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- EMDB-21015: Two-pore channel 3 -

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Basic information

Entry
Database: EMDB / ID: EMD-21015
TitleTwo-pore channel 3
Map data
SampleDimer of two-pore channel 3 subunits:
Two pore channel 3 / ligand
Function / homologyIon transport domain / Voltage-dependent channel domain superfamily / ion channel activity / integral component of membrane / Two pore channel 3
Function and homology information
Biological speciesDanio rerio (zebrafish)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.11 Å
AuthorsDickinson MS / Stroud RM
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R01 GM24485 United States
CitationJournal: Proc Natl Acad Sci U S A / Year: 2020
Title: Resting state structure of the hyperdepolarization activated two-pore channel 3.
Authors: Miles Sasha Dickinson / Alexander Myasnikov / Jacob Eriksen / Nicole Poweleit / Robert M Stroud /
Abstract: Voltage-gated ion channels endow membranes with excitability and the means to propagate action potentials that form the basis of all neuronal signaling. We determined the structure of a voltage-gated ...Voltage-gated ion channels endow membranes with excitability and the means to propagate action potentials that form the basis of all neuronal signaling. We determined the structure of a voltage-gated sodium channel, two-pore channel 3 (TPC3), which generates ultralong action potentials. TPC3 is distinguished by activation only at extreme membrane depolarization (V ∼ +75 mV), in contrast to other TPCs and Na channels that activate between -20 and 0 mV. We present electrophysiological evidence that TPC3 voltage activation depends only on voltage sensing domain 2 (VSD2) and that each of the three gating arginines in VSD2 reduces the activation threshold. The structure presents a chemical basis for sodium selectivity, and a constricted gate suggests a closed pore consistent with extreme voltage dependence. The structure, confirmed by our electrophysiology, illustrates the configuration of a bona fide resting state voltage sensor, observed without the need for any inhibitory ligand, and independent of any chemical or mutagenic alteration.
Validation ReportPDB-ID: 6v1q

SummaryFull reportAbout validation report
History
DepositionNov 20, 2019-
Header (metadata) releaseDec 4, 2019-
Map releaseDec 4, 2019-
UpdateFeb 12, 2020-
Current statusFeb 12, 2020Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.863
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 0.863
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6v1q
  • Surface level: 0.863
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

FileDownload / File: emd_21015.map.gz / Format: CCP4 / Size: 83.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.26 Å/pix.
x 280 pix.
= 351.68 Å
1.26 Å/pix.
x 280 pix.
= 351.68 Å
1.26 Å/pix.
x 280 pix.
= 351.68 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.256 Å
Density
Contour LevelBy AUTHOR: 0.863 / Movie #1: 0.863
Minimum - Maximum-0.64165455 - 2.344392
Average (Standard dev.)-0.0008597384 (±0.0817106)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions280280280
Spacing280280280
CellA=B=C: 351.68002 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.2561.2561.256
M x/y/z280280280
origin x/y/z0.0000.0000.000
length x/y/z351.680351.680351.680
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ300300300
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS280280280
D min/max/mean-0.6422.344-0.001

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Supplemental data

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Additional map: Locally sharpened map from non-uniform refinement in cryoSPARC...

Fileemd_21015_additional.map
AnnotationLocally sharpened map from non-uniform refinement in cryoSPARC v2, used for atomic modeling.
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Sample components

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Entire Dimer of two-pore channel 3 subunits

EntireName: Dimer of two-pore channel 3 subunits / Number of components: 3

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Component #1: protein, Dimer of two-pore channel 3 subunits

ProteinName: Dimer of two-pore channel 3 subunits / Recombinant expression: No
SourceSpecies: Danio rerio (zebrafish)
Source (engineered)Expression System: Homo sapiens (human)

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Component #2: protein, Two pore channel 3

ProteinName: Two pore channel 3 / Number of Copies: 2 / Recombinant expression: No
MassTheoretical: 88.774164 kDa
SourceSpecies: Danio rerio (zebrafish)
Source (engineered)Expression System: Homo sapiens (human)

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Component #3: ligand, SODIUM ION

LigandName: SODIUM IONSodium / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 2.29905 MDa

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Experimental details

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Sample preparation

SpecimenSpecimen state: Particle / Method: cryo EM
Sample solutionSpecimen conc.: 3 mg/mL / pH: 7.4
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Temperature: 283 K / Humidity: 100 %

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
ImagingMicroscope: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Electron dose: 92 e/Å2 / Illumination mode: OTHER
LensCs: 2.7 mm / Imaging mode: BRIGHT FIELD
Specimen HolderModel: FEI TITAN KRIOS AUTOGRID HOLDER
CameraDetector: GATAN K2 SUMMIT (4k x 4k)

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Image processing

ProcessingMethod: single particle reconstruction / Applied symmetry: C2 (2 fold cyclic) / Number of projections: 213328
3D reconstructionSoftware: cryoSPARC / Resolution: 3.11 Å / Resolution method: FSC 0.143 CUT-OFF

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Atomic model buiding

Output model

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