scholarly journals Activation and inhibition of microsomal glutathione transferase from mouse liver

1988 ◽  
Vol 249 (3) ◽  
pp. 819-823 ◽  
Author(s):  
C Andersson ◽  
M Söderström ◽  
B Mannervik
Keyword(s):  
Molecular Mass ◽  
Mouse Liver ◽  
Glutathione Transferase ◽  
Cross Reactivity ◽  
Steep Slope ◽  
Glutathione Transferases ◽  
Inhibitor Concentration ◽  
Cytosolic Glutathione ◽  
Concentration Curves ◽  
Microsomal Glutathione

Mouse liver microsomal glutathione transferase was purified in an N-ethylmaleimide-activated as well as an unactivated form. The enzyme had a molecular mass of 17 kDa and a pI of 8.8. It showed cross-reactivity with antibodies raised against rat liver microsomal glutathione transferase, but not with any of the available antisera raised against cytosolic glutathione transferases. The fully N-ethylmaleimide-activated enzyme could be further activated 1.5-fold by inclusion of 1 microM-bromosulphophthalein in the assay system. The latter effect was reversible, which was not the case for the N-ethylmaleimide activation. At 20 microM-bromosulphophthalein the activated microsomal glutathione transferase was strongly inhibited, while the unactivated form was activated 2.5-fold. Inhibitors of the microsomal glutathione transferase from mouse liver showed either about the same I50 values for the activated and the unactivated form of the enzyme, or significantly lower I50 values for the activated form compared with the unactivated form. The low I50 values and the steep slope of the activity-versus-inhibitor-concentration curves for the latter group of inhibitors tested on the activated enzyme indicate a co-operative effect involving conversion of activated enzyme into the unactivated form, as well as conventional inhibition of the enzyme.

scholarly journals Effects of inducers of drug metabolism on basic hepatic forms of mouse glutathione transferase

1989 ◽  
Vol 263 (3) ◽  
pp. 679-685 ◽  
Author(s):  
P Di Simplicio ◽  
H Jensson ◽  
B Mannervik
Keyword(s):  
Mouse Liver ◽  
Glutathione Transferase ◽  
Major Effect ◽  
Glutathione Transferases ◽  
Control Group ◽  
Glyoxalase I ◽  
Protein Profiles ◽  
Trans Stilbene ◽  
Cytosolic Glutathione ◽  
Gst Activity

The cytosolic glutathione transferases (GSTs) with basic pI values have been studied in mouse liver after treatment with 2,3-t-butylhydroxyanisole (BHA), cafestol palmitate (CAF), phenobarbital (PB), 3-methylcholanthrene (3-MC) and trans-stilbene oxide (t-SBO). The cytosolic GST activity was induced by all compounds except for 3-MC. Three forms of GST were isolated by means of affinity chromatography and f.p.l.c. The examination of protein profiles and enzymic activities with specific substrates showed that the three GSTs correspond to those found in control animals, i.e. GSTs MI, MII and MIII. The class Mu GST MIII accounted for the major effect of induction, whereas the class Alpha GST MI and the class Pi GST MII were unchanged or somewhat down-regulated. The greatest induction was obtained with BHA, PB and CAF. The activities of other glutathione-dependent enzymes were also studied. An increase in glutathione reductase and thioltransferase activities was observed after BHA, PB or CAF treatment; glyoxalase I and Se-dependent glutathione peroxidase were depressed in comparison with the control group in all cases studied.

scholarly journals Leukotriene C synthase in mouse mastocytoma cells. An enzyme distinct from cytosolic and microsomal glutathione transferases

1988 ◽  
Vol 250 (3) ◽  
pp. 713-718 ◽  
Author(s):  
M Söderström ◽  
S Hammarström ◽  
B Mannervik
Keyword(s):  
Microsomal Fraction ◽  
Glutathione Transferase ◽  
Specific Activity ◽  
Glutathione Transferases ◽  
Leukotriene C4 ◽  
Cytosol Fraction ◽  
Mastocytoma Cells ◽  
Specific Activities ◽  
Leukotriene A4 ◽  
Microsomal Glutathione

