HDL-like discs for assaying membrane proteins in drug discovery

• Published in: Biophysical chemistry 2012-05, Vol.165-166, p.56-61
• Main Authors: Fiez-Vandal, Cédric, Leder, Lukas, Freuler, Felix, Sykes, David, Charlton, Steven J., Siehler, Sandra, Schopfer, Ulrich, Duckely, Myriam
• Format: Article
• Language: English
• Subjects: Adrenergic beta-2 Receptor Antagonists - metabolism; Animals; Apolipoprotein A-I - metabolism; Beta-2-adrenoceptor; Drug Discovery; G-protein coupled receptor; HEK293 Cells; Humans; Kinetics; Lipid Bilayers - chemistry; Lipid Bilayers - metabolism; Lipoproteins, HDL - metabolism; Membrane protein; Models, Molecular; Protein Binding; Protein Conformation; Receptors, Adrenergic, beta-2 - chemistry; Receptors, Adrenergic, beta-2 - metabolism; Recombinant high-density lipoprotein; Recombinant Proteins - chemistry; Recombinant Proteins - metabolism; Scintillation proximity assay
• ISSN: 0301-4622; 1873-4200
• DOI: 10.1016/j.bpc.2012.03.005

To broaden the use of the recombinant high-density lipoprotein (rHDL) approach to the characterization of lead compounds, we investigated the pharmacology of the human beta-2-adrenoceptor in nanolipid bilayers (rHDL) with a broad set of beta-adrenoceptor antagonists. To that end, we developed a homogeneous copper-chelate scintillation proximity binding assay (SPA) in order to compare receptor-ligand binding affinities before and after reconstitution into rHDLs. Our results clearly show that the beta-2-adrenoceptor reconstituted in rHDLs display the same pharmacology as that in cell membranes and that rHDLs can be used not only to measure affinities for a range of ligands but also to study binding kinetics. [Display omitted] ► A SPA-based homogeneous assay allowing the study of ligand binding to the β2AR. ► β2AR has the same pharmacology in rHDLs than in cell membranes for a set of antagonists. ► Antagonist binding is right-shifted to lower affinities for the solubilized receptor. ► rHDL-like discs can be used for compound screening and lead optimization.