Decrease in accessible thiols as an index of oxidative damage to membrane proteins.

The effect of several oxidative agents (hydrogen peroxide, tert-butyl hydroperoxide, menadione, AAPH, peroxynitrite and ionizing radiation) on the ratio of weakly to strongly immobilized residues of erythrocyte membrane-bound maleimide-tempo spin label (h(w)/h(s) ratio) was studied in order to test the hypothesis that a decrease in the h(w)/h(s) ratio may be a general index of oxidative damage to membrane proteins. Most of the agents studied decreased though H2O2 almost did not affect and ionizing radiation increased the h(w)/h(s) ratio. In parallel, the ratio of DTNB accessible/DTNB inaccessible membrane protein-SH groups was determined from membrane-SH group measurements with the Ellman reagent in the absence and in the presence of sodium dodecyl sulfate. This ratio decreased in all cases studied and seems to be a more universal and easy to measure parameter to describe the oxidative damage to membrane proteins.