Developmental aspects of glutathione S-transferase B (ligandin) in rat liver.
Ontology highlight
ABSTRACT:
SUBMITTER: Hales BF
PROVIDER: S-EPMC1164227 | biostudies-other | 1976 Nov
REPOSITORIES: biostudies-other
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Project description:The unique glutathione S-transferase (GST) subunit Yx, which is undetectable in normal adult rat liver cytosol, was shown to occur in the liver cytosol of rats with hereditary hyperbilirubinuria (EHB). The Yx subunit is a member of the Alpha-class GST subunits, and is immunologically closely related to the Yc subunit. The Yx subunit has an apparent M(r) of 26,400, different from those of Ya (M(r) 25,800), Yb1 and Yb2 (both M(r) 27,200) and Yc (M(r) 28,400). During postnatal development in livers of EHB rats, the Yx subunit concentration in either sex was highest during the first week post partum and declined rapidly with age. Although the concentration of subunit Yx at 8 weeks of age accounted for about 60% in females and 40% in males of that observed in 1-week-old 'neonatal' male EHB rats, concentrations in females thereafter increased gradually to almost the neonatal level and remained at this high level at least up to 37 weeks of age, whereas the concentration in males did not increase again. Thus the post-pubertal Yx subunit concentration was 2-fold higher in females than in males. In contrast, in normal Sprague-Dawley rat liver, the Yfetus subunit, with the same M(r) as the Yx subunit, had the highest concentration in 10-day-old animals, declined rapidly thereafter, and was not detectable in the post-pubertal period. The Yfetus subunit was also immunoreactive with an antibody against GST YcYc. The analysis of GST subunits by reverse-phase h.p.l.c. revealed that the Yx subunit was eluted at a retention time different from other known subunits, but coincided with that of Yfetus. The N-terminal amino acid sequence of the Yx subunit displayed a high degree of sequence similarity to that of the Yfetus subunit. These data suggest that the Yx subunit in EHB rats may be very similar to, if not identical with, the Yfetus subunit.
| S-EPMC1131029 | biostudies-other
Project description:Glutathione S-transferases (GSTs) form a superfamily of multifunctional proteins with essential roles in cellular detoxification processes. A new fungal specific class of GST has been highlighted by genomic approaches. The biochemical and structural characterization of one isoform of this class in Phanerochaete chrysosporium revealed original properties. The three-dimensional structure showed a new dimerization mode and specific features by comparison with the canonical GST structure. An additional ?-hairpin motif in the N-terminal domain prevents the formation of the regular GST dimer and acts as a lid, which closes upon glutathione binding. Moreover, this isoform is the first described GST that contains all secondary structural elements, including helix ?4' in the C-terminal domain, of the presumed common ancestor of cytosolic GSTs (i.e. glutaredoxin 2). A sulfate binding site has been identified close to the glutathione binding site and allows the binding of 8-anilino-1-naphtalene sulfonic acid. Competition experiments between 8-anilino-1-naphtalene sulfonic acid, which has fluorescent properties, and various molecules showed that this GST binds glutathionylated and sulfated compounds but also wood extractive molecules, such as vanillin, chloronitrobenzoic acid, hydroxyacetophenone, catechins, and aldehydes, in the glutathione pocket. This enzyme could thus function as a classical GST through the addition of glutathione mainly to phenethyl isothiocyanate, but alternatively and in a competitive way, it could also act as a ligandin of wood extractive compounds. These new structural and functional properties lead us to propose that this GST belongs to a new class that we name GSTFuA, for fungal specific GST class A.
