Modeling Cytochrome Oxidase:  A Quantum Chemical Study of the O−O Bond Cleavage Mechanism

The mechanism of O−O bond cleavage at the binuclear center in cytochrome oxidase has been investigated by using hybrid density functional theory (B3LYP). A potential energy surface for the reaction step from compound A, with the O 2 molecule coordinated to heme a 3, to the bond-cleaved compound P is constructed. The features of the calculated potential surface agree well with experimental information on this reaction step. First, a free energy of activation of 15 kcal/mol is obtained, reasonably close to the value of 12 kcal/mol corresponding to the observed lifetime of compound A. Second, the calculations give a large entropy effect on the reaction rate, which explains the weak temperature dependence observed for the P formation reaction. Third, the calculated potential surface has no stable intermediate between A and P, in agreement with the experimental observation that compound A decays with the same rate as compound P forms. Fourth, the calculations show that the oxo−ferryl compound P together with a tyrosyl radical and a cupric−hydroxyl can be formed in a close to thermoneutral reaction, which gives support to the suggestion that the source of the fourth electron in the O−O bond cleavage reaction is the cross-linked tyrosine residue near the binuclear center. The calculations indicate, however, that a direct hydrogen-atom transfer from the tyrosine to the iron-coordinated O 2 molecule produces too high an energy barrier. We attribute this to weak electronic coupling between the tyrosine and the iron center. Instead, our work suggests that the rate-limiting step involves a hydrogen-atom transfer from a water molecule in the vicinity of the copper center, and that the actual O−O bond cleavage step requires an extra proton to be available in the vicinity of the tyrosine residue, possibly on the hydroxyl group of the heme a 3 farnesyl side chain. The reason the additional proton is needed for the O−O bond cleavage is discussed.