Naturally Occurring Disease-Related Mutations in the 40–57 Ω-Loop of Human Cytochrome c Control Triggering of the Alkaline Isomerization

Naturally occurring mutations found in one of the two Ω-loop substructures in human cytochrome c are associated with low blood platelet count (thrombocytopenia). Both Ω-loops participate in the formation of conformers associated with cytochrome c peroxidase activity and apoptotic function. At alkaline pH values, the Met80 ligand to the ferric heme iron dissociates, and a lysine residue in the 71–85 Ω-loop coordinates to the iron. The alkaline isomerization has been the focus of extensive kinetic studies, and it is established that a deprotonation triggers the release of the Met80 ligand (pKtrigger). A second deprotonation stabilizes a pentacoordinate heme form (pKa2). In this study, site-directed variants at the 41 and 48 positions in the 40–57 Ω-loop and at the 81 and 83 positions in the 71–85 Ω-loop reveal that conformational transitions in the 71–85 Ω-loop, leading to the alkaline or peroxidatic conformers, are controlled by the 40–57 Ω-loop. We find that the variants causing thrombocytopenia, G41S and...