MOLECULAR DETERMINANT OF ION SELECTIVITY OF A (NA-COUPLED RAT-BRAIN GLUTAMATE TRANSPORTER(+K+))

Glutamate transporters remove this neurotransmitter from the synaptic cleft by a two-stage electrogenic process, in which glutamate is first cotransported with three sodium ions and a proton, Subsequently, the cycle is completed by translocation of a potassium ion in the opposite direction, Recently, we have identified an amino acid residue of the glutamate transporter GLT-1 (Glu-404) that influences potassium coupli ng, We have now analyzed the effect of seven other amino acid residues in the highly conserved region surrounding this site, One of these re sidues, Tyr-403, also proved important for potassium coupling, because mutation to Phe (Y403F) resulted in an electroneutral obligate exchan ge mode of glutamate transport, This mutation in the transporter also caused an approximately 8-fold increase in the apparent sodium affinit y, with no change in the apparent affinity for L-glutamate or D-aspart ate. Strikingly, although exchange catalyzed by the mild-type transpor ter is strictly dependent on sodium, the selectivity of Y403F mutant t ransporters is altered so that sodium can be replaced by other alkalin e metal cations including lithium and cesium, These results indicate t he presence of interacting sites in or near the transporter pore that control selectivity for sodium and potassium.