TRPV1 structures in nanodiscs reveal mechanisms of ligand and lipid action

Abstract

When integral membrane proteins are visualized in detergents or other artificial systems, an important layer of information is lost regarding lipid interactions and their effects on protein structure. This is especially relevant to proteins for which lipids have both structural and regulatory roles. Here we demonstrate the power of combining electron cryo-microscopy with lipid nanodisc technology to ascertain the structure of the rat TRPV1 ion channel in a native bilayer environment. Using this approach, we determined the locations of annular and regulatory lipids and showed that specific phospholipid interactions enhance binding of a spider toxin to TRPV1 through formation of a tripartite complex. Furthermore, phosphatidylinositol lipids occupy the binding site for capsaicin and other vanilloid ligands, suggesting a mechanism whereby chemical or thermal stimuli elicit channel activation by promoting the release of bioactive lipids from a critical allosteric regulatory site.

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Figure 1: TRPV1 structures determined in lipid nanodisc.
Figure 2: Structural details of tripartite toxin–channel–lipid complex.
Figure 3: Movement of protein and lipids associated with toxin binding.
Figure 4: Shared binding pocket for phosphatidylinositol lipids and vanilloid ligands.
Figure 5: Structural rearrangements associated with vanilloid binding.
Figure 6: Mechanistic models for TRPV1 activation.

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Electron Microscopy Data Bank

Protein Data Bank

Data deposits

The three-dimensional cryo-EM density maps of the TRPV1–nanodisc complexes without low-pass filter and amplitude modification have been deposited in the Electron Microscopy Data Bank under accession numbers EMD-8118 (TRPV1–nanodisc), EMD-8117 (TRPV1–RTX/DkTx–nanodisc), EMD-8119 (TRPV1–capsazepine–nanodisc) and EMD-8120 (TRPV1–capsazepine in amphipol). Particle image stacks after motion correction related to TRPV1–nanodisc and TRPV1–RTX/DkTx–nanodisc have been deposited in the Electron Microscopy Pilot Image Archive (http://www.ebi.ac.uk/pdbe/emdb/empiar/) under accession number EMPIAR-10059. Atomic coordinates for the atomic model of TRPV1 in nanodisc, TRPV1–RTX/DkTx in nanodisc and TRPV1–capsazepine in nanodisc have been deposited in the Protein Data Bank under accession numbers 5IRZ, 5IRX and 5IS0.

References

  1. 1

    Hilgemann, D. W. Getting ready for the decade of the lipids. Annu. Rev. Physiol. 65, 697–700 (2003)

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  2. 2

    Hille, B., Dickson, E. J., Kruse, M., Vivas, O. & Suh, B. C. Phosphoinositides regulate ion channels. Biochim. Biophys. Acta 1851, 844–856 (2015)

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  3. 3

    Lee, A. G. Biological membranes: the importance of molecular detail. Trends Biochem. Sci. 36, 493–500 (2011)

    CAS  Article  Google Scholar 

  4. 4

    Caffrey, M. A lipid’s eye view of membrane protein crystallization in mesophases. Curr. Opin. Struct. Biol. 10, 486–497 (2000)

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  5. 5

    Landau, E. M. & Rosenbusch, J. P. Lipidic cubic phases: a novel concept for the crystallization of membrane proteins. Proc. Natl Acad. Sci. USA 93, 14532–14535 (1996)

    CAS  PubMed  Article  ADS  PubMed Central  Google Scholar 

  6. 6

    Gonen, T. et al. Lipid-protein interactions in double-layered two-dimensional AQP0 crystals. Nature 438, 633–638 (2005)

    CAS  PubMed  PubMed Central  Article  ADS  Google Scholar 

  7. 7

    Wang, L. & Sigworth, F. J. Structure of the BK potassium channel in a lipid membrane from electron cryomicroscopy. Nature 461, 292–295 (2009)

    CAS  PubMed  PubMed Central  Article  ADS  Google Scholar 

  8. 8

    Bayburt, T. H., Grinkova, Y. V. & Sligar, S. G. Self-assembly of discoidal phospholipid bilayer nanoparticles with membrane scaffold proteins. Nano Lett. 2, 853–856 (2002)

