Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

Agonist-bound adenosine A2A receptor structures reveal common features of GPCR activation

Nature volume 474, pages 521–525 (2011)Cite this article

Subjects

Abstract

Adenosine receptors and β-adrenoceptors are G-protein-coupled receptors (GPCRs) that activate intracellular G proteins on binding the agonists adenosine1 or noradrenaline2, respectively. GPCRs have similar structures consisting of seven transmembrane helices that contain well-conserved sequence motifs, indicating that they are probably activated by a common mechanism3,4. Recent structures of β-adrenoceptors highlight residues in transmembrane region 5 that initially bind specifically to agonists rather than to antagonists, indicating that these residues have an important role in agonist-induced activation of receptors5,6,7. Here we present two crystal structures of the thermostabilized human adenosine A2A receptor (A2AR-GL31) bound to its endogenous agonist adenosine and the synthetic agonist NECA. The structures represent an intermediate conformation between the inactive and active states, because they share all the features of GPCRs that are thought to be in a fully activated state, except that the cytoplasmic end of transmembrane helix 6 partially occludes the G-protein-binding site. The adenine substituent of the agonists binds in a similar fashion to the chemically related region of the inverse agonist ZM241385 (ref. 8). Both agonists contain a ribose group, not found in ZM241385, which extends deep into the ligand-binding pocket where it makes polar interactions with conserved residues in H7 (Ser 2777.42 and His 2787.43; superscripts refer to Ballesteros–Weinstein numbering9) and non-polar interactions with residues in H3. In contrast, the inverse agonist ZM241385 does not interact with any of these residues and comparison with the agonist-bound structures indicates that ZM241385 sterically prevents the conformational change in H5 and therefore it acts as an inverse agonist. Comparison of the agonist-bound structures of A2AR with the agonist-bound structures of β-adrenoceptors indicates that the contraction of the ligand-binding pocket caused by the inward motion of helices 3, 5 and 7 may be a common feature in the activation of all GPCRs.

This is a preview of subscription content, access via your institution

Relevant articles

Open Access articles citing this article.

Access options

Rent or buy this article

Prices vary by article type

from$1.95

to$39.95

Learn more

Prices may be subject to local taxes which are calculated during checkout

Figure 1: Structure of the adenosine A 2A receptor bound to NECA compared to other GPCR structures.
Figure 2: Comparison of receptor–ligand interactions for A 2A R bound to the inverse agonist ZM241385 and the agonists NECA and adenosine.
Figure 3: Positions of adenosine and ZM241385 in the A 2A R ligand-binding pocket.
Figure 4: Comparison of the positions of agonists in the binding pockets of the A 2A R and β 1 -AR.

Accession codes

Primary accessions

Protein Data Bank

Data deposits

Co-ordinates and structure factors have been submitted to the PDB database under accession codes 2YDO and 2YDV for A2AR-GL31 bound to adenosine or NECA, respectively.

References

  1. Fredholm, B. B. et al. International Union of Basic and Clinical Pharmacology. LXXXI. Nomenclature and classification of adenosine receptors—an update. Pharmacol. Rev. 63, 1–34 (2011)

    Article  CAS  Google Scholar 

  2. Evans, B. A. et al. Ligand-directed signalling at β-adrenoceptors. Br. J. Pharmacol. 159, 1022–1038 (2010)

    Article  CAS  Google Scholar 

  3. Hofmann, K. P. et al. A G protein-coupled receptor at work: the rhodopsin model. Trends Biochem. Sci. 34, 540–552 (2009)

    Article  CAS  Google Scholar 

  4. Rosenbaum, D. M., Rasmussen, S. G. & Kobilka, B. K. The structure and function of G-protein-coupled receptors. Nature 459, 356–363 (2009)

    Article  ADS  CAS  Google Scholar 

  5. Rasmussen, S. G. et al. Structure of a nanobody-stabilized active state of the β2 adrenoceptor. Nature 469, 175–180 (2011)

    Article  ADS  CAS  Google Scholar 

  6. Rosenbaum, D. M. et al. Structure and function of an irreversible agonist–β2 adrenoceptor complex. Nature 469, 236–240 (2011)

    Article  ADS  CAS  Google Scholar 

  7. Warne, T. et al. The structural basis for agonist and partial agonist action on a β1-adrenergic receptor. Nature 469, 241–244 (2011)

    Article  ADS  CAS  Google Scholar 

  8. Jaakola, V. P. et al. The 2.6 Ångstrom crystal structure of a human A2A adenosine receptor bound to an antagonist. Science 322, 1211–1217 (2008)

    Article  ADS  CAS  Google Scholar 

  9. Ballesteros, J. A. & Weinstein, H. Integrated methods for the construction of three dimensional models and computational probing of structure function relations in G protein-coupled receptors. Methods Neurosci. 25, 366–428 (1995)

    Article  CAS  Google Scholar 

  10. Kobilka, B. K. & Deupi, X. Conformational complexity of G-protein-coupled receptors. Trends Pharmacol. Sci. 28, 397–406 (2007)

