Cytochrome c oxidase: the mechanistic significance of structural H+ in energy transduction.
Changes in the bulk-phase concentration of O2 and H+ associated with the reduction of O2 to water are simultaneously determined in reactions catalyzed by fully reduced cytochrome c oxidase both isolated and embedded in liposomes. Consistent with the polyphasic kinetics of electron transfer through the oxidase, the time course of O2 consumption and H+ translocation exhibit the following novel characteristics: (1) The uptake of scalar protons (Hm+), the ejection of vectorial protons (Hv+), and the consumption of O2, all proceed in a kinetically polyphasic process. (2) During thefirst phase of the reaction the rates of O2 uptake and H+ transfer are extremely fast and compatible with the rates of electron flow through the oxidase. (3) The Km of the oxidase for O2 is close to 75 microM, the same for O2 consumption and scalar H+ uptake. The Vmax of O2 reduction to water in reactions catalyzed by the isolated enzyme is, at least, 0.5 x 10(4) s(-1). (4) The extent of vectorial H+ ejection by cytochrome c oxidase embedded in liposomes is an exponential function dependent on both enzyme concentration and extent of O2 consumption. (5) The H+/O stoichiometry of H+ ejection is a variable that may reach a maximum value of 4.0 only when the enzyme undergoes net oxidation at extremely high enzyme/O2 molar ratios. It is postulated that the generation of useful energy at the level of cytochrome c oxidase depends not only on the number of molecules of O2 reduced to water but also on the extent and state of reduction and/or protonation of the enzyme.
Reynafarje
Baltazar
B
Department of Biological Chemistry, The Johns Hopkins University School of Medicine, Baltimore, Maryland 21205, USA.
Ferreira
Jorge
J
D000818:Animals; D002417:Cattle; D003576:Electron Transport Complex IV / Q000378:metabolism*; D004735:Energy Transfer; D006736:Horses; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008081:Liposomes; D008956:Models, Chemical; D010100:Oxygen / Q000378:metabolism; D011522:Protons; D014867:Water / Q000378:metabolism
D008081:Liposomes; D011522:Protons; D014867:Water; D003576:Electron Transport Complex IV; D010100:Oxygen