    CAS  Article  ADS  Google Scholar 

  9. 9

    Banerjee, S., Huber, T. & Sakmar, T. P. Rapid incorporation of functional rhodopsin into nanoscale apolipoprotein bound bilayer (NABB) particles. J. Mol. Biol. 377, 1067–1081 (2008)

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  10. 10

    Ritchie, T. K. et al. Reconstitution of membrane proteins in phospholipid bilayer nanodiscs. Methods Enzymol. 464, 211–231 (2009)

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  11. 11

    Efremov, R. G., Leitner, A., Aebersold, R. & Raunser, S. Architecture and conformational switch mechanism of the ryanodine receptor. Nature 517, 39–43 (2015)

    CAS  PubMed  Article  ADS  PubMed Central  Google Scholar 

  12. 12

    Frauenfeld, J. et al. Cryo-EM structure of the ribosome–SecYE complex in the membrane environment. Nature Struct. Mol. Biol. 18, 614–621 (2011)

    CAS  Article  Google Scholar 

  13. 13

    Bai, X. C., McMullan, G. & Scheres, S. H. How cryo-EM is revolutionizing structural biology. Trends Biochem. Sci. 40, 49–57 (2015)

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  14. 14

    Cheng, Y. Single-particle cryo-EM at crystallographic resolution. Cell 161, 450–457 (2015)

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  15. 15

    Kühlbrandt, W. Cryo-EM enters a new era. eLife 3, e03678 (2014)

    PubMed  PubMed Central  Article  CAS  Google Scholar 

  16. 16

    Bevan, S., Quallo, T. & Andersson, D. A. Trpv1. Handb. Exp. Pharmacol. 222, 207–245 (2014)

    CAS  Google Scholar 

  17. 17

    Julius, D. TRP channels and pain. Annu. Rev. Cell Dev. Biol. 29, 355–384 (2013)

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  18. 18

    Cao, E., Liao, M., Cheng, Y. & Julius, D. TRPV1 structures in distinct conformations reveal activation mechanisms. Nature 504, 113–118 (2013)

    CAS  PubMed  PubMed Central  Article  ADS  Google Scholar 

  19. 19

    Liao, M., Cao, E., Julius, D. & Cheng, Y. Structure of the TRPV1 ion channel determined by electron cryo-microscopy. Nature 504, 107–112 (2013)

    CAS  PubMed  PubMed Central  Article  ADS  Google Scholar 

  20. 20

    Long, S. B., Tao, X., Campbell, E. B. & MacKinnon, R. Atomic structure of a voltage-dependent K+ channel in a lipid membrane-like environment. Nature 450, 376–382 (2007)

    CAS  PubMed  PubMed Central  Article  ADS  Google Scholar 

  21. 21

    Szallasi, A. & Blumberg, P. M. Resiniferatoxin, a phorbol-related diterpene, acts as an ultrapotent analog of capsaicin, the irritant constituent in red pepper. Neuroscience 30, 515–520 (1989)

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  22. 22

    Bohlen, C. J. et al. A bivalent tarantula toxin activates the capsaicin receptor, TRPV1, by targeting the outer pore domain. Cell 141, 834–845 (2010)

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  23. 23

    Chou, M. Z., Mtui, T., Gao, Y. D., Kohler, M. & Middleton, R. E. Resiniferatoxin binds to the capsaicin receptor (TRPV1) near the extracellular side of the S4 transmembrane domain. Biochemistry 43, 2501–2511 (2004)

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  24. 24

    Gavva, N. R. et al. Molecular determinants of vanilloid sensitivity in TRPV1. J. Biol. Chem. 279, 20283–20295 (2004)

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  25. 25

    Hanson, S. M., Newstead, S., Swartz, K. J. & Sansom, M. S. P. Capsaicin interaction with TRPV1 channels in a lipid bilayer: molecular dynamics simulation. Biophys. J. 108, 1425–1434 (2015)

    CAS  PubMed  PubMed Central  Article  ADS  Google Scholar 

  26. 26

    Jordt, S. E. & Julius, D. Molecular basis for species-specific sensitivity to “hot” chili peppers. Cell 108, 421–430 (2002)

