Abstract

Bifunctional μ- and δ-opioid receptor (OR) ligands are potential therapeutic alternatives, with diminished side effects, to alkaloid opiate analgesics. We solved the structure of human δ-OR bound to the bifunctional δ-OR antagonist and μ-OR agonist tetrapeptide H-Dmt-Tic-Phe-Phe-NH2 (DIPP-NH2) by serial femtosecond crystallography, revealing a cis-peptide bond between H-Dmt and Tic. The observed receptor-peptide interactions are critical for understanding of the pharmacological profiles of opioid peptides and for development of improved analgesics.

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Acknowledgements

This work was supported by US National Institutes of Health (NIH), National Institute of General Medical Sciences grants U54 GM094618 (R.C.S., V.C. and V.K.), R01 GM108635 (V.C.), U54 GM094599 (P.F.), R01 GM095583 (P.F.) and P41 GM103393 (S. Boutet); US National Institute of Drug Abuse grants P01 DA035764 (V.C., V.K., B.L.R. and R.C.S.) and R01 DA017204 (B.L.R.); the US National Institute of Mental Health Psychoactive Drug Screening Program (P.G. and B.L.R.); the Michael Hooker Chair for Protein Therapeutics and Translational Proteomics to B.L.R.; and US National Science Foundation Science and Technology Center award 1231306 (J.C.H.S., P.F. and U.W.). Parts of this work were supported by the Helmholtz Association, the German Research Foundation (DFG) Cluster of Excellence ‘Center for Ultrafast Imaging’ and the German Federal Ministry of Education and Research (BMBF) projects FKZ 05K12CH1 (H.N.C., A.B., C.G., O.M.Y., T.A.W., D.O. and M. Metz) and 05K2012 (D.O. and H.N.C.). C.G. thanks the PIER Helmholtz Graduate School and the Helmholtz Association for financial support. M. Metz. acknowledges support from the Marie Curie Initial Training Network NanoMem (grant no. 317079). C.B., S. Ballet, D.T. and P.W.S. were supported by a collaboration convention between the Ministère du Développement Economique, de l'Innovation et de l'Exportation du Québec (PSR-SIIRI-417) and the Research Foundation–Flanders (FWO Vlaanderen, grant FWOAL570) and by grants to P.W.S. from the Canadian Institutes of Health Research (CIHR) (MOP-89716) and the NIH (DA-004443). We thank J. Velasquez, T. Trinh, M. Chu and A. Walker. Parts of this research were carried out at the Linac Coherent Light Source (LCLS), a US National User Facility operated by Stanford University on behalf of the US Department of Energy, Office of Basic Energy Sciences and at the GM/CA CAT, beamline 23ID-B, Advanced Photon Source, which is supported by the US National Cancer Institute grant Y1-CO-1020 and the US National Institute of General Medical Sciences grant Y1-GM-1104.

Author information

Author notes

    • Gustavo Fenalti
    • , Gye Won Han
    • , Andrii Ishchenko
    • , Wei Liu
    • , Haitao Zhang
    • , Marc Messerschmidt
    • , Vsevolod Katritch
    • , Raymond C Stevens
    •  & Vadim Cherezov

    Present addresses: Celgene Corporation, San Diego, California, USA (G.F.), The Bridge Institute, University of Southern California, Los Angeles, California, USA (G.W.H., A.I., H.Z., V.K., R.C.S. and V.C.), Department of Chemistry, University of Southern California, Los Angeles, California, USA (G.W.H., A.I., R.C.S. and V.C.), Department of Biological Sciences, University of Southern California, Los Angeles, California, USA (H.Z., V.K. and R.C.S.), Center for Applied Structural Discovery at the Biodesign Institute, Arizona State University, Tempe, Arizona, USA (W.L.) and National Science Foundation BioXFEL Science and Technology Center, Buffalo, New York, USA (M. Messerschmidt).

Affiliations

  1. Department of Integrative Structural and Computational Biology, The Scripps Research Institute, La Jolla, California, USA.

    • Gustavo Fenalti
    • , Gye Won Han
    • , Andrii Ishchenko
    • , Wei Liu
    • , Haitao Zhang
    • , Vsevolod Katritch
    • , Raymond C Stevens
    •  & Vadim Cherezov
  2. Department of Physics, Arizona State University, Tempe, Arizona, USA.

    • Nadia A Zatsepin
    • , Daniel James
    • , Dingjie Wang
    • , Uwe Weierstall
    •  & John C H Spence
  3. Department of Chemistry, Vrije Universiteit Brussel, Brussels, Belgium.

