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- EMDB-21538: Structure of TRPA1 (ligand-free and calcium-free), PMAL-C8, class 2 -

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Open data


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Basic information

Entry
Database: EMDB / ID: EMD-21538
TitleStructure of TRPA1 (ligand-free and calcium-free), PMAL-C8, class 2
Map dataTRPA1 (apo) in PMAL-C8, class 2 (open)
Sample
  • Complex: TRPA1 (ligand-free and calcium free), solubilized in PMAL-C8, class 2
    • Protein or peptide: Transient receptor potential A1 (TRPA1)
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 4.3 Å
AuthorsZhao J / Lin King JV / Paulsen CE / Cheng Y / Julius D
Funding support United States, 2 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of Neurological Disorders and Stroke (NIH/NINDS)R35 NS105038 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R01 GM098672 United States
CitationJournal: 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.
History
DepositionMar 11, 2020-
Header (metadata) releaseApr 8, 2020-
Map releaseMay 6, 2020-
UpdateJul 29, 2020-
Current statusJul 29, 2020Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 4
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 4
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_21538.png

Downloads & links

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Map

FileDownload / File: emd_21538.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationTRPA1 (apo) in PMAL-C8, class 2 (open)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.22 Å/pix.
x 256 pix.
= 311.194 Å		1.22 Å/pix.
x 256 pix.
= 311.194 Å		1.22 Å/pix.
x 256 pix.
= 311.194 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.2156 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 4 / Movie #1: 4
Minimum - Maximum-14.989781 - 22.766779
Average (Standard dev.)-0.00000000277 (±1)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 311.1936 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.21560156251.21560156251.2156015625
M x/y/z256256256
origin x/y/z0.0000.0000.000
length x/y/z311.194311.194311.194
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ400400400
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS256256256
D min/max/mean-14.99022.767-0.000

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Supplemental data

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Mask #1

Fileemd_21538_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_21538_half_map_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #2

Fileemd_21538_half_map_2.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : TRPA1 (ligand-free and calcium free), solubilized in PMAL-C8, class 2

EntireName: TRPA1 (ligand-free and calcium free), solubilized in PMAL-C8, class 2
Components
  • Complex: TRPA1 (ligand-free and calcium free), solubilized in PMAL-C8, class 2
    • Protein or peptide: Transient receptor potential A1 (TRPA1)

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Supramolecule #1: TRPA1 (ligand-free and calcium free), solubilized in PMAL-C8, class 2

SupramoleculeName: TRPA1 (ligand-free and calcium free), solubilized in PMAL-C8, class 2
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Homo sapiens (human) / Recombinant cell: HEK293F

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Macromolecule #1: Transient receptor potential A1 (TRPA1)

MacromoleculeName: Transient receptor potential A1 (TRPA1) / type: protein_or_peptide / ID: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MGSSHHHHHH HHGSSMKIEE GKLVIWINGD KGYNGLAEVG KKFEKDTGIK VTVEHPDKLE EKFPQVAATG DGPDIIFWAH DRFGGYAQSG LLAEITPDKA FQDKLYPFTW DAVRYNGKLI AYPIAVEALS LIYNKDLLPN PPKTWEEIPA LDKELKAKGK SALMFNLQEP ...String:
MGSSHHHHHH HHGSSMKIEE GKLVIWINGD KGYNGLAEVG KKFEKDTGIK VTVEHPDKLE EKFPQVAATG DGPDIIFWAH DRFGGYAQSG LLAEITPDKA FQDKLYPFTW DAVRYNGKLI AYPIAVEALS LIYNKDLLPN PPKTWEEIPA LDKELKAKGK SALMFNLQEP YFTWPLIAAD GGYAFKYENG KYDIKDVGVD NAGAKAGLTF LVDLIKNKHM NADTDYSIAE AAFNKGETAM TINGPWAWSN IDTSKVNYGV TVLPTFKGQP SKPFVGVLSA GINAASPNKE LAKEFLENYL LTDEGLEAVN KDKPLGAVAL KSYEEELAKD PRIAATMENA QKGEIMPNIP QMSAFWYAVR TAVINAASGR QTVDEALKDA QTNSSSNNNN NNNNNNLGIE ENLYFQGAMG SMKRSLRKMW RPGEKKEPQG VVYEDVPDDT EDFKESLKVV FEGSAYGLQN FNKQKKLKRC DDMDTFFLHY AAAEGQIELM EKITRDSSLE VLHEMDDYGN TPLHCAVEKN QIESVKFLLS RGANPNLRNF NMMAPLHIAV QGMNNEVMKV LLEHRTIDVN LEGENGNTAV IIACTTNNSE ALQILLKKGA KPCKSNKWGC FPIHQAAFSG SKECMEIILR FGEEHGYSRQ LHINFMNNGK ATPLHLAVQN GDLEMIKMCL DNGAQIDPVE KGRCTAIHFA ATQGATEIVK LMISSYSGSV DIVNTTDGCH ETMLHRASLF DHHELADYLI SVGADINKID SEGRSPLILA TASASWNIVN LLLSKGAQVD IKDNFGRNFL HLTVQQPYGL KNLRPEFMQM QQIKELVMDE DNDGCTPLHY ACRQGGPGSV NNLLGFNVSI HSKSKDKKSP LHFAASYGRI NTCQRLLQDI SDTRLLNEGD LHGMTPLHLA AKNGHDKVVQ LLLKKGALFL SDHNGWTALH HASMGGYTQT MKVILDTNLK CTDRLDEDGN TALHFAAREG HAKAVALLLS HNADIVLNKQ QASFLHLALH NKRKEVVLTI IRSKRWDECL KIFSHNSPGN KCPITEMIEY LPECMKVLLD FCMLHSTEDK SCRDYYIEYN FKYLQCPLEF TKKTPTQDVI YEPLTALNAM VQNNRIELLN HPVCKEYLLM KWLAYGFRAH MMNLGSYCLG LIPMTILVVN IKPGMAFNST GIINETSDHS EILDTTNSYL IKTCMILVFL SSIFGYCKEA GQIFQQKRNY FMDISNVLEW IIYTTGIIFV LPLFVEIPAH LQWQCGAIAV YFYWMNFLLY LQRFENCGIF IVMLEVILKT LLRSTVVFIF LLLAFGLSFY ILLNLQDPFS SPLLSIIQTF SMMLGDINYR ESFLEPYLRN ELAHPVLSFA QLVSFTIFVP IVLMNLLIGL AVGDIADVQK HASLKRIAMQ VELHTSLEKK LPLWFLRKVD QKSTIVYPNK PRSGGMLFHI FCFLFCTGEI RQEIPNADKS LEMEILKQKY RLKDLTFLLE KQHELIKLII QKMEIISETE DDDSHCSFQD RFKKEQMEQR NSRWNTVLRA VKAKTHHLEP

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

BufferpH: 8
GridDetails: unspecified
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI POLARA 300
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: SUPER-RESOLUTION / Average electron dose: 40.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD
Experimental equipment
Model: Tecnai Polara / Image courtesy: FEI Company

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Image processing

Final reconstructionApplied symmetry - Point group: C4 (4 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 4.3 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 28558
Initial angle assignmentType: NOT APPLICABLE
Final angle assignmentType: PROJECTION MATCHING
FSC plot (resolution estimation)

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