    Article  CAS  Google Scholar 

  11. Yao, X. J. et al. The effect of ligand efficacy on the formation and stability of a GPCR-G protein complex. Proc. Natl Acad. Sci. USA 106, 9501–9506 (2009)

    Article  ADS  CAS  Google Scholar 

  12. Vauquelin, G. & Van Liefde, I. G protein-coupled receptors: a count of 1001 conformations. Fundam. Clin. Pharmacol. 19, 45–56 (2005)

    Article  CAS  Google Scholar 

  13. Palczewski, K. et al. Crystal structure of rhodopsin: a G protein-coupled receptor. Science 289, 739–745 (2000)

    Article  ADS  CAS  Google Scholar 

  14. Li, J. et al. Structure of bovine rhodopsin in a trigonal crystal form. J. Mol. Biol. 343, 1409–1438 (2004)

    Article  CAS  Google Scholar 

  15. Park, J. H. et al. Crystal structure of the ligand-free G-protein-coupled receptor opsin. Nature 454, 183–187 (2008)

    Article  ADS  CAS  Google Scholar 

  16. Scheerer, P. et al. Crystal structure of opsin in its G-protein-interacting conformation. Nature 455, 497–502 (2008)

    Article  ADS  CAS  Google Scholar 

  17. Cherezov, V. et al. High-resolution crystal structure of an engineered human β2-adrenergic G protein-coupled receptor. Science 318, 1258–1265 (2007)

    Article  ADS  CAS  Google Scholar 

  18. Rasmussen, S. G. et al. Crystal structure of the human β2 adrenergic G-protein-coupled receptor. Nature 450, 383–387 (2007)

    Article  ADS  CAS  Google Scholar 

  19. Warne, T. et al. Structure of a β1-adrenergic G-protein-coupled receptor. Nature 454, 486–491 (2008)

    Article  ADS  CAS  Google Scholar 

  20. Wu, B. et al. Structures of the CXCR4 chemokine GPCR with small-molecule and cyclic peptide antagonists. Science 330, 1066–1071 (2010)

    Article  ADS  CAS  Google Scholar 

  21. Chien, E. Y. et al. Structure of the human dopamine D3 receptor in complex with a D2/D3 selective antagonist. Science 330, 1091–1095 (2010)

    Article  ADS  CAS  Google Scholar 

  22. Oldham, W. M. & Hamm, H. E. Heterotrimeric G protein activation by G-protein-coupled receptors. Nature Rev. Mol. Cell Biol. 9, 60–71 (2008)

    Article  CAS  Google Scholar 

  23. Murphree, L. J. et al. Human A2A adenosine receptors: high-affinity agonist binding to receptor-G protein complexes containing Gβ4 . Mol. Pharmacol. 61, 455–462 (2002)

    Article  CAS  Google Scholar 

  24. Serrano-Vega, M. J., Magnani, F., Shibata, Y. & Tate, C. G. Conformational thermostabilization of the β1-adrenergic receptor in a detergent-resistant form. Proc. Natl Acad. Sci. USA 105, 877–882 (2008)

    Article  ADS  CAS  Google Scholar 

  25. Magnani, F., Shibata, Y., Serrano-Vega, M. J. & Tate, C. G. Co-evolving stability and conformational homogeneity of the human adenosine A2A receptor. Proc. Natl Acad. Sci. USA 105, 10744–10749 (2008)

    Article  ADS  CAS  Google Scholar 

  26. Shibata, Y. et al. Thermostabilization of the neurotensin receptor NTS1. J. Mol. Biol. 390, 262–277 (2009)

    Article  CAS  Google Scholar 

  27. Kim, S. K. et al. Modeling the adenosine receptors: comparison of the binding domains of A2A agonists and antagonists. J. Med. Chem. 46, 4847–4859 (2003)

    Article  CAS  Google Scholar 

  28. Dal Ben, D. et al. Adenosine receptor modeling: what does the A2A crystal structure tell us? Curr. Top. Med. Chem. 10, 993–1018 (2010)

    Article  CAS  Google Scholar 

  29. Wacker, D. et al. Conserved binding mode of human β2 adrenergic receptor inverse agonists and antagonist revealed by X-ray crystallography. J. Am. Chem. Soc. 132, 11443–11445 (2010)

    Article  CAS  Google Scholar 

  30. Xu, F. et al. Structure of an agonist-bound human A2A adenosine receptor. Science (2011)

  31. Potterton, L. et al. Developments in the CCP4 molecular-graphics project. Acta Crystallogr. D 60, 2288–2294 (2004)

    Article  Google Scholar 

  32. Lebon, G. Bennett, K. Jazayeri, A. & Tate, C. G. Thermostabilization of an agonist-bound conformation of the human adenosine A2A receptor. J. Mol. Biol. 10.1016/j.jmb.2011.03.075 (in the press)

