Structural aspects of the copper sites in cytochrome c oxidase. An x-ray absorption spectroscopic investigation of the resting-state enzyme

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Abstract
Copper K-edge X-ray absorption spectroscopy (XAS) has been used to investigate the structural details of the coordination environment of the copper sites in eight resting-state samples of beef heart cytochrome c oxidase prepared by different methods. The unusual position and structure of the resting-state copper edge spectrum can be adequately explained by the presence of sulfur-containing ligands, with a significant amount of S .fwdarw. Cu(II) charge transfer (i.e., a covalent site). Quantitative curve-fitting analysis of the copper extended X-ray absorption fine structure (EXAFS) data indicates similar average first coordination spheres for all resting-state samples, regardless of preparation method. The average coordination sphere (per 2 coppers) mainly consists of 6 .+-. 1 nitrogens or oxygens at an average Cu.sbd.(N,O) distance of 1.99 .+-. 0.03 .ANG. and 2 .+-. 1 sulfurs at an averaged Cu.sbd.S distance of 2.28 .+-. 0.02 .ANG.. Quantitative curve-fitting analysis of the outer shell of the copper EXAFS indicates the presence of a Cu.cntdot..cntdot..cntdot.Fe interaction at a distance of 3.00 .+-. 0.03 .ANG.. Proposed structures of the two copper sites based on these and other spectroscopic results are presented, and differences between our results and those of other published copper XAS studies [Powers, L., Chance, B., Ching, Y., and Angiolillo, P. (1981) Biophys. J. 34, 465-498] are discussed.