Cytochrome c Oxidase Components III. SPECTRAL PROPERTIES OF CYTOCHROMES a AND us*

The spectral properties of cytochrome c oxidase have been the subject of a number of studies (1-14). Although these studies have not, in themselves, convinced all investigators that more than one hemoprotein was involved, they have confirmed the original observations which formed the basis for distinguishing the cytochromes a and u3. These observations by Keilin and Hartree (1) indicated that the absorption at 605 and 444 rnp was due to two components. One of these, which they termed cytochrome u3, was autoxidizable and combined with carbon monoxide and cyanide, causing a spectral shift. The other, cytochrome a, which was not autoxidizable and did not combine with these reagents, showed no spectral alteration. Some investigators (15-19) have interpreted the spectral data obtained with purified cytochrome c oxidase as indicating that only a single cytochrome is present. Wainio (15,16) has further suggested that the copper present in the purified preparations may account for the spectral changes observed with carbon monoxide, cyanide, and nitric oxide. It has been held that only separation of the cytochromes a and a3 would offer satisfactory evidence for the existence of two cytochromes. However, in any study designed to isolate the cytochromes a and u3, it would be necessary to correlate the properties of isolated components with the properties of the cytochromes in the intact cytochrome c oxidase system. It is, therefore, essential to establish the properties of the cytochromes while they are still a part of that system. Yonetani (13) has recently made an effort to do this by detailed study of his purified preparation. By the use of difference spectra, he was able to distinguish quantitatively the absorption due to cytochrome a and that due to cytochrome US. In a continuation of our efforts to characterize the components (2, 3, 8, 20) of the cytochrome c oxidase, we have attempted to define further the spectral properties of cytochrome a and cytochrome u3. With this information, we should have added criteria useful in comparing preparations which contain only cytochrome a or cytochrome ~3.