Transmembrane domain I contributes to the permeation pathway for serotoninand ions in the serotonin transporter

Mutation of a conserved Asp (D98) in the rat serotonin (5HT) transporter (r SERT) to Glu (D98E) led to decreased 5HT transport capacity, diminished cou pling to extracellular Na+ and Cl-, and a selective loss of antagonist pote ncies (cocaine, imipramine, and citalopram but not paroxetine or mazindol) with no change in 5HT K-m value. D98E, which extends the acidic side chain by one carbon, affected the rank-order potency of substrate analogs for inh ibition of 5HT transport, selectively increasing the potency of two analogs with shorter alkylamine side chains, gramine, and dihydroxybenzylamine. D9 8E also increased the efficacy of gramine relative to 5HT for inducing subs trate-activated currents in Xenopus laevis oocytes, but these currents were noticeably dependent on extracellular medium acidification. I-V profiles f or substrate-independent and -dependent currents indicated that the mutatio n selectively impacts ion permeation coupled to 5HT occupancy. The ability of the D98E mutant to modulate selective aspects of substrate recognition, to perturb ion dependence as well as modify substrate-induced currents, sug gests that transmembrane domain I plays a critical role in defining the per meation pathway of biogenic amine transporters.