χ-Coflotoxifl and Tricyclic Antidepressant Interactions at the Norepinephrine Transporter Define a New Transporter Model.

Monoamine neurotransmitter transporters for norepinephrine (NE), dopamine and serotonin are important targets for antidepressants and analgesics. The conopeptide χ-MrIA is a noncompetitive and highly selective inhibitor of the NE transporter (NET) and is being developed as a novel intrathecal analgesic. We used site-directed mutagenesis to generate a suite of mutated transporters to identify two amino acids (Leu469 and Glu382) that affected the affinity of χ-MrIA to inhibit [³H]NE uptake through human NET. Residues that increased the Kd of a tricyclic antidepressant (nisoxetine) were also identified (Phe207, Ser225, His296, Thr381, and Asp473), Phe207, Ser225, His296, and Thr381 also affected the rate of NE transport without affecting NE Km. In a new model of NET constructed from the bLeuT crystal structure, χ-MrIA-interacting residues were located at the mouth of the transporter near residues affecting the binding of small molecule inhibitors. [ABSTRACT FROM AUTHOR]