Letter | Published:

Conformational dynamics of a class C G-protein-coupled receptor

Nature volume 524, pages 497501 (27 August 2015) | Download Citation

Abstract

G-protein-coupled receptors (GPCRs) constitute the largest family of membrane receptors in eukaryotes. Crystal structures have provided insight into GPCR interactions with ligands and G proteins1,2, but our understanding of the conformational dynamics of activation is incomplete. Metabotropic glutamate receptors (mGluRs) are dimeric class C GPCRs that modulate neuronal excitability, synaptic plasticity, and serve as drug targets for neurological disorders3,4. A ‘clamshell’ ligand-binding domain (LBD), which contains the ligand-binding site, is coupled to the transmembrane domain via a cysteine-rich domain, and LBD closure seems to be the first step in activation5,6. Crystal structures of isolated mGluR LBD dimers led to the suggestion that activation also involves a reorientation of the dimer interface from a ‘relaxed’ to an ‘active’ state7,8, but the relationship between ligand binding, LBD closure and dimer interface rearrangement in activation remains unclear. Here we use single-molecule fluorescence resonance energy transfer to probe the activation mechanism of full-length mammalian group II mGluRs. We show that the LBDs interconvert between three conformations: resting, activated and a short-lived intermediate state. Orthosteric agonists induce transitions between these conformational states, with efficacy determined by occupancy of the active conformation. Unlike mGluR2, mGluR3 displays basal dynamics, which are Ca2+-dependent and lead to basal protein activation. Our results support a general mechanism for the activation of mGluRs in which agonist binding induces closure of the LBDs, followed by dimer interface reorientation. Our experimental strategy should be widely applicable to study conformational dynamics in GPCRs and other membrane proteins.

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Acknowledgements

We thank Z. Fu and H. Okada for technical assistance, J. P. Pin for generously providing the SNAP- and CLIP-tagged mGluRs and advice on their properties, and J. P. Pin, E. Margeat, P. Rondard, A. Jain, A. Reiner and members of the Isacoff laboratory for discussions. Funding was provided by the National Institutes of Health Nanomedicine Development Center for the Optical Control of Biological Function (2PN2EY018241) and the National Science Foundation (EAGER: IOS-1451027). R.V. is a Merck fellow of the Life Science Research Foundation.

Author information

Author notes

    • Reza Vafabakhsh
    •  & Joshua Levitz

    These authors contributed equally to this work.

Affiliations

  1. Department of Molecular and Cell Biology, University of California, Berkeley, California 94720, USA

    • Reza Vafabakhsh
    • , Joshua Levitz
    •  & Ehud Y. Isacoff
  2. Helen Wills Neuroscience Institute, University of California, Berkeley, California 94720, USA

    • Ehud Y. Isacoff
  3. Physical Bioscience Division, Lawrence Berkeley National Laboratory, Berkeley, California 94720, USA

    • Ehud Y. Isacoff

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Contributions

R.V., J.L. and E.Y.I. designed the research. R.V. set up, performed and analysed single-molecule FRET experiments. J.L. performed and analysed ensemble FRET and electrophysiology experiments and contributed to single-molecule FRET experiments. R.V., J.L. and E.IY. wrote the paper.

Competing interests

The authors declare no competing financial interests.

Corresponding author

Correspondence to Ehud Y. Isacoff.

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DOI

https://doi.org/10.1038/nature14679

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