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- EMDB-2594: Deep classification of a large cryo-EM dataset defines the confor... -
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Basic information
Entry | Database: EMDB / ID: EMD-2594 | |||||||||
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Title | Deep classification of a large cryo-EM dataset defines the conformational landscape of the 26S proteasome | |||||||||
![]() | Reconstruction of 26S proteasome in low-energy state (s1) | |||||||||
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![]() | Proteasome / AAA-ATPase / ATP-analog / classification | |||||||||
Function / homology | ![]() SAGA complex localization to transcription regulatory region / Metalloprotease DUBs / peroxisome fission / proteasome storage granule assembly / transcription export complex 2 / proteasome regulatory particle assembly / protein deneddylation / maintenance of DNA trinucleotide repeats / filamentous growth / COP9 signalosome ...SAGA complex localization to transcription regulatory region / Metalloprotease DUBs / peroxisome fission / proteasome storage granule assembly / transcription export complex 2 / proteasome regulatory particle assembly / protein deneddylation / maintenance of DNA trinucleotide repeats / filamentous growth / COP9 signalosome / proteasome regulatory particle / cytosolic proteasome complex / proteasome regulatory particle, lid subcomplex / protein-containing complex localization / proteasome-activating activity / mitochondrial fission / proteasome regulatory particle, base subcomplex / metal-dependent deubiquitinase activity / K48-linked polyubiquitin modification-dependent protein binding / nonfunctional rRNA decay / proteasome core complex assembly / nuclear outer membrane-endoplasmic reticulum membrane network / protein deubiquitination / Cross-presentation of soluble exogenous antigens (endosomes) / TNFR2 non-canonical NF-kB pathway / proteasomal ubiquitin-independent protein catabolic process / Ubiquitin Mediated Degradation of Phosphorylated Cdc25A / Regulation of PTEN stability and activity / KEAP1-NFE2L2 pathway / proteasome binding / CDK-mediated phosphorylation and removal of Cdc6 / Neddylation / FBXL7 down-regulates AURKA during mitotic entry and in early mitosis / peptide catabolic process / regulation of protein catabolic process / Orc1 removal from chromatin / MAPK6/MAPK4 signaling / proteasome storage granule / Antigen processing: Ubiquitination & Proteasome degradation / endopeptidase activator activity / proteasome assembly / polyubiquitin modification-dependent protein binding / proteasome endopeptidase complex / positive regulation of RNA polymerase II transcription preinitiation complex assembly / proteasome core complex, beta-subunit complex / proteasome core complex, alpha-subunit complex / Ub-specific processing proteases / threonine-type endopeptidase activity / mRNA export from nucleus / enzyme regulator activity / ERAD pathway / protein folding chaperone / Neutrophil degranulation / proteasome complex / ubiquitin binding / nucleotide-excision repair / positive regulation of transcription elongation by RNA polymerase II / double-strand break repair via homologous recombination / positive regulation of protein catabolic process / metallopeptidase activity / peroxisome / protein-macromolecule adaptor activity / ubiquitin-dependent protein catabolic process / proteasome-mediated ubiquitin-dependent protein catabolic process / endopeptidase activity / ubiquitinyl hydrolase 1 / cysteine-type deubiquitinase activity / molecular adaptor activity / regulation of cell cycle / chromatin remodeling / protein domain specific binding / mRNA binding / ubiquitin protein ligase binding / endoplasmic reticulum membrane / structural molecule activity / endoplasmic reticulum / positive regulation of transcription by RNA polymerase II / ATP hydrolysis activity / mitochondrion / ATP binding / identical protein binding / nucleus / metal ion binding / cytosol / cytoplasm Similarity search - Function | |||||||||
Biological species | ![]() ![]() | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 7.7 Å | |||||||||
![]() | Unverdorben P / Beck F / Sledz P / Schweitzer A / Pfeifer G / Plitzko JM / Baumeister W / Foerster F | |||||||||
