Inhibition of the lepidopteran amino acid co-transporter KAAT1 by phenylglyoxal : role of arginine 76

Phenylglyoxal (PGO), an arginine-modifying reagent, is an irreversible inhibitor of KAAT1-mediated leucine transport, expressed in Xenopus oocytes. The PGO effect was dose-dependent and 5 mM PGO determined a Vmax reduction to 24% of the control, consistent with the covalent binding to transporter arginine residues not located in the leucine binding site. The use of labelled [14C]PGO confirmed that the inhibitor binds KAAT1. The protein membrane domain contains seven arginine residues one of which, arginine 76, is conserved in the family of GABA transporters. Using site-directed mutagenesis we showed that only arginine 76 is crucial for KAAT1 activity and is involved in PGO binding.