Specific cation binding site in mammalian cytochrome oxidase.

Calcium ion binds reversibly with cytochrome c oxidase from beef heart mitochondria (Kd approximately 2 microM) shifting alpha- and gamma-absorption bands of heme a to the red. Two sodium ions compete with one Ca2+ for the binding site with an average dissociation constant square root[K1(Na) x K2(Na)] approximately 3.6 mM. The Ca2+-induced spectral shift of heme a is specific for mammalian cytochrome c oxidase and is not observed in bacterial or yeast aa3 oxidases although the Ca2+-binding site has been revealed in the bacterial enzyme [Ostermeier, C., Harrenga, A., Ermler, U. and Michel, H. (1997) Proc. Natl. Acad. Sci. USA 94, 10547-10553]. As His-59 and Gln-63 involved in Ca2+ binding with Subunit I of P. denitrificans oxidase are not conserved in bovine oxidase, these residues have to be substituted by alternative ligands in mammalian enzyme, which is indeed the case as shown by refined structure of bovine heart cytochrome oxidase (S. Yoshikawa, personal communication). We propose that it is interaction of Ca2+ with the species-specific ligand(s) in bovine oxidase that accounts for perturbation of heme a. The Ca2+/Na2+-binding site may be functionally associated with the exit part of 'pore B' proton channel in subunit I of mammalian cytochrome c oxidase.