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Structure determination of the seven-helix transmembrane receptor sensory rhodopsin II by solution NMR spectroscopy

Abstract

Seven-helix membrane proteins represent a challenge for structural biology. Here we report the first NMR structure determination of a detergent-solubilized seven-helix transmembrane (7TM) protein, the phototaxis receptor sensory rhodopsin II (pSRII) from Natronomonas pharaonis, as a proof of principle. The overall quality of the structure ensemble is good (backbone r.m.s. deviation of 0.48 Å) and agrees well with previously determined X-ray structures. Furthermore, measurements in more native-like small phospholipid bicelles indicate that the protein structure is the same as in detergent micelles, suggesting that environment-specific effects are minimal when using mild detergents. We use our case study as a platform to discuss the feasibility of similar solution NMR studies for other 7TM proteins, including members of the family of G protein–coupled receptors.

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Figure 1: Side chain assignment and NOE-derived distance restraints in pSRII.
Figure 2: Solution-NMR structure of pSRII in DHPC micelles.
Figure 3: Mapping of hydrophobic and solvent-exposed regions of pSRII through titration with the spin labels 16-DSA and gadoteridol.
Figure 4: Comparison of small-bicelle versus micelle reconstitution of pSRII.

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Acknowledgements

We thank Y. Ito for discussions and assistance with NMR experiments, D. Moskau and R. Kuemmerle for help with NMR measurements, T. Warne for the preparation of β1-AR and E.D. Laue and C.-W. Chung for their support in the initial stages of the project. The use of the CamGrid computing resource is acknowledged for the structure calculations. This work was supported by a grant from the Royal Society (RG 2005/R1 and Biotechnology and Biological Sciences Research Council (BB/G011915/1). A.G. was supported by a Marie Curie Actions studentship sponsored by the European Union. The NMR Facility of the Department of Biochemistry is supported by the Biotechnology and Biological Sciences Research Council, Cancer Research UK and the Wellcome Trust.

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A.G. and J.P.K. carried out protein expression, purification and detergent reconstitution; A.G. and H.R.M. assigned resonances and carried out structure calculations; M.J.B. did the spin label experiments and bicelle studies; D.N. wrote the manuscript and designed and conducted research including NMR data acquisition, processing, assignment of spectra and data analysis.

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Correspondence to Daniel Nietlispach.

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The authors declare no competing financial interests.

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Gautier, A., Mott, H., Bostock, M. et al. Structure determination of the seven-helix transmembrane receptor sensory rhodopsin II by solution NMR spectroscopy. Nat Struct Mol Biol 17, 768–774 (2010). https://doi.org/10.1038/nsmb.1807

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