REGULATION OF THE ACTIVITY OF GLYCERALDEHYDE-3-PHOSPHATE DEHYDROGENASE BY GLUTATHIONE AND H2O2

The activity lost during storage of a solution of muscle glyceraldehyd e 3-phosphate dehydrogenase was rapidly restored on adding a thiol com pound, but not arsenite or azide. On treatment with H2O2, the enzyme w as partially inactivated and complete loss of activity occurred in the presence of glutathione. Samples of the enzyme pretreated with glutat hione followed by removal of the thiol compound by filtration on a Sep hadex column showed both full activity and its complete loss on adding H2O2, in the absence of added glutathione. Most of the activity was r estored when the H2O2-inactivated enzyme was incubated with glutathion e (25 mM) or dithiothreitol (5 mM) whereas arsenite or azide were part ly effective and ascorbate was ineffective. The need for incubation fo r a long time with a strong reducing agent for restoration of activity suggests that the oxidized group (disulfide or sulfenate) must be in a masked state in the H2O2-inactivated enzyme. Analysis by SDS-PAGE ga ve evidence for the formation of a small quantity of glutathione-rever sible disulfide-form of the enzyme. Circular dichroic spectra indicate d a decrease in alpha-helical content in the inactivated form of the e nzyme. The evidence suggest that glutathione and H2O2 can regulate the active state of this enzyme.