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Open data
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Basic information
Entry | Database: EMDB / ID: EMD-2563 | |||||||||
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Title | Cryo electron microscopy of E. coli ClpB | |||||||||
![]() | Cryo-EM reconstruction of E.coli ClpB. Six fold symmetry applied. | |||||||||
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![]() | chaperone / disaggregase / ClpB / coiled-coil domain | |||||||||
Biological species | ![]() ![]() | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 29.0 Å | |||||||||
![]() | Carroni M / Kummer E / Oguchi Y / Clare DK / Wendler P / Sinning I / Kopp J / Mogk A / Bukau B / Saibil HR | |||||||||
![]() | ![]() Title: Head-to-tail interactions of the coiled-coil domains regulate ClpB activity and cooperation with Hsp70 in protein disaggregation. Authors: Marta Carroni / Eva Kummer / Yuki Oguchi / Petra Wendler / Daniel K Clare / Irmgard Sinning / Jürgen Kopp / Axel Mogk / Bernd Bukau / Helen R Saibil / ![]() ![]() Abstract: The hexameric AAA+ chaperone ClpB reactivates aggregated proteins in cooperation with the Hsp70 system. Essential for disaggregation, the ClpB middle domain (MD) is a coiled-coil propeller that binds ...The hexameric AAA+ chaperone ClpB reactivates aggregated proteins in cooperation with the Hsp70 system. Essential for disaggregation, the ClpB middle domain (MD) is a coiled-coil propeller that binds Hsp70. Although the ClpB subunit structure is known, positioning of the MD in the hexamer and its mechanism of action are unclear. We obtained electron microscopy (EM) structures of the BAP variant of ClpB that binds the protease ClpP, clearly revealing MD density on the surface of the ClpB ring. Mutant analysis and asymmetric reconstructions show that MDs adopt diverse positions in a single ClpB hexamer. Adjacent, horizontally oriented MDs form head-to-tail contacts and repress ClpB activity by preventing Hsp70 interaction. Tilting of the MD breaks this contact, allowing Hsp70 binding, and releasing the contact in adjacent subunits. Our data suggest a wavelike activation of ClpB subunits around the ring.DOI: http://dx.doi.org/10.7554/eLife.02481.001. | |||||||||
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 | ![]() | 7.2 MB | ![]() | |
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Header (meta data) | ![]() ![]() | 11.1 KB 11.1 KB | Display Display | ![]() |
FSC (resolution estimation) | ![]() | 5.4 KB | Display | ![]() |
Images | ![]() | 328.2 KB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Validation report
Summary document | ![]() | 227.1 KB | Display | ![]() |
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Full document | ![]() | 226.2 KB | Display | |
Data in XML | ![]() | 7.6 KB | Display | |
Arichive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 2555C ![]() 2556C ![]() 2557C ![]() 2558C ![]() 2559C ![]() 2560C ![]() 2561C ![]() 2562C ![]() 4ciuC ![]() 4d2qC ![]() 4d2uC ![]() 4d2xC 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 | Cryo-EM reconstruction of E.coli ClpB. Six fold symmetry applied. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 2 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 : ClpB with ATPgammaS.
Entire | Name: ClpB with ATPgammaS. |
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Components |
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-Supramolecule #1000: ClpB with ATPgammaS.
Supramolecule | Name: ClpB with ATPgammaS. / type: sample / ID: 1000 / Oligomeric state: Homohexamer. / Number unique components: 1 |
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Molecular weight | Theoretical: 500 KDa |
-Macromolecule #1: ClpB
Macromolecule | Name: ClpB / type: protein_or_peptide / ID: 1 / Number of copies: 6 / Oligomeric state: Hexamer / Recombinant expression: Yes |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Experimental: 80 KDa / Theoretical: 80 KDa |
Recombinant expression | Organism: ![]() ![]() |
-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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Buffer | pH: 7.5 Details: 20 mM Tris-HCl, pH 7.5, 20 mM KCl, 15 mM MgCl2, 1 mM DTT, 2 mM ATPgammaS |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 77 K / Instrument: FEI VITROBOT MARK II |
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Electron microscopy
Microscope | FEI TECNAI F20 |
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Temperature | Min: 77 K / Max: 88 K / Average: 83 K |
Alignment procedure | Legacy - Astigmatism: Objective lens astigmatism was corrected at 150,000 x magnification |
Date | Feb 4, 2011 |
Image recording | Category: CCD / Film or detector model: GATAN ULTRASCAN 4000 (4k x 4k) / Number real images: 200 / Average electron dose: 15 e/Å2 |
Electron beam | Acceleration voltage: 200 kV / Electron source: ![]() |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2 mm / Nominal defocus max: 4.0 µm / Nominal defocus min: 1.5 µm / Nominal magnification: 80000 |
Sample stage | Specimen holder model: SIDE ENTRY, EUCENTRIC |
Experimental equipment | ![]() Model: Tecnai F20 / Image courtesy: FEI Company |
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Image processing
-Atomic model buiding 1
Initial model | PDB ID: Chain - Chain ID: A |
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Software | Name: ![]() |
Details | Fitting of separate domains was performed manually and locally optimised using Chimera. Known domain interfaces were used to guide the fit. |
Refinement | Space: REAL / Protocol: RIGID BODY FIT |
-Atomic model buiding 2
Initial model | PDB ID: Chain - Chain ID: A |
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Software | Name: ![]() |
Details | Fitting of separate domains was performed manually and locally optimised using Chimera. Known domain interfaces were used to guide the fit. |
Refinement | Space: REAL / Protocol: RIGID BODY FIT |