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- EMDB-20143: Cryo-EM structure of the C4-symmetric TRPV2/RTx complex in amphip... -

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Basic information

Entry
Database: EMDB / ID: EMD-20143
TitleCryo-EM structure of the C4-symmetric TRPV2/RTx complex in amphipol resolved to 2.9 A
Map data
SampleTRPV2:
ligand
Function / homology
Function and homology information


growth cone membrane / positive regulation of calcium ion import / positive regulation of axon extension / calcium channel activity / axonal growth cone / cell body / positive regulation of cold-induced thermogenesis / response to heat / cell surface / integral component of membrane / identical protein binding
Ankyrin repeat / Ankyrin repeat-containing domain / Ankyrin repeat region circular profile. / Ankyrin repeat profile. / Ankyrin repeats (3 copies) / Ion transport protein / Ankyrin repeat-containing domain superfamily / Transient receptor potential channel, vanilloid 2 / Transient receptor potential cation channel subfamily V / Ion transport domain / Transient receptor potential channel, vanilloid 1-4
Transient receptor potential cation channel subfamily V member 2
Biological speciesOryctolagus cuniculus (rabbit)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.9 Å
AuthorsZubcevic L / Hsu AL / Borgnia MJ / Lee S-Y
CitationJournal: Elife / Year: 2019
Title: Symmetry transitions during gating of the TRPV2 ion channel in lipid membranes.
Authors: Lejla Zubcevic / Allen L Hsu / Mario J Borgnia / Seok-Yong Lee /
Abstract: The Transient Receptor Potential Vanilloid 2 (TRPV2) channel is a member of the temperature-sensing thermoTRPV family. Recent advances in cryo-electronmicroscopy (cryo-EM) and X-ray crystallography ...The Transient Receptor Potential Vanilloid 2 (TRPV2) channel is a member of the temperature-sensing thermoTRPV family. Recent advances in cryo-electronmicroscopy (cryo-EM) and X-ray crystallography have provided many important insights into the gating mechanisms of thermoTRPV channels. Interestingly, crystallographic studies of ligand-dependent TRPV2 gating have shown that the TRPV2 channel adopts two-fold symmetric arrangements during the gating cycle. However, it was unclear if crystal packing forces played a role in stabilizing the two-fold symmetric arrangement of the channel. Here, we employ cryo-EM to elucidate the structure of full-length rabbit TRPV2 in complex with the agonist resiniferatoxin (RTx) in nanodiscs and amphipol. We show that RTx induces two-fold symmetric conformations of TRPV2 in both environments. However, the two-fold symmetry is more pronounced in the native-like lipid environment of the nanodiscs. Our data offers insights into a gating pathway in TRPV2 involving symmetry transitions.
Validation ReportPDB-ID: 6oo3

SummaryFull reportAbout validation report
DateDeposition: Apr 22, 2019 / Header (metadata) release: May 1, 2019 / Map release: May 29, 2019 / Update: May 29, 2019

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.08
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 0.08
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: : PDB-6oo3
  • Surface level: 0.08
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

FileDownload / File: emd_20143.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.08 Å/pix.
x 256 pix.
= 276.48 Å
1.08 Å/pix.
x 256 pix.
= 276.48 Å
1.08 Å/pix.
x 256 pix.
= 276.48 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.08 Å
Density
Contour LevelBy AUTHOR: 0.08 / Movie #1: 0.08
Minimum - Maximum-0.3009168 - 0.43747014
Average (Standard dev.)-0.000030957082 (±0.014828271)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 276.48 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.081.081.08
M x/y/z256256256
origin x/y/z0.0000.0000.000
length x/y/z276.480276.480276.480
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS256256256
D min/max/mean-0.3010.437-0.000

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Supplemental data

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Sample components

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Entire TRPV2

EntireName: TRPV2
Details: TRPV2 in complex with RTx reconstituted into amphipol A8-35
Number of components: 3

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Component #1: protein, TRPV2

ProteinName: TRPV2
Details: TRPV2 in complex with RTx reconstituted into amphipol A8-35
Recombinant expression: No
MassTheoretical: 300 kDa
SourceSpecies: Oryctolagus cuniculus (rabbit)
Source (engineered)Expression System: Spodoptera frugiperda (fall armyworm) / Cell of expression system: Sf9

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Component #2: protein, TRPV2

ProteinName: TRPV2 / Number of Copies: 4 / Recombinant expression: No
MassTheoretical: 88.72268 kDa
SourceSpecies: Oryctolagus cuniculus (rabbit)
Source (engineered)Expression System: Spodoptera frugiperda (fall armyworm)

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Component #3: ligand, resiniferatoxin

LigandName: resiniferatoxin / Number of Copies: 4 / Recombinant expression: No
MassTheoretical: 0.628708 kDa

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Experimental details

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Sample preparation

SpecimenSpecimen state: Particle / Method: cryo EM
Sample solutionSpecimen conc.: 2 mg/mL / pH: 8
VitrificationInstrument: LEICA EM GP / Cryogen name: ETHANE / Temperature: 296 K / Humidity: 90 % / Details: Blotted 3 seconds before plunging.

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
ImagingMicroscope: FEI TITAN KRIOS
Electron gunElectron source: OTHER / Accelerating voltage: 300 kV / Electron dose: 42 e/Å2 / Illumination mode: OTHER
LensImaging mode: OTHER
Specimen HolderModel: OTHER
CameraDetector: FEI FALCON III (4k x 4k)

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Image acquisition

Image acquisitionNumber of digital images: 1293

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Image processing

ProcessingMethod: single particle reconstruction / Applied symmetry: C4 (4 fold cyclic) / Number of projections: 101570
3D reconstructionSoftware: RELION / Resolution: 2.9 Å / Resolution method: FSC 0.143 CUT-OFF
FSC plot (resolution estimation)

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Atomic model buiding

Modeling #1Refinement protocol: rigid body / Refinement space: REAL
Input PDB model: 5AN8
Chain ID: A
Output model

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