Résumé
The γ-amino-n-butyric acid type B (GABAB) receptor is composed of two subunits, GABAB1 and GABAB2, belonging to the family 3 heptahelix receptors. These proteins possess two domains, a seven transmembrane core and an extracellular domain containing the agonist binding site. This binding domain is likely to fold like bacterial periplasmic binding proteins that are constituted of two lobes that close upon ligand binding. Here, using molecular modeling and site-directed mutagenesis, we have identified residues in the GABAB1 subunit that are critical for agonist binding and activation of the heteromeric receptor. Our data suggest that two residues (Ser246 and Asp471) located within lobe I form H bonds and a salt bridge with carboxylic and amino groups of GABA, respectively, demonstrating the pivotal role of lobe I in agonist binding. Interestingly, our data also suggest that a residue within lobe II (Tyr366) interacts with the agonists in a closed form model of the binding domain, and its mutation into Ala converts the agonist baclofen into an antagonist. These data demonstrate the pivotal role played by the GABAB1 subunit in the activation of the heteromeric GABAB receptor and are consistent with the idea that a closed state of the binding domain of family 3 receptors is required for their activation.