Cytochrome c oxidase: The influence of the "open-closed" transition on cytochrome a₃

Cytochrome c oxidase is known to display a conformational transition from a "closed" to an "open" structure upon partial reduction. In the past exogenous ligand binding kinetic differences and differences in tryptophan fluorescence have been associated with this structural change. We report changes in the resonance Raman spectrum of NO-bound cytochrome a_s upon this transition. We detect changes in both the heme vibrational modes and in the Fe-N-O bending mode. Furthermore, we have been able to identify a hydroxide-bound intermediate in the enzyme. Analysis of the conditions for its generation lead to the proposal that it is stable only in the open conformation of the protein. In both of these series of experiments advantage was taken of the photoreducibility of the heme groups by the incident laser beam.