Specific nitration of tyrosines 46 and 48 makes cytochrome c assemble a non-functional apoptosome

► Solvent-exposed tyrosines 46 and 48 of cytochrome c are nitrated in vitro. ► Nitration of cytochrome c does not affect the reaction with cytochrome c oxidase. ► Nitration of cytochrome c impairs its ability to activate caspase-9. Under nitroxidative stress, a minor fraction of cytochrome c can be modified by tyrosine nitration. Here we analyze the specific effect of nitration of tyrosines 46 and 48 on the dual role of cytochrome c in cell survival and cell death. Our findings reveal that nitration of these two solvent-exposed residues has a negligible effect on the rate of electron transfer from cytochrome c to cytochrome c oxidase, but impairs the ability of the heme protein to activate caspase-9 by assembling a non-functional apoptosome. It seems that cytochrome c nitration under cellular stress counteracts apoptosis in light of the small amount of modified protein. We conclude that other changes such as increased peroxidase activity prevail and allow the execution of apoptosis.