile acids are formed by the 7?-dehydroxylation of primary bile acids carried out by intestinal anaerobic bacteria. We have previously described a multistep biochemical pathway in Clostridium scindens that is responsible for bile acid 7?-dehydroxylation. We have identified a large (12 kb) bile acid inducible (bai) operon in this bacterium that encodes eight genes involved in bile acid 7?-dehydroxylation. However, the function of the baiF gene product in this operon has not been elucidated. In the current study, we cloned and expressed the baiF gene in E. coli and discovered it has bile acid CoA transferase activity. In addition, we discovered a second bai operon encoding three genes. The baiK gene in this operon was expressed in E. coli and found to encode a second bile acid CoA transferase. Both bile acid CoA transferases were determined to be members of the type III family by amino acid sequence comparisons. Both bile acid CoA transferases had broad substrate specificity, except the baiK gene product, which failed to use lithocholyl-CoA as a CoA donor. Primary bile acids are ligated to CoA via an ATP-dependent mechanism during the initial steps of 7?-dehydroxylation. The bile acid CoA transferases conserve the thioester bond energy, saving the cell ATP molecules during bile acid 7?-dehydroxylation. ATP-dependent CoA ligation is likely quickly supplanted by ATP-independent CoA transfer.
| S-EPMC3243482 | biostudies-literature