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

Schihada, Hannes; Vandenabeele, Sylvie; Zabel, Ulrike; Frank, Monika; Lohse, Martin J.; Maiellaro, Isabella

Abstract

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 α2A-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 β2-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.

Citation

Schihada, H., Vandenabeele, S., Zabel, U., Frank, M., Lohse, M. J., & Maiellaro, I. (2018). A universal bioluminescence resonance energy transfer sensor design enables high-sensitivity screening of GPCR activation dynamics. Communications Biology, 1, Article 105. https://doi.org/10.1038/s42003-018-0072-0

• Journal Article Type: Article
• Acceptance Date: May 11, 2018
• Online Publication Date: Aug 7, 2018
• Publication Date: Dec 1, 2018
• Deposit Date: Apr 1, 2020
• Publicly Available Date: Apr 1, 2020
• Journal: Communications Biology
• Electronic ISSN: 2399-3642
• Publisher: Nature Publishing Group
• Peer Reviewed: Peer Reviewed
• Volume: 1
• Article Number: 105
• DOI: https://doi.org/10.1038/s42003-018-0072-0
• Public URL: https://nottingham-repository.worktribe.com/output/4230650
• Publisher URL: https://www.nature.com/articles/s42003-018-0072-0
• Additional Information: Received: 28 November 2017; Accepted: 11 May 2018; First Online: 7 August 2018; : The authors declare that The University of Würzburg holds a patent on this technology: WO2004057333 A1.