Functional GhrR was site-specifically tagged with the unnatural amino acid, azF
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GhrR azF mutants were labeled bioorthogonally with a fluorophore using SpAAC
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Novel assays were developed to probe binding of fluorescent ligands to labeled GhrR
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Ligand-induced conformational changes could be detected in fluorescent GhrR
Summary
Ghrelin receptor (GhrR) is a promising drug target because of its central role in energy homeostasis. GhrR, known for high constitutive activity, is thought to display multi-state conformations during activation and signaling. We used genetically encoded unnatural amino acids and bioorthogonal labeling reactions to engineer multiple fluorescent donor-acceptor pairs to probe ligand-directed structural changes in GhrR. We demonstrate how conformational dynamics of a G-protein-coupled receptor can be measured in reconstituted systems.