| S-EPMC3493941 | biostudies-literature
Project description:Glutathione transferases (GSTs) are the main detoxification enzymes in schistosomes. These parasitic enzymes tend to be upregulated during drug treatment, with Schistosoma haematobium being one of the species that mainly affect humans. There is a lack of complete sequence information on the closely related bovis and haematobium 26-kDa GST isoforms in any database. Consequently, we engineered a pseudo-26-kDa S. bovis/haematobium GST (Sbh26GST) to understand structure-function relations and ligandin activity towards selected potential ligands. Sbh26GST was overexpressed in Escherichia coli as an MBP-fusion protein, purified to homogeneity and catalyzed 1-chloro-2,4-dinitrobenzene-glutathione (CDNB-GSH) conjugation activity, with a specific activity of 13 μmol/min/mg. This activity decreased by ~95% in the presence of bromosulfophthalein (BSP), which showed an IC50 of 27 µM. Additionally, enzyme kinetics revealed that BSP acts as a non-competitive inhibitor relative to GSH. Spectroscopic studies affirmed that Sbh26GST adopts the canonical GST structure, which is predominantly α-helical. Further extrinsic 8-anilino-1-naphthalenesulfonate (ANS) spectroscopy illustrated that BSP, praziquantel (PZQ), and artemisinin (ART) might preferentially bind at the dimer interface or in proximity to the hydrophobic substrate-binding site of the enzyme. The Sbh26GST-BSP interaction is both enthalpically and entropically driven, with a stoichiometry of one BSP molecule per Sbh26GST dimer. Enzyme stability appeared enhanced in the presence of BSP and GSH. Induced fit ligand docking affirmed the spectroscopic, thermodynamic, and molecular modelling results. In conclusion, BSP is a potent inhibitor of Sbh26GST and could potentially be rationalized as a treatment for schistosomiasis.
| S-EPMC8699318 | biostudies-literature
Project description:CM-cellulose chromatography of rat liver and kidney cytosol at pH6 reveals the presence of a second Ya-subunit dimer of glutathione S-transferase (GST-F) in addition to the recently described GST-YaYa (GST-L; our nomenclature) [Hayes & Clarkson (1982) Biochem. J. 207, 459-470]. The two forms are structurally similar (by the criteria of CNBr- and Staphylococcus-V8-proteinase-cleavage peptide maps), and both are sensitive to inhibition by haemin. However, their kinetic parameters with 1-chloro-2,4-dinitrobenzene are quite distinct, and they show differential inducibility by phenobarbitone. These results suggest a similar heterogeneity in Ya-subunits to that previously described for Yb-subunits of glutathione S-transferase and indicate that significant gene duplication may have occurred in these multifunctional intracellular binding proteins.
| S-EPMC1153420 | biostudies-other
Project description:production of glutathione s-transferase from Lactobacillus plantarum Ku720558
| PRJEB33504 | ENA
Project description:The ontogeny of rat liver glutathione S-transferase (EC 2.5.1.18) (GSTs) during foetal and postnatal development was investigated. The GSTs are dimers, the subunits of which belong to three multigene families, Alpha (subunits 1, 2, 8 and 10), Mu (subunits 3, 4, 6, 9 and 11) and Pi (subunit 7) [Mannervik, Alin, Guthenberg, Jennsson, Tahir, Warholm & Jörnvall (1985) Proc. Natl. Acad. Sci. U.S.A. 82, 7202-7206; Kispert, Meyer, Lalor, Coles & Ketterer (1989) Biochem. J. 260, 789-793]. There is considerable structural homology within each gene family, with the result that whereas reverse-phase h.p.l.c. successfully differentiates individual subunits, immunocytochemical and Northern-blotting analyses may only differentiate families. Enzymic activity, h.p.l.c. and Northern blotting indicated that expression of GST increased from very low levels at 12 days of foetal growth to substantial amounts at day 21. At birth, GST concentrations underwent a dramatic decline and remained low until 5-10 days post partum, after which they increased to adult levels. During the period under study, GST subunits underwent differential expression. The Mu family had a lower level of expression than the Alpha family, and, within the Alpha family, subunit 1 was more dominant in the adult than the foetus. Subunit 2 is the major form in the foetus. Most noteworthy were subunits 7 and 10, which were prominent in the foetus, but present at low levels post partum. Immunocytochemical analysis of the 17-day foetal and newborn rat livers showed marked differences in the distribution of GSTs in hepatocytes. In the 17-day foetal liver Pi greater than Alpha greater than Mu whereas in the newborns Alpha greater than Mu much greater than Pi. Erythropoietic cells were not stained for any of the three GST families. Steady-state mRNA concentrations in the foetus correlated with the relative transcription of the Alpha, Mu and Pi class genes. However, in those genes expressed post partum, namely the Alpha and Mu class, low transcriptional activity was associated with high concentrations of mRNA. This suggests that there is a switch from transcriptional control to post-transcriptional control at birth. GST 7-7 appears to be regulated predominantly by transcription throughout the period of liver development under observation.
