Snapshot of an oxygen intermediate in the catalytic reaction of cytochrome <i>c</i> oxidase.

Abstract

Cytochrome <i>c</i> oxidase (C<i>c</i>O) reduces dioxygen to water and harnesses the chemical energy to drive proton translocation across the inner mitochondrial membrane by an unresolved mechanism. By using time-resolved serial femtosecond crystallography, we identified a key oxygen intermediate of bovine C<i>c</i>O. It is assigned to the P_R-intermediate, which is characterized by specific redox states of the metal centers and a distinct protein conformation. The heme <i>a</i>₃ iron atom is in a ferryl (Fe⁴⁺ = O^2-) configuration, and heme <i>a</i> and Cu_B are oxidized while Cu_A is reduced. A Helix-X segment is poised in an open conformational state; the heme <i>a</i> farnesyl sidechain is H-bonded to S382, and loop-I-II adopts a distinct structure. These data offer insights into the mechanism by which the oxygen chemistry is coupled to unidirectional proton translocation.