Redox-linked protolytic reactions in soluble cytochrome-c oxidase from beef-heart mitochondria: Redox Bohr effects

A study is presented of co-operative redox-linked protolytic reactions (redox Bohr effects) in soluble cytochrome-c oxidase purified from bovine-heart mitochondria. Bohr effects were analyzed by direct measurement, with accurate spectrophotometric and potentiometric methods, of H+ uptake and release by the oxidase associated with reduction and oxidation of hemes a and a(3), Cu-A and Cu-B in the unliganded and in the CN- or CO-liganded enzyme. The results show that there are in the bovine oxidase four protolytic groups undergoing reversible pK shifts upon oxide-reduction of the electron transfer metals. Two groups with pK(ox) and pK(red) values around 7 and > 12 respectively appear to be linked to redox transitions of heme a(3). One group with pK(ox) and pK(red) around 6 and 7 is apparently linked to Cu-B, a fourth one with pK(ox) and pK(red) of 6 and 9 appears to be linked to heme a. The possible nature of the amino acids involved in the redox Bohr effects and their role in H+ translocation is discussed.