+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-2656 | |||||||||
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Title | The 15S precursor complex of the yeast 20S proteasome | |||||||||
Map data | 3D reconstruction of the 15S proteasomal precursor complex from S. cerevisiae | |||||||||
Sample |
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Keywords | 15S precursor / Pba1 / Pba2 / proteasome / yeast | |||||||||
Biological species | Saccharomyces cerevisiae (brewer's yeast) | |||||||||
Method | single particle reconstruction / negative staining / Resolution: 19.3 Å | |||||||||
Authors | Kock M / Nunes MM / Hemann M / Kube S / Dohmen JR / Herzog F / Ramos PC / Wendler P | |||||||||
Citation | Journal: Nat Commun / Year: 2015 Title: Proteasome assembly from 15S precursors involves major conformational changes and recycling of the Pba1-Pba2 chaperone. Authors: Malte Kock / Maria M Nunes / Matthias Hemann / Sebastian Kube / R Jürgen Dohmen / Franz Herzog / Paula C Ramos / Petra Wendler / Abstract: The chaperones Ump1 and Pba1-Pba2 promote efficient biogenesis of 20S proteasome core particles from its subunits via 15S intermediates containing alpha and beta subunits, except beta7. Here we ...The chaperones Ump1 and Pba1-Pba2 promote efficient biogenesis of 20S proteasome core particles from its subunits via 15S intermediates containing alpha and beta subunits, except beta7. Here we elucidate the structural role of these chaperones in late steps of core particle biogenesis using biochemical, electron microscopy, cross-linking and mass spectrometry analyses. In 15S precursor complexes, Ump1 is largely unstructured, lining the inner cavity of the complex along the interface between alpha and beta subunits. The alpha and beta subunits form loosely packed rings with a wider alpha ring opening than in the 20S core particle, allowing for the Pba1-Pba2 heterodimer to be partially embedded in the central alpha ring cavity. During biogenesis, the heterodimer is expelled from the alpha ring by a restructuring event that organizes the beta ring and leads to tightening of the alpha ring opening. In this way, the Pba1-Pba2 chaperone is recycled for a new round of proteasome assembly. | |||||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | EM map: SurfViewMolmilJmol/JSmol |
Supplemental images |
-Downloads & links
-EMDB archive
Map data | emd_2656.map.gz | 74.2 KB | EMDB map data format | |
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Header (meta data) | emd-2656-v30.xml emd-2656.xml | 9.3 KB 9.3 KB | Display Display | EMDB header |
Images | emd_2656.tif | 364.2 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-2656 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-2656 | HTTPS FTP |
-Related structure data
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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-Map
File | Download / File: emd_2656.map.gz / Format: CCP4 / Size: 3.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | 3D reconstruction of the 15S proteasomal precursor complex from S. cerevisiae | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 2.9 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Sample components
-Entire : 15S proteasomal precursor complex from S. cerevisiae containing t...
Entire | Name: 15S proteasomal precursor complex from S. cerevisiae containing the dedicated assembly chaperones Pba1, Pba2 and Ump1 |
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Components |
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-Supramolecule #1000: 15S proteasomal precursor complex from S. cerevisiae containing t...
Supramolecule | Name: 15S proteasomal precursor complex from S. cerevisiae containing the dedicated assembly chaperones Pba1, Pba2 and Ump1 type: sample / ID: 1000 / Oligomeric state: Monomer / Number unique components: 1 |
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Molecular weight | Experimental: 440 KDa / Theoretical: 440 KDa / Method: Sedimentation |
-Macromolecule #1: 15S proteasomal precursor complex
Macromolecule | Name: 15S proteasomal precursor complex / type: protein_or_peptide / ID: 1 / Name.synonym: 15S / Number of copies: 1 / Oligomeric state: Monomer / Recombinant expression: No |
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Source (natural) | Organism: Saccharomyces cerevisiae (brewer's yeast) / Strain: MO27 / synonym: Bakers' Yeast |
Molecular weight | Experimental: 440 KDa / Theoretical: 440 KDa |
-Experimental details
-Structure determination
Method | negative staining |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 0.05 mg/mL |
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Buffer | pH: 7.5 Details: 50 mM Tris-HCl pH 7.5, 5 mM MgCl2, 2 mM ATP, 150 mM NaCl, 15 % (v/v) glycerol, 10 mM imidazole |
Staining | Type: NEGATIVE Details: Grids with adsorbed protein floated on 4 drops of 2 % (w/v) uranyl acetate for 10 s each |
Grid | Details: 400 mesh copper grid with carbon support, glow discharged in ambient atmosphere |
Vitrification | Cryogen name: NONE / Instrument: OTHER |
-Electron microscopy
Microscope | FEI TECNAI SPIRIT |
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Electron beam | Acceleration voltage: 120 kV / Electron source: LAB6 |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.0 mm / Nominal defocus max: 1.0 µm / Nominal defocus min: 0.3 µm / Nominal magnification: 103448 |
Sample stage | Specimen holder model: SIDE ENTRY, EUCENTRIC |
Alignment procedure | Legacy - Astigmatism: Objective lens astigmatism was corrected at 96000 times magnification |
Date | Apr 30, 2012 |
Image recording | Category: CCD / Film or detector model: FEI EAGLE (2k x 2k) / Number real images: 218 / Average electron dose: 20 e/Å2 |
Experimental equipment | Model: Tecnai Spirit / Image courtesy: FEI Company |
-Image processing
CTF correction | Details: Micrograph-based phase flipping |
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Final reconstruction | Applied symmetry - Point group: C1 (asymmetric) / Algorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 19.3 Å / Resolution method: OTHER / Software - Name: IMAGIC, Spider / Number images used: 10156 |
Details | Particles were selected manually |