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  27. 27

    Phillips, E., Reeve, A., Bevan, S. & McIntyre, P. Identification of species-specific determinants of the action of the antagonist capsazepine and the agonist PPAHV on TRPV1. J. Biol. Chem. 279, 17165–17172 (2004)

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  28. 28

    Yang, F. et al. Structural mechanism underlying capsaicin binding and activation of the TRPV1 ion channel. Nat. Chem. Biol. 11, 518–524 (2015)

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  29. 29

    Bevan, S. et al. Capsazepine: a competitive antagonist of the sensory neurone excitant capsaicin. Br. J. Pharmacol. 107, 544–552 (1992)

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  30. 30

    Boukalova, S., Marsakova, L., Teisinger, J. & Vlachova, V. Conserved residues within the putative S4-S5 region serve distinct functions among thermosensitive vanilloid transient receptor potential (TRPV) channels. J. Biol. Chem. 285, 41455–41462 (2010)

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  31. 31

    Lee, S. Y. & MacKinnon, R. A membrane-access mechanism of ion channel inhibition by voltage sensor toxins from spider venom. Nature 430, 232–235 (2004)

    CAS  PubMed  Article  ADS  PubMed Central  Google Scholar 

  32. 32

    Milescu, M. et al. Interactions between lipids and voltage sensor paddles detected with tarantula toxins. Nature Struct. Mol. Biol. 16, 1080–1085 (2009)

    CAS  Article  Google Scholar 

  33. 33

    Milescu, M. et al. Tarantula toxins interact with voltage sensors within lipid membranes. J. Gen. Physiol. 130, 497–511 (2007)

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  34. 34

    Bae, C. et al. Structural insights into the mechanism of activation of the TRPV1 channel by a membrane-bound tarantula toxin. eLife 5, e11273 (2016)

    PubMed  PubMed Central  Article  Google Scholar 

  35. 35

    Hardie, R. C. TRP channels and lipids: from Drosophila to mammalian physiology. J. Physiol. 578, 9–24 (2007)

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  36. 36

    Qin, F. Regulation of TRP ion channels by phosphatidylinositol-4,5-bisphosphate. Handb. Exp. Pharmacol. 179, 509–525 (2007)

    CAS  Article  Google Scholar 

  37. 37

    Rohacs, T. Phosphoinositide regulation of TRPV1 revisited. Pflugers Arch. 467, 1851–1869 (2015)

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  38. 38

    Cao, E., Cordero-Morales, J. F., Liu, B., Qin, F. & Julius, D. TRPV1 channels are intrinsically heat sensitive and negatively regulated by phosphoinositide lipids. Neuron 77, 667–679 (2013)

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  39. 39

    Prescott, E. D. & Julius, D. A modular PIP2 binding site as a determinant of capsaicin receptor sensitivity. Science 300, 1284–1288 (2003)

    CAS  PubMed  Article  ADS  PubMed Central  Google Scholar 

  40. 40

    Ufret-Vincenty, C. A. et al. Mechanism for phosphoinositide selectivity and activation of TRPV1 ion channels. J. Gen. Physiol. 145, 431–442 (2015)

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  41. 41

    Ufret-Vincenty, C. A., Klein, R. M., Hua, L., Angueyra, J. & Gordon, S. E. Localization of the PIP2 sensor of TRPV1 ion channels. J. Biol. Chem. 286, 9688–9698 (2011)

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  42. 42

    Hansen, S. B., Tao, X. & MacKinnon, R. Structural basis of PIP2 activation of the classical inward rectifier K+ channel Kir2.2. Nature 477, 495–498 (2011)

    CAS  PubMed  PubMed Central  ADS  Google Scholar 

  43. 43

    Booth, D. S., Avila-Sakar, A. & Cheng, Y. Visualizing proteins and macromolecular complexes by negative stain EM: from grid preparation to image acquisition. J. Vis. Exp. 58, 3227 (2011)

    Google Scholar 

  44. 44

    Li, X. et al. Electron counting and beam-induced motion correction enable near-atomic-resolution single-particle cryo-EM. Nature Methods 10, 584–590 (2013)

