Mechanistic insights into G-protein coupling with an agonist-bound G-protein-coupled receptor.

G-protein-coupled receptors (GPCRs) activate heterotrimeric G proteins by promoting guanine nucleotide exchange. Here, we investigate the coupling of G proteins with GPCRs and describe the events that ultimately lead to the ejection of GDP from its binding pocket in the Gα subunit, the rate-limiting step during G-protein activation. Using molecular dynamics simulations, we investigate the temporal progression of structural rearrangements of GDP-bound G(s) protein (G(s)·GDP; hereafter G(s)(GDP)) upon coupling to the β(2)-adrenergic receptor (β(2)AR) in atomic detail. The binding of G(s)(GDP) to the β(2)AR is followed by long-range allosteric effects that significantly reduce the energy needed for GDP release: the opening of α1-αF helices, the displacement of the αG helix and the opening of the α-helical domain. Signal propagation to the G(s) occurs through an extended receptor interface, including a lysine-rich motif at the intracellular end of a kinked transmembrane helix 6, which was confirmed by site-directed mutagenesis and functional assays. From this β(2)AR-G(s)(GDP) intermediate, G(s) undergoes an in-plane rotation along the receptor axis to approach the β(2)AR-G(s)(empty) state. The simulations shed light on how the structural elements at the receptor-G-protein interface may interact to transmit the signal over 30 à to the nucleotide-binding site. Our analysis extends the current limited view of nucleotide-free snapshots to include additional states and structural features responsible for signaling and G-protein coupling specificity.