|Entry||Database: EMDB / ID: 2095|
|Title||Cryo-electron microscopy reconstruction of doublecortin-stabilised microtubules in absence of kinesin|
|Keywords||doublecortin / microtubule / Microtubule-Associated Protein|
|Source||Bos taurus / mammal / cattle / ウシ / |
Homo sapiens / human
|Map data||Reconstruction of doublecortin-stabilised microtubules in absence of kinesin|
|Method||helical reconstruction, at 8.3 Å resolution|
|Authors||Liu JS / Schubert CR / Fu X / Fourniol FJ / Jaiswal JK / Houdusse A / Stultz CM / Moores CA / Walsh CA|
|Citation||Mol. Cell, 2012, 47, 707-721|
Mol. Cell, 2012, 47, 707-721 Yorodumi Papers
|Validation Report||PDB-ID: 4atu|
SummaryFull reportAbout validation report
|Date||Deposition: May 9, 2012 / Header (metadata) release: May 17, 2012 / Map release: Aug 15, 2012 / Last update: Sep 26, 2012|
Downloads & links
|File||emd_2095.map.gz (map file in CCP4 format, 1376 KB)|
|Projections & slices|
Images are generated by Spider package.
|Voxel size||X=Y=Z: 2.8 Å|
CCP4 map header:
-Entire Doublecortin-stabilised microtubules
|Entire||Name: Doublecortin-stabilised microtubules / Number of components: 3 / Oligomeric State: 13-protofilament microtubule|
-Component #1: protein, tubulin alpha-1D chain
|Protein||Name: tubulin alpha-1D chain / Oligomeric Details: heterodimer / Recombinant expression: No|
|Mass||Theoretical: 50 kDa|
|Source||Species: Bos taurus / mammal / cattle / ウシ /|
|Source (natural)||Location in cell: cytoplasm / Organ or tissue: brain|
|External references||InterPro: InterPro: 002452|
-Component #2: protein, doublecortin
|Protein||Name: doublecortin / a.k.a: DCX / Recombinant expression: Yes|
|Mass||Theoretical: 40 kDa|
|Source||Species: Homo sapiens / human|
|Source (engineered)||Expression System: Spodoptera frugiperda / arthropod / Vector: pFastBac|
|External references||InterPro: InterPro: 003533|
-Component #3: protein, tubulin beta-2B chain
|Helical parameters||Hand: LEFT HANDED / Delta z: 9.2 Å / Delta phi: 27.7 deg.|
|Sample solution||Buffer solution: 20mM PIPES, 1mM EGTA, 3mM MgCl2, 1mM TCEP, 0.5mM GTP|
|Support film||300 mesh lacey carbon grids|
|Vitrification||Instrument: FEI VITROBOT MARK I / Cryogen name: ETHANE / Humidity: 100 %|
-Electron microscopy imaging
Model: Tecnai F20 / Image courtesy: FEI Company
|Imaging||Microscope: FEI TECNAI F20 / Date: Oct 1, 2010|
|Electron gun||Electron source: FIELD EMISSION GUN / Accelerating voltage: 200 kV / Electron dose: 17 e/Å2 / Illumination mode: FLOOD BEAM|
|Lens||Magnification: 50000 X (nominal)|
Astigmatism: Objective lens astigmatism was corrected at 150,000 times magnification
Cs: 2 mm / Imaging mode: BRIGHT FIELD / Defocus: 900 - 3600 nm
|Specimen Holder||Model: SIDE ENTRY, EUCENTRIC / Temperature: 93 K|
|Camera||Detector: KODAK SO-163 FILM|
|Image acquisition||Scanner: ZEISS SCAI / Bit depth: 8|
|Processing||Method: helical reconstruction|
Details: Particles were aligned using Spider and Frealign (Sindelar and Downing, 2010)
|3D reconstruction||Algorithm: Single particle / Software: SPIDER, FREALIGN / CTF correction: done with FREALIGN|
Details: A sub-selection of approximately 146,000 tubulin dimers (75% of total dataset) went into the final reconstruction
Resolution: 8.3 Å / Resolution method: FSC 0.5
-Atomic model buiding
|Modeling #1||Software: Chimera / Refinement protocol: rigid body / Target criteria: cross correlation / Refinement space: REAL / Details: Protocol: Rigid body|
Input PDB model: 2XRP
Chain ID: A, B, C, D, E, F, G, H
|Modeling #2||Software: Chimera, Flex-EM / Refinement protocol: flexible / Target criteria: cross correlation / Refinement space: REAL|
Details: Protocol: flexible fitting. The EM map was zoned around the doublecortin density. The N-DC atomic model (1MJD model 11, aa 38-150) was extended at its C-terminus (aa 151-156) using Chimera. Flexible fitting was performed into the zoned around EM map. The core DC domain (aa 53-129) and the docked W146 region (aa 145-146) were treated as a single rigid body.
Input PDB model: 1MJD
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External links: The 2017 Nobel Prize in Chemistry - Press Release
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