A Role for the Protein in Internal Electron Transfer to the Catalytic Center of CytochromecOxidase

Publisher Website
Google Scholar
Abstract
Internal electron transfer (ET) to heme a3 during anaerobic reduction of oxidized bovine heart cytochrome c oxidase (CcO) was studied under conditions where heme a and CuA were fully reduced by excess hexaamineruthenium. The data show that ET to heme a3 is controlled by the state of ionization of a single protolytic residue with a pKa of 6.5 ± 0.2. On the basis of the view that ET to the catalytic site is limited by coupled proton transfer, this pKa was attributed to Glu60 which is located at the entrance of the proton-conducting K channel on the matrix side of CcO. It is proposed that Glu60 controls proton entry into the channel. However, even with this channel open, there is the second factor that regulates ET, and this is ascribed to the rate of proton diffusion in the channel. In addition, it is concluded that proton transfer in the K channel is reversibly inhibited by the detergent Triton X-100. It is also found that the rate of ET to heme a3 in the as-isolated resting enzyme and in CcO “activated” by reaction of fully reduced enzyme with O2 is the same, implying that the catalytic sites of these two forms of oxidized enzyme are essentially identical.