Leukotriene C4 synthesis was studied in preparations from mouse mastocytoma cells. Enzymic conjugation of leukotriene A4 with glutathione was catalysed by both the cytosol and the microsomal fraction. The specific activity of the microsomal fraction (7.8 nmol/min per mg of protein) was 17 times that of the cytosol fraction. The cytosol fraction of the mastocytoma cells contained two glutathione transferases, which were purified to homogeneity and characterized. A microsomal glutathione transferase was purified from mouse liver; this enzyme was shown by immunoblot analysis to be present in the mastocytoma microsomal fraction at a concentration one-tenth or less of that in the liver microsomal fraction. Both the cytosolic and the microsomal glutathione transferases in the mastocytoma cells were identified with enzymes previously characterized, by determining specific activities with various substrates, sensitivities to inhibitors, reactions with antibodies, and physical properties. The purified microsomal glutathione transferase from liver was inactive with leukotriene A4 or its methyl ester as substrate. The cytosolic enzymes displayed activity with leukotriene A4, but their specific activities and intracellular concentrations were too low to account for the leukotriene C4 formation in the mastocytoma cells. The microsomal fraction of the cells contained an enzyme distinguishable by various criteria from the previously studied glutathione transferases. This membrane-bound enzyme, leukotriene C synthase (leukotriene A4:glutathione S-leukotrienyltransferase), appears to carry the main responsibility for the biosynthesis of leukotriene C4.

scholarly journals Cytosolic glutathione transferases from rat liver. Primary structure of class alpha glutathione transferase 8-8 and characterization of low-abundance class Mu glutathione transferases

1989 ◽  
Vol 261 (2) ◽  
pp. 531-539 ◽  
Author(s):  
P Alin ◽  
H Jensson ◽  
E Cederlund ◽  
H Jörnvall ◽  
B Mannervik
Keyword(s):  
Rat Liver ◽  
Glutathione Transferase ◽  
Glutathione Transferases ◽  
Rat Lung ◽  
Cytosol Fraction ◽  
Methionine Residue ◽  
Isoelectric Points ◽  
Cytosolic Glutathione ◽  
Lung And Kidney

Six GSH transferases with neutral/acidic isoelectric points were purified from the cytosol fraction of rat liver. Four transferases are class Mu enzymes related to the previously characterized GSH transferases 3-3, 4-4 and 6-6, as judged by structural and enzymic properties. Two additional GSH transferases are distinguished by high specific activities with 4-hydroxyalk-2-enals, toxic products of lipid peroxidation. The most abundant of these two enzymes, GSH transferase 8-8, a class Alpha enzyme, has earlier been identified in rat lung and kidney. The amino acid sequence of subunit 8 was determined and showed a typical class Alpha GSH transferase structure including an N-acetylated N-terminal methionine residue.

scholarly journals Kinetic independence of the subunits of cytosolic glutathione transferase from the rat

1985 ◽  
Vol 231 (2) ◽  
pp. 263-267 ◽  
Author(s):  
U H Danielson ◽  
B Mannervik
Keyword(s):  
Glutathione Transferase ◽  
Ethacrynic Acid ◽  
Glutathione Transferases ◽  
Kinetic Properties ◽  
Subunit Interactions ◽  
Steady State Kinetics ◽  
Binding Isotherms ◽  
Cytosolic Glutathione ◽  
Kinetics Of ◽  
Rate Behaviour

The steady-state kinetics of the dimeric glutathione transferases deviate from Michaelis-Menten kinetics, but have hyperbolic binding isotherms for substrates and products of the enzymic reaction. The possibility of subunit interactions during catalysis as an explanation for the rate behaviour was investigated by use of rat isoenzymes composed of subunits 1, 2, 3 and 4, which have distinct substrate specificities. The kinetic parameter kcat./Km was determined with 1-chloro-2,4-dinitrobenzene, 4-hydroxyalk-2-enals, ethacrynic acid and trans-4-phenylbut-3-en-2-one as electrophilic substrates for six isoenzymes: rat glutathione transferases 1-1, 1-2, 2-2, 3-3, 3-4 and 4-4. It was found that the kcat./Km values for the heterodimeric transferases 1-2 and 3-4 could be predicted from the kcat./Km values of the corresponding homodimers. Likewise, the initial velocities determined with transferases 3-3, 3-4 and 4-4 at different degrees of saturation with glutathione and 1-chloro-2,4-dinitrobenzene demonstrated that the kinetic properties of the subunits are additive. These results show that the subunits of glutathione transferase are kinetically independent.

scholarly journals Characterization of Dog Glutathione Transferase P1-1, an Enzyme Relevant to Veterinary Medicine

2021 ◽  
Vol 22 (8) ◽  
pp. 4079
Author(s):  
Aram Ismail ◽  
Elizabeth Lewis ◽  
Birgitta Sjödin ◽  
Bengt Mannervik
Keyword(s):  
Canis Lupus ◽  
Glutathione Transferase ◽  
Biological Significance ◽  
Cross Reactivity ◽  
Glutathione Transferases ◽  
Human Homolog ◽  
Canis Lupus Familiaris ◽  
Ic50 Value ◽  
Active Substrate

