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- EMDB-6343: Cryo-EM study of a channel -

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Basic information

Entry
Database: EMDB / ID: EMD-6343
TitleCryo-EM study of a channel
Map dataReconstruction of channel
Sample
  • Sample: Piezo-type mechanosensitive ion channel component 1
  • Protein or peptide: Piezo-type mechanosensitive ion channel component 1
KeywordsCryo-EM / single particle
Function / homology
Function and homology information


mechanosensitive monoatomic cation channel activity / cuticular plate / positive regulation of cell-cell adhesion mediated by integrin / detection of mechanical stimulus / positive regulation of integrin activation / mechanosensitive monoatomic ion channel activity / stereocilium / positive regulation of myotube differentiation / monoatomic cation transport / lamellipodium membrane ...mechanosensitive monoatomic cation channel activity / cuticular plate / positive regulation of cell-cell adhesion mediated by integrin / detection of mechanical stimulus / positive regulation of integrin activation / mechanosensitive monoatomic ion channel activity / stereocilium / positive regulation of myotube differentiation / monoatomic cation transport / lamellipodium membrane / monoatomic cation channel activity / endoplasmic reticulum-Golgi intermediate compartment membrane / regulation of membrane potential / cellular response to mechanical stimulus / endoplasmic reticulum membrane / endoplasmic reticulum / identical protein binding / plasma membrane
Similarity search - Function
Piezo family / Piezo non-specific cation channel, R-Ras-binding domain / Piezo domain / Piezo non-specific cation channel, R-Ras-binding domain / Piezo
Similarity search - Domain/homology
Piezo-type mechanosensitive ion channel component 1
Similarity search - Component
Biological speciesMus musculus (house mouse)
Methodsingle particle reconstruction / cryo EM / negative staining / Resolution: 4.8 Å
AuthorsGe J / Li W / Zhao Q / Li N / Chen M / Zhi P / Li R / Gao N / Xiao B / Yang M
CitationJournal: Nature / Year: 2015
Title: Architecture of the mammalian mechanosensitive Piezo1 channel.
Authors: Jingpeng Ge / Wanqiu Li / Qiancheng Zhao / Ningning Li / Maofei Chen / Peng Zhi / Ruochong Li / Ning Gao / Bailong Xiao / Maojun Yang /
Abstract: Piezo proteins are evolutionarily conserved and functionally diverse mechanosensitive cation channels. However, the overall structural architecture and gating mechanisms of Piezo channels have ...Piezo proteins are evolutionarily conserved and functionally diverse mechanosensitive cation channels. However, the overall structural architecture and gating mechanisms of Piezo channels have remained unknown. Here we determine the cryo-electron microscopy structure of the full-length (2,547 amino acids) mouse Piezo1 (Piezo1) at a resolution of 4.8 Å. Piezo1 forms a trimeric propeller-like structure (about 900 kilodalton), with the extracellular domains resembling three distal blades and a central cap. The transmembrane region has 14 apparently resolved segments per subunit. These segments form three peripheral wings and a central pore module that encloses a potential ion-conducting pore. The rather flexible extracellular blade domains are connected to the central intracellular domain by three long beam-like structures. This trimeric architecture suggests that Piezo1 may use its peripheral regions as force sensors to gate the central ion-conducting pore.
History
DepositionJun 5, 2015-
Header (metadata) releaseSep 16, 2015-
Map releaseSep 23, 2015-
UpdateNov 25, 2015-
Current statusNov 25, 2015Processing site: PDBj / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.0273
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 0.0273
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-3jac
  • Surface level: 0.0273
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

400_6343.gif

Downloads & links

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Map

FileDownload / File: emd_6343.map.gz / Format: CCP4 / Size: 51.5 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationReconstruction of channel
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.32 Å/pix.
x 240 pix.
= 316.8 Å		1.32 Å/pix.
x 240 pix.
= 316.8 Å		1.32 Å/pix.
x 240 pix.
= 316.8 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.32 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.0273 / Movie #1: 0.0273
Minimum - Maximum-0.02647332 - 0.06010438
Average (Standard dev.)0.0007118 (±0.00467585)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions240240240
Spacing240240240
CellA=B=C: 316.80002 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.321.321.32
M x/y/z240240240
origin x/y/z0.0000.0000.000
length x/y/z316.800316.800316.800
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS240240240
D min/max/mean-0.0260.0600.001

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Supplemental data

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Sample components

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Entire : Piezo-type mechanosensitive ion channel component 1

EntireName: Piezo-type mechanosensitive ion channel component 1
Components
  • Sample: Piezo-type mechanosensitive ion channel component 1
  • Protein or peptide: Piezo-type mechanosensitive ion channel component 1

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Supramolecule #1000: Piezo-type mechanosensitive ion channel component 1

SupramoleculeName: Piezo-type mechanosensitive ion channel component 1 / type: sample / ID: 1000 / Number unique components: 1
Molecular weightTheoretical: 900 KDa

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Macromolecule #1: Piezo-type mechanosensitive ion channel component 1

MacromoleculeName: Piezo-type mechanosensitive ion channel component 1 / type: protein_or_peptide / ID: 1 / Number of copies: 3 / Recombinant expression: Yes
Source (natural)Organism: Mus musculus (house mouse) / synonym: mouse / Location in cell: Plasma membrane
Recombinant expressionOrganism: Homo sapiens (human) / Recombinant cell: HEK 293T / Recombinant plasmid: pCDNA3.1(-)
SequenceUniProtKB: Piezo-type mechanosensitive ion channel component 1

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Experimental details

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Structure determination

Methodnegative staining, cryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

BufferpH: 7.2
Details: 25 mM NaPIPES, 140 mM NaCl, 2 mM DTT and 0.026% (w/v) C12E10
StainingType: NEGATIVE
Details: Grids with adsorbed protein floated on 2% w/v uranyl acetate for 30 seconds
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
DateApr 25, 2014
Image recordingCategory: CCD / Film or detector model: GATAN K2 (4k x 4k) / Digitization - Sampling interval: 4 µm / Number real images: 2164 / Average electron dose: 16 e/Å2
Details: Every image is the average of 14 frames recorded by the direct electron detector
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal magnification: 22500
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

CTF correctionDetails: CTFFIND
Final reconstructionResolution.type: BY AUTHOR / Resolution: 4.8 Å / Resolution method: OTHER / Software - Name: RELION / Number images used: 30021

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Atomic model buiding 1

Initial modelPDB ID:

4rax


Chain - Chain ID: A
RefinementSpace: REAL
Output model

PDB-3jac:
Cryo-EM study of a channel

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