  33. Warne, T., Chirnside, J. & Schertler, G. F. Expression and purification of truncated, non-glycosylated turkey β-adrenergic receptors for crystallization. Biochim. Biophys. Acta 1610, 133–140 (2003)

    Article  CAS  Google Scholar 

  34. Schaffner, W. & Weissmann, C. A rapid, sensitive, and specific method for the determination of protein in dilute solution. Anal. Biochem. 56, 502–514 (1973)

    Article  CAS  Google Scholar 

  35. Gorrec, F., Palmer, C., Lebon, G. & Warne, T. Pi sampling: a methodical and flexible approach to macromolecular crystallization initial screening. Acta Crystallogr. D 67, 463–470 (2011)

    Article  CAS  Google Scholar 

  36. Leslie, A. G. The integration of macromolecular diffraction data. Acta Crystallogr. D 62, 48–57 (2006)

    Article  Google Scholar 

  37. Evans, P. Scaling and assessment of data quality. Acta Crystallogr. D 62, 72–82 (2006)

    Article  Google Scholar 

  38. McCoy, A. J. et al. Phaser crystallographic software. J. Appl. Cryst. 40, 658–674 (2007)

    Article  CAS  Google Scholar 

  39. Murshudov, G. N., Vagin, A. A. & Dodson, E. J. Refinement of macromolecular structures by the maximum-likelihood method. Acta Crystallogr. D 53, 240–255 (1997)

    Article  CAS  Google Scholar 

  40. Emsley, P., Lohkamp, B., Scott, W. G. & Cowtan, K. Features and development of Coot. Acta Crystallogr. D 66, 486–501 (2010)

    Article  CAS  Google Scholar 

  41. McDonald, I. K. & Thornton, J. M. Satisfying hydrogen bonding potential in proteins. J. Mol. Biol. 238, 777–793 (1994)

    Article  CAS  Google Scholar 

  42. Jones, T. A., Zou, J. Y., Cowan, S. W. & Kjeldgaard, M. Improved methods for building protein models in electron-density maps and the location of errors in these models. Acta Crystallogr. A 47, 110–119 (1991)

    Article  Google Scholar 

  43. Davis, I. W. et al. MolProbity: all-atom contacts and structure validation for proteins and nucleic acids. Nucleic Acids Res. 35, W375–W383 (2007)

    Article  ADS  Google Scholar 

  44. Robertson, N. et al. The properties of thermostabilised G protein-coupled receptors (StaRs) and their use in drug discovery. Neuropharmacology 60, 36–44 (2011)

    Article  CAS  Google Scholar 

Download references

Acknowledgements

This work was supported by core funding from the Medical Research Council, and grants from Heptares Therapeutics Ltd and from the Biotechnology and Biological Sciences Research Council (BB/G003653/1). We would like to thank F. Magnani for technical help at the start of the project and F. Gorrec for developing the crystallization screen. We also thank the beamline staff at the European Synchrotron Radiation Facility, particularly at (beamline ID23-2; D. Flot and A. Popov), the Swiss Light Source (beamline X06SA) and at the Diamond Light Source (beamline I24; G. Evans, D. Axford and R. Owen). F. Marshall, M. Weir, M. Congreve and R. Henderson are thanked for their comments on the manuscript.

Author information

Authors and Affiliations

  1. MRC Laboratory of Molecular Biology, Hills Road, Cambridge CB2 0QH, UK

    Guillaume Lebon, Tony Warne, Patricia C. Edwards, Andrew G. W. Leslie & Christopher G. Tate

  2. Heptares Therapeutics, BioPark, Broadwater Road, Welwyn Garden City AL7 3AX, UK

    Kirstie Bennett & Christopher J. Langmead

Authors
  1. Guillaume Lebon
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Tony Warne
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Patricia C. Edwards
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Kirstie Bennett
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Christopher J. Langmead
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Andrew G. W. Leslie
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Christopher G. Tate
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

G.L. devised and performed receptor expression, purification, crystallization, cryo-cooling of the crystals, data collection, data processing and structure refinement. T.W. and P.C.E. helped with expression, crystal cryo-cooling and data collection. K.B. performed the radioligand binding assays and pharmacological analyses on receptor mutants in whole cells and C.J.L. was involved in data analysis and experimental design. A.G.W.L. was involved in data processing and structure refinement. Manuscript preparation was performed by G.L., A.G.W.L. and C.G.T. Overall project management was by C.G.T.

Corresponding author

Correspondence to Christopher G. Tate.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Information

This file contains Supplementary Figures 1-8 with legends, Supplementary Table 1 and Supplementary Text. (PDF 4588 kb)

PowerPoint slides

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Lebon, G., Warne, T., Edwards, P. et al. Agonist-bound adenosine A2A receptor structures reveal common features of GPCR activation. Nature 474, 521–525 (2011). https://doi.org/10.1038/nature10136

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nature10136

This article is cited by

Comments

By submitting a comment you agree to abide by our Terms and Community Guidelines. If you find something abusive or that does not comply with our terms or guidelines please flag it as inappropriate.

Search

Advanced search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing