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Yorodumi- EMDB-2447: Electron microscopy of the complex formed by chaperones TBCE and ... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-2447 | |||||||||
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Title | Electron microscopy of the complex formed by chaperones TBCE and TBCB and alpha-tubulin | |||||||||
Map data | Reconstruction of the complex formed by TBCE, TBCB and alpha-tubulin | |||||||||
Sample |
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Keywords | chaperones / protein folding / protein degradation / tubulin proteostasis | |||||||||
Function / homology | Function and homology information peripheral nervous system neuron axonogenesis / post-chaperonin tubulin folding pathway / muscle atrophy / Post-chaperonin tubulin folding pathway / tubulin complex assembly / developmental growth / alpha-tubulin binding / mitotic spindle organization / adult locomotory behavior / post-embryonic development ...peripheral nervous system neuron axonogenesis / post-chaperonin tubulin folding pathway / muscle atrophy / Post-chaperonin tubulin folding pathway / tubulin complex assembly / developmental growth / alpha-tubulin binding / mitotic spindle organization / adult locomotory behavior / post-embryonic development / Hydrolases; Acting on acid anhydrides; Acting on GTP to facilitate cellular and subcellular movement / structural constituent of cytoskeleton / microtubule cytoskeleton organization / microtubule cytoskeleton / protein folding / mitotic cell cycle / nervous system development / protein-folding chaperone binding / microtubule / cell differentiation / GTPase activity / GTP binding / metal ion binding / cytoplasm / cytosol Similarity search - Function | |||||||||
Biological species | Homo sapiens (human) / Bos taurus (cattle) | |||||||||
Method | single particle reconstruction / negative staining / Resolution: 21.0 Å | |||||||||
Authors | Serna M / Carranza G / Martin-Benito J / Janowsk R / Canals A / Coll M / Zabala JC / Valpuesta JM | |||||||||
Citation | Journal: J Cell Sci / Year: 2015 Title: The structure of the complex between α-tubulin, TBCE and TBCB reveals a tubulin dimer dissociation mechanism. Authors: Marina Serna / Gerardo Carranza / Jaime Martín-Benito / Robert Janowski / Albert Canals / Miquel Coll / Juan Carlos Zabala / José María Valpuesta / Abstract: Tubulin proteostasis is regulated by a group of molecular chaperones termed tubulin cofactors (TBC). Whereas tubulin heterodimer formation is well-characterized biochemically, its dissociation ...Tubulin proteostasis is regulated by a group of molecular chaperones termed tubulin cofactors (TBC). Whereas tubulin heterodimer formation is well-characterized biochemically, its dissociation pathway is not clearly understood. Here, we carried out biochemical assays to dissect the role of the human TBCE and TBCB chaperones in α-tubulin-β-tubulin dissociation. We used electron microscopy and image processing to determine the three-dimensional structure of the human TBCE, TBCB and α-tubulin (αEB) complex, which is formed upon α-tubulin-β-tubulin heterodimer dissociation by the two chaperones. Docking the atomic structures of domains of these proteins, including the TBCE UBL domain, as we determined by X-ray crystallography, allowed description of the molecular architecture of the αEB complex. We found that heterodimer dissociation is an energy-independent process that takes place through a disruption of the α-tubulin-β-tubulin interface that is caused by a steric interaction between β-tubulin and the TBCE cytoskeleton-associated protein glycine-rich (CAP-Gly) and leucine-rich repeat (LRR) domains. The protruding arrangement of chaperone ubiquitin-like (UBL) domains in the αEB complex suggests that there is a direct interaction of this complex with the proteasome, thus mediating α-tubulin degradation. | |||||||||
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_2447.map.gz | 119.2 KB | EMDB map data format | |
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Header (meta data) | emd-2447-v30.xml emd-2447.xml | 14.9 KB 14.9 KB | Display Display | EMDB header |
Images | EMD-2447.png | 91.2 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-2447 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-2447 | HTTPS FTP |
-Validation report
Summary document | emd_2447_validation.pdf.gz | 213.6 KB | Display | EMDB validaton report |
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Full document | emd_2447_full_validation.pdf.gz | 212.7 KB | Display | |
Data in XML | emd_2447_validation.xml.gz | 4.5 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-2447 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-2447 | HTTPS FTP |
-Related structure data
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_2447.map.gz / Format: CCP4 / Size: 126 KB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | Reconstruction of the complex formed by TBCE, TBCB and alpha-tubulin | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 4.66 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 : Ternary complex of alpha-tubulin, TBCE and TBCB
Entire | Name: Ternary complex of alpha-tubulin, TBCE and TBCB |
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Components |
