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- EMDB-5411: The Human Nuclear Pore Complex as Revealed by Cryo-Electron Tomography -

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Basic information

Entry
Database: EMDB / ID: EMD-5411
TitleThe Human Nuclear Pore Complex as Revealed by Cryo-Electron Tomography
Map dataAveraged structure of the human nuclear pore complex asymmetric unit
Sample
  • Sample: Asymmetric unit of the human nuclear pore complex
  • Organelle or cellular component: nuclear pore complexNuclear pore
KeywordsCryo-electron tomography / human nuclear pore complex / nuclear envelope / nucleocytoplasmic transport
Biological speciesHomo sapiens (human)
Methodsubtomogram averaging / cryo EM / Resolution: 66.0 Å
AuthorsMaimon T / Elad N / Dahan I / Medalia O
CitationJournal: Structure / Year: 2012
Title: The human nuclear pore complex as revealed by cryo-electron tomography.
Authors: Tal Maimon / Nadav Elad / Idit Dahan / Ohad Medalia /
Abstract: Nuclear pore complexes (NPCs) are the sole passage through the nuclear envelope, connecting the cytoplasm to the nucleoplasm. These gigantic molecular machines, over 100 MDa in molecular weight, ...Nuclear pore complexes (NPCs) are the sole passage through the nuclear envelope, connecting the cytoplasm to the nucleoplasm. These gigantic molecular machines, over 100 MDa in molecular weight, allow free diffusion of small molecules and ions while mediating selective energy-dependent nucleocytoplasmic transport of large macromolecules. Here, we applied cryo-electron tomography to human fibroblast cells, reconstructing their nuclear envelopes without applying any purification steps. From these reconstructions, we extracted subtomograms containing individual NPCs and utilized in silico subtomogram averaging procedures to determine the structure of the mammalian pore complex at a resolution of ∼6.6 nm. Beyond revealing the canonical features of the human NPC, our analysis identified inner lateral channels and fusing bridge-like structures, suggesting alternative routes of peripheral nuclear passage. Finally, we concluded from our structural analysis that the human NPC is structurally distinct from that of lower eukaryotes in terms of dimension and organization but resembles its amphibian (frog) counterpart.
History
DepositionApr 7, 2012-
Header (metadata) releaseApr 17, 2013-
Map releaseApr 17, 2013-
UpdateSep 25, 2013-
Current statusSep 25, 2013Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.115
  • Imaged by UCSF Chimera
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  • Surface view colored by height
  • Surface level: 0.115
  • Imaged by UCSF Chimera
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Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_5411.jpg

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Map

FileDownload / File: emd_5411.map.gz / Format: CCP4 / Size: 7.8 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationAveraged structure of the human nuclear pore complex asymmetric unit
Voxel sizeX=Y=Z: 8.2 Å
Density
Contour LevelBy AUTHOR: 0.115 / Movie #1: 0.115
Minimum - Maximum-0.37820762 - 0.48475263
Average (Standard dev.)0.00469575 (±0.07388216)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions128128128
Spacing128128128
CellA=B=C: 1049.6 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z8.28.28.2
M x/y/z128128128
origin x/y/z0.0000.0000.000
length x/y/z1049.6001049.6001049.600
α/β/γ90.00090.00090.000
start NX/NY/NZ-62-62-62
NX/NY/NZ125125125
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS128128128
D min/max/mean-0.3780.4850.005

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Supplemental data

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Sample components

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Entire : Asymmetric unit of the human nuclear pore complex

EntireName: Asymmetric unit of the human nuclear pore complex
Components
  • Sample: Asymmetric unit of the human nuclear pore complex
  • Organelle or cellular component: nuclear pore complexNuclear pore

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Supramolecule #1000: Asymmetric unit of the human nuclear pore complex

SupramoleculeName: Asymmetric unit of the human nuclear pore complex / type: sample / ID: 1000
Oligomeric state: octamer (actual refined structure represents an asymmetric unit of the NPC)
Number unique components: 1

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Supramolecule #1: nuclear pore complex

SupramoleculeName: nuclear pore complex / type: organelle_or_cellular_component / ID: 1 / Oligomeric state: octamer / Recombinant expression: No / Database: NCBI
Source (natural)Organism: Homo sapiens (human) / synonym: human / Cell: U2OS / Organelle: Nucleus / Location in cell: Nuclear envelope

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Experimental details

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Structure determination

Methodcryo EM
Processingsubtomogram averaging
Aggregation stateparticle

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Sample preparation

BufferDetails: Dulbecco phosphate buffer saline
GridDetails: 200 mesh gold grids, silicon coated
VitrificationCryogen name: ETHANE / Instrument: OTHER / Method: Blot for 4 seconds before plunging

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Electron microscopy

MicroscopeFEI POLARA 300
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 1.2 µm / Nominal defocus min: 1.2 µm / Nominal magnification: 18000
Specialist opticsEnergy filter - Name: GIF2002
Sample stageSpecimen holder: nitrogen cooled / Specimen holder model: GATAN LIQUID NITROGEN / Tilt series - Axis1 - Min angle: -60 ° / Tilt series - Axis1 - Max angle: 60 °
TemperatureAverage: 90 K
DateJan 1, 2010
Experimental equipment
Model: Tecnai Polara / Image courtesy: FEI Company

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Image processing

Final reconstructionAlgorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 66.0 Å / Resolution method: FSC 0.5 CUT-OFF / Software - Name: TOM_toolbox_software_package / Number subtomograms used: 1696
Detailsmanual selection

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