G protein β 4 as a structural determinant of enhanced nucleotide exchange in the A 2A AR-Gs complex.

Adenosine A 2A receptors (A 2A AR) evoke pleiotropic intracellular signaling events via activation of the stimulatory heterotrimeric G protein, Gs. Here, we used cryoEM to solve the agonist-bound structure of A 2A AR in a complex with full-length Gs α and Gβ 4 γ 2 (A 2A AR-Gs α:β 4 γ 2 ). The orthosteric binding site of A 2A AR-Gs α:β 4 γ 2 was similar to other structures of agonist-bound A 2A AR, with or without Gs. Unexpectedly, the solvent accessible surface area within the interior of the complex was substantially larger for the complex with Gβ 4 versus the closest analog, A 2A AR-miniGs α:β 1 γ 2 . Consequently, there are fewer interactions between the switch II in Gs α and the Gβ 4 torus. In reconstitution experiments Gβ 4 γ 2 displayed a ten-fold higher efficiency over Gβ 1 γ 2 in catalyzing A 2A AR dependent GTPγS binding to Gs α. We propose that the less constrained switch II in A 2A AR-Gs α:β 4 γ 2 accounts for this increased efficiency. These results suggest that Gβ 4 functions as a positive allosteric enhancer versus Gβ 1 .