A universal bioluminescence resonance energy transfer sensor design enables high-sensitivity screening of GPCR activation dynamics

G-protein-coupled receptors (GPCRs) represent one of the most important classes of drug targets. The discovery of new GCPR therapeutics would greatly benefit from the development of a generalizable high-throughput assay to directly monitor their activation or de-activation. Here we screened a variety of labels inserted into the third intracellular loop and the C-terminus of the α 2A -adrenergic receptor and used fluorescence (FRET) and bioluminescence resonance energy transfer (BRET) to monitor ligand-binding and activation dynamics. We then developed a universal intramolecular BRET receptor sensor design to quantify efficacy and potency of GPCR ligands in intact cells and real time. We demonstrate the transferability of the sensor design by cloning β 2 -adrenergic and PTH1-receptor BRET sensors and monitored their efficacy and potency. For all biosensors, the Z factors were well above 0.5 showing the suitability of such design for microtiter plate assays. This technology will aid the identification of novel types of GPCR ligands. Hannes Schihada et al. report the design of 11 BRET-based biosensors that allow for quantification of GPCR ligand-binding dynamics in a micro-titer format. The biosensors achieve higher dynamic range and sensitivity than FRET-based biosensors and their design can be extended to the study of other receptor types.