Kazuhiro Ohta

Bovine cytochrome c oxidase structures enable O2 reduction with minimization of reactive oxygens and provide a proton-pumping gate

The O2 reduction site of cytochrome c oxidase (CcO), comprising iron (Fea3) and copper (CuB) ions, is probed by x-ray structural analyses of CO, NO, and CN- derivatives to investigate the mechanism of the complete reduction of O2. Formation of the derivative contributes to the trigonal planar coordination of and displaces one of its three coordinated imidazole groups while a water molecule becomes hydrogen bonded to both the CN- ligand and the hydroxyl group of Tyr244. When O2 is bound to , it is negatively polarized (), and expected to induce the same structural change induced by CN-. This structural change allows to receive three electron equivalents nonsequentially from , , and Tyr-OH, providing complete reduction of O2 with minimization of production of active oxygen species. The proton-pumping pathway of bovine CcO comprises a hydrogen-bond network and a water channel which extend to the positive and negative side surfaces, respectively. Protons transferred through the water channel are pumped through the hydrogen-bond network electrostatically with positive charge created at the Fea center by electron donation to the O2 reduction site. Binding of CO or NO to induces significant narrowing of a section of the water channel near the hydrogen-bond network junction, which prevents access of water molecules to the network. In a similar manner, O2 binding to is expected to prevent access of water molecules to the hydrogen-bond network. This blocks proton back-leak from the network and provides an efficient gate for proton-pumping.

X-ray structure of the NO-bound Cu(B) in bovine cytochrome c oxidase.

The X-ray crystallographic structure of nitric oxide-treated bovine heart cytochrome c oxidase (CcO) in the fully reduced state has been determined at 50 K under light illumination. In this structure, nitric oxide (NO) is bound to the CcO oxygen-reduction site, which consists of haem and a Cu atom (the haem a(3)-Cu(B) site). Electron density for the NO molecule was observed close to Cu(B). The refined structure indicates that NO is bound to Cu(B) in a side-on manner.