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Conformational dynamics in TRPV1 channels reported by an encoded coumarin amino acid
Journal article   Open access   Peer reviewed

Conformational dynamics in TRPV1 channels reported by an encoded coumarin amino acid

Ximena Steinberg, Marina A Kasimova, Deny Cabezas-Bratesco, Jason D Galpin, Ernesto Ladron-de-Guevara, Federica Villa, Vincenzo Carnevale, Leon Islas, Christopher A Ahern and Sebastian E Brauchi
eLife, Vol.6, e28626
12/05/2017
DOI: 10.7554/eLife.28626
PMCID: PMC5716661
PMID: 29206105
url
https://doi.org/10.7554/eLife.28626 View
Published (Version of record) Open Access

Abstract

TRPV1 channels support the detection of noxious and nociceptive input. Currently available functional and structural data suggest that TRPV1 channels have two gates within their permeation pathway: one formed by a 'bundle-crossing' at the intracellular entrance and a second constriction at the selectivity filter. To describe conformational changes associated with channel gating, the fluorescent non-canonical amino acid coumarin-tyrosine was genetically encoded at Y671, a residue proximal to the selectivity filter. Total internal reflection fluorescence microscopy was performed to image the conformational dynamics of the channels in live cells. Photon counts and optical fluctuations from coumarin encoded within TRPV1 tetramers correlates with channel activation by capsaicin, providing an optical marker of conformational dynamics at the selectivity filter. In agreement with the fluorescence data, molecular dynamics simulations display alternating solvent exposure of Y671 in the closed and open states. Overall, the data point to a dynamic selectivity filter that may serve as a gate for permeation.
Capsaicin - metabolism Humans Models, Molecular TRPV Cation Channels - chemistry Tyrosine - analysis Molecular Dynamics Simulation TRPV Cation Channels - genetics TRPV Cation Channels - metabolism Coumarins - analysis Staining and Labeling HEK293 Cells Protein Conformation Intravital Microscopy Microscopy, Fluorescence

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