Site specific phosphorylation of cytochrome c oxidase subunits I, IVi1 and Vb in rabbit hearts subjected to ischemia/reperfusion

We have mapped the sites of ischemia/reperfusion-induced phosphorylation of cytochrome c oxidase (CcO) subunits in rabbit hearts by using a combination of Blue Native gel/Tricine gel electrophoresis and nano-LC-MS/MS approaches. We used precursor ion scanning combined with neutral loss scanning and found that mature CcO subunit I was phosphorylated at tandem Ser115/Ser116 positions, subunit IVi1 at Thr52 and subunit Vb at Ser40. These sites are highly conserved in mammalian species. Molecular modeling suggests that phosphorylation sites of subunit I face the inter membrane space while those of subunits IVi1 and Vb face the matrix side.