Global insights into the fine tuning of human A(2A)AR conformational dynamics in a ternary complex with an engineered G protein viewed by NMR

G protein-coupled receptor (GPCR) conformational plasticity enables formation of ternary signaling complexes with intracellular proteins in response to binding extracellular ligands. We investigate the dynamic process of GPCR complex formation in solution with the human A(2A) adenosine receptor (A(2A)AR) and an engineered G(s) protein, mini-G(s). 2D nuclear magnetic resonance (NMR) data with uniform stable isotope-labeled A(2A)AR enabled a global comparison of A(2A)AR conformations between complexes with an agonist and mini-G(s) and with an agonist alone. The two conformations are similar and show subtle differences at the receptor intracellular surface, supporting a model whereby agonist binding alone is sufficient to populate a conformation resembling the active state. However, an A(2A)AR “hot spot” connecting the extracellular ligand-binding pocket to the intracellular surface is observed to be highly dynamic in the ternary complex, suggesting a mechanism for allosteric connection between the bound G protein and the drug-binding pocket involving structural plasticity of the “toggle switch” tryptophan.