![]() | ![]() Title: Deep classification of a large cryo-EM dataset defines the conformational landscape of the 26S proteasome. Authors: Pia Unverdorben / Florian Beck / Paweł Śledź / Andreas Schweitzer / Günter Pfeifer / Jürgen M Plitzko / Wolfgang Baumeister / Friedrich Förster / ![]() Abstract: The 26S proteasome is a 2.5 MDa molecular machine that executes the degradation of substrates of the ubiquitin-proteasome pathway. The molecular architecture of the 26S proteasome was recently ...The 26S proteasome is a 2.5 MDa molecular machine that executes the degradation of substrates of the ubiquitin-proteasome pathway. The molecular architecture of the 26S proteasome was recently established by cryo-EM approaches. For a detailed understanding of the sequence of events from the initial binding of polyubiquitylated substrates to the translocation into the proteolytic core complex, it is necessary to move beyond static structures and characterize the conformational landscape of the 26S proteasome. To this end we have subjected a large cryo-EM dataset acquired in the presence of ATP and ATP-γS to a deep classification procedure, which deconvolutes coexisting conformational states. Highly variable regions, such as the density assigned to the largest subunit, Rpn1, are now well resolved and rendered interpretable. Our analysis reveals the existence of three major conformations: in addition to the previously described ATP-hydrolyzing (ATPh) and ATP-γS conformations, an intermediate state has been found. Its AAA-ATPase module adopts essentially the same topology that is observed in the ATPh conformation, whereas the lid is more similar to the ATP-γS bound state. Based on the conformational ensemble of the 26S proteasome in solution, we propose a mechanistic model for substrate recognition, commitment, deubiquitylation, and translocation into the core particle. | |||||||||
History |
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Structure visualization
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Structure viewer | EM map: ![]() ![]() ![]() |
Supplemental images |
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Downloads & links
-EMDB archive
Map data | ![]() | 78.5 MB | ![]() | |
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Header (meta data) | ![]() ![]() | 8.8 KB 8.8 KB | Display Display | ![]() |
Images | ![]() | 182.3 KB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 4cr2MC ![]() 2595C ![]() 2596C ![]() 4cr3C ![]() 4cr4C M: atomic model generated by this map C: citing same article ( |
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Similar structure data |
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Links
EMDB pages | ![]() ![]() |
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Related items in Molecule of the Month |
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Map
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Annotation | Reconstruction of 26S proteasome in low-energy state (s1) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.99 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
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Sample components
-Entire : 26S Proteasome from Saccharomyces cerevisiae
Entire | Name: 26S Proteasome from Saccharomyces cerevisiae |
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Components |
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-Supramolecule #1000: 26S Proteasome from Saccharomyces cerevisiae
Supramolecule | Name: 26S Proteasome from Saccharomyces cerevisiae / type: sample / ID: 1000 / Number unique components: 1 |
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Molecular weight | Experimental: 2.5 MDa / Theoretical: 2.5 MDa |
-Macromolecule #1: 26S Proteasome
Macromolecule | Name: 26S Proteasome / type: protein_or_peptide / ID: 1 / Recombinant expression: No |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Experimental: 2.5 MDa / Theoretical: 2.5 MDa |
-Experimental details
-Structure determination
Method | cryo EM |
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![]() | single particle reconstruction |
Aggregation state | particle |
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Sample preparation
Concentration | 0.3 mg/mL |
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Vitrification | Cryogen name: ETHANE / Instrument: OTHER |
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Electron microscopy
Microscope | FEI TITAN KRIOS |
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Alignment procedure | Legacy - Electron beam tilt params: 0 |
Date | Dec 24, 2013 |
Image recording | Category: CCD / Film or detector model: TVIPS TEMCAM-F816 (8k x 8k) / Number real images: 30000 / Average electron dose: 25 e/Å2 / Bits/pixel: 14 |
Tilt angle min | 0 |
Tilt angle max | 0 |
Electron beam | Acceleration voltage: 200 kV / Electron source: ![]() |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2 mm / Nominal defocus max: 3.5 µm / Nominal defocus min: 1.0 µm |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |
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Image processing
Details | The particles were selected using an automatic selection program. Each physical 26S particles was considered as two particles for processing according to pseudo-C2 symmetry. |
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CTF correction | Details: micrograph |
Final reconstruction | Applied symmetry - Point group: C1 (asymmetric) / Algorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 7.7 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: xmipp / Number images used: 1300000 |