Analysis of β(2)AR-G(s) and β(2)AR-G(i) complex formation by NMR spectroscopy

The β(2)-adrenergic receptor (β(2)AR) is a prototypical G protein-coupled receptor (GPCR) that preferentially couples to the stimulatory G protein G(s) and stimulates cAMP formation. Functional studies have shown that the β(2)AR also couples to inhibitory G protein G(i), activation of which inhibits cAMP formation [R. P. Xiao, Sci. STKE 2001, re15 (2001)]. A crystal structure of the β(2)AR-G(s) complex revealed the interaction interface of β(2)AR-G(s) and structural changes upon complex formation [S. G. Rasmussen et al., Nature 477, 549–555 (2011)], yet, the dynamic process of the β(2)AR signaling through G(s) and its preferential coupling to G(s) over G(i) 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 β(2)AR in response to the full agonist BI-167107 and G(s) and G(i1). 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 β(2)AR are qualitatively the same for G(s) and G(i1), we detected distinct differences between the β(2)AR-G(s) and the β(2)AR-G(i1) complex in intracellular loop 2 (ICL2). Interactions with ICL2 are essential for activation of G(s). These differences between the β(2)AR-G(s) and β(2)AR-G(i1) complexes in ICL2 may be key determinants for G protein coupling selectivity.