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-Supramolecule #1000: Ternary complex of alpha-tubulin, TBCE and TBCB
Supramolecule | Name: Ternary complex of alpha-tubulin, TBCE and TBCB / type: sample / ID: 1000 / Oligomeric state: heterotrimer / Number unique components: 3 |
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Molecular weight | Theoretical: 137 KDa |
-Macromolecule #1: Tubulin binding cofactor E
Macromolecule | Name: Tubulin binding cofactor E / type: protein_or_peptide / ID: 1 / Name.synonym: Tubulin-specific chaperone E / Number of copies: 1 / Oligomeric state: Monomer / Recombinant expression: Yes |
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Source (natural) | Organism: Homo sapiens (human) / synonym: Human / Location in cell: Cytoplasm |
Molecular weight | Theoretical: 59 KDa |
Recombinant expression | Organism: Spodoptera frugiperda (fall armyworm) / Recombinant cell: Sf9 / Recombinant plasmid: pFastBac-1 |
Sequence | UniProtKB: Tubulin-specific chaperone E / InterPro: CAP Gly-rich domain |
-Macromolecule #2: Tubulin binding cofactor B
Macromolecule | Name: Tubulin binding cofactor B / type: protein_or_peptide / ID: 2 Name.synonym: Tubulin-specific chaperone B, Tubulin-folding cofactor B Number of copies: 1 / Oligomeric state: Monomer / Recombinant expression: Yes |
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Source (natural) | Organism: Homo sapiens (human) / synonym: Human / Location in cell: Cytoplasm |
Molecular weight | Theoretical: 27 KDa |
Recombinant expression | Organism: Escherichia coli (E. coli) / Recombinant strain: BL21 (DE3) pLysS / Recombinant plasmid: pET3a |
Sequence | UniProtKB: Tubulin-folding cofactor B / InterPro: CAP Gly-rich domain |
-Macromolecule #3: alpha-tubulin
Macromolecule | Name: alpha-tubulin / type: protein_or_peptide / ID: 3 / Name.synonym: Tubulin alpha-1B chain / Number of copies: 1 / Oligomeric state: Monomer / Recombinant expression: No |
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Source (natural) | Organism: Bos taurus (cattle) / synonym: Bovin / Tissue: Brain / Location in cell: Cytoplasm |
Molecular weight | Theoretical: 50 KDa |
Sequence | UniProtKB: Tubulin alpha-1B chain / InterPro: Alpha tubulin |
-Experimental details
-Structure determination
Method | negative staining |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 0.02 mg/mL |
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Buffer | pH: 6.7 / Details: 100mM MES-NaOH, 25mM KCl, 1mM MgCl2, 1mM EGTA |
Staining | Type: NEGATIVE Details: Grids with adsorbed protein floated on 2% w/v uranyl acetate for 1 min. |
Grid | Details: 300 mesh copper grid with thin carbon support, glow discharged in air atmosphere |
Vitrification | Cryogen name: NONE / Instrument: OTHER |
-Electron microscopy
Microscope | JEOL 1200EXII |
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Alignment procedure | Legacy - Astigmatism: Objective lens astigmatism was corrected at 100,000 times magnification |
Date | Nov 1, 2010 |
Image recording | Category: FILM / Film or detector model: KODAK SO-163 FILM / Digitization - Scanner: ZEISS SCAI / Digitization - Sampling interval: 14 µm / Number real images: 85 / Average electron dose: 10 e/Å2 |
Electron beam | Acceleration voltage: 100 kV / Electron source: TUNGSTEN HAIRPIN |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 5.6 mm / Nominal defocus max: 1.5 µm / Nominal defocus min: 1.0 µm / Nominal magnification: 60000 |
Sample stage | Specimen holder model: JEOL |
-Image processing
Details | The particles were manually selected using the XMIPP program |
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CTF correction | Details: N. Grigorieff CTFFIND3 |
Final reconstruction | Applied symmetry - Point group: C1 (asymmetric) / Algorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 21.0 Å / Resolution method: FSC 0.33 CUT-OFF / Software - Name: EMAN, Spider, XMIPP Details: Individual particles were manually selected using XMIPP software package. Number images used: 26129 |
-Atomic model buiding 1
Initial model | PDB ID: Chain - Chain ID: A |
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Software | Name: Chimera |
Details | The domain was fitted by manual docking using Chimera |
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: Chimera |
Details | The domain was fitted by manual docking using Chimera |
Refinement | Space: REAL / Protocol: RIGID BODY FIT |
-Atomic model buiding 3
Initial model | PDB ID: Chain - Chain ID: A |
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Software | Name: Chimera |
Details | The domain was fitted by manual docking using Chimera |
Refinement | Space: REAL / Protocol: RIGID BODY FIT |
-Atomic model buiding 4
Initial model | PDB ID: Chain - Chain ID: A |
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Software | Name: Chimera |
Details | The domain was fitted by manual docking using Chimera |
Refinement | Space: REAL / Protocol: RIGID BODY FIT |
-Atomic model buiding 5
Initial model | PDB ID: Chain - Chain ID: A |
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Software | Name: Chimera |
Details | The domain was fitted by manual docking using Chimera |
Refinement | Space: REAL / Protocol: RIGID BODY FIT |