Molecular basis for the interaction of the mammalian amino acid transporters B0AT1 and B0AT3 with their ancillary protein collectrin

Many solute carrier 6 (SLC6) family transporters require ancillary subunits to modify their expression and activity. The main apical membrane neutral amino acid transporters in mouse intestine and kidney, B0AT1 and B0AT3, require the ancillary protein collectrin or ACE2 for plasma membrane expression. Expression and activity of SLC6 neurotransmitter transporters are modulated by interaction with syntaxin 1A. Utilizing monocarboxylate-B⁰AT1/3 fusion constructs, we discovered that collectrin is also necessary for B⁰AT1 and B⁰AT3 catalytic function. Syntaxin 1A and syntaxin 3 inhibit the membrane expression of B0AT1 by competing with collectrin for access. A mutagenesis screening approach identified residues on trans-membrane domains 1α, 5, and 7 on one face of B0AT3 as a key region involved in interaction with collectrin. Mutant analysis established residues that were involved in collectrin-de-pendent functions as follows: plasma membrane expression of B0AT3, catalytic activation, or both. These results identify a potential binding site for collectrin and other SLC6 ancillary proteins.