Subjects

Abstract

Prostanoids are a series of bioactive lipid metabolites that function in an autacoid manner via activation of cognate G-protein-coupled receptors (GPCRs). Here, we report the crystal structure of human prostaglandin (PG) E receptor subtype EP3 bound to endogenous ligand PGE2 at 2.90 Å resolution. The structure reveals important insights into the activation mechanism of prostanoid receptors and provides a molecular basis for the binding modes of endogenous ligands.

Access optionsAccess options

Rent or Buy article

Get time limited or full article access on ReadCube.

from$8.99

All prices are NET prices.

Data availability

The atomic coordinates and structure factor files have been deposited in the Protein Data Bank with accession codes 6AK3.

Additional information

Publisher’s note: Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

References

  1. 1.

    Woodward, D. F., Jones, R. L. & Narumiya, S. Pharmacol. Rev. 63, 471–538 (2011).

  2. 2.

    Toyoda, Y. et al. Nat. Chem. Biol. https://doi.org/10.1038/s41589-018-0131-3 (2018).

  3. 3.

    Ballesteros, J. A. & Weinstein, H. in Methods in Neurosciences. Vol. 25, (ed. Sealfon, S.) 366–428 (Elsevier, Amsterdam, 1995).

  4. 4.

    Jin, J., Mao, G. F. & Ashby, B. Br. J. Pharmacol. 121, 317–323 (1997).

  5. 5.

    Zhang, H. et al. Nature 544, 327–332 (2017).

  6. 6.

    Kedzie, K. M., Donello, J. E., Krauss, H. A., Regan, J. W. & Gil, D. W. Mol. Pharmacol. 54, 584–590 (1998).

  7. 7.

    Neuschäfer-Rube, F., Engemaier, E., Koch, S., Böer, U. & Püschel, G. P. Biochem. J. 371, 443–449 (2003).

  8. 8.

    Stitham, J., Stojanovic, A., Merenick, B. L., O’Hara, K. A. & Hwa, J. J. Biol. Chem. 278, 4250–4257 (2003).

  9. 9.

    Funk, C. D., Furci, L., Moran, N. & Fitzgerald, G. A. Mol. Pharmacol. 44, 934–939 (1993).

  10. 10.

    Scheerer, P. et al. Nature 455, 497–502 (2008).

  11. 11.

    Koehl, A. et al. Nature 558, 547–552 (2018).

  12. 12.

    Draper-Joyce, C. J. et al. Nature 558, 559–563 (2018).

  13. 13.

    García-Nafría, J., Nehmé, R., Edwards, P. C. & Tate, C. G. Nature 558, 620–623 (2018).

  14. 14.

    Katritch, V. et al. Trends. Biochem. Sci. 39, 233–244 (2014).

  15. 15.

    Hanson, M. A. et al. Science 335, 851–855 (2012).

  16. 16.

    Chrencik, J. E. et al. Cell 161, 1633–1643 (2015).

  17. 17.

    Hua, T. et al. Cell 167, 750–762.e14 (2016).

  18. 18.

    Hua, T. et al. Nature 547, 468–471 (2017).

  19. 19.

    Choe, H. W. et al. Nature 471, 651–655 (2011).

  20. 20.

    Shiroishi, M. et al. Microb. Cell. Fact. 11, 78 (2012).

  21. 21.

    Hattori, M., Hibbs, R. E. & Gouaux, E. Structure 20, 1293–1299 (2012).

  22. 22.

    Chu, R. et al. J. Mol. Biol. 323, 253–262 (2002).

  23. 23.

    Caffrey, M. & Cherezov, V. Nat. Protoc. 4, 706–731 (2009).

  24. 24.

    Ueno, G. et al. AIP Conf. Proc. 1741, 050021 (2016).

  25. 25.

    Hirata, K., Foadi, J., Evans, G., Hasegawa, K. & Zeldin, O. B. in Advanced Methods in Structural Biology. (eds. Senda, T. & Maenaka, K.) 241–273 (Springer Protocol Handbooks, 2016).

  26. 26.

    Yamashita, K., Hirata, K. & Yamamoto, M. Acta Crystallogr. D Struct. Biol. 74, 441–449 (2018).

  27. 27.

    Kabsch, W. Acta Crystallogr, D. Biol. Crystallogr. 66, 125–132 (2010).

  28. 28.

    Foadi, J. et al. Acta Crystallogr. D Biol. Crystallogr. 69, 1617–1632 (2013).

  29. 29.

