Allosteric interactions between GABA_B1 subunits control orthosteric binding sites occupancy within GABA_B oligomers

The GABA_B receptor was the first G protein-coupled receptor identified as an obligate heterodimer. It is composed of two subunits, GABA_B1 containing the agonist binding site and GABA_B2 responsible for G protein activation. The GABA_B receptor was found to associate into larger complexes through GABA_B1-GABA_B1 interactions, both in transfected cells and in brain membranes. Here we assessed the possible allosteric interactions between GABA_B heterodimers by analyzing the effect of mutations located at the putative interface between the extracellular binding domains. These mutations decrease, but do not suppress, the Förster resonance energy transfer (FRET) signal measured between GABA_B1 subunits. Further analysis of one of these mutations revealed an increase in G protein-coupling efficacy and in the maximal antagonist binding by approximately two-fold. Hypothesizing that a tetramer is an elementary unit within oligomers, additional FRET data using fluorescent ligands and tagged subunits suggest that adjacent binding sites within the GABA_B oligomers are not simultaneously occupied. Our data show a strong negative effect between GABA_B1 binding sites within GABA_B oligomers. Accordingly, GABA_B receptor assembly appears to limit receptor signaling to G proteins, a property that may offer novel regulatory mechanism for this important neuronal receptor. This article is part of the "Special Issue Dedicated to Norman G. Bowery".