Mutagenesis analysis of hydrogen bond network formed in bovine heart cytochrome c oxidase

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We carried out mutagenesis analyses of a hydrogen bond network, designated as D-pathway, formed in the bovine heart cytochrome c oxidase employing HeLa cell bovine/human hybrid enzyme expression system developed by us. A pathway corresponding to this D-pathway is also found in the bacterial enzymes, and has been proposed to convey both pumping and water forming protons. The structures of these two pathways are almost identical to each other and amino acid residues essential to the bacterial enzymes are conserved in the bovine enzyme. Hence, it has been thought that these two have the same function, thereby opposing to the proton-pumping pathway of the bovine enzyme proposed by us. The results of this study strongly suggest D-pathway is not for pumping proton but for water forming proton.