Transient receptor potential ankyrin 1 (TRPA1) is an excitatory ion channel involved in pain, inflammation and itching. This channel gates in response to many irritant and proalgesic agents, and can be modulated by calcium and depolarizing voltage. While the closed-state structure of TRPA1 has been recently resolved, also having its open state is essential for understanding how this channel works. Here we use molecular dynamics simulations combined with electrophysiological measurements and systematic mutagenesis to predict and explore the conformational changes coupled to the expansion of the presumptive channel's lower gate. We show that, upon opening, the upper part of the sensor module approaches the pore domain of an adjacent subunit and the conformational dynamics of the first extracellular flexible loop may govern the voltage-dependence of multimodal gating, thereby serving to stabilize the open state of the channel. These results are generally important in understanding the structure and function of TRPA1 and offer new insights into the gating mechanism of TRPA1 and related channels.

Department of Cellular Neurophysiology Institute of Physiology Czech Academy of Sciences Prague Czechia

Division of Biomolecular Physics Faculty of Mathematics and Physics Institute of Physics Charles University Prague Czechia

See more in PubMed

Abraham M. J., Murtola T., Schulz R., Páll S., Smith J. C., Hess B., et al. (2015). GROMACS: High performance molecular simulations through multi-level parallelism from laptops to supercomputers. SoftwareX 1–2, 19–25. 10.1016/j.softx.2015.06.001 DOI

Bae C., Anselmi C., Kalia J., Jara-Oseguera A., Schwieters C. D., Krepkiy D., et al. . (2016). Structural insights into the mechanism of activation of the TRPV1 channel by a membrane-bound tarantula toxin. eLife 5:e11273. 10.7554/eLife.11273 PubMed DOI PMC

Benedikt J., Teisinger J., Vyklicky L., Vlachova V. (2007). Ethanol inhibits cold-menthol receptor TRPM8 by modulating its interaction with membrane phosphatidylinositol 4,5-bisphosphate. J. Neurochem. 100, 211–224. 10.1111/j.1471-4159.2006.04192.x PubMed DOI

Best R. B., Zhu X., Shim J., Lopes P. E. M., Mittal J., Feig M., et al. . (2012). Optimization of the Additive CHARMM All-Atom Protein Force Field Targeting Improved Sampling of the Backbone phi, psi and Side-Chain chi(1) and chi(2) Dihedral Angles. J. Chem. Theory Comput. 8, 3257–3273. 10.1021/ct300400x PubMed DOI PMC

Brewster M. S., Gaudet R. (2015). How the TRPA1 receptor transmits painful stimuli: inner workings revealed by electron cryomicroscopy. Bioessays 37, 1184–1192. 10.1002/bies.201500085 PubMed DOI PMC

Cao E., Liao M., Cheng Y., Julius D. (2013). TRPV1 structures in distinct conformations reveal activation mechanisms. Nature 504, 113–118. 10.1038/nature12823 PubMed DOI PMC

Chacón P., Wriggers W. (2002). Multi-resolution contour-based fitting of macromolecular structures. J. Mol. Biol. 317, 375–384. 10.1006/jmbi.2002.5438 PubMed DOI

Chang Q., Hoefs S., van der Kemp A. W., Topala C. N., Bindels R. J., Hoenderop J. G. (2005). The β-glucuronidase klotho hydrolyzes and activates the TRPV5 channel. Science 310, 490–493. 10.1126/science.1114245 PubMed DOI

Chen X., Wang Q., Ni F., Ma J. (2010). Structure of the full-length Shaker potassium channel Kv1.2 by normal-mode-based X-ray crystallographic refinement. Proc. Natl. Acad. Sci. U.S.A. 107, 11352–11357. 10.1073/pnas.1000142107 PubMed DOI PMC

Christensen A. P., Akyuz N., Corey D. P. (2016). The outer pore and selectivity filter of TRPA1. PLoS ONE 11:e0166167. 10.1371/journal.pone.0166167 PubMed DOI PMC

Crooks G. E., Hon G., Chandonia J. M., Brenner S. E. (2004). WebLogo: a sequence logo generator. Genome Res. 14, 1188–1190. 10.1101/gr.849004 PubMed DOI PMC

Deering-Rice C. E., Shapiro D., Romero E. G., Stockmann C., Bevans T. S., Phan Q. M., et al. . (2015). Activation of transient receptor potential ankyrin-1 by insoluble particulate material and association with asthma. Am. J. Respir. Cell Mol. Biol. 53, 893–901. 10.1165/rcmb.2015-0086OC PubMed DOI PMC

Dietrich A., Mederos y Schnitzler M., Emmel J., Kalwa H., Hofmann T., Gudermann T. (2003). N-linked protein glycosylation is a major determinant for basal TRPC3 and TRPC6 channel activity. J. Biol. Chem. 278, 47842–47852. 10.1074/jbc.M302983200 PubMed DOI

