Neutral amino acid transporter ASCT2 displays substrate-induced Na+ exchange and a substrate-gated anion conductance

The neutral amino acid transporter ASCT2 mediates electroneutral obligatory antiport but at the same time requires Na+ for its function. To elucidate the mechanism, ASCT2 was expressed in Xenopus laevis oocytes and transport was analysed by flux studies and two-electrode voltage clamp recordings. Fl ux studies with (NaCl)-Na-22 indicated that the uptake of one molecule of g lutamine or alanine is accompanied by the uptake of four to seven Na+ ions. Similarly to the transport of amino acids, the Na+ uptake was mediated by an obligatory Na+ exchange mechanism that depended on the presence of amino acids but was not stoichiometrically coupled to the amino acid transport. Other cations could not replace Na+ in this transport mechanism. When NaCl was replaced by NaSCN in the transport buffer, the superfusion of oocytes w ith amino acid substrates resulted in large inward currents, indicating the presence of a substrate-gated anion channel in the ASCT2 transporter. The K-m for glutamine derived from these experiments is in good agreement with the K-m, derived from flux studies; it varied between 40 and 90 mu M at hol ding potentials of - 60 and - 20 mV respectively. The permeability of the s ubstrate-gated anion conductance decreased in the order SCN- much greater t han NO3- > I- > Cl- and also required the presence of Na+.