HYDROGEN PEROXIDE-MEDIATED ALTERATION OF THE HEME PROSTHETIC GROUP OFMETMYOGLOBIN TO AN IRON CHLORIN PRODUCT - EVIDENCE FOR A NOVEL OXIDATIVE PATHWAY

Treatment of metmyoglobin with H2O2 is known to lead to the crosslinki ng of an active site tyrosine residue to the heme [Catalano, C. E., Y. S. Choe, and P. R. Ortiz de Montellano (1989) J. Biol. Chem. 264, 105 34-10541]. We have found in this study that this reaction also leads t o an altered heme product not covalently bound to the protein. This pr oduct was characterized by visible absorption, infrared absorption, an d mass and NMR spectrometry as an iron chlorin product formed from the saturation of the double bond between carbon atoms at positions 17 an d 18 of pyrrole ring D with concomitant addition of a hydroxyl group o n the carbon atom at position 18 and lactonization of the propionic ac id to the carbon atom at position 17. Studies with the use of O-18-lab eled H2O2, O-2, and H2O clearly indicate that the source of the added oxygen on the heme is water. Evidently, water adds regiospecifically t o a cationic site formed on a carbon atom at position 18 after oxidati on of the ferric heme prosthetic group with peroxide. Prolonged incuba tion of the reaction mixture containing the iron hydroxychlorin produc t led to the formation of an iron dihydroxychlorin product, presumably from a slow addition of water to the initial iron hydroxychlorin. The iron chlorin products characterized in this study are distinct from t he meso-oxyheme species, which is thought to be formed during peroxide -mediated degradation of metmyoglobin, cytochrome P450, ferric heme, a nd model ferric hemes, and give further insight into the mechanism of H2O2-induced heme alterations.