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Yorodumi- EMDB-1152: Electron cryotomography of the E. coli pyruvate and 2-oxoglutarat... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-1152 | |||||||||
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Title | Electron cryotomography of the E. coli pyruvate and 2-oxoglutarate dehydrogenase complexes. | |||||||||
Map data | Volume of an extracted E. coli 2-oxoglutarate multienzyme complex particle from a dual-tilt tomogram. | |||||||||
Sample |
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Function / homology | Dihydrolipoamide succinyltransferase / 2-oxoglutarate dehydrogenase E1 component / Dihydrolipoamide dehydrogenase Function and homology information | |||||||||
Biological species | Escherichia coli (E. coli) | |||||||||
Method | subtomogram averaging / cryo EM / Resolution: 55.0 Å | |||||||||
Authors | Murphy GE / Jensen GJ | |||||||||
Citation | Journal: Structure / Year: 2005 Title: Electron cryotomography of the E. coli pyruvate and 2-oxoglutarate dehydrogenase complexes. Authors: Gavin E Murphy / Grant J Jensen / Abstract: The E. coli pyruvate and 2-oxoglutarate dehydrogenases are two closely related, large complexes that exemplify a growing number of multiprotein "machines" whose domains have been studied extensively ...The E. coli pyruvate and 2-oxoglutarate dehydrogenases are two closely related, large complexes that exemplify a growing number of multiprotein "machines" whose domains have been studied extensively and modeled in atomic detail, but whose quaternary structures have remained unclear for lack of an effective imaging technology. Here, electron cryotomography was used to show that the E1 and E3 subunits of these complexes are flexibly tethered approximately 11 nm away from the E2 core. This result demonstrates unambiguously that electron cryotomography can reveal the relative positions of features as small as 80 kDa in individual complexes, elucidating quaternary structure and conformational flexibility. | |||||||||
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_1152.map.gz | 227 KB | EMDB map data format | |
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Header (meta data) | emd-1152-v30.xml emd-1152.xml | 12.9 KB 12.9 KB | Display Display | EMDB header |
Images | 1152.gif | 23.6 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-1152 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-1152 | HTTPS FTP |
-Validation report
Summary document | emd_1152_validation.pdf.gz | 269 KB | Display | EMDB validaton report |
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Full document | emd_1152_full_validation.pdf.gz | 268.1 KB | Display | |
Data in XML | emd_1152_validation.xml.gz | 3.5 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-1152 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-1152 | HTTPS FTP |
-Related structure data
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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-Map
File | Download / File: emd_1152.map.gz / Format: CCP4 / Size: 977.5 KB / Type: IMAGE STORED AS SIGNED INTEGER (2 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | Volume of an extracted E. coli 2-oxoglutarate multienzyme complex particle from a dual-tilt tomogram. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 8.2 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 : Unengineered E. coli 2-oxoglutarate dehydrogenase
Entire | Name: Unengineered E. coli 2-oxoglutarate dehydrogenase |
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Components |
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-Supramolecule #1000: Unengineered E. coli 2-oxoglutarate dehydrogenase
Supramolecule | Name: Unengineered E. coli 2-oxoglutarate dehydrogenase / type: sample / ID: 1000 Details: The sample was thawed from storage at -80 degrees Celcius before being loaded onto the grid. Oligomeric state: Up to 24 E1o 2-oxoglutarate dehydrogenase and E3 dihydrolipoamide dehydrogenase dimers together bind to the 24 dihydrolipoamide succinate transferase E2o octahedral core. Number unique components: 3 |
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Molecular weight | Theoretical: 5.2 MDa |
-Macromolecule #1: E2o octahedral core
Macromolecule | Name: E2o octahedral core / type: protein_or_peptide / ID: 1 / Name.synonym: dihydrolipoamide succinate transferase / Number of copies: 24 / Oligomeric state: 24mer / Recombinant expression: Yes |
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Source (natural) | Organism: Escherichia coli (E. coli) / synonym: E. coli / Location in cell: cytoplasm |
Molecular weight | Experimental: 44 KDa |
Recombinant expression | Organism: Escherichia coli (E. coli) |
Sequence | InterPro: Dihydrolipoamide succinyltransferase |
-Macromolecule #2: E1o
Macromolecule | Name: E1o / type: protein_or_peptide / ID: 2 / Name.synonym: 2-oxoglutarate dehydrogenase / Oligomeric state: Dimer / Recombinant expression: Yes |
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Source (natural) | Organism: Escherichia coli (E. coli) / synonym: E. coli / Location in cell: cytoplasm |
Molecular weight | Experimental: 200 KDa |
Recombinant expression | Organism: Escherichia coli (E. coli) |
Sequence | InterPro: 2-oxoglutarate dehydrogenase E1 component |
-Macromolecule #3: E3
Macromolecule | Name: E3 / type: protein_or_peptide / ID: 3 / Name.synonym: dihydrolipoamide dehydrogenase / Oligomeric state: Dimer / Recombinant expression: Yes |
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Source (natural) | Organism: Escherichia coli (E. coli) / synonym: E. coli / Location in cell: cytoplasm |
Molecular weight | Experimental: 100 KDa |
Recombinant expression | Organism: Escherichia coli (E. coli) |
Sequence | InterPro: Dihydrolipoamide dehydrogenase |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | subtomogram averaging |
-Sample preparation
Concentration | 2 mg/mL |
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Buffer | pH: 7 / Details: 20 mM Potassium Phosphate |
Grid | Details: R 1.5/1.3 Quantifoil |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Instrument: OTHER Details: Vitrification instrument: Vitrobot. 4 ul of sample placed on grid at 22 C in air before plunging. Method: Blot for 3.5 seconds with an offset of -3 before plunging. |
-Electron microscopy
Microscope | FEI TECNAI F30 |
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Temperature | Min: 82 K / Max: 82 K / Average: 82 K |
Alignment procedure | Legacy - Electron beam tilt params: 0 |
Specialist optics | Energy filter - Name: GIF 3000 / Energy filter - Lower energy threshold: 0.0 eV / Energy filter - Upper energy threshold: 20.0 eV |
Date | Sep 7, 2004 |
Image recording | Category: CCD / Film or detector model: GATAN ULTRASCAN 1000 (2k x 2k) / Average electron dose: 120 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Calibrated magnification: 36600 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2 mm / Nominal defocus max: 10.0 µm / Nominal defocus min: 10.0 µm / Nominal magnification: 27500 |
Sample stage | Specimen holder: FEI Polara / Specimen holder model: GATAN HELIUM / Tilt series - Axis1 - Min angle: -63 ° / Tilt series - Axis1 - Max angle: 63 ° |
Experimental equipment | Model: Tecnai F30 / Image courtesy: FEI Company |
-Image processing
Details | Dual-axis tilt series, with 44 sections on one axis, and 44 on the other. Average number of tilts used in the 3D reconstructions: 88. Average tomographic tilt angle increment: 3. |
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Final reconstruction | Algorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 55.0 Å / Resolution method: FSC 0.5 CUT-OFF / Software - Name: IMOD Details: The individual, orthogonal tomograms were filtered at their first CTF zero and then merged. |
-Atomic model buiding 1
Initial model | PDB ID: |
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Details | The octameric E2o core (1E2O) was fit into the core density using bfind from the Bsoft package. |