Conformationally sensitive proximity of extracellular loops 2 and 4 of the γ-aminobutyric acid (GABA) transporter GAT-1 inferred from paired cysteine mutagenesis.

ABSTRACT: The sodium- and chloride-coupled GABA transporter GAT-1 is a member of the neurotransmitter:sodium:symporters, which are crucial for synaptic transmission. Structural work on the bacterial homologue LeuT suggests that extracellular loop 4 closes the extracellular solvent pathway when the transporter becomes inward-facing. To test whether this model can be extrapolated to GAT-1, cysteine residues were introduced at positions 359 and 448 of extracellular loop 4 and transmembrane helix 10, respectively. Treatment of HeLa cells, expressing the double cysteine mutant S359C/K448C with the oxidizing reagent copper(II)(1,10-phenantroline)3, resulted in a significant inhibition of [(3)H]GABA transport. However, transport by the single cysteine mutant S359C was also inhibited by the oxidant, whereas

SUBMITTER: Hilwi M 

PROVIDER: S-EPMC4256357 | biostudies-literature | 2014 Dec

REPOSITORIES: biostudies-literature