Dittert I., Benedikt J., Vyklický L., Zimmermann K., Reeh P. W., Vlachova V. (2006). Improved superfusion technique for rapid cooling or heating of cultured cells under patch-clamp conditions. J. Neurosci. Methods 151, 178–185. 10.1016/j.jneumeth.2005.07.005 PubMed DOI

Doerner J. F., Gisselmann G., Hatt H., Wetzel C. H. (2007). Transient receptor potential channel A1 is directly gated by calcium ions. J. Biol. Chem. 282, 13180–13189. 10.1074/jbc.M607849200 PubMed DOI

Egan T. J., Acuña M. A., Zenobi-Wong M., Zeilhofer H. U., Urech D. (2016). Effects of N-glycosylation of the human cation channel TRPA1 on agonist-sensitivity. Biosci. Rep. 36:e00390 10.1042/BSR20160149 PubMed DOI PMC

Füll Y., Seebohm G., Lerche H., Maljevic S. (2013). A conserved threonine in the S1-S2 loop of KV7.2 and K V7.3 channels regulates voltage-dependent activation. Pflugers Arch. 465, 797–804. 10.1007/s00424-012-1184-x PubMed DOI

Gao Y., Cao E., Julius D., Cheng Y. (2016). TRPV1 structures in nanodiscs reveal mechanisms of ligand and lipid action. Nature 534, 347–351. 10.1038/nature17964 PubMed DOI PMC

Gui J., Liu B., Cao G., Lipchik A. M., Perez M., Dekan Z., et al. . (2014). A tarantula-venom peptide antagonizes the TRPA1 nociceptor ion channel by binding to the S1-S4 gating domain. Curr. Biol. 24, 473–483. 10.1016/j.cub.2014.01.013 PubMed DOI PMC

Hinman A., Chuang H. H., Bautista D. M., Julius D. (2006). TRP channel activation by reversible covalent modification. Proc. Natl. Acad. Sci. U.S.A. 103, 19564–19568. 10.1073/pnas.0609598103 PubMed DOI PMC

Ho B. K., Gruswitz F. (2008). HOLLOW: generating accurate representations of channel and interior surfaces in molecular structures. BMC Struct. Biol. 8:49. 10.1186/1472-6807-8-49 PubMed DOI PMC

Humphrey W., Dalke A., Schulten K. (1996). VMD: visual molecular dynamics. J. Mol. Graph. 14, 33–38. PubMed

Jorgensen W. L., Chandrasekhar J., Madura J. D., Impey R. W., Klein M. L. (1983). Comparison of simple potential functions for simulating liquid water. J. Chem. Phys. 79, 926–935. 10.1063/1.445869 DOI

Karashima Y., Prenen J., Talavera K., Janssens A., Voets T., Nilius B. (2010). Agonist-induced changes in Ca2+ permeation through the nociceptor cation channel TRPA1. Biophys. J. 98, 773–783. 10.1016/j.bpj.2009.11.007 PubMed DOI PMC

Kremeyer B., Lopera F., Cox J. J., Momin A., Rugiero F., Marsh S., et al. . (2010). A gain-of-function mutation in TRPA1 causes familial episodic pain syndrome. Neuron 66, 671–680. 10.1016/j.neuron.2010.04.030 PubMed DOI PMC

Lee S. Y., Banerjee A., MacKinnon R. (2009). Two separate interfaces between the voltage sensor and pore are required for the function of voltage-dependent K+ channels. PLoS Biol. 7:e47. 10.1371/journal.pbio.1000047 PubMed DOI PMC

Liao M., Cao E., Julius D., Cheng Y. (2013). Structure of the TRPV1 ion channel determined by electron cryo-microscopy. Nature 504, 107–112. 10.1038/nature12822 PubMed DOI PMC

Lindorff-Larsen K., Piana S., Palmo K., Maragakis P., Klepeis J. L., Dror R. O., et al. . (2010). Improved side-chain torsion potentials for the Amber ff99SB protein force field. Proteins 78, 1950–1958. 10.1002/prot.22711 PubMed DOI PMC

Long S. B., Tao X., Campbell E. B., MacKinnon R. (2007). Atomic structure of a voltage-dependent K+ channel in a lipid membrane-like environment. Nature 450, 376–382. 10.1038/nature06265 PubMed DOI

Macpherson L. J., Dubin A. E., Evans M. J., Marr F., Schultz P. G., Cravatt B. F., et al. . (2007). Noxious compounds activate TRPA1 ion channels through covalent modification of cysteines. Nature 445, 541–545. 10.1038/nature05544 PubMed DOI

Nilius B., Appendino G., Owsianik G. (2012). The transient receptor potential channel TRPA1: from gene to pathophysiology. Pflugers Archiv. 464, 425–458. 10.1007/s00424-012-1158-z PubMed DOI

