scholarly journals Nonhistone protein BA is a glutathione S-transferase localized to interchromatinic regions of the cell nucleus.

1986 ◽  
Vol 102 (2) ◽  
pp. 600-609 ◽  
Author(s):  
C F Bennett ◽  
D L Spector ◽  
L C Yeoman
Keyword(s):  
Rat Liver ◽  
Cell Nucleus ◽  
Polyacrylamide Gel Electrophoresis ◽  
Rnase A ◽  
Transferase Activity ◽  
Immunoperoxidase Technique ◽  
Glutathione S Transferase ◽  
Nonhistone Protein ◽  
Glutathione S Transferases ◽  
Nuclear Domains

A DNA-binding nonhistone protein, protein BA, was previously demonstrated to co-localize with U-snRNPs within discrete nuclear domains (Bennett, F. C., and L. C. Yeoman, 1985, Exp. Cell Res., 157:379-386). To further define the association of protein BA and U-snRNPs within these discrete nuclear domains, cells were fractionated in situ and the localization of the antigens determined by double-labeled immunofluorescence. Protein BA was extracted from the nucleus with the 2.0 M NaCl soluble chromatin fraction, while U-snRNPs were only partially extracted from the 2.0 M NaCl-resistant nuclear structures. U-snRNPs were extracted from the residual nuclear material by combined DNase I/RNase A digestions. Using an indirect immunoperoxidase technique and electron microscopy, protein BA was localized to interchromatinic regions of the cell nucleus. Protein BA was noted to share a number of chemical and physical properties with a family of cytoplasmic enzymes, the glutathione S-transferases. Comparison of the published amino acid composition of protein BA and glutathione S-transferases showed marked similarities. Nonhistone protein BA isolated from saline-EDTA nuclear extracts exhibited glutathione S-transferase activity with a variety of substrates. Substrate specificity and subunit analysis by SDS polyacrylamide gel electrophoresis revealed that it was a mixture of several glutathione S-transferase isoenzymes. Protein BA isolated from rat liver chromatin was shown by immunoblotting and peptide mapping techniques to be two glutathione S-transferase isoenzymes composed of the Yb and Yb' subunits. Glutathione S-transferase Yb subunits were demonstrated to be both nuclear and cytoplasmic proteins by indirect immunolocalization on rat liver cryosections. The identification of protein BA as glutathione S-transferase suggests that this family of multifunctional enzymes may play an important role in those nuclear domains containing U-snRNPs.

scholarly journals Purification and characterization of a new cytosolic glutathione S-transferase (glutathione S-transferase X) from rat liver

1983 ◽  
Vol 215 (3) ◽  
pp. 617-625 ◽  
Author(s):  
T Friedberg ◽  
U Milbert ◽  
P Bentley ◽  
T M Guenther ◽  
F Oesch
Keyword(s):  
Rat Liver ◽  
Sodium Dodecyl Sulphate ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Deae Cellulose ◽  
Polyacrylamide Gel ◽  
Glutathione S Transferase ◽  
Cytosolic Glutathione ◽  
Glutathione S Transferases

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)

scholarly journals A sex difference in hepatic glutathione S-transferase B and the effect of hypophysectomy

1976 ◽  
Vol 160 (2) ◽  
pp. 223-229 ◽  
Author(s):  
B F Hales ◽  
A H Neims
Keyword(s):  
Sex Difference ◽  
Rat Liver ◽  
Total Protein ◽  
Specific Activity ◽  
Male Rat ◽  
Transferase Activity ◽  
Female Rat ◽  
Glutathione S Transferase ◽  
Catalytic Activities ◽  
Glutathione S Transferases

The glutathione S-transferases are a group of proteins with overlapping substrate specificities and ligand-binding capacities. This report examines certain approaches to the measurement of transferase B (ligandin) in the rat liver. The ratio of catalytic activities toward 1-chloro-2,4-dinitrobenzene and 1,2-dichloro-4-nitrobenzene gives some indication of the relative proportions of the various transferases present in 100 000 g supernatants. The fraction of catalytic activity towards 1-chloro-2,4-dinitrobenzene, due to transferase B, was best measured by immunoprecipitation with anti-(transferase B). Male rat liver exhibited three times more activity towards 1,2-dichloro-4-nitrobenzene than female tissue; however, the activities towards 1-chloro-2,4-dinitrobenzene were almost identical. By assuming a specific activity of 11 mumol/min per mg, immunoprecipitable transferase B comprised 4.5 +/- 0.2% of total protein in the 100 000 g supernatant of female rat liver, and 70% of the transferase activity towards 1-chloro-2,4-dinitrobenzene. The amount of transferase B in the 100 000 g supernatant from male rat liver is significantly lower with respect to both fraction of total protein (3.3 +/- 0.2%) and overall transferase activity towards 1-chloro-2,4-dinitrobenzene (48%). Hypophysectomy eliminated this sex difference in the hepatic concentration of glutathione S-transferase B.

