A dynamic and screening-compatible nanoluciferase-based complementation assay enables profiling of individual GPCR-G...

G protein-coupled receptors (GPCRs) are currently the target of more than 30% of the marketed medicines. However, there is an important medical need for ligands with improved pharmacological activities on validated drug targets. Moreover, most of these ligands remain poorly characterized, notably because of a lack of pharmacological tools. Thus, there is an important demand for innovative assays that can detect and drive the design of compounds with novel or improved pharmacological properties. In particular, a functional and screening-compatible GPCR-G protein interaction assay is still unavailable. Here, we report on a nanoluciferase-based complementation technique to detect ligands that promote a GPCR-G protein interaction. We demonstrate that our system can be used to profile compounds with regard to the G proteins they activate through a given GPCR. Furthermore, we established a proof of applicability of screening for distinct G proteins on dopamine receptor D₂ whose differential coupling to Gα_i/o family members has been extensively studied. In a D₂-Gα_i1versus D₂-Gα_o screening, we retrieved five agonists that are currently being used in antiparkinsonian medications. We determined that in this assay, piribedil and pergolide are full agonists for the recruitment of Gα_i1 but are partial agonists for Gα_o, that the agonist activity of ropinirole is biased in favor of Gα_i1 recruitment, and that the agonist activity of apomorphine is biased for Gα_o We propose that this newly developed assay could be used to develop molecules that selectively modulate a particular G protein pathway.