Glutathione transferases (GSTs) form a family of detoxication enzymes instrumental in the inactivation and elimination of electrophilic mutagenic and carcinogenic compounds. The Pi class GST P1-1 is present in most tissues and is commonly overexpressed in neoplastic cells. GST P1-1 in the dog, Canis lupus familiaris, has merits as a marker for tumors and as a target for enzyme-activated prodrugs. We produced the canine enzyme CluGST P1-1 by heterologous bacterial expression and verified its cross-reactivity with antihuman-GST P1-1 antibodies. The catalytic activity with alternative substrates of biological significance was determined, and the most active substrate found was benzyl isothiocyanate. Among established GST inhibitors, Cibacron Blue showed positive cooperativity with an IC50 value of 43 nM. Dog GST P1-1 catalyzes activation of the prodrug Telcyta, but the activity is significantly lower than that of the human homolog.

scholarly journals Structure-activity relationships of 4-hydroxyalkenals in the conjugation catalysed by mammalian glutathione transferases

1987 ◽  
Vol 247 (3) ◽  
pp. 707-713 ◽  
Author(s):  
U H Danielson ◽  
H Esterbauer ◽  
B Mannervik
Keyword(s):  
Chain Length ◽  
Active Site ◽  
Glutathione Transferase ◽  
Physiological Role ◽  
Catalytic Efficiency ◽  
Glutathione Transferases ◽  
Side Chain ◽  
Structure Activity ◽  
Cytosolic Glutathione ◽  
Free Energy Of Binding

The substrate specificities of 15 cytosolic glutathione transferases from rat, mouse and man have been explored by use of a homologous series of 4-hydroxyalkenals, extending from 4-hydroxypentenal to 4-hydroxypentadecenal. Rat glutathione transferase 8-8 is exceptionally active with the whole range of 4-hydroxyalkenals, from C5 to C15. Rat transferase 1-1, although more than 10-fold less efficient than transferase 8-8, is the second most active transferase with the longest chain length substrates. Other enzyme forms showing high activities with these substrates are rat transferase 4-4 and human transferase mu. The specificity constants, kcat./Km, for the various enzymes have been determined with the 4-hydroxyalkenals. From these constants the incremental Gibbs free energy of binding to the enzyme has been calculated for the homologous substrates. The enzymes responded differently to changes in the length of the hydrocarbon side chain and could be divided into three groups. All glutathione transferases displayed increased binding energy in response to increased hydrophobicity of the substrate. For some of the enzymes, steric limitations of the active site appear to counteract the increase in binding strength afforded by increased chain length of the substrate. Comparison of the activities with 4-hydroxyalkenals and other activated alkenes provides information about the active-site properties of certain glutathione transferases. The results show that the ensemble of glutathione transferases in a given species may serve an important physiological role in the conjugation of the whole range of 4-hydroxyalkenals. In view of its high catalytic efficiency with all the homologues, rat glutathione transferase 8-8 appears to have evolved specifically to serve in the detoxication of these reactive compounds of oxidative metabolism.

scholarly journals Glutathione transferases in rat hepatoma cells. Effects of ascites cells on the isoenzyme pattern in liver and induction of glutathione transferases in the tumour cells

1989 ◽  
Vol 257 (1) ◽  
pp. 215-220 ◽  
Author(s):  
M K Tahir ◽  
C Guthenberg ◽  
B Mannervik
Keyword(s):  
Glutathione Transferase ◽  
Hepatoma Cell ◽  
Normal Liver ◽  
Hepatoma Cells ◽  
Glutathione Transferases ◽  
Trans Stilbene ◽  
Cytosolic Glutathione ◽  
Fold Induction ◽  
Rat Hepatoma ◽  
Rat Hepatoma Cells

Rat hepatoma cells grown intraperitoneally as an ascites tumour were analysed with respect to their contents of cytosolic glutathione transferases. In contrast with normal liver tissue, the hepatoma cells were dominated by the class Pi glutathione transferase 7-7. All the major hepatic enzyme forms were down-regulated to almost undetectable concentrations. Livers of rats bearing ascites-hepatoma cells expressed low, but significant, amounts of protein which, by electrophoretic and immunochemical properties, appeared identical with transferase 7-7. This enzyme is not detectable in normal hepatocytes. Treatment of rats with trans-stilbene oxide induced the expression of transferase 7-7 in the livers of normal rats as well as in hepatoma-cell-bearing animals. In addition, a 2-fold induction of transferase 7-7 was measured in the hepatoma ascites cells. No significant elevation of any other enzyme forms in the hepatoma cells was noted.

Parameters for the Two-Dimensional Crystallization of the Membrane Protein Microsomal Glutathione Transferase

1998 ◽  
Vol 123 (2) ◽  
pp. 87-96 ◽  
Author(s):  
Ingeborg Schmidt-Krey ◽  
Gerd Lundqvist ◽  
Ralf Morgenstern ◽  
Hans Hebert
Keyword(s):  
Membrane Protein ◽  
Glutathione Transferase ◽  
Two Dimensional ◽  
Microsomal Glutathione
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