Rapid reactions of peroxynitrite with heme-thiolate proteins as the basis for protection of prostacyclin synthase from inactivation by nitration

Prostacyclin (PGI(2)) synthase is a heme-thiolate (P450) protein which reac ts with low levels of peroxynitrite (PN) under tyrosine nitration and inact ivation. Studying heme proteins as models, we have found the hemethiolate p rotein NADH-NO reductase (P450(NOR)) to be highly efficient in decomposing PN under concomitant nitration of phenol. The present study investigates tw o other P450 proteins, P450(BM-3) and chloroperoxidase, in order to test fo r the specific role of the thiolate ligand in the reaction with PN,A compar ison with horseradish peroxidase and microperoxidase gives evidence of kine tic differences that classify heme-thiolate proteins, but not other heme pr oteins, as effective inhibitors of PGI(2) synthase nitration and inactivati on. P450(BM-3) with PN catalyzes phenol nitration and nitration of its own tyrosine below 10 mu M PN, whereas chloroperoxidase and P450(NOR) at such c oncentrations also nitrate phenol but not enzyme-bound tyrosine residues. W e conclude that heme-thiolate proteins in general exhibit high reactivity w ith PN and turnover, probably due to the special electronic structure of th e presumed thiolate-ferryl intermediate. (C) 2000 Academic Press.