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Yorodumi- EMDB-5252: electron cryo-tomography reconstruction and subvolume averaging o... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-5252 | |||||||||
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Title | electron cryo-tomography reconstruction and subvolume averaging of the basal body triplet from Chlamydomonas reinhardtii | |||||||||
Map data | averaged microtubule triplet from basal body from Chlamydomonas reinhardtii | |||||||||
Sample |
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Keywords | basal body / centriole / electron cryo-tomography / three-dimensional reconstruction | |||||||||
Biological species | Chlamydomonas reinhardtii (plant) | |||||||||
Method | subtomogram averaging / cryo EM / Resolution: 33.0 Å | |||||||||
Authors | Li S / Fernandez JJ / Marshall WF / Agard DA | |||||||||
Citation | Journal: EMBO J / Year: 2012 Title: Three-dimensional structure of basal body triplet revealed by electron cryo-tomography. Authors: Sam Li / Jose-Jesus Fernandez / Wallace F Marshall / David A Agard / Abstract: Basal bodies and centrioles play central roles in microtubule (MT)-organizing centres within many eukaryotes. They share a barrel-shaped cylindrical structure composed of nine MT triplet blades. ...Basal bodies and centrioles play central roles in microtubule (MT)-organizing centres within many eukaryotes. They share a barrel-shaped cylindrical structure composed of nine MT triplet blades. Here, we report the structure of the basal body triplet at 33 Å resolution obtained by electron cryo-tomography and 3D subtomogram averaging. By fitting the atomic structure of tubulin into the EM density, we built a pseudo-atomic model of the tubulin protofilaments at the core of the triplet. The 3D density map reveals additional densities that represent non-tubulin proteins attached to the triplet, including a large inner circular structure in the basal body lumen, which functions as a scaffold to stabilize the entire basal body barrel. We found clear longitudinal structural variations along the basal body, suggesting a sequential and coordinated assembly mechanism. We propose a model in which δ-tubulin and other components participate in the assembly of the basal body. | |||||||||
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_5252.map.gz | 4.4 MB | EMDB map data format | |
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Header (meta data) | emd-5252-v30.xml emd-5252.xml | 10.7 KB 10.7 KB | Display Display | EMDB header |
Images | emd_5252_1.tif | 235.2 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-5252 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-5252 | HTTPS FTP |
-Validation report
Summary document | emd_5252_validation.pdf.gz | 78.1 KB | Display | EMDB validaton report |
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Full document | emd_5252_full_validation.pdf.gz | 77.2 KB | Display | |
Data in XML | emd_5252_validation.xml.gz | 493 B | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-5252 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-5252 | HTTPS FTP |
-Related structure data
Similar structure data |
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-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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-Map
File | Download / File: emd_5252.map.gz / Format: CCP4 / Size: 6 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | averaged microtubule triplet from basal body from Chlamydomonas reinhardtii | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Projections & slices | Image control
Images are generated by Spider. generated in cubic-lattice coordinate | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 6.5 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 : basal body microtubule triplet
Entire | Name: basal body microtubule triplet |
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Components |
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-Supramolecule #1000: basal body microtubule triplet
Supramolecule | Name: basal body microtubule triplet / type: sample / ID: 1000 / Number unique components: 1 |
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Molecular weight | Theoretical: 15.1 MDa Method: estimation based on the volume assuming the protein density is 1.41g per cm3 |
-Supramolecule #1: basal body
Supramolecule | Name: basal body / type: organelle_or_cellular_component / ID: 1 / Name.synonym: basal body / Number of copies: 2 / Oligomeric state: singlet / Recombinant expression: No / Database: NCBI |
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Source (natural) | Organism: Chlamydomonas reinhardtii (plant) / Strain: cc3491 / synonym: green algea / Cell: Chlamydomonas reinhardtii / Organelle: basal body / Location in cell: flagellum apparatus |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | subtomogram averaging |
-Sample preparation
Buffer | pH: 8 / Details: 10mM Tris 1mM EDTA |
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Grid | Details: 300 mesh copper grid |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 90 % / Chamber temperature: 93 K / Instrument: OTHER / Details: Vitrification instrument: Vitrobot / Method: Blot for 3 seconds before plunging |
-Electron microscopy
Microscope | FEI POLARA 300 |
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Temperature | Min: 91 K / Max: 93 K / Average: 93 K |
Specialist optics | Energy filter - Name: Gatan GIF2000 / Energy filter - Lower energy threshold: 0.0 eV / Energy filter - Upper energy threshold: 25.0 eV |
Details | defocus maximum value is 24000nm |
Image recording | Category: CCD / Film or detector model: GATAN ULTRASCAN 1000 (2k x 2k) / Average electron dose: 80 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Calibrated magnification: 46153 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.0 mm / Nominal defocus min: 9.0 µm / Nominal magnification: 34000 |
Sample stage | Specimen holder: Eucentric / Specimen holder model: OTHER / Tilt series - Axis1 - Min angle: -60 ° / Tilt series - Axis1 - Max angle: 60 ° |
Experimental equipment | Model: Tecnai Polara / Image courtesy: FEI Company |
-Image processing
Details | The tomographic tilt series were taken by UCSF Tomography program. Average number of tilts used in the 3D reconstructions: 82. Average tomographic tilt angle increment: 2. |
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Final reconstruction | Algorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 33.0 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: Priism / Details: Final map was average of 1644 subvolumes |
CTF correction | Details: each image, Wiener filter |
-Atomic model buiding 1
Initial model | PDB ID: |
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Software | Name: Chimera |
Details | Protocol: rigid body. The atomic structure was first manually fitting into EM density by program O and the fitting was refined by Chimera |
Refinement | Space: REAL / Protocol: RIGID BODY FIT / Target criteria: cross correlation coefficiet |