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Nature. 2017 Jul 6;547(7661):68-73. doi: 10.1038/nature22354. Epub 2017 Jun 7.

Single-molecule analysis of ligand efficacy in β2AR-G-protein activation.

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Department of Physiology and Biophysics, Weill Cornell Medicine, New York, New York, USA.
Department of Molecular and Cellular Physiology, Stanford University School of Medicine, Stanford, California, USA.
Department of Psychiatry, Columbia University College of Physicians and Surgeons, New York, New York, USA.
Division of Molecular Therapeutics, New York State Psychiatric Institute, New York, New York, USA.
Laboratory for Molecular Pharmacology, Department of Neuroscience and Pharmacology, University of Copenhagen, Blegdamsvej 3, Copenhagen 2200, Denmark.
NNF Center for Basic Metabolic Research, University of Copenhagen, Blegdamsvej 3, Copenhagen 2200, Denmark.
Department of Pharmacology, Columbia University College of Physicians and Surgeons, New York, New York, USA.
The HRH Prince Alwaleed Bin Talal Bin Abdulaziz Alsaud Institute for Computational Biomedicine, Weill Cornell Medical College of Cornell University, New York, New York, USA.


G-protein-coupled receptor (GPCR)-mediated signal transduction is central to human physiology and disease intervention, yet the molecular mechanisms responsible for ligand-dependent signalling responses remain poorly understood. In class A GPCRs, receptor activation and G-protein coupling entail outward movements of transmembrane helix 6 (TM6). Here, using single-molecule fluorescence resonance energy transfer imaging, we examine TM6 movements in the β2 adrenergic receptor (β2AR) upon exposure to orthosteric ligands with different efficacies, in the absence and presence of the Gs heterotrimer. We show that partial and full agonists differentially affect TM6 motions to regulate the rate at which GDP-bound β2AR-Gs complexes are formed and the efficiency of nucleotide exchange leading to Gs activation. These data also reveal transient nucleotide-bound β2AR-Gs species that are distinct from known structures, and provide single-molecule perspectives on the allosteric link between ligand- and nucleotide-binding pockets that shed new light on the G-protein activation mechanism.

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