Structural basis of mitochondrial dysfunction in response to cytochrome c phosphorylation at tyrosine 48

Regulation of mitochondrial activity allows cells to adapt to changing conditions and to control oxidative stress, and its dysfunction can lead to hypoxia-dependent pathologies such as ischemia and cancer. Although cytochrome c phosphorylation—in particular, at tyrosine 48—is a key modulator of mitochondrial signaling, its action and molecular basis remain unknown. Here we mimic phosphorylation of cytochrome c by replacing tyrosine 48 with p-carboxy-methyl-L-phenylalanine (pCMF). The NMR structure of the resulting mutant reveals significant conformational shifts and enhanced dynamics around pCMF that could explain changes observed in its functionality: The phosphomimetic mutation impairs cytochrome c diffusion between respiratory complexes, enhances hemeprotein peroxidase and reactive oxygen species scavenging activities, and hinders caspase-dependent apoptosis. Our findings provide a framework to further investigate the modulation of mitochondrial activity by phosphorylated cytochrome c and to develop novel therapeutic approaches based on its prosurvival effects.