Inactivation of Glutathione Peroxidase 1 and Peroxiredoxin 2 by Peroxides in Red Blood Cells

Abstract

Glutathione peroxidase 1 (GPx1), peroxiredoxin II (Prx II), and catalase are the principal enzymes responsible for peroxide elimination in red blood cells (RBCs). GPx1, which contains a selenocysteine (Sec) residue at its active site, is irreversibly inactivated by its own substrate as the result of the oxidation of selenium atom followed by the conversion of oxidized Sec to dehydroalanine (DHA). Prx II is inactivated when its catalytic cysteine (Cys) is hyperoxidized to cysteine sulfinic acid during catalysis. The hyperoxidation can be reversed by sulfiredoxin. The activity of sulfiredoxin in RBCs is sufficient to counteract the Prx II hyperoxidation that occurs during elimination of H₂O₂ molecules resulting from hemoglobin (Hb) autoxidation. We developed a blot method for detection of DHA-containing proteins, with the use of which we observed that the amount of DHA-containing GPx1 increases with aging of RBCs as well as in RBCs exposed to H₂O₂ generated either externally by glucose oxidase or internally as a result of aniline-induced Hb autoxidation. Given that the conversion of Sec to DHA is irreversible and that protein turnover mechanism is lacking in RBCs, the content of DHA–GPx1 in each RBC likely reflects total oxidative stress experienced by the cell during its lifetime. Therefore, DHA–GPx1 in RBCs might be a suitable surrogate marker for evaluation of oxidative stress in the body.