    • Cecilia Betti
    • , Karel Guillemyn
    • , Dirk Tourwé
    •  & Steven Ballet
  4. Department of Bioengineering Sciences, Vrije Universiteit Brussel, Brussels, Belgium.

    • Cecilia Betti
    • , Karel Guillemyn
    • , Dirk Tourwé
    •  & Steven Ballet
  5. National Institute of Mental Health Psychoactive Drug Screening Program, University of North Carolina Chapel Hill Medical School, Chapel Hill, North Carolina, USA.

    • Patrick Giguere
    •  & Bryan L Roth
  6. Department of Pharmacology, University of North Carolina Chapel Hill Medical School, Chapel Hill, North Carolina, USA.

    • Patrick Giguere
    •  & Bryan L Roth
  7. Division of Chemical Biology and Medicinal Chemistry, University of North Carolina Chapel Hill Medical School, Chapel Hill, North Carolina, USA.

    • Patrick Giguere
    •  & Bryan L Roth
  8. Linac Coherent Light Source, SLAC National Accelerator Laboratory, Menlo Park, California, USA.

    • Sébastien Boutet
    • , Marc Messerschmidt
    •  & Garth J Williams
  9. Center for Free Electron Laser Science, Deutsches Elektronen-Synchrotron (DESY), Hamburg, Germany.

    • Cornelius Gati
    • , Oleksandr M Yefanov
    • , Thomas A White
    • , Dominik Oberthuer
    • , Markus Metz
    • , Chun Hong Yoon
    • , Anton Barty
    •  & Henry N Chapman
  10. Institute of Biochemistry and Molecular Biology, University of Hamburg, Hamburg, Germany.

    • Dominik Oberthuer
  11. Department of Physics, University of Hamburg, Hamburg, Germany.

    • Markus Metz
    •  & Henry N Chapman
  12. European X-ray Free-Electron Laser Facility (XFEL GmbH), Hamburg, Germany.

    • Chun Hong Yoon
  13. Department of Chemistry and Biochemistry, Arizona State University, Tempe, Arizona, USA.

    • Shibom Basu
    • , Jesse Coe
    • , Chelsie E Conrad
    • , Raimund Fromme
    •  & Petra Fromme
  14. Center for Applied Structural Discovery at the Biodesign Institute, Arizona State University, Tempe, Arizona, USA.

    • Shibom Basu
    • , Jesse Coe
    • , Chelsie E Conrad
    • , Raimund Fromme
    •  & Petra Fromme
  15. Laboratory of Chemical Biology and Peptide Research, Clinical Research Institute of Montreal, Montreal, Quebec, Canada.

    • Peter W Schiller

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Contributions

G.F. designed, optimized and purified δ-OR receptor constructs for structural studies, crystallized the receptor in LCP, collected and processed synchrotron diffraction data, determined the synchrotron and XFEL structures, analyzed the data and wrote the paper; N.A.Z. collected and processed XFEL data; C.B. synthesized peptide ligands for structural and signaling studies; P.G. performed signaling studies, analyzed the data and wrote the paper; G.W.H. helped with structure refinement and analysis; A.I., H.Z. and W.L. collected XFEL data and helped with sample preparation; K.G. synthesized peptide ligands for structural and signaling studies; O.M.Y. refined the detector geometry and contributed to XFEL data processing; D.J., D.W., U.W. and J.C.H.S. designed the LCP injector and controlled it during XFEL data collection; S. Boutet, M. Messerschmidt and G.J.W. operated the CXI beamline at LCLS and contributed to XFEL data collection and processing; C.G., T.A.W., D.O., M. Metz, C.H.Y., A.B., H.N.C. and S. Basu participated in XFEL data collection and contributed to XFEL data processing; J.C., C.E.C., R.F. and P.F. collected and analyzed XFEL data and helped with biophysical characterization of crystals at LCLS; D.T. and P.W.S. helped with manuscript preparation; B.L.R. supervised the pharmacology studies, analyzed the data and wrote the paper; S. Ballet supervised the peptide synthesis and screening studies, synthesized peptide ligands for structural studies and wrote the paper; V.K. analyzed the data and wrote the paper; R.C.S. determined the overall project strategy, analyzed the data and wrote the paper; V.C. determined the overall project strategy and provided management, supervised XFEL data collection, analyzed the data and wrote the paper with contributions from all other coauthors.

Competing interests

The authors declare no competing financial interests.

Corresponding author

Correspondence to Vadim Cherezov.

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https://doi.org/10.1038/nsmb.2965

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