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- EMDB-5407: Cryo-EM map of a yeast minimal preinitiation complex interacting ... -
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Open data
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Basic information
Entry | Database: EMDB / ID: EMD-5407 | |||||||||
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Title | Cryo-EM map of a yeast minimal preinitiation complex interacting with the Mediator Head module | |||||||||
![]() | Cryo-EM map of a complex including RNA polymerase II, Mediator Head module, TFIIF, TFIIB, TBP, and promoter DNA. TFIIB, TBP, and DNA are not visible in the map and are presumed to be disordered | |||||||||
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![]() | transcription / RNA polymerase II / Mediator / Head module / preinitiation complex | |||||||||
Function / homology | ![]() RNA polymerase II complex recruiting activity / core mediator complex / negative regulation of ribosomal protein gene transcription by RNA polymerase II / transcription open complex formation at RNA polymerase II promoter / RPB4-RPB7 complex / TFIIH-class transcription factor complex binding / mediator complex / nuclear-transcribed mRNA catabolic process, deadenylation-dependent decay / RNA Polymerase I Transcription Initiation / Processing of Capped Intron-Containing Pre-mRNA ...RNA polymerase II complex recruiting activity / core mediator complex / negative regulation of ribosomal protein gene transcription by RNA polymerase II / transcription open complex formation at RNA polymerase II promoter / RPB4-RPB7 complex / TFIIH-class transcription factor complex binding / mediator complex / nuclear-transcribed mRNA catabolic process, deadenylation-dependent decay / RNA Polymerase I Transcription Initiation / Processing of Capped Intron-Containing Pre-mRNA / RNA Polymerase III Transcription Initiation From Type 2 Promoter / RNA Pol II CTD phosphorylation and interaction with CE / Formation of the Early Elongation Complex / mRNA Capping / RNA polymerase II transcribes snRNA genes / TP53 Regulates Transcription of DNA Repair Genes / RNA Polymerase II Promoter Escape / RNA Polymerase II Transcription Pre-Initiation And Promoter Opening / RNA Polymerase II Transcription Initiation / RNA Polymerase II Transcription Initiation And Promoter Clearance / termination of RNA polymerase II transcription / RNA-templated transcription / termination of RNA polymerase III transcription / RNA Polymerase II Pre-transcription Events / : / Formation of TC-NER Pre-Incision Complex / : / termination of RNA polymerase I transcription / transcription initiation at RNA polymerase III promoter / RNA Polymerase I Promoter Escape / nucleolar large rRNA transcription by RNA polymerase I / Gap-filling DNA repair synthesis and ligation in TC-NER / maintenance of transcriptional fidelity during transcription elongation by RNA polymerase II / positive regulation of nuclear-transcribed mRNA poly(A) tail shortening / transcription initiation at RNA polymerase I promoter / transcription by RNA polymerase I / TFIID-class transcription factor complex binding / Estrogen-dependent gene expression / transcription by RNA polymerase III / Dual incision in TC-NER / transcription elongation by RNA polymerase I / positive regulation of transcription initiation by RNA polymerase II / RNA polymerase II core promoter sequence-specific DNA binding / positive regulation of translational initiation / RNA polymerase I complex / RNA polymerase III complex / transcription-coupled nucleotide-excision repair / RNA polymerase III activity / RNA polymerase II, core complex / tRNA transcription by RNA polymerase III / RNA polymerase I activity / translesion synthesis / RNA polymerase II activity / RNA polymerase II preinitiation complex assembly / translation initiation factor binding / TBP-class protein binding / DNA-templated transcription initiation / transcription elongation by RNA polymerase II / transcription initiation at RNA polymerase II promoter / positive regulation of transcription elongation by RNA polymerase II / transcription coregulator activity / P-body / ribonucleoside binding / DNA-directed 5'-3' RNA polymerase activity / cytoplasmic stress granule / DNA-directed RNA polymerase / transcription corepressor activity / peroxisome / ribosome biogenesis / protein-macromolecule adaptor activity / single-stranded DNA binding / DNA-binding transcription factor binding / transcription by RNA polymerase II / nucleic acid binding / transcription coactivator activity / single-stranded RNA binding / protein dimerization activity / RNA polymerase II cis-regulatory region sequence-specific DNA binding / protein domain specific binding / nucleotide binding / mRNA binding / nucleolus / regulation of transcription by RNA polymerase II / negative regulation of transcription by RNA polymerase II / positive regulation of transcription by RNA polymerase II / mitochondrion / DNA binding / zinc ion binding / nucleoplasm / nucleus / metal ion binding / cytoplasm Similarity search - Function | |||||||||
Biological species | ![]() ![]() | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 16.0 Å | |||||||||
![]() | Cai G / Chaban Y / Imasaki T / Kovacs JA / Calero G / Penczek PA / Takagi Y / Asturias FJ | |||||||||
