Fully and partially reduced forms of isolated bovine cytochrome c oxidase undergo a two-electron oxidation-reduction process with added peroxynitrite, leading to catalytic oxidation of ferrocytochrome c to ferricytochrome c. The other major reaction product is nitrite ion, 86% of the added peroxynitrite being measurably converted to this species. The reaction is inhibited in the presence of cyanide, implicating the heme a(3)-Cu(B) binuclear pair as the active site. Moreover, provided peroxynitrite is not added to excess, the reductase activity of the enzyme toward this oxidant efficiently protects other protein and detergent molecules in vitro from nitration of tyrosine residues and oxidative damage. If the enzyme is exposed to approximately 10(2)-fold excesses of peroxynitrite, then significant irreversible loss of electron transfer activity results, and the heme a(3)-Cu(B) binuclear pair no longer undergo a characteristic carbon monoxide-driven reduction. The accompanying rather small changes in the observed electronic absorption spectrum are suggestive of a modification in the vicinity of one or both hemes but probably not to the cofactors themselves.