Studying Ligand Efficacy at G Protein-Coupled Receptors Using FRET

Drug “ligands” that bind G protein-coupled receptors (GPCRs) can either stimulate, fully (full agonists) or partially (partial agonists), or reduce (inverse agonists) basal receptor activity, by stabilizing different receptor conformations. The term “intrinsic efficacy” was introduced as a parameter to express the ability of a ligand to activate its receptor and to differentiate the varying signaling capacity of diverse ligands when they occupy the same fraction of a single receptor. Most methods use downstream biochemical and physiological responses as proxies of “intrinsic efficacy” but cannot measure it directly at the level of the receptor. Here I describe the development of a F�rster resonance energy transfer (FRET) approach that permits the rigorous measurement of the intrinsic efficacy of a ligand directly at the level of a GPCR and independent from variation in experimental conditions. This approach also allows intrinsic efficacies of ligands to be linked with the effects of receptor polymorphisms or receptor heterodimerization.