Direct analysis of a GPCR-agonist interaction by surface plasmon resonance
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Despite their clinical importance, detailed analysis of ligand binding at G-protein coupled receptors (GPCRs) has proved difficult. Here we successfully measure the binding of a GPCR, neurotensin receptor-1 (NTS-1), to its ligand, neurotensin (NT), using surface plasmon resonance (SPR). Specific responses were observed between NT and purified, detergent-solublised, recombinant NTS-1, using a novel configuration where the biotinylated NT ligand was immobilised on the biosensor surface. This SPR approach shows promise as a generic approach for the study of ligand interactions with other suitable GPCRs.
KeywordsGPCR Neurotensin receptor Surface plasmon resonance Ligand-binding High-throughput screening
GTP-binding protein-coupled receptor
High pressure liquid chromatography
Escherichia coli maltose binding protein
Surface plasmon resonance
We acknowledge the Medical Research Council (MRC), Biotechnology and Biological Sciences Research Council (BBSRC) and the UK Bionanotechnology IRC for funding this work. We thank GlaxoSmithKine (GSK), specifically Claus Spitzfaden, Martin Hibbs, Emma Jones, and Mark Hessey for allowing access to large scale fermentation, purification and SPR facilities. We would like to thank Dr. Simon Davis (Nuffield Department of Medicine, Oxford) for the use of his SPR machine.
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