  Neurotensin receptor 1 (NTSR1) is a G protein-coupled receptor that is
important for signaling in the brain and the gut. Its agonist ligand
neurotensin (NTS), a 13-amino-acid peptide, binds with nanomolar affinity from
the extracellular side to NTSR1 and induces conformational changes that trigger
intracellular signaling processes. Our goal is to monitor the conformational
dynamics of single fluorescently labeled NTSR1. For this, we fused the
fluorescent protein mNeonGreen to the C terminus of NTSR1, purified the
receptor fusion protein from E. coli membranes, and reconstituted NTSR1 into
liposomes with E. coli polar lipids. Using single-molecule anisotropy
measurements, NTSR1 was found to be monomeric in liposomes, with a small
fraction being dimeric and oligomeric, showing homoFRET. Similar results were
obtained for NTSR1 in detergent solution. Furthermore, we demonstrated agonist
binding to NTSR1 by time-resolved single-molecule F\"orster resonance energy
transfer (smFRET), using neurotensin labeled with the fluorophore ATTO594.
