Analysis of βAR-G and βAR-G complex formation by NMR spectroscopy.

The β-adrenergic receptor (βAR) is a prototypical G protein-coupled receptor (GPCR) that preferentially couples to the stimulatory G protein G and stimulates cAMP formation. Functional studies have shown that the βAR also couples to inhibitory G protein G, activation of which inhibits cAMP formation [R. P. Xiao, Sci. STKE , re15 (2001)]. A crystal structure of the βAR-G complex revealed the interaction interface of βAR-G and structural changes upon complex formation [S. G. Rasmussen et al., Nature 477, 549-555 (2011)], yet, the dynamic process of the βAR signaling through G and its preferential coupling to G over G is still not fully understood. Here, we utilize solution nuclear magnetic resonance (NMR) spectroscopy and supporting molecular dynamics (MD) simulations to monitor the conformational changes in the G protein coupling interface of the βAR in response to the full agonist BI-167107 and G and G These results show that BI-167107 stabilizes conformational changes in four transmembrane segments (TM4, TM5, TM6, and TM7) prior to coupling to a G protein, and that the agonist-bound receptor conformation is different from the G protein coupled state. While most of the conformational changes observed in the βAR are qualitatively the same for G and G, we detected distinct differences between the βAR-G and the βAR-G complex in intracellular loop 2 (ICL2). Interactions with ICL2 are essential for activation of G These differences between the βAR-G and βAR-G complexes in ICL2 may be key determinants for G protein coupling selectivity.