TOP-DOWN CONTROL ANALYSIS OF TEMPERATURE EFFECT ON OXIDATIVE-PHOSPHORYLATION

effects of temperature on the control of respiration rate, phosphoryla tion rate, proton leakage rate, the protonmotive force and the effecti ve ATP/O ratio were determined in isolated rat liver mitochondria over a range of respiratory conditions by applying top-down elasticity and control analyses. Simultaneous measurements of membrane potential, ox idation and phosphorylation rates were performed under various ATP tur nover rates, ranging from state 4 to state 3. Although the activities of the three subsystems decreased with temperature (over 30-fold betwe en 37 and 4 degrees C), the effective ATP/O ratio exhibited a maximum at 25 degrees C, far below the physiological value. Top-down elasticit y analysis revealed that maximal membrane potential was maintained ove r the range of temperature studied, and that the proton leakage rate w as considerably reduced at 4 degrees C. These results definitely rule out a possible uncoupling of mitochondria at low temperature. At 4 deg rees C, the decrease in ATP/O ratio is explained by the relative decre ase in phosphorylation processes revealed by the decrease in depolariz ation after ADP addition [Diolez and Moreau (1985) Biochim. Biophys. A cta 806, 56-63]. The change in depolarization between 37 and 25 degree s C was too small to explain the decrease in ATP/O ratio. This result is best explained by the changes in the elasticity of proton leakage t o membrane potential between 37 and 25 degrees C, leading to a higher leak rate at 37 degrees C for the same value of membrane potential. To p-down control analysis showed that despite the important changes in a ctivities of the three subsystems between 37 and 25 degrees C, the pat terns of the control distribution are very similar. However, a differe nt pattern was obtained at 4 degrees C under all phosphorylating condi tions. Surprisingly, control by the proton leakage subsystem was almos t unchanged, although both control patterns by substrate oxidation and phosphorylation subsystems were affected at 4 degrees C. In compariso n with results for 25 and 37 degrees C, at 4 degrees C there was evide nce for increased control by the phosphorylation subsystem over both f luxes of oxidation and phosphorylation as well as on the ATP/O ratio w hen the system is close to state 3. However, the pattern of control co efficients as a function of mitochondrial activity also showed enhance d control exerted by the substrate oxidation subsystem under all inter mediate conditions. These results suggest that passive membrane permea bility to protons is not involved in the effect of temperature on the control of oxidative phosphorylation.