| S-EPMC1130909 | biostudies-other
Project description:A hitherto unknown cytosolic glutathione S-transferase from rat liver was discovered and a method developed for its purification to apparent homogeneity. This enzyme had several properties that distinguished it from other glutathione S-transferases, and it was named glutathione S-transferase X. The purification procedure involved DEAE-cellulose chromatography, (NH4)2SO4 precipitation, affinity chromatography on Sepharose 4B to which glutathione was coupled and CM-cellulose chromatography, and allowed the isolation of glutathione S-transferases X, A, B and C in relatively large quantities suitable for the investigation of the toxicological role of these enzymes. Like glutathione S-transferase M, but unlike glutathione S-transferases AA, A, B, C, D and E, glutathione S-transferase X was retained on DEAE-cellulose. The end product, which was purified from rat liver 20 000 g supernatant about 50-fold, as determined with 1-chloro-2,4-dinitrobenzene as substrate and about 90-fold with the 1,2-dichloro-4-nitrobenzene as substrate, was judged to be homogeneous by several criteria, including sodium dodecyl sulphate/polyacrylamide-gel electrophoresis, isoelectric focusing and immunoelectrophoresis. Results from sodium dodecyl sulphate/polyacrylamide-gel electrophoresis and gel filtration indicated that transferase X was a dimer with Mr about 45 000 composed of subunits with Mr 23 500. The isoelectric point of glutathione S-transferase X was 6.9, which is different from those of most of the other glutathione S-transferases (AA, A, B and C). The amino acid composition of transferase X was similar to that of transferase C. Immunoelectrophoresis of glutathione S-transferases A, C and X and precipitation of various combinations of these antigens by antisera raised against glutathione S-transferase X or C revealed that the glutathione S-transferases A, C and X have different electrophoretic mobilities, and indicated that transferase X is immunologically similar to transferase C, less similar to transferase A and not cross-reactive to transferases B and E. In contrast with transferases B and AA, glutathione S-transferase X did not bind cholic acid, which, together with the determination of the Mr, shows that it does not possess subunits Ya or Yc. Glutathione S-transferase X did not catalyse the reaction of menaphthyl sulphate with glutathione, and was in this respect dissimilar to glutathione S-transferase M; however, it conjugated 1,2-dichloro-4-nitrobenzene very rapidly, in contrast with transferases AA, B, D and E, which were nearly inactive towards that substrate.(ABSTRACT TRUNCATED AT 400 WORDS)
| S-EPMC1152444 | biostudies-other
Project description:The activity of rat liver microsomal glutathione transferase is increased by limited tryptic proteolysis; the membrane-bound and purified forms of the enzyme are activated about 5- and 10-fold respectively. The cleavage sites that correlate with this activation were determined by amino acid sequence analysis to be located after Lys-4 and Lys-41. Differences in the relative extent of cleavage at these two sites did not consistently affect the degree of activation. Thus the data support the conclusion that cleavage at either site results in activation. The trypsin-activated enzyme was compared with the form activated with N-ethylmaleimide, which modifies Cys-49. These two differently activated forms were found to have similar kinetic parameters, which differ from those of the unactivated enzyme. The relatedness of the two types of activation is also demonstrated by the observation that microsomal glutathione transferase fully activated by N-ethylmaleimide is virtually resistant to further activation by trypsin. This is the case despite the fact that the N-ethylmaleimide-activated enzyme is much more susceptible to trypsin cleavage at Lys-41 than is the untreated enzyme. The latter observation indicates that activation with N-ethylmaleimide is accompanied by a conformational change involving Lys-41.
