Diamide effect on the ouabain-insensitive APTase activity of red cell membrane

Membranes from human erythrocytes exhibit a marked decrease of the ouabain-insensitive ATPase activity and of the total membrane thiol content after treatment with diazenedicarboxylic acid bis(N,N-dimethylamide) (diamide). These effects increase with diamide concentrations up to 2-2.5 mM and are persistent after removal of the reagent. Treatment with 2,3-dihydroxy-1,4-dithiolbutane (dithioerythritol or DTE) reduced glutathione or 2-mercaptoethanol partially but significantly restores at about the same extent the ouabain-insensitive ATPase activity. These results indicate that the perturbation of the ATPase microenvironment caused by membrane thiol oxidation is at good extent responsible for alterations of the divalent cation-dependent ATPase activity.