scholarly journals The distribution, induction and isoenzyme profile of glutathione S-transferase and glutathione peroxidase in isolated rat liver parenchymal, Kupffer and endothelial cells

1989 ◽  
Vol 264 (3) ◽  
pp. 737-744 ◽  
Author(s):  
P Steinberg ◽  
H Schramm ◽  
L Schladt ◽  
L W Robertson ◽  
H Thomas ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Glutathione Peroxidase ◽  
Rat Liver ◽  
Peroxidase Activity ◽  
Cell Types ◽  
Transferase Activity ◽  
Glutathione Peroxidase Activity ◽  
Glutathione S Transferase ◽  
Liver Parenchymal ◽  
Parenchymal Cells

The distribution and inducibility of cytosolic glutathione S-transferase (EC 2.5.1.18) and glutathione peroxidase (EC 1.11.1.19) activities in rat liver parenchymal, Kupffer and endothelial cells were studied. In untreated rats glutathione S-transferase activity with 1-chloro-2,4-dinitrobenzene and 4-hydroxynon-2-trans-enal as substrates was 1.7-2.2-fold higher in parenchymal cells than in Kupffer and endothelial cells, whereas total, selenium-dependent and non-selenium-dependent glutathione peroxidase activities were similar in all three cell types. Glutathione S-transferase isoenzymes in parenchymal and non-parenchymal cells isolated from untreated rats were separated by chromatofocusing in an f.p.l.c. system: all glutathione S-transferase isoenzymes observed in the sinusoidal lining cells were also detected in the parenchymal cells, whereas Kupffer and endothelial cells lacked several glutathione S-transferase isoenzymes present in parenchymal cells. At 5 days after administration of Arocolor 1254 glutathione S-transferase activity was only enhanced in parenchymal cells; furthermore, selenium-dependent glutathione peroxidase activity decreased in parenchymal and non-parenchymal cells. At 13 days after a single injection of Aroclor 1254 a strong induction of glutathione S-transferase had taken place in all three cell types, whereas selenium-dependent glutathione peroxidase activity remained unchanged (endothelial cells) or was depressed (parenchymal and Kupffer cells). Hence these results clearly establish that glutathione S-transferase and glutathione peroxidase are differentially regulated in rat liver parenchymal as well as non-parenchymal cells. The presence of glutathione peroxidase and several glutathione S-transferase isoenzymes capable of detoxifying a variety of compounds in Kupffer and endothelial cells might be crucial to protect the liver from damage by potentially hepatotoxic substances.

scholarly journals The presence and longitudinal distribution of the glutathione S-transferase in rat epididymis and vas deferens

1980 ◽  
Vol 189 (1) ◽  
pp. 135-142 ◽  
Author(s):  
Barbara F. Hales ◽  
Christiane Hachey ◽  
Bernard Robaire
Keyword(s):  
Substrate Specificity ◽  
Isoelectric Focusing ◽  
Vas Deferens ◽  
Longitudinal Distribution ◽  
Transferase Activity ◽  
Glutathione S Transferase ◽  
Potential Significance ◽  
Glutathione S Transferases ◽  
Rat Epididymis ◽  
First Time

The presence of the glutathione S-transferases, enzymes that catalyse the conjugation of glutathione with a variety of compounds, is reported here, for the first time, in the mammalian epididymis–vas deferens. These glutathione S-transferases, approx. 50% of those from rat liver on a per-mg-of-protein basis, are resolved by isoelectric focusing into six peaks, each with a characteristic isoelectric point and substrate specificity. By these same criteria, the first three peaks (pI 8.9, 8.2 and 7.8) can be identified as transferases B, A and C respectively. The fifth peak (pI7.2) may correspond to transferase M; the fourth (pI7.5) and sixth (pI7.0) peaks do not correspond to previously described transferases. The distribution of transferase activity towards any one substrate studied differs in sequential sections of the epididymis and vas deferens; in addition, the longitudinal-distribution pattern differs for each of the three substrates studied. Isoelectric focusing of the cytosol fractions of the different sections further substantiates these observations. The potential significance of these enzymes and of their distribution in terms of epididymal function, maturation of spermatozoa, is discussed.

