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Yorodumi- EMDB-2309: Structural visualization of key steps in human transcription init... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-2309 | |||||||||
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Title | Structural visualization of key steps in human transcription initiation | |||||||||
Map data | apo TFIIH | |||||||||
Sample |
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Keywords | human transcription initiation | |||||||||
Biological species | Homo sapiens (human) | |||||||||
Method | single particle reconstruction / negative staining / Resolution: 20.0 Å | |||||||||
Authors | He Y / Fang J / Taatjes DJ / Nogales E | |||||||||
Citation | Journal: Nature / Year: 2013 Title: Structural visualization of key steps in human transcription initiation. Authors: Yuan He / Jie Fang / Dylan J Taatjes / Eva Nogales / Abstract: Eukaryotic transcription initiation requires the assembly of general transcription factors into a pre-initiation complex that ensures the accurate loading of RNA polymerase II (Pol II) at the ...Eukaryotic transcription initiation requires the assembly of general transcription factors into a pre-initiation complex that ensures the accurate loading of RNA polymerase II (Pol II) at the transcription start site. The molecular mechanism and function of this assembly have remained elusive due to lack of structural information. Here we have used an in vitro reconstituted system to study the stepwise assembly of human TBP, TFIIA, TFIIB, Pol II, TFIIF, TFIIE and TFIIH onto promoter DNA using cryo-electron microscopy. Our structural analyses provide pseudo-atomic models at various stages of transcription initiation that illuminate critical molecular interactions, including how TFIIF engages Pol II and promoter DNA to stabilize both the closed pre-initiation complex and the open-promoter complex, and to regulate start--initiation complexes, combined with the localization of the TFIIH helicases XPD and XPB, support a DNA translocation model of XPB and explain its essential role in promoter opening. | |||||||||
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_2309.map.gz | 7.1 MB | EMDB map data format | |
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Header (meta data) | emd-2309-v30.xml emd-2309.xml | 8.6 KB 8.6 KB | Display Display | EMDB header |
Images | EMD-2309.png | 126.7 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-2309 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-2309 | HTTPS FTP |
-Related structure data
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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-Map
File | Download / File: emd_2309.map.gz / Format: CCP4 / Size: 7.8 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | apo TFIIH | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 2.56 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 : apo TFIIH
Entire | Name: apo TFIIH |
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Components |
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-Supramolecule #1000: apo TFIIH
Supramolecule | Name: apo TFIIH / type: sample / ID: 1000 / Number unique components: 1 |
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Molecular weight | Theoretical: 450 KDa |
-Macromolecule #1: General transcription factor IIH
Macromolecule | Name: General transcription factor IIH / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Recombinant expression: No |
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Source (natural) | Organism: Homo sapiens (human) / synonym: Human / Cell: Hela / Organelle: Nucleus |
Molecular weight | Theoretical: 450 KDa |
-Experimental details
-Structure determination
Method | negative staining |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 0.04 mg/mL |
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Buffer | pH: 7.9 Details: 10 mM HEPES, 5% glycerol, 10 mM MgCl2, 50 mM KCl, 1 mM DTT, 0.05% NP-40 |
Staining | Type: NEGATIVE Details: Grids with adsorbed protein floated on 2% w/v uranyl formate for 1 minute. |
Grid | Details: 200 mesh Cu grid |
Vitrification | Cryogen name: NONE / Instrument: OTHER |
-Electron microscopy
Microscope | FEI TECNAI F20 |
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Electron beam | Acceleration voltage: 120 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Calibrated magnification: 80000 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.2 mm / Nominal defocus max: 1.2 µm / Nominal defocus min: 0.5 µm / Nominal magnification: 80000 |
Sample stage | Specimen holder: Room temp single tilt / Specimen holder model: SIDE ENTRY, EUCENTRIC |
Alignment procedure | Legacy - Astigmatism: objective lens astigmatism was corrected at 250,000 times magnification. |
Details | Data acquired using Leginon |
Date | Apr 17, 2012 |
Image recording | Category: CCD / Film or detector model: GATAN ULTRASCAN 4000 (4k x 4k) / Number real images: 450 / Average electron dose: 20 e/Å2 |
Experimental equipment | Model: Tecnai F20 / Image courtesy: FEI Company |
-Image processing
CTF correction | Details: whole micrograph |
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Final reconstruction | Applied symmetry - Point group: C1 (asymmetric) / Algorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 20.0 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: EMAN2, SPARX Details: Image processing was performed in the Appion processing environment. 3D reconstruction was performed using EMAN2 and SPARX libraries. Final map was filtered to local resolution using the ...Details: Image processing was performed in the Appion processing environment. 3D reconstruction was performed using EMAN2 and SPARX libraries. Final map was filtered to local resolution using the blocres function in Bsoft package. Number images used: 13023 |