| S-EPMC1138707 | biostudies-other
Project description:Liver fluke infection of livestock causes economic losses of over US$ 3 billion worldwide per annum. The disease is increasing in livestock worldwide and is a re-emerging human disease. There are currently no commercial vaccines, and only one drug with significant efficacy against adult worms and juveniles. A liver fluke vaccine is deemed essential as short-lived chemotherapy, which is prone to resistance, is an unsustainable option in both developed and developing countries. Protein superfamilies have provided a number of leading liver fluke vaccine candidates. A new form of glutathione transferase (GST) family, Sigma class GST, closely related to a leading Schistosome vaccine candidate (Sm28), has previously been revealed by proteomics in the liver fluke but not functionally characterised.In this manuscript we show that a purified recombinant form of the F. hepatica Sigma class GST possesses prostaglandin synthase activity and influences activity of host immune cells. Immunocytochemistry and western blotting have shown the protein is present near the surface of the fluke and expressed in eggs and newly excysted juveniles, and present in the excretory/secretory fraction of adults. We have assessed the potential to use F. hepatica Sigma class GST as a vaccine in a goat-based vaccine trial. No significant reduction of worm burden was found but we show significant reduction in the pathology normally associated with liver fluke infection.We have shown that F. hepatica Sigma class GST has likely multi-functional roles in the host-parasite interaction from general detoxification and bile acid sequestration to PGD synthase activity.
| S-EPMC3362645 | biostudies-literature
Project description:Dichloroacetate (DCA) has potential for treating mitochondrial disorders and cancer by activating the mitochondrial pyruvate dehydrogenase complex. Repeated dosing of DCA results in reduced drug clearance due to inactivation of glutathione transferase ζ1 (GSTZ1), its metabolizing enzyme. We investigated the time-course of inactivation of GSTZ1 in hepatic cytosol and mitochondria after one oral dose of 100 mg/kg DCA to female Sprague-Dawley rats aged 4 weeks (young) and 52 weeks (adult) as models for children and adults, respectively. GSTZ1 activity with both DCA and an endogenous substrate, maleylacetone (MA), as well as GSTZ1 protein expression were rapidly reduced in cytosol from both ages following DCA treatment. In mitochondria, loss of GSTZ1 protein and activity with DCA were even more rapid. The cytosolic in vivo half-lives of the loss of GSTZ1 activity with DCA were 1.05 ± 0.03 and 0.82 ± 0.02 h (mean ± S.D., n = 6) for young and adult rats, respectively, with inactivation significantly more rapid in adult rats, p < 0.001. The mitochondrial inactivation half-lives were similar in young (0.57 ± 0.02 h) and adult rats (0.54 ± 0.02 h) and were significantly (p < 0.0001) shorter than cytosolic inactivation half-lives. By 24 h after DCA administration, activity and expression remained at 10% or less than control values. The in vitro GSTZ1 inactivation half-lives following incubation with 2 mM DCA in the presence of physiological chloride (Cl-) concentrations (cytosol = 44 mM, mitochondria = 1-2 mM) exhibited marked differences between subcellular fractions, being 3 times longer in the cytosol than in the mitochondria, regardless of age, suggesting that the lower Cl- concentration in mitochondria explained the faster degradation of GSTZ1. These results demonstrate for the first time that rat mitochondrial GSTZ1 is more readily inactivated by DCA than cytosolic GSTZ1, and cytosolic GSTZ1 is inactivated more rapidly in adult than young rats.
| S-EPMC8582596 | biostudies-literature