Letter | Published:

Propagation of conformational changes during μ-opioid receptor activation

Nature volume 524, pages 375378 (20 August 2015) | Download Citation

Abstract

µ-Opioid receptors (µORs) are G-protein-coupled receptors that are activated by a structurally diverse spectrum of natural and synthetic agonists including endogenous endorphin peptides, morphine and methadone. The recent structures of the μOR in inactive1 and agonist-induced active states (Huang et al., ref. 2) provide snapshots of the receptor at the beginning and end of a signalling event, but little is known about the dynamic sequence of events that span these two states. Here we use solution-state NMR to examine the process of μOR activation using a purified receptor (mouse sequence) preparation in an amphiphile membrane-like environment. We obtain spectra of the μOR in the absence of ligand, and in the presence of the high-affinity agonist BU72 alone, or with BU72 and a G protein mimetic nanobody. Our results show that conformational changes in transmembrane segments 5 and 6 (TM5 and TM6), which are required for the full engagement of a G protein, are almost completely dependent on the presence of both the agonist and the G protein mimetic nanobody, revealing a weak allosteric coupling between the agonist-binding pocket and the G-protein-coupling interface (TM5 and TM6), similar to that observed for the β2-adrenergic receptor3. Unexpectedly, in the presence of agonist alone, we find larger spectral changes involving intracellular loop 1 and helix 8 compared to changes in TM5 and TM6. These results suggest that one or both of these domains may play a role in the initial interaction with the G protein, and that TM5 and TM6 are only engaged later in the process of complex formation. The initial interactions between the G protein and intracellular loop 1 and/or helix 8 may be involved in G-protein coupling specificity, as has been suggested for other family A G-protein-coupled receptors.

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Acknowledgements

We acknowledge support from INSERM (S.G.) and CNRS (H.D.) and from the National Institutes of Health Grant (NIDA-DA036246 to B.K.K. and S.G.). We also acknowledge the National Institute of Drug Abuse Drug Supply Program for providing [Dmt1]DALDA.

Author information

Affiliations

  1. Institut de Genomique Fonctionnelle, CNRS UMR-5203 INSERM U1191, University of Montpellier, F-34000 Montpellier, France

    • Rémy Sounier
    • , Camille Mas
    •  & Sébastien Granier
  2. Structural Biology Brussels, Vrije Universiteit Brussel, Pleinlaan 2, B-1050 Brussels, Belgium

    • Jan Steyaert
    •  & Toon Laeremans
  3. Structural Biology Research Center, VIB, Pleinlaan 2, B-1050 Brussels, Belgium

    • Jan Steyaert
    •  & Toon Laeremans
  4. Department of Molecular and Cellular Physiology, Stanford University School of Medicine, Stanford, California 94305, USA

    • Aashish Manglik
    • , Weijiao Huang
    •  & Brian K. Kobilka
  5. Centre de Biochimie Structurale, CNRS UMR 5048-INSERM 1054- University of Montpellier, 29 rue de Navacelles, 34090 Montpellier Cedex, France

    • Héléne Déméné

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Contributions

R.S., H.D., A.M., W.H., B.K.K. and S.G. designed experiments, performed research and analysed data. C.M. expressed, purified and characterized receptor and nanobody preparations. J.S. and T.L. developed the G protein mimetic nanobodies. H.D. supervised NMR data analysis. S.G. and R.S. prepared the manuscript with the help of H.D. and B.K.K. S.G. supervised the overall project.

Competing interests

A.M., T.L., J.S. and B.K.K. have filed a patent for active-state stabilizing nanobodies for opioid receptors.

Corresponding authors

Correspondence to Héléne Déméné or Sébastien Granier.

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DOI

https://doi.org/10.1038/nature14680

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