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- EMDB-2499: Substrate Recruitment Pathways in the Yeast Exosome by Electron M... -
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Open data
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Basic information
Entry | Database: EMDB / ID: EMD-2499 | |||||||||
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Title | Substrate Recruitment Pathways in the Yeast Exosome by Electron Microscopy | |||||||||
![]() | Exosome incubated with Streptavidin labeled ssRNA( 47 nucleotides in length) | |||||||||
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![]() | exosome / RNA degradation route | |||||||||
Biological species | ![]() ![]() ![]() ![]() | |||||||||
Method | single particle reconstruction / negative staining / Resolution: 26.0 Å | |||||||||
![]() | Liu J-J / Bratkowski MA / Liu XQ / Niu C-Y / Ke AL / Wang H-W | |||||||||
![]() | ![]() Title: Visualization of distinct substrate-recruitment pathways in the yeast exosome by EM. Authors: Jun-Jie Liu / Matthew A Bratkowski / Xueqi Liu / Chu-Ya Niu / Ailong Ke / Hong-Wei Wang / ![]() ![]() Abstract: The eukaryotic exosome is a multisubunit complex typically composed of a catalytically inactive core and the Rrp44 protein, which contains 3'-to-5' exo- and endo-RNase activities. RNA substrates have ...The eukaryotic exosome is a multisubunit complex typically composed of a catalytically inactive core and the Rrp44 protein, which contains 3'-to-5' exo- and endo-RNase activities. RNA substrates have been shown to be recruited through the core to reach Rrp44's exo-RNase (EXO) site. Using single-particle EM and biochemical analysis, we provide visual evidence that two distinct substrate-recruitment pathways exist. In the through-core route, channeling of the single-stranded substrates from the core to Rrp44 induces a characteristic conformational change in Rrp44. In the alternative direct-access route, this conformational change does not take place, and the RNA substrate is visualized to avoid the core and enter Rrp44's EXO site directly. Our results provide mechanistic explanations for several RNA processing scenarios by the eukaryotic exosome and indicate substrate-specific modes of degradation by this complex. | |||||||||
History |
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Structure visualization
Movie |
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Structure viewer | EM map: ![]() ![]() ![]() |
Supplemental images |
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Downloads & links
-EMDB archive
Map data | ![]() | 2.6 MB | ![]() | |
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Header (meta data) | ![]() ![]() | 16.6 KB 16.6 KB | Display Display | ![]() |
Images | ![]() | 55 KB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Validation report
Summary document | ![]() | 211.6 KB | Display | ![]() |
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Full document | ![]() | 210.8 KB | Display | |
Data in XML | ![]() | 4.7 KB | Display | |
Arichive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 2491C ![]() 2492C ![]() 2493C ![]() 2494C ![]() 2495C ![]() 2496C ![]() 2497C ![]() 2498C ![]() 2500C ![]() 2501C ![]() 2502C ![]() 2522C C: citing same article ( |
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Similar structure data |
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Links
EMDB pages | ![]() ![]() |
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Map
File | ![]() | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | Exosome incubated with Streptavidin labeled ssRNA( 47 nucleotides in length) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 3 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 : Rrp44-Exosome with Streptavidin labeled RNA47
+Supramolecule #1000: Rrp44-Exosome with Streptavidin labeled RNA47
+Macromolecule #1: Rrp44
+Macromolecule #2: Rrp43
+Macromolecule #3: Rrp4
+Macromolecule #4: Csl4
+Macromolecule #5: Rrp45
+Macromolecule #6: Rrp46-TAP
+Macromolecule #7: Rrp41
+Macromolecule #8: Rrp42
+Macromolecule #9: Mtr3
+Macromolecule #10: Rrp40
+Macromolecule #12: Steptavidin
+Macromolecule #11: RNA47
-Experimental details
-Structure determination
Method | negative staining |
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![]() | single particle reconstruction |
Aggregation state | particle |
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Sample preparation
Concentration | 1 mg/mL |
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Buffer | pH: 8 / Details: 150mM NaCl, 50mM Tris-HCL,1mM DTT |
Staining | Type: NEGATIVE Details: All samples were diluted at a final concentration of ~80 nM of the exosome in the non-digestive buffer and negatively stained in 2% (w/v) uranyl acetate solution following the standard deep ...Details: All samples were diluted at a final concentration of ~80 nM of the exosome in the non-digestive buffer and negatively stained in 2% (w/v) uranyl acetate solution following the standard deep stain procedure on holey-carbon coated EM copper grids covered with a thin layer of continuous carbon |
Grid | Details: 300 mesh continus carbon |
Vitrification | Cryogen name: NONE / Instrument: OTHER |
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Electron microscopy
Microscope | FEI TECNAI F20 |
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Specialist optics | Energy filter - Name: FEI |
Date | Oct 10, 2012 |
Image recording | Category: CCD / Film or detector model: GENERIC GATAN (4k x 4k) / Number real images: 300 / Average electron dose: 30 e/Å2 |
Electron beam | Acceleration voltage: 200 kV / Electron source: ![]() |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD |
Sample stage | Specimen holder model: OTHER |
Experimental equipment | ![]() Model: Tecnai F20 / Image courtesy: FEI Company |
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Image processing
Final reconstruction | Applied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 26.0 Å / Resolution method: FSC 0.5 CUT-OFF / Software - Name: SPIDER / Number images used: 25004 |
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Final angle assignment | Details: SPIDER: theta 90 degrees, phi 359.9 degrees |
-Atomic model buiding 1
Initial model | PDB ID: |
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Software | Name: ![]() |
Refinement | Space: REAL / Protocol: RIGID BODY FIT |