Control of mitochondrial respiration: A quantitative evaluation of the roles of cytochrome c and oxygen

The dependence of the mitochondrial respiratory rate on the reduction of cytochrome c has been measured as a function of the exogenous [ATP] [ADP][P_i] ratio and pH. The respiratory rate at [ADP] [ADP][P_i] values of less than 10^-1 m^-1 is proportional to the reduction of cytochrome c and independent of pH from pH 6.5 to pH 8.O. The maximal turnover number (at 100% reduction) for cytochrome c is approximately 70 s^-1. As the [ATP] [ADP][P_i] ratio is increased from 10^-1 m^-1 to 10⁴ m^-1, the respiration at any given level of reduction of cytochrome c is progressively inhibited. Greater inhibition is observed at more oxidized levels of cytochorme c with respiratory control values for oxidation of reduced cytochrome c exceeding 10. The behavior of mitochondrial respiratory control is shown to be quantitatively consistent with a proposed mechanism in which the regulation occurs in the reaction of oxygen with cytochrome oxidase. A steady-state rate expression is derived which fits the mitochondrial respiratory rate dependence on (i) the extramitochondrial [ATP] [ADP][P_i] ratio; (ii) the level of reduction of cytochrome c (or the intramitochondrial [NAD⁺] [NADH]) at different [ATP] [ADP][P_i] values; (iii) the pH of the suspending medium. This rate expression appears to correctly predict the relationships of the cytoplasmic [ATP] [ADP][P_i] ratio, the mitochondrial [NAD⁺] [NADH] ratio, and the mitochondrial respiratory rate in intact cells as well as suspensions of isolated mitochondria.