    McCoy, A. J. et al. J. Appl. Crystallogr. 40, 658–674 (2007).

  30. 30.

    Emsley, P., Lohkamp, B., Scott, W. G. & Cowtan, K. Acta Crystallogr. D Biol. Crystallogr. 66, 486–501 (2010).

  31. 31.

    Murshudov, G. N. et al. Acta Crystallogr. D Biol. Crystallogr. 67, 355–367 (2011).

  32. 32.

    Adams, P. D. et al. Acta Crystallogr. D Biol. Crystallogr. 66, 213–221 (2010).

  33. 33.

    Inoue, A. et al. Nat. Methods 9, 1021–1029 (2012).

Download references

Acknowledgements

This work was supported by AMED under grant Numbers JP18gm0910007 (CREST; T.K.) and JP18am0101079 (S.I.), JP18am0101070 (M.Y.) (Platform Project for Supporting Drug Discovery and Life Science Research; Basis for Supporting Innovative Drug Discovery and Life Science Research (BINDS)) and JSPS KAKENHI grant Number 15J00102 (K.M.). The authors acknowledge support from the Toray Science Foundation (T.K.), Takeda Science Foundation (R.S. and T.K.), Naito Foundation (T.K.), and Koyanagi Foundation (T.K.). The synchrotron radiation experiments were performed at BL32XU of SPring-8 with the approval of the Japan Synchrotron Radiation Research Institute (JASRI) (Proposal No. 2017A2524, 2017B2524, and BINDS0483). We thank the staff members of BL32XU for help with X-ray data collection. DNA sequencing analysis was performed at the Medical Research Support Center, Graduate School of Medicine, Kyoto University. We are grateful to A. Inoue for providing instruction on the TGFα shedding assay, M. Shiroishi for providing instruction on FSEC-TS, D. Im and T. Shimamura (Kyoto University) for providing the plasmid encoding mbIIG2, S. Horita for support for structure refinement, and H. Tsujimoto and M. Sasanuma for technical assistance.

Author information

Affiliations

  1. Department of Cell Biology, Graduate School of Medicine, Kyoto University, Konoe-cho, Yoshida, Sakyo-ku, Kyoto, Japan

    • Kazushi Morimoto
    • , Ryoji Suno
    • , Yunhong Hotta
    • , So Iwata
    •  & Takuya Kobayashi
  2. Japan Agency for Medical Research and Development (AMED), Core Research for Evolutional Science and Technology (CREST), Chiyoda-ku, Tokyo, Japan

    • Kazushi Morimoto
    • , Ryoji Suno
    •  & Takuya Kobayashi
  3. RIKEN SPring-8 Center, Sayo-cho, Sayo-gun, Hyogo, Japan

    • Keitaro Yamashita
    • , Kunio Hirata
    •  & Masaki Yamamoto
  4. Medical Innovation Center, Graduate School of Medicine, Kyoto University, Sakyo-ku, Kyoto, Japan

    • Shuh Narumiya
  5. Department of Medical Chemistry, Kansai Medical University, Hirakata, Osaka, Japan

    • Takuya Kobayashi

Authors

  1. Search for Kazushi Morimoto in:

  2. Search for Ryoji Suno in:

  3. Search for Yunhong Hotta in:

  4. Search for Keitaro Yamashita in:

  5. Search for Kunio Hirata in:

  6. Search for Masaki Yamamoto in:

  7. Search for Shuh Narumiya in:

  8. Search for So Iwata in:

  9. Search for Takuya Kobayashi in:

Contributions

K.M. initiated the project; optimized the construct; developed the purification procedure; purified the EP3 protein for crystallization; performed crystallization trials, ligand-binding assay, and TGFα shedding assay; and wrote the manuscript. Y.H. helped with construct optimization. R.S. collected X-ray diffraction data and solved the structure. K.Y. and K.H. helped with data collection. M.Y. oversaw data collection. S.N. helped with interpretation and edited the manuscript. S.I. helped with structure analysis and interpretation and edited the manuscript. T.K. supervised the research.

Competing interests

The authors declare no competing interests.

Corresponding authors

Correspondence to Kazushi Morimoto or So Iwata or Takuya Kobayashi.

Supplementary information

  1. Supplementary Text and Figures

    Supplementary Figures 1–15, Supplementary Tables 1–2

  2. Reporting Summary

About this article

Publication history

Received

Accepted

Published

DOI

https://doi.org/10.1038/s41589-018-0171-8

Further reading