Although the present methodology concerns the measurement of ADP-ATP exchange rate mediated by the ANT (i.e. the ANT activity), mindful that the ANT is a reversible transporter, I outline a few details regarding its directionality. To put it in the most naive way, the ANT can transport ADP inside and ATP outside the mitochondria (forward mode), or ADP outside and ATP inside the mitochondria (reverse mode). The membrane potential value at which there is no net transfer of adenine nucleotides is called the reversal potential of the ANT, designated as Erev_ANT. Erev_ANT depends exclusively on the concentration of the ANT reactants, i.e. ADP and ATP across the inner mitochondrial membrane. By thermodynamic deduction, Erev_ANT is equal to:
Just like Erev_ANT, F0-F1 ATP synthase has its own reversal potential, designated as Erev_ATPase. By thermodynamic deduction, Erev_ATPase is equal to:
where “in” and “out” signify inside and outside the mitochondrial matrix, respectively, R is the universal gas constant 8.31 J*mol-1*K-1, F is the Faraday constant 9.64*104 C·mol-1, and T is temperature (in Kelvin).
PLEASE DO NOT CONFUSE ANT ACTIVITY WITH ANT DIRECTIONALITY.
where “n” is the H+/ATP coupling ratio (Watt et al., 2010), and [P-] is the free phosphate concentration (in Molar), where pKa2=7.2 for phosphoric acid.
PLEASE DO NOT CONFUSE ATPase ACTIVITY WITH ATPase DIRECTIONALITY.