|Entry||Database: EMDB / ID: EMD-21538|
|Title||Structure of TRPA1 (ligand-free and calcium-free), PMAL-C8, class 2|
|Sample||TRPA1 (ligand-free and calcium free), solubilized in PMAL-C8, class 2:|
Transient receptor potential A1 (TRPA1)
|Biological species||Homo sapiens (human)|
|Method||single particle reconstruction / cryo EM / Resolution: 4.3 Å|
|Authors||Zhao J / Lin King JV / Paulsen CE / Cheng Y / Julius D|
|Funding support|| United States, 2 items |
|Citation||Journal: Nature / Year: 2020|
Title: Irritant-evoked activation and calcium modulation of the TRPA1 receptor.
Authors: Jianhua Zhao / John V Lin King / Candice E Paulsen / Yifan Cheng / David Julius /
Abstract: The transient receptor potential ion channel TRPA1 is expressed by primary afferent nerve fibres, in which it functions as a low-threshold sensor for structurally diverse electrophilic irritants, ...The transient receptor potential ion channel TRPA1 is expressed by primary afferent nerve fibres, in which it functions as a low-threshold sensor for structurally diverse electrophilic irritants, including small volatile environmental toxicants and endogenous algogenic lipids. TRPA1 is also a 'receptor-operated' channel whose activation downstream of metabotropic receptors elicits inflammatory pain or itch, making it an attractive target for novel analgesic therapies. However, the mechanisms by which TRPA1 recognizes and responds to electrophiles or cytoplasmic second messengers remain unknown. Here we use strutural studies and electrophysiology to show that electrophiles act through a two-step process in which modification of a highly reactive cysteine residue (C621) promotes reorientation of a cytoplasmic loop to enhance nucleophilicity and modification of a nearby cysteine (C665), thereby stabilizing the loop in an activating configuration. These actions modulate two restrictions controlling ion permeation, including widening of the selectivity filter to enhance calcium permeability and opening of a canonical gate at the cytoplasmic end of the pore. We propose a model to explain functional coupling between electrophile action and these control points. We also characterize a calcium-binding pocket that is highly conserved across TRP channel subtypes and accounts for all aspects of calcium-dependent TRPA1 regulation, including potentiation, desensitization and activation by metabotropic receptors. These findings provide a structural framework for understanding how a broad-spectrum irritant receptor is controlled by endogenous and exogenous agents that elicit or exacerbate pain and itch.
|Validation Report||Summary, Full report, XML, About validation report|
|Structure viewer||EM map: |
Downloads & links
|File||Download / File: emd_21538.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)|
|Projections & slices|
Images are generated by Spider.
|Voxel size||X=Y=Z: 1.2156 Å|
|Symmetry||Space group: 1|
CCP4 map header:
|Projections & Slices|
-Half map: #1
|Projections & Slices|
-Half map: #2
-Entire TRPA1 (ligand-free and calcium free), solubilized in PMAL-C8, class 2
|Entire||Name: TRPA1 (ligand-free and calcium free), solubilized in PMAL-C8, class 2|
Number of components: 2
-Component #1: protein, TRPA1 (ligand-free and calcium free), solubilized in PMA...
|Protein||Name: TRPA1 (ligand-free and calcium free), solubilized in PMAL-C8, class 2|
Recombinant expression: No
|Source||Species: Homo sapiens (human)|
|Source (engineered)||Expression System: Homo sapiens (human) / Cell of expression system: HEK293F|
-Component #2: protein, Transient receptor potential A1 (TRPA1)
|Protein||Name: Transient receptor potential A1 (TRPA1) / Recombinant expression: No|
|Source||Species: Homo sapiens (human)|
|Source (engineered)||Expression System: Homo sapiens (human)|
|Specimen||Specimen state: Particle / Method: cryo EM|
|Sample solution||pH: 8|
|Vitrification||Cryogen name: ETHANE|
-Electron microscopy imaging
Model: Tecnai Polara / Image courtesy: FEI Company
|Imaging||Microscope: FEI POLARA 300|
|Electron gun||Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Electron dose: 40 e/Å2 / Illumination mode: FLOOD BEAM|
|Lens||Imaging mode: BRIGHT FIELD|
|Specimen Holder||Model: OTHER|
|Camera||Detector: GATAN K2 SUMMIT (4k x 4k)|
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