The circular dichroic spectra of cytochrome c oxidase in the visible-wavelength region

Abstract

Cytochrome oxidase purified from beef heart shows distinct visible circular dichroic spectra which are oxidation- and ligand-state dependent. The oxidized enzyme and its cyanide complex exhibit broad negative extrema at 570 nm ([θ] = −4 × 103 degrees-cm2∙dmol−1) and 620 nm. A single Cotton effect was resolved for each of these extrema, except in the case of the cyanide complex where two Cotton effects (558.9 and 572.4 nm) were located within the 570-nm extremum.The reduced enzyme displays a major positive α-elliptic band at 608 nm and minor ones at 570 and 518 nm in accordance with its absorption spectrum. Carbon monoxide and cyanide cause the α-elliptic band of the reduced enzyme to shift to 604 nm ([θ] = 26 × 103 degrees-cm2∙dmol−1) and 602 nm ([θ] = 33 × 103 degrees-cm2∙dmol−1) respectively. Computer-assisted resolution of these three spectra suggests a common profile whereby two major transitions (Cotton effects II and III) of about equal rotational strength constitute the main α-elliptic band observed between 602 and 608 nm. Carbon monoxide induces Cotton effect II of the reduced enzyme to shift from 612.6 to 601.6 nm but not Cotton effect III at 608.4 nm. In contrast, cyanide alters the band position, band width, amplitudes, and the rotational strength of these two. These results permit identifying Cotton effect II as arising from the heme chromophore of cytochrome a3, and Cotton effect III from cytochrome a. Carbon monoxide also induces a red shift to the β-elliptic band at 520 nm of the reduced enzyme. On the other hand, cyanide causes a blue shift to this same band. The dependency of this elliptic band upon the extrinsic ligand of cytochrome a3 is consistent with the charge transfer nature of this band.