The stoichiometry of H+ pumping in cytochrome oxidase and the mechanism of uncoupling.

It is suggested that loose coupling in free energy transducing organelles is due partly to leaks through the phospholipid bilayer (extrinsic uncoupling) and partly to "slipping" of the proton pumps (intrinsic uncoupling). The flow ratio of the redox pumps (JH/JO) measured at level flow is not affected by extrinsic uncoupling, but it will be lower the higher the extent of intrinsic uncoupling. During operation of cytochrome oxidase with ferrocyanide or N,N,N',N'-tetraphenyl-p-phenylenediamine as substrates, the rate of resting respiration depends on substrate concentration and does not exhibit control by delta muH; the available data strongly suggest that the enzyme is intrinsically uncoupled to a high and variable (substrate concentration-dependent) extent. It is concluded that flow ratios (at level flows) provide underestimates of the cytochrome oxidase pump stoichiometry.