Ligand-Dependent Perturbation of the Conformational Ensemble for the GPCR β2 Adrenergic Receptor Revealed by HDX

Mechanism of G protein-coupled receptor (GPCR) activation and their modulation by functionally distinct ligands remains elusive. Using the technique of amide hydrogen/deuterium exchange coupled with mass spectrometry, we examined the ligand-induced changes in conformational states and stability within the beta-2-adrenergic receptor (β2AR). Differential HDX reveals ligand-specific alterations in the energy landscape of the receptor's conformational ensemble. The inverse agonists timolol and carazolol were found to be most stabilizing even compared with the antagonist alprenolol, notably in intracellular regions where G proteins are proposed to bind, while the agonist isoproterenol induced the largest degree of conformational mobility. The partial agonist clenbuterol displayed conformational effects found in both the inverse agonists and the agonist. This study highlights the regional plasticity of the receptor and characterizes unique conformations spanning the entire receptor sequence stabilized by functionally selective ligands, all of which differ from the profile for the apo receptor. [Display omitted] ► Comprehensive HDX study comparing apo β2AR to β2AR in complex with five ligands ► Each complex affords a unique HDX fingerprint ► Inverse agonists and an antagonist show varying degrees of increased stability ► Full agonists increase conformational mobility in helix VIII on long timescales