STRUCTURAL BASIS OF RECEPTOR G-PROTEIN COUPLING SELECTIVITY STUDIED WITH MUSCARINIC RECEPTORS AS MODEL SYSTEMS

Different muscarinic acetylcholine receptor subtypes were used as mode l systems to study the structural basis of receptor/G protein coupling selectivity. Extensive mutagenesis studies have previously led to the identification of single amino acids on the m3 muscarinic receptor pr otein (located in the second intracellular loop (i2) and at the N- and C-terminus of the third intracellular loop (i3)) that dictate selecti ve recognition of Gq/11 proteins by this receptor subtype. Based on th ese results, we proposed a model of the intracellular m3 receptor surf ace in which the functionally critical residues project into the inter ior of the transmembrane receptor core. To identify specific regions o n the G protein(s) that are contacted by these different, functionally critical receptor sites, we recently employed a novel experimental st rategy involving the coexpression of hybrid m2/m3 muscarinic receptors with hybrid G alpha-subunits. Using this approach, we could demonstra te that the C-terminus of G protein alpha(i)/(0)-subunits is recognize d by a short sequence element in the m2 muscarinic receptor (''VTIL'') that is located at the junction between the sixth transmembrane domai n (TM VI) and the i3 loop. We could show that this interaction is crit ically involved in determining coupling selectivity and triggering G p rotein activation. By using a similar strategy (coexpression of mutant muscarinic receptors with hybrid G alpha-subunits), other major recep tor/G protein contact sites are currently being identified. These stud ies, complemented by biochemical and biophysical approaches, should ev entually lead to a detailed structural model of the ligand-receptor-G protein complex.