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  45. 45

    Li, X., Zheng, S., Agard, D. A. & Cheng, Y. Asynchronous data acquisition and on-the-fly analysis of dose fractionated cryoEM images by UCSFImage. J. Struct. Biol. 192, 174–178 (2015)

    PubMed  PubMed Central  Article  Google Scholar 

  46. 46

    Rohou, A. & Grigorieff, N. CTFFIND4: fast and accurate defocus estimation from electron micrographs. J. Struct. Biol. 192, 216–221 (2015)

    PubMed  PubMed Central  Article  Google Scholar 

  47. 47

    Frank, J. et al. SPIDER and WEB: processing and visualization of images in 3D electron microscopy and related fields. J. Struct. Biol. 116, 190–199 (1996)

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  48. 48

    Grant, T. & Grigorieff, N. Measuring the optimal exposure for single particle cryo-EM using a 2.6 Å reconstruction of rotavirus VP6. eLife 4, e06980 (2015)

    PubMed  PubMed Central  Article  Google Scholar 

  49. 49

    Scheres, S. H. W. RELION: implementation of a Bayesian approach to cryo-EM structure determination. J. Struct. Biol. 180, 519–530 (2012)

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  50. 50

    Scheres, S. H. W. & Chen, S. Prevention of overfitting in cryo-EM structure determination. Nature Methods 9, 853–854 (2012)

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  51. 51

    Kucukelbir, A., Sigworth, F. J. & Tagare, H. D. Quantifying the local resolution of cryo-EM density maps. Nature Methods 11, 63–65 (2014)

    CAS  Article  Google Scholar 

  52. 52

    Hohn, M. et al. SPARX, a new environment for cryo-EM image processing. J. Struct. Biol. 157, 47–55 (2007)

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  53. 53

    Tang, G. et al. EMAN2: an extensible image processing suite for electron microscopy. J. Struct. Biol. 157, 38–46 (2007)

    CAS  PubMed  PubMed Central  Google Scholar 

  54. 54

    Pettersen, E. F. et al. UCSF Chimera—a visualization system for exploratory research and analysis. J. Comput. Chem. 25, 1605–1612 (2004)

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  55. 55

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

    CAS  PubMed  PubMed Central  Google Scholar 

  56. 56

    Moriarty, N. W., Grosse-Kunstleve, R. W. & Adams, P. D. electronic Ligand Builder and Optimization Workbench (eLBOW): a tool for ligand coordinate and restraint generation. Acta Crystallogr. D 65, 1074–1080 (2009)

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  57. 57

    Adams, P. D. et al. PHENIX: a comprehensive Python-based system for macromolecular structure solution. Acta Crystallogr. D 66, 213–221 (2010)

    CAS  PubMed  PubMed Central  Google Scholar 

  58. 58

    van Aalten, D. M. F. et al. PRODRG, a program for generating molecular topologies and unique molecular descriptors from coordinates of small molecules. J. Comput. Aided Mol. Des. 10, 255–262 (1996)

    CAS  PubMed  Article  ADS  PubMed Central  Google Scholar 

  59. 59

    Afonine, P. V., Headd, J. J., Terwilliger, T. C. & Adams, P. D. New tool: phenix. real_space_refine. Computational Crystallography Newsletter 4, 43–44 (2013)

    Google Scholar 

  60. 60

    Amunts, A. et al. Structure of the yeast mitochondrial large ribosomal subunit. Science 343, 1485–1489 (2014)

    CAS  PubMed  PubMed Central  Article  ADS  Google Scholar 

  61. 61

    Chen, V. B. et al. MolProbity: all-atom structure validation for macromolecular crystallography. Acta Crystallogr. D 66, 12–21 (2010)

    CAS  PubMed  PubMed Central  Article  Google Scholar 

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Acknowledgements

We thank our laboratory colleagues, past and present, for many helpful discussions and manuscript critiques, C. Paulsen and E. Green for helping with initial screening for nanodisc reconstitution, and E. Palovcak for providing scripts for focused classification. This work was supported by grants from the National Institutes of Health (R01NS047723, R37NS065071 and R01NS055299 to D.J., S10OD020054, R01GM098672, P01GM111126 and P50GM082250 to Y.C.). Y.C. is an Investigator with the Howard Hughes Medical Institute.

