Interactions of heme proteins with hydrogen peroxide: Protein crosslinking and covalent binding of benzo[a]pyrene and 17β-estradiol

This work reveals two biochemical effects of hydrogen peroxide treatment on hemoglobin, myoglobin, and cytochrome c. First, these heme proteins rapidly formed covalently crosslinked dimers and polymers detectable by detergent gel electrophoresis. Second, when treated in the presence of radioactive benzo[a]pyrene or 17β-estradiol, the heme proteins became covalently labeled. Nonheme proteins exhibited both cross-linking and radioactive labeling upon peroxide treatment in the presence but not the absence of heme protein or free hemin. Benzoyl peroxide or glucose and glucose oxidase effectively replaced direct addition of hydrogen peroxide. These results indicate that adventitious peroxidase activity expressed by oxygen carrying and electron transport proteins yields active oxygen species that can damage these heme proteins and nearby macromolecules, a possible biochemical mechanism for the lethal and other deleterious intracellular effects of perioxide.