Protein S-thiolation and redox regulation of membrane-bound glutathione transferase.
Membrane-bound GST transferase (GSTm) occurs in hepatic microsomal and plasma membranes as well as in the outer mitochondrial membrane, and it is known to be activated by N-ethylmaleimide. We recently analysed the activation by GSSG in some detail. The approximately 5-fold stimulation is reversed upon reduction of GSSG by GSSG reductase. In steady-state experiments, the Kox value was determined to be 0.05, i.e. 20 times more GSSG than GSH produces half-maximal activation. Kox is independent of the total glutathione concentration, indicating that S-thiolation by mixed disulfide formation, rather than interchain or intrachain disulfide bridge formation, is responsible for activation. In Western blots, a 17.7 kDa band, in addition to the 17.3 kDa band, was detected upon treatment with GSSG or with GSH plus t-butyl hydroperoxide. We suggest that under oxidative stress, GSTm is activated through direct S-thiolation of the enzyme. Dethiolation occurs via thiol disulfide exchange governed by the cellular glutathione redox state.
Sies
H
H
Institut für Physiologische Chemie I, Heinrich-Heine-Universität Düsseldorf, Germany. helmut.sies@uni-duesseldorf.de
Dafré
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Ji
Y
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Akerboom
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TP
D000818:Animals; D002462:Cell Membrane / Q000201:enzymology; D004789:Enzyme Activation / Q000187:drug effects; D019803:Glutathione Disulfide / Q000494:pharmacology; D005982:Glutathione Transferase / Q000378:metabolism*; D007425:Intracellular Membranes / Q000201:enzymology; D015227:Lipid Peroxidation; D008099:Liver / Q000201:enzymology; D008862:Microsomes, Liver / Q000201:enzymology; D008930:Mitochondria, Liver / Q000201:enzymology; D010084:Oxidation-Reduction; D018384:Oxidative Stress; D051381:Rats; D013438:Sulfhydryl Compounds / Q000378:metabolism
D013438:Sulfhydryl Compounds; D005982:Glutathione Transferase; D019803:Glutathione Disulfide