Palovcak E., Delemotte L., Klein M. L., Carnevale V. (2015). Comparative sequence analysis suggests a conserved gating mechanism for TRP channels. J. Gen. Physiol. 146, 37–50. 10.1085/jgp.201411329 PubMed DOI PMC

Paulsen C. E., Armache J. P., Gao Y., Cheng Y., Julius D. (2015). Structure of the TRPA1 ion channel suggests regulatory mechanisms. Nature 520, 511–517. 10.1038/nature14367 PubMed DOI PMC

Phillips J. C., Braun R., Wang W., Gumbart J., Tajkhorshid E., Villa E., et al. . (2005). Scalable molecular dynamics with NAMD. J. Comput. Chem. 26, 1781–1802. 10.1002/jcc.20289 PubMed DOI PMC

Saotome K., Singh A. K., Yelshanskaya M. V., Sobolevsky A. I. (2016). Crystal structure of the epithelial calcium channel TRPV6. Nature 534, 506–511. 10.1038/nature17975 PubMed DOI PMC

Schwetz T. A., Norring S. A., Bennett E. S. (2010). N-glycans modulate Kv1.5 gating but have no effect on Kv1.4 gating. Biochim. Biophys. Acta 1798, 367–375. 10.1016/j.bbamem.2009.11.018 PubMed DOI

Story G. M., Peier A. M., Reeve A. J., Eid S. R., Mosbacher J., Hricik T. R., et al. . (2003). ANKTM1, a TRP-like channel expressed in nociceptive neurons, is activated by cold temperatures. Cell 112, 819–829. 10.1016/S0092-8674(03)00158-2 PubMed DOI

Sura L., Zima V., Marsakova L., Hynkova A., Barvik I., Vlachova V. (2012). C-terminal Acidic Cluster Is Involved in Ca2+-induced regulation of human transient receptor potential ankyrin 1 channel. J. Biol. Chem. 287, 18067–18077. 10.1074/jbc.M112.341859 PubMed DOI PMC

Trabuco L. G., Villa E., Mitra K., Frank J., Schulten K. (2008). Flexible fitting of atomic structures into electron microscopy maps using molecular dynamics. Structure 16, 673–683. 10.1016/j.str.2008.03.005 PubMed DOI PMC

Trabuco L. G., Villa E., Schreiner E., Harrison C. B., Schulten K. (2009). Molecular dynamics flexible fitting: a practical guide to combine cryo-electron microscopy and X-ray crystallography. Methods 49, 174–180. 10.1016/j.ymeth.2009.04.005 PubMed DOI PMC

Vriens J., Held K., Janssens A., Tóth B. I., Kerselaers S., Nilius B., et al. . (2014). Opening of an alternative ion permeation pathway in a nociceptor TRP channel. Nat. Chem. Biol. 10, 188–195. 10.1038/nchembio.1428 PubMed DOI

Wang Y. Y., Chang R. B., Waters H. N., McKemy D. D., Liman E. R. (2008). The nociceptor ion channel trpa1 is potentiated and inactivated by permeating calcium ions. J. Biol. Chem. 283, 32691–32703. 10.1074/jbc.M803568200. PubMed DOI PMC

Wriggers W., Milligan R. A., McCammon J. A. (1999). Situs: a package for docking crystal structures into low-resolution maps from electron microscopy. J. Struct. Biol. 125, 185–195. 10.1006/jsbi.1998.4080 PubMed DOI

Zhang X., Ren W., DeCaen P., Yan C., Tao X., Tang L., et al. . (2012). Crystal structure of an orthologue of the NaChBac voltage-gated sodium channel. Nature 486, 130–134. 10.1038/nature11054 PubMed DOI PMC

Zhu J., Watanabe I., Poholek A., Koss M., Gomez B., Yan C., et al. . (2003). Allowed N-glycosylation sites on the Kv1.2 potassium channel S1-S2 linker: implications for linker secondary structure and the glycosylation effect on channel function. Biochem. J. 375(Pt 3), 769–775. 10.1042/BJ20030517 PubMed DOI PMC

Zubcevic L., Herzik M. A., Jr., Chung B. C., Liu Z., Lander G. C., Lee S. Y. (2016). Cryo-electron microscopy structure of the TRPV2 ion channel. Nat. Struct. Mol. Biol. 23, 180–186. 10.1038/nsmb.3159 PubMed DOI PMC

Zurborg S., Yurgionas B., Jira J. A., Caspani O., Heppenstall P. A. (2007). Direct activation of the ion channel TRPA1 by Ca2+. Nat. Neurosci. 10, 277–279. 10.1038/nn1843 PubMed DOI

Zygmunt P. M., Hogestatt E. D. (2014). Trpa1. Handb. Exp. Pharmacol. 222, 583–630. 10.1007/978-3-642-54215-2_23 PubMed DOI

Human Transient Receptor Potential Ankyrin 1 Channel: Structure, Function, and Physiology

Human and Mouse TRPA1 Are Heat and Cold Sensors Differentially Tuned by Voltage