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Yorodumi- EMDB-25817: Cryo-EM map of protomer of the cytoplasmic ring of the nuclear po... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-25817 | |||||||||
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Title | Cryo-EM map of protomer of the cytoplasmic ring of the nuclear pore complex from Xenopus laevis | |||||||||
Map data | Cryo-EM map of protomer of the cytoplasmic ring of the Nuclear pore complex from Xenopus laevis. | |||||||||
Sample |
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Keywords | Nuclear pore complex / NUCLEAR PROTEIN | |||||||||
Function / homology | Function and homology information macromolecule localization / nitrogen compound transport / GATOR2 complex / nephron development / organic substance transport / macromolecule metabolic process / nuclear pore organization / nuclear pore outer ring / attachment of mitotic spindle microtubules to kinetochore / structural constituent of nuclear pore ...macromolecule localization / nitrogen compound transport / GATOR2 complex / nephron development / organic substance transport / macromolecule metabolic process / nuclear pore organization / nuclear pore outer ring / attachment of mitotic spindle microtubules to kinetochore / structural constituent of nuclear pore / nucleocytoplasmic transport / mitotic metaphase chromosome alignment / cellular response to nutrient levels / ribosomal small subunit export from nucleus / ribosomal large subunit export from nucleus / mRNA transport / intracellular transport / nuclear pore / positive regulation of TORC1 signaling / kinetochore / protein transport / nuclear membrane / lysosomal membrane / cell division / structural molecule activity / metal ion binding / nucleus / cytosol Similarity search - Function | |||||||||
Biological species | Xenopus laevis (African clawed frog) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 6.9 Å | |||||||||
Authors | Fontana P / Wu H | |||||||||
Funding support | 1 items
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Citation | Journal: Science / Year: 2022 Title: Structure of cytoplasmic ring of nuclear pore complex by integrative cryo-EM and AlphaFold. Authors: Pietro Fontana / Ying Dong / Xiong Pi / Alexander B Tong / Corey W Hecksel / Longfei Wang / Tian-Min Fu / Carlos Bustamante / Hao Wu / Abstract: INTRODUCTION The nuclear pore complex (NPC) is the molecular conduit in the nuclear membrane of eukaryotic cells that regulates import and export of biomolecules between the nucleus and the cytosol, ...INTRODUCTION The nuclear pore complex (NPC) is the molecular conduit in the nuclear membrane of eukaryotic cells that regulates import and export of biomolecules between the nucleus and the cytosol, with vertebrate NPCs ~110 to 125 MDa in molecular mass and ~120 nm in diameter. NPCs are organized into four main rings: the cytoplasmic ring (CR) at the cytosolic side, the inner ring and the luminal ring on the plane of the nuclear membrane, and the nuclear ring facing the nucleus. Each ring possesses an approximate eightfold symmetry and is composed of multiple copies of different nucleoporins. NPCs have been implicated in numerous biological processes, and their dysfunctions are associated with a growing number of serious human diseases. However, despite pioneering studies from many groups over the past two decades, we still lack a full understanding of NPCs' organization, dynamics, and complexity. RATIONALE We used the oocyte as a model system for the structural characterization because each oocyte possesses a large number of NPC particles that can be visualized on native nuclear membranes without the aid of detergent extraction. We used single-particle cryo-electron microscopy (cryo-EM) analysis on data collected at different stage tilt angles for three-dimensional reconstruction and structure prediction with AlphaFold for model building. RESULTS We reconstructed the CR map of NPC at 6.9 and 6.7 Å resolutions for the full CR protomer and a core region, respectively, and predicted the structures of the individual nucleoporins using AlphaFold because no high-resolution models of Nups were available. For any ambiguous subunit interactions, we also predicted complex structures, which further guided model fitting of the CR protomer. We placed the nucleoporin or complex structures into the CR density to obtain an almost full CR atomic model, composed of the inner and outer Y-complexes, two copies of Nup205, two copies of the Nup214-Nup88-Nup62 complex, one Nup155, and five copies of Nup358. In particular, we predicted the largest protein in the NPC, Nup358, as having an S-shaped globular domain, a coiled-coil domain, and a largely disordered C-terminal region containing phenylalanine-glycine (FG) repeats previously shown to form a gel-like condensate phase for selective cargo passage. Four of the Nup358 copies clamp around the inner and outer Y-complexes to stabilize the CR, and the fifth Nup358 situates in the center of the cluster of clamps. AlphaFold also predicted a homo-oligomeric, likely specifically pentameric, coiled-coil structure of Nup358 that may provide the avidity for Nup358 recruitment to the NPC and for lowering the threshold for Nup358 condensation in NPC biogenesis. CONCLUSION Our studies offer an example of integrative cryo-EM and structure prediction as a general approach for attaining more precise models of megadalton protein complexes from medium-resolution density maps. The more accurate and almost complete model of the CR presented here expands our understanding of the molecular interactions in the NPC and represents a substantial step forward toward the molecular architecture of a full NPC, with implications for NPC function, biogenesis, and regulation. [Figure: see text]. | |||||||||
History |
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-Structure visualization
Supplemental images |
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-Downloads & links
-EMDB archive
Map data | emd_25817.map.gz | 49.3 MB | EMDB map data format | |
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Header (meta data) | emd-25817-v30.xml emd-25817.xml | 7.9 KB 7.9 KB | Display Display | EMDB header |
Images | emd_25817.png | 69.3 KB | ||
Filedesc metadata | emd-25817.cif.gz | 3.6 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-25817 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-25817 | HTTPS FTP |
-Related structure data
Related structure data | 7tdzMC M: atomic model generated by this map C: citing same article (ref.) |
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Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_25817.map.gz / Format: CCP4 / Size: 103 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||
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Annotation | Cryo-EM map of protomer of the cytoplasmic ring of the Nuclear pore complex from Xenopus laevis. | ||||||||||||||||||||||||||||||||||||
Projections & slices | Image control
Images are generated by Spider. | ||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 2.8 Å | ||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Sample components
-Entire : Cytoplasmic ring of Nuclear Pore Complex
Entire | Name: Cytoplasmic ring of Nuclear Pore Complex |
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Components |
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-Supramolecule #1: Cytoplasmic ring of Nuclear Pore Complex
Supramolecule | Name: Cytoplasmic ring of Nuclear Pore Complex / type: complex / ID: 1 / Parent: 0 |
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Source (natural) | Organism: Xenopus laevis (African clawed frog) |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Buffer | pH: 7.5 |
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Vitrification | Cryogen name: ETHANE |
-Electron microscopy
Microscope | TFS KRIOS |
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Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 3.0 µm / Nominal defocus min: 1.0 µm |
Image recording | Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Average electron dose: 1.25 e/Å2 |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
-Image processing
Startup model | Type of model: EMDB MAP EMDB ID: |
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Initial angle assignment | Type: MAXIMUM LIKELIHOOD |
Final angle assignment | Type: MAXIMUM LIKELIHOOD |
Final reconstruction | Resolution.type: BY AUTHOR / Resolution: 6.9 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 333214 |