The B°AT1 amino acid transporter from rat kidney reconstituted in liposomes: Kinetics and inactivation by methylmercury

The neutral amino acid transporter B°-like from rat kidney, previously reconstituted in liposomes, was identified as B°AT1 by a specific antibody. Collectrin was present in the brush-border extract but not in functionally active proteoliposomes, indicating that it was not required for the transport function. Neutral amino acids behaved as competitive inhibitors of the glutamine transport mediated by B°AT1 with half saturation constants ranging from 0.13 to 4.74 mM. The intraliposomal half saturation constant for glutamine was 2.0 mM. By a bisubstrate kinetic analysis of the glutamine-Na⁺ cotransport, a random simultaneous mechanism was found. Methylmercury and HgCl₂ inhibited the transporter; the inhibition was reversed by dithioerythritol, Cys and, at a lower extent, N-acetylcysteine but not by S-carboxymethylcysteine. The IC₅₀ of the transporter for methylmercury and HgCl₂ was 1.88 and 1.75 μM, respectively. The reagents behaved as non-competitive inhibitors toward both glutamine and Na ⁺ and no protection by glutamine or Na⁺ was found for the two inhibitors.