Ligand Environment of the Active Site Copper in Cytochrome Oxidase

The studies of the copper redox sites of cytochrome oxidase have been continued to include the preparation preferentially depleted of eh EPR-detectable copper (Cua) by means of a short exposure to bathocuproine sulfonate at pH5.0(1). This preparation of the oxidase retains at least 75% of the EPR-undetectable copper (cUa3) but has no near IR absorption band and, of course, no EPR signal attributable to copper indicating that the residual copper remains spin-coupled to the iron of cytochrome a3 X-ray absorption spectroscopy of the remaining copper atom shows that 1) the Cua3 is 3- or 4- coordinate with at least one and perhaps two of the ligands being S and the others being N, or possibly 0; 2) the Cu- Fe distance of 3.75 + 0.05 A that is observed with the intact oxidase is preserved;3) the geometry of the Cua3 is less tetrahedryl than in the oxidized "resting" state of the untreated enzyme. These results are consistent with those reported previously (2).