A role for subunit III in proton uptake into the D pathway and a possible proton exit pathway in Rhodobacter sphaeroides cytochrome c oxidase.
Protons are transferred from the inner surface of cytochrome c oxidase to the active site by the D and K pathways, as well as from the D pathway to the outer surface by a largely undefined proton exit route. Alteration of the initial proton acceptor of the D pathway, D132, to alanine has previously been shown to greatly inhibit oxidase turnover and slow proton uptake into the D pathway. Here it is shown that the removal of subunit III restores a substantial rate of O(2) reduction to D132A. Presumably an alternative proton acceptor for the D pathway becomes active in the absence of subunit III and D132. Thus, in the absence of subunit III cytochrome oxidase shows greater flexibility in terms of proton entry into the D pathway. In the presence of DeltaPsi and DeltapH, turnover of the wild-type oxidase or D132A is slower in the absence of subunit III. Comparison of the turnover rates of subunit III-depleted wild-type oxidase to those of the zinc-inhibited wild-type oxidase containing subunit III, both reconstituted into vesicles, leads to the hypothesis that the absence of subunit III inhibits the ability of the normal proton exit pathway to take up protons from the outside in the presence of DeltaPsi and DeltapH. Thus, subunit III appears to affect the transfer of protons from both the inner and outer surfaces of cytochrome oxidase, perhaps accounting for the long-observed lower efficiency of proton pumping by the subunit III-depleted oxidase.
Mills
Denise A
DA
Department of Biochemistry, Michigan State University, East Lansing, Michigan 48824, USA.
Tan
Zi
Z
Ferguson-Miller
Shelagh
S
Hosler
Jonathan
J
D019943:Amino Acid Substitution; D001665:Binding Sites; D003576:Electron Transport Complex IV / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005231:Fatty Acids, Unsaturated / Q000737:chemistry / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D010084:Oxidation-Reduction; D010743:Phospholipids / Q000737:chemistry / Q000378:metabolism; D021122:Protein Subunits; D011522:Protons; D012242:Rhodobacter sphaeroides / Q000201:enzymology*; D013053:Spectrophotometry / Q000379:methods; D015032:Zinc / Q000494:pharmacology
D004791:Enzyme Inhibitors; D005231:Fatty Acids, Unsaturated; D010743:Phospholipids; D021122:Protein Subunits; D011522:Protons; D003576:Electron Transport Complex IV; D015032:Zinc