Spectral and kinetic equivalence of oxidized cytochrome c oxidase as isolated and "activated" by reoxidation

The spectral and kinetic characteristics of two oxidized states of bovine heart cytochrome c oxidase (CcO) have been compared. The first is the oxidized state of enzyme isolated in the fast form (O) and the second is the form that is obtained immediately after oxidation of fully reduced CcO with O₂ (O_H). No observable differences were found between O and OH states in: (i) the rate of anaerobic reduction of heme a₃ for both the detergent-solubilized enzyme and for enzyme embedded in its natural membraneous environment, (ii) the one-electron distribution between heme a₃ and Cu_B in the course of the full anaerobic reduction, (iii) the optical and (iv) EPR spectra. Within experimental error of these characteristics both forms are identical. Based on these observations it is concluded that the reduction potentials and the ligation states of heme a₃ and Cu _B are the same for CcO in the O and O_H states.