title: Suicide inactivation of cytochrome c oxidase: catalytic turnover in the absence of subunit III alters the active site.
abstract: The catalytic core of cytochrome c oxidase is composed of three subunits: I, II, and III. Subunit III is a highly hydrophobic membrane protein that contains no redox centers; its role in cytochrome oxidase function is not obvious. Here, subunit III has been removed from the three-subunit mitochondrial-like oxidase of Rhodobacter sphaeroides by detergent washing. The resulting two-subunit oxidase, subunit III (-), is highly active. Ligand-binding analyses and resonance Raman spectroscopy show that its heme a(3)-Cu(B) active site is normal. However, subunit III (-) spontaneously and irreversibly inactivates during O(2) reduction. At pH 7.5, its catalytic lifetime is only 2% that of the normal oxidase. This suicide inactivation event primarily alters the active site. Its ability to form specific O(2) reduction intermediates is lost, and CO binding experiments suggest that the access of O(2) to reduced heme a(3) is inhibited. Reduced heme a accumulates in response to a decrease in the redox potential of heme a(3); electron transfer between the hemes is inhibited. Ligand-binding experiments and resonance Raman analysis show that increased flexibility in the structure of the active site accompanies inactivation. Cu(B) is partially lost. It is proposed that suicide inactivation results from the dissociation of a ligand of Cu(B) and that subunit III functions to prevent suicide inactivation by maintaining the structural integrity of the Cu(B) center via long-range interactions.
authors.affiliation: Department of Biochemistry, University of Mississippi Medical Center, Jackson 39216, USA.
authors.forename: M R
authors.initials: MR
authors.lastname: Bratton
authors.forename: M A
authors.initials: MA
authors.lastname: Pressler
authors.forename: J P
authors.initials: JP
authors.lastname: Hosler
mesh_terms: D001665:Binding Sites; D002384:Catalysis; D003300:Copper / Q000378:metabolism; D003576:Electron Transport Complex IV / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000378:metabolism*; D004789:Enzyme Activation; D006418:Heme / Q000031:analogs & derivatives / Q000378:metabolism; D006358:Hot Temperature; D006861:Hydrogen Peroxide / Q000378:metabolism; D017830:Octoxynol; D010084:Oxidation-Reduction; D010446:Peptide Fragments / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000172:deficiency* / Q000378:metabolism*; D010743:Phospholipids / Q000378:metabolism; D011522:Protons; D012242:Rhodobacter sphaeroides / Q000201:enzymology; D013059:Spectrum Analysis, Raman; D013481:Superoxides / Q000378:metabolism
chemical_list: D010446:Peptide Fragments; D010743:Phospholipids; D011522:Protons; D013481:Superoxides; C027728:heme a; D006418:Heme; D003300:Copper; D017830:Octoxynol; D006861:Hydrogen Peroxide; D003576:Electron Transport Complex IV