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Contributions

Y.G. carried out protein purification, nanodisc reconstitution, and detailed cryo-EM experiments, including data acquisition, image processing, atomic model building and refinement of TRPV1–nanodisc complexes. E.C. carried out cryo-EM experiments of the TRPV1–capsazepine complex solubilized in amphipol. All authors contributed to experimental design, data analysis, and manuscript preparation.

Corresponding authors

Correspondence to David Julius or Yifan Cheng.

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The authors declare no competing financial interests.

Extended data figures and tables

Extended Data Figure 1 Reconstitution of TRPV1 into lipid nanodisc.

a, Size-exclusion chromatography of TRPV1 channel reconstituted into lipid nanodisc using MSP2N2. Void volume and peaks corresponding to TRPV1 and cleaved MBP are indicated. b, SDS–polyacrylamide gel electrophoresis (SDS–PAGE) of detergent-solubilized MBP–TRPV1 fusion protein and material from nanodisc reconstituted with TRPV1 following MBP cleavage (middle peak in a). Note the presence of both bands for TRPV1 and MSP2N2. c, Representative micrograph of negative-stained TRPV1–nanodisc sample showing mono-dispersed and homogeneous particles. d, Reference-free two-dimensional class averages of particles in c, revealing band-like density contributed by the lipid disc (side view) and tetrameric arrangement of channel subunits (top view). e, Two-dimensional class averages of the same protein reconstituted into MSP1E3 nanodisc, which is smaller in diameter. Note the extra space within the disc offered by MSP2N2 scaffold protein in d. The size of the class average window is 258 Å.

Extended Data Figure 2 Single-particle cryo-EM of unliganded TRPV1 in lipid nanodisc.

a, Representative raw micrograph of apo TRPV1 in nanodisc. b, Fourier transform of image in a. Note that Thon rings are visible at up to 3 Å. c, Gallery of two-dimensional class averages, with size of window as 233 Å. d, Slices through the unsharpened density map at different levels along the channel symmetry axis (numbers start from extracellular side). e, Euler angle distribution of all particles included in the calculation of the final three-dimensional reconstruction. Position of each sphere (grey) relative to the density map (green) represents its angle assignment and the radius of the sphere is proportional to the amount of particles in this specific orientation. f, Final three-dimensional density map coloured with local resolution in side and top views. g, Fourier shell coefficient (FSC) curves between two independently refined half maps before (blue) and after (red) the post-processing in RELION. h, FSC curves for cross-validation: model versus summed map (purple), model versus half map 1 (used in test refinement, cyan), model versus half map 2 (not used in test refinement, orange). Small differences between the ‘work’ and ‘free’ curves indicate little effect of over-fitting.

Extended Data Figure 3 Single-particle cryo-EM studies of agonist-bound TRPV1 in lipid nanodisc.

a, Representative raw micrograph of TRPV1–RTX/DkTx in nanodisc. b, Fourier transform of image in a. c, Gallery of two-dimensional class averages, with size of window as 233 Å. d, Slices through the unsharpened density map at different levels along the channel symmetry axis (numbers start from extracellular side). e, Euler angle distribution of all particles included in the calculation of the final three-dimensional reconstruction. Position of each sphere (grey) relative to the density map (green) represents its angle assignment and the radius of the sphere is proportional to the amount of particles in this specific orientation. f, Final three-dimensional density map coloured with local resolution in side and top views. g, FSC curves between two independently refined half maps before (blue) and after (red) the post-processing in RELION. h, FSC curves for cross-validation: model versus summed map (purple), model versus half map 1 (used in test refinement, cyan), model versus half map 2 (not used in test refinement, orange). Small differences between the ‘work’ and ‘free’ curves indicate little effect of over-fitting.

