Mutagenesis of the GABA ρ1 receptor alters agonist affinity and channel gating

Seventeen site-directed mutations were constructed in the GABA ρ1 receptor with the aim of finding agonist binding domains common to ρ1 and ρ2 receptors but distinct from those identified in members of the family of homologous, ligand gated ion channels. Mutated cDNAs were expressed in Xenopusoocytes and tested by voltage clamp experiments. Five of the mutations abolished responsiveness to GABA. Mutation Q189H, in the conserved cystein loop, diminished apparent GABA affinity to about 1/10 of wild type values in a manner consistent with decreased allosteric cooperativity among agonist recognition sites. Mutation R316A, located in the extracellular loop between transmembrane domains II and III, increased the Hill coefficient to 3.9 in a fashion consistent with enhanced open probability of a receptor multimer.