Substrate-charge dependence of stoichiometry shows membrane potential is the driving force for proton-peptide cotransport in rat renal cortex

The proton dependence of the transport of three labelled, hydrolysis-resistant synthetic dipeptides carrying a net charge of −1, 0 or +1 has been investigated in a brush border membrane vesicle preparation obtained from rat renal cortex. Cross-inhibition studies are consistent with the transport of all peptides studied being through a single system. The extent and time course of uptake in response to an inwardly directed electrochemical gradient of protons differed for each peptide. For the cationic peptide D-Phe-L-Lys this gradient did not stimulate the initial rate of uptake, while for the neutral dipeptide D-Phe-L-Ala and the anionic peptide D-Phe-L-Glu stimulation was observed. However, the effect on D-Phe-L-Glu was more marked than that on D-Phe-L-Ala and the proton activation differed for these two peptides. The calculated Hill coefficients for the two proton-dependent peptides were 1.14±0.16 and 2.15±0.10 for D-Phe-L-Ala and D-Phe-L-Glu, respectively, providing evidence that the stoichiometry of proton: peptide cotransport is different for each peptide (0∶1, 1∶1 and 2∶1 for D-Phe-L-Lys, D-Phe-L-Ala and D-Phe-L-Glu respectively); studies on energetics are compatible with this conclusion. The physiological and molecular implications of this model are discussed, as are the applicability of the conclusions to secondary active transport systems more generally.