Thermodynamic aspects of translocation of reducing equivalents by mitochondria

The "poise" in which there is no net transport of reducing equivalents mediated by glutamate and aspartate movements across the mitochondrial membrane has been studied using nonrespiring liver mitochondria, supplied with a given extramitochondrial ATP/ADP ratio. Absorbance shifts in safranine have been used as a probe to estimate the membrane potential generated under these conditions. Measurements of the asymmetric distribution of substrate anions and of reducing potential across the limiting membrane gave estimated differences of between 2.2 and 3.3 kcal when the ATP/ADP ratio in the suspending medium was about 30. The estimated membrane potential under these conditions was about 140 mV. All of the above parameters were decreased in a corresponding manner when the ATP/ADP ratio imposed from outside was decreased. A strong inflection in the slope of correlated parameters was obtained when the ATP/ADP ratio was below about 0.5. On the other hand, there was a linear relationship between the membrane potential and the asymmetric poise of reducing equivalents and of carbon metabolites. It is concluded that the poise of reducing equivalents mediated by the malate-aspartate shuttle is determined principally by the membrane potential through its effect on the electrogenic exchange of glutamate and aspartate.