D-Enantiomers Take a Close Look at the Functioning of a Cardiac Cationic L-Amino Acid Transporter

Cationic amino acid transporters are highly selective for L-enantiomers such as L-arginine (L-Arg). Because of this stereoselectivity, little is known about the interaction of these transporters with D-isomers. To study whether these compounds can provide information on the molecular mechanism of transport, inward currents activated by L-Arg with low apparent affinity were measured in whole-cell voltage-clamped cardiomyocytes as a function of extracellular L-Arg and D-Arg concentrations. D-Arg inhibited L-Arg currents in a membrane-potential (V-M)-dependent competitive manner, indicating the presence of D-Arg binding sites in the carrier. Analysis of these steady-state currents showed that L- and D-Arg binding reactions dissipate a similar small fraction of the membrane electric field. Since D-Arg is not transported, these results suggest that enantiomer recognition occurs at conformational transitions that initiate amino acid translocation. The V-M dependence of maximal current levels suggests that inward currents arise from the slow outward movement of negative charges in the unliganded transporter. Translocation of the L-Arg-bound complex, on the other hand, appears to be electroneutral. D-Arg-dependent transient charge movements, also detected in these cells, displayed a Vm-dependent charge distribution and kinetics that are consistent with amino acid binding in an ion well in a shallow, water-filled extracellular binding pocket.