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Structure of the human smoothened receptor bound to an antitumour agent

Abstract

The smoothened (SMO) receptor, a key signal transducer in the hedgehog signalling pathway, is responsible for the maintenance of normal embryonic development and is implicated in carcinogenesis. It is classified as a class frizzled (class F) G-protein-coupled receptor (GPCR), although the canonical hedgehog signalling pathway involves the GLI transcription factors and the sequence similarity with class A GPCRs is less than 10%. Here we report the crystal structure of the transmembrane domain of the human SMO receptor bound to the small-molecule antagonist LY2940680 at 2.5 Å resolution. Although the SMO receptor shares the seven-transmembrane helical fold, most of the conserved motifs for class A GPCRs are absent, and the structure reveals an unusually complex arrangement of long extracellular loops stabilized by four disulphide bonds. The ligand binds at the extracellular end of the seven-transmembrane-helix bundle and forms extensive contacts with the loops.

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Figure 1: Overall structure of the human SMO receptor 7TM domain in complex with LY2940680.
Figure 2: Comparison of the 7TM bundle of the SMO receptor with class A GPCRs.
Figure 3: Ligand-binding pocket for LY2940680.
Figure 4: The structure of the ECD linker domain and ECLs.
Figure 5: Comparison of ECL2 of the SMO receptor with class A GPCRs.
Figure 6: Structural insight for the FZD receptors on the basis of their sequence homology with the SMO receptor.

Accession codes

Primary accessions

Protein Data Bank

Data deposits

The coordinates and the structure factors have been deposited in the Protein Data Bank under the accession code 4JKV.

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Acknowledgements

This work was supported by the National Institutes of Health Common Fund grant P50 GM073197 for technology development (V.C. and R.C.S.), PSI:Biology grant U54 GM094618 for biological studies and structure production (target GPCR-131) (V.K., V.C. and R.C.S.); F32 DK088392 (F.Y.S.); R01 MH61887, U19 MH82441, R01 DA27170 and the NIMH Psychoactive Drug Screening Program (X.-P.H. and B.L.R.) and the Michael Hooker Chair of Pharmacology (B.L.R.). We thank J. Velasquez for help with molecular biology, T. Trinh and M. Chu for help with baculovirus expression, K. Kadyshevskaya for assistance with figure preparation, A. Walker for assistance with manuscript preparation, D. Wacker for assistance with SAD data collection and processing and J. Smith, R. Fischetti and N. Sanishvili for assistance in development and use of the minibeam and beamtime at beamline 23-ID at the Advanced Photon Source, which is supported by National Cancer Institute grant Y1-CO-1020 and National Institute of General Medical Sciences grant Y1-GM-1104.

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C.W. designed and made the constructs, purified and crystallized the receptor in LCP, optimized crystallization conditions, grew crystals for data collection, assisted with crystal collection and heavy-atom soaking experiment, collected the diffraction data and prepared the manuscript. H.W. performed baculovirus expression of the receptor, purified the receptor, optimized crystallization conditions and prepared the manuscript. V.K. prepared the manuscript. G.W.H. solved and refined the structure and assisted with preparation of the manuscript. X.-P.H. performed the radioligand binding assay. W.L. assisted with data collection. F.Y.S. provided material and conditions for heavy-atom soaking experiment. B.L.R. supervised the radioligand binding experiment and assisted with preparing the manuscript. V.C. collected crystals, performed the heavy-atom soaking experiment, collected and processed diffraction data and assisted with preparation of the manuscript. R.C.S. was responsible for the overall project strategy and management and wrote the manuscript.

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Correspondence to Raymond C. Stevens.

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The authors declare no competing financial interests.

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Wang, C., Wu, H., Katritch, V. et al. Structure of the human smoothened receptor bound to an antitumour agent. Nature 497, 338–343 (2013). https://doi.org/10.1038/nature12167

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