scholarly journals Inhibition by inorganic anions of glutathione S-transferases from insect and mammalian sources

1991 ◽  
Vol 278 (1) ◽  
pp. 193-198 ◽  
Author(s):  
A G Clark ◽  
J F Hamilton ◽  
S N Marshall
Keyword(s):  
Rat Liver ◽  
Galleria Mellonella ◽  
Limited Range ◽  
Inorganic Anions ◽  
Glutathione S Transferase ◽  
Inhibitory Potency ◽  
Glutathione S Transferases ◽  
Wax Moth

Glutathione S-transferases 1-1, 3-3, 3-4 and 4-4 from rat liver and the major glutathione S-transferase from the wax moth (Galleria mellonella) are all inhibited by several simple inorganic anions. For each of 3-3, 3-4 and the insect enzyme, the order of inhibitory potency was ClO4- greater than or equal to SCN- greater than I- greater than NO3- greater than Br-. A more limited range of anions was tested on the isoenzymes 1-1 and 4-4, but the same trend was apparent. Values for Ki ranged from about 200 mM for Cl- to 6 mM for SCN- in the case of the insect enzyme and from 50 mM for Br- to 0.3 mM for SCN- for the rat isoform 3-3. Acetate, F-, SO4(2-) and PO4(3-) were not found to have significant inhibitory properties. The mode of inhibition was characterized as non-competitive in the case of the insect enzyme and rat transferase 1-1, whereas the mode of inhibition was partially non-competitive in the case of the rat isoforms 3-3, 3-4 and 4-4.

Effect of inhibitors of glutathione S-transferase on glyceryl trinitrate activity in isolated rat aorta

1993 ◽  
Vol 71 (2) ◽  
pp. 179-184 ◽  
Author(s):  
Rita Nigam ◽  
Tracy Whiting ◽  
Brian M. Bennett
Keyword(s):  
Glyceryl Trinitrate ◽  
Cyclic Gmp ◽  
Guanylyl Cyclase ◽  
Rat Aorta ◽  
Supernatant Fraction ◽  
Organic Nitrates ◽  
Transferase Activity ◽  
Glutathione S Transferase ◽  
Glutathione S Transferases

We investigated the role of glutathione S-transferases (enzymes known to biotransform organic nitrates) in the vascular action of glyceryl trinitrate (GTN). Relaxation of phenylephrine-contracted rat aortic strips was assessed in the presence or absence of the glutathione S-transferase inhibitors Basilen Blue, bromosulfophthalein, Rose Bengal, hematin, chlorotriphenyltin, and (octyloxy)benzoylvinylglutathione. Whereas none of the inhibitors increased the EC50 for GTN relaxation, glutathione S-transferase activity in the 100 000 × g supernatant fraction of rat aorta was inhibited markedly by most of the inhibitors. In addition, GTN-stimulated activation of aortic guanylyl cyclase in broken-cell preparations was attenuated by all of the glutathione S-transferase inhibitors, suggesting a direct inhibitory action on guanylyl cyclase. In other experiments using aortic strips preexposed to phenylephrine, the inhibitors had no effect on GTN-induced cyclic GMP accumulation or on vascular biotransformation of GTN. In contrast, both Basilen Blue and bromosulfophthalein significantly inhibited GTN-induced relaxation of K+-contracted aortic strips, and Basilen Blue significantly inhibited GTN biotransformation in aortic strips preexposed to 25 mM K+. This may be due to a more favourable electrochemical gradient for entry of the inhibitors into membrane-depolarized tissues. We conclude that vascular glutathione S-transferases play a role in mediating the vasodilator actions of GTN in intact tissues in vitro, but that this appears to depend upon the nature of the contractile agent used in such studies.Key words: glyceryl trinitrate, glutathione S-transferase, cyclic GMP, vascular smooth muscle, biotransformation.

Circadian rhythms in glutathione and glutathione-S transferase activity of rat liver

1992 ◽  
Vol 100 (1) ◽  
pp. 83-87 ◽  
Author(s):  
M. J. Tuñón ◽  
P. González ◽  
P. López ◽  
G. M. Salido ◽  
J. A. Madrid
Keyword(s):  
Circadian Rhythms ◽  
Rat Liver ◽  
Transferase Activity ◽  
Glutathione S Transferase
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