Extended Data Figure 4 Single-particle cryo-EM studies of antagonist-bound TRPV1 in lipid nanodisc.

a, Representative raw micrograph of TRPV1–capsazepine complex in nanodisc. b, Fourier transform of image in a. c, Gallery of two-dimensional class averages, with size of window as 233 Å. d, Slices through the unsharpened density map at different levels along the channel symmetry axis (numbers start from extracellular side). e, Euler angle distribution of all particles included in the calculation of the final three-dimensional reconstruction. Position of each sphere (grey) relative to the density map (green) represents its angle assignment and the radius of the sphere is proportional to the amount of particles in this specific orientation. f, Final three-dimensional density map coloured with local resolution in side and top views. g, FSC curves between two independently refined half maps before (blue) and after (red) the post-processing in RELION. h, FSC curves for cross-validation: model versus summed map (purple), model versus half map 1 (used in test refinement, cyan), model versus half map 2 (not used in test refinement, orange). Small differences between the ‘work’ and ‘free’ curves indicate little effect of over-fitting.

Extended Data Figure 5 Improved resolution for structures determined in nanodisc.

Comparison of density maps (blue mesh) determined from nanodisc- and amphipol-stabilized TRPV1 at various regions of the channel facing the lipid bilayer or at the bilayer surface. Refined atomic models (gold, nanodisc; grey, amphipol) are fit to corresponding densities. Side-chain densities were considerably improved in nanodisc-stabilized TRPV1–DkTx/RTX structure (a, b), and notable improvement was also seen for unliganded (c, d) and capsazepine-bound (e, f) channels in nanodisc.

Extended Data Figure 6 Newly resolved TRPV1 cytoplasmic region in nanodisc-stabilized structure.

a, A region in the TRPV1 C terminus, previously unresolved in amphipol-stabilized structures (blue) is clearly resolved in the nanodisc-stabilized structure. b, Enlarged view of boxed region in a showing density map (blue mesh) and superimposed model (gold). Previously resolved TRP domain and N-terminal β-strands are depicted in ribbon diagram format (cyan).

Extended Data Figure 7 Categories of lipid densities observed in TRPV1 structures.

a, Two continuous layers of density (blue) contributed by lipid head groups of bilayer within nanodisc are shown for apo channel (left) and channel in complex with RTX–DkTx (right). b, Atomic model of annular lipids could be built into well-resolved densities (blue mesh) surrounding the channel protein. DkTx is shown as ribbon diagram (pink). Top-down views show distribution of resolved annular lipids (blue) in inter-subunit crevices at the outer leaflet of the membrane. c, Well-resolved densities (blue mesh) in the structures representing a phosphatidylcholine molecule (left) and a phosphatidylinositol molecule (right). Transmembrane helices of TRPV1 close to the binding site are also shown as ribbon diagrams (grey).

Extended Data Figure 8 Focused analysis of DkTx density map.

a, Flow-chart showing procedures of focused three-dimensional classification of DkTx and proximal regions (see Methods for details). b, Atomic models for both knots of DkTx are superimposed on density maps (pink mesh).

Extended Data Figure 9 Lipid co-factor and vanilloids at the vanilloid binding site of TRPV1.

a, Chemical structure of phosphatidylinositol. b, Local environment of the phosphatidylinositol-binding site may accommodate multiple phosphatidylinositide species with phosphate substituents at the 3, 4 and/or 5 positions of the inositol ring (drawn in red). Adjacent regions of the channel are shown as ribbon diagram (grey). c, Tyr511 assumes two possible orientations that differ in apo versus agonist-bound states of the TRPV1 channel. In the apo state, one acyl chain of the resident phosphatidylinositol lipid (blue mesh superimposed with atomic model) prevents the Tyr511 side chain from assuming the upward rotamer position. d, Density maps of vanilloids (resiniferatoxin, red mesh; capsazepine, gold mesh) superimposed with density of the bound phosphatidylinositol lipid (blue mesh), suggesting that they occupy overlapping, but not identical sites. Atomic models for both drugs and their chemical structures are also shown. e, Overlap of transmembrane region of one TRPV1 subunit corresponding to apo (blue) and RTX/DkTx-bound (orange) states. Note the relatively small conformational change of the voltage sensor-like domain (S1–S4, boxed region). f, Overlap of transmembrane region of one TRPV1 subunit corresponding to apo (blue) and capsazepine-bound (gold) states.

Extended Data Table 1 Summary of data sets and statistics

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Gao, Y., Cao, E., Julius, D. et al. TRPV1 structures in nanodiscs reveal mechanisms of ligand and lipid action. Nature 534, 347–351 (2016). https://doi.org/10.1038/nature17964

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