Conformational Complexity and Dynamics in a Muscarinic Receptor Revealed by NMR Spectroscopy

• DOI: 10.1016/j.molcel.2019.04.028
• Publication Date: 2019-05-15T14:33:30Z
• Authors: Jun Xu; Yunfei Hu; Jonas Kaindl; Philipp Risel; Harald Hübner; Shoji Maeda; Xiaogang Niu; Hongwei Li; Peter Gmeiner; Changwen Jin; Brian K. Kobilka
• Keywords: Protein Conformation, alpha-Helical; 0301 basic medicine; Binding Sites; Magnetic Resonance Spectroscopy; Pyridines; Genetic Vectors; Pilocarpine; Gene Expression; Isoxazoles; Molecular Dynamics Simulation; Ligands; Acetylcholine; Kinetics; 03 medical and health sciences; Animals; Humans; Carbachol; Protein Conformation, beta-Strand; Protein Interaction Domains and Motifs; Cloning, Molecular; Baculoviridae; Protein Binding

The M2 muscarinic acetylcholine receptor (M2R) is a prototypical GPCR that plays important roles in regulating heart rate and CNS functions. Crystal structures provide snapshots of the M2R in inactive and active states, but the allosteric link between the ligand binding pocket and cytoplasmic surface remains poorly understood. Here we used solution NMR to examine the structure and dynamics of the M2R labeled with 13CH3-ε-methionine upon binding to various orthosteric and allosteric ligands having a range of efficacy for both G protein activation and arrestin recruitment. We observed ligand-specific changes in the NMR spectra of 13CH3-ε-methionine probes in the M2R extracellular domain, transmembrane core, and cytoplasmic surface, allowing us to correlate ligand structure with changes in receptor structure and dynamics. We show that the M2R has a complex energy landscape in which ligands with different efficacy profiles stabilize distinct receptor conformations.