![]() | ![]() Title: Interaction of the mediator head module with RNA polymerase II. Authors: Gang Cai / Yuriy L Chaban / Tsuyoshi Imasaki / Julio A Kovacs / Guillermo Calero / Pawel A Penczek / Yuichiro Takagi / Francisco J Asturias / ![]() Abstract: Mediator, a large (21 polypeptides, MW ∼1 MDa) complex conserved throughout eukaryotes, plays an essential role in control of gene expression by conveying regulatory signals that influence the ...Mediator, a large (21 polypeptides, MW ∼1 MDa) complex conserved throughout eukaryotes, plays an essential role in control of gene expression by conveying regulatory signals that influence the activity of the preinitiation complex. However, the precise mode of interaction between Mediator and RNA polymerase II (RNAPII), and the mechanism of regulation by Mediator remain elusive. We used cryo-electron microscopy and reconstituted in vitro transcription assays to characterize a transcriptionally-active complex including the Mediator Head module and components of a minimum preinitiation complex (RNAPII, TFIIF, TFIIB, TBP, and promoter DNA). Our results reveal how the Head interacts with RNAPII, affecting its conformation and function. | |||||||||
History |
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Structure visualization
Movie |
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Structure viewer | EM map: ![]() ![]() ![]() |
Supplemental images |
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Downloads & links
-EMDB archive
Map data | ![]() | 1.3 MB | ![]() | |
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Header (meta data) | ![]() ![]() | 15.6 KB 15.6 KB | Display Display | ![]() |
Images | ![]() | 229.8 KB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Validation report
Summary document | ![]() | 299.3 KB | Display | ![]() |
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Full document | ![]() | 298.9 KB | Display | |
Data in XML | ![]() | 5 KB | Display | |
Arichive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 3j1nMC ![]() 3j1oMC M: atomic model generated by this map C: citing same article ( |
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Similar structure data |
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Links
EMDB pages | ![]() ![]() |
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Related items in Molecule of the Month |
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Map
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Annotation | Cryo-EM map of a complex including RNA polymerase II, Mediator Head module, TFIIF, TFIIB, TBP, and promoter DNA. TFIIB, TBP, and DNA are not visible in the map and are presumed to be disordered | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 3.83 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
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Sample components
-Entire : Complex of RNA polymerase II, Mediator Head module, TFIIF, TFIIB,...
Entire | Name: Complex of RNA polymerase II, Mediator Head module, TFIIF, TFIIB, TBP, and promoter DNA |
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Components |
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-Supramolecule #1000: Complex of RNA polymerase II, Mediator Head module, TFIIF, TFIIB,...
Supramolecule | Name: Complex of RNA polymerase II, Mediator Head module, TFIIF, TFIIB, TBP, and promoter DNA type: sample / ID: 1000 Details: Although biochemical and functional evidence indicates the presence of all 6 original components (RNA polymerase II, Mediator Head, TFIIF, TFIIB, TBP, DNA), only 3 components (RNA polymerase ...Details: Although biochemical and functional evidence indicates the presence of all 6 original components (RNA polymerase II, Mediator Head, TFIIF, TFIIB, TBP, DNA), only 3 components (RNA polymerase II, Mediator Head, TFIIF) can be clearly identified in the cryo-EM map. Number unique components: 6 |
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Molecular weight | Theoretical: 900 KDa / Method: sequence |
-Macromolecule #1: RNA polymerase II
Macromolecule | Name: RNA polymerase II / type: protein_or_peptide / ID: 1 / Name.synonym: pol II / Number of copies: 1 / Recombinant expression: No / Database: NCBI |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 540 KDa |
-Macromolecule #2: Head module
Macromolecule | Name: Head module / type: protein_or_peptide / ID: 2 / Name.synonym: Head / Number of copies: 1 / Recombinant expression: Yes |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 240 KDa |
Recombinant expression | Organism: ![]() |
-Macromolecule #3: TFIIF
Macromolecule | Name: TFIIF / type: protein_or_peptide / ID: 3 / Name.synonym: IIF / Number of copies: 1 / Recombinant expression: No / Database: NCBI |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 150 KDa |
-Experimental details
-Structure determination
Method | cryo EM |
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![]() | single particle reconstruction |
Aggregation state | particle |
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Sample preparation
Concentration | 0.3 mg/mL |
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Buffer | pH: 8 / Details: 25 mM KCl, 25 mM Tris-HCl, 10 mM DTT |
Grid | Details: 400 mesh Cu/Rh grids, coated with a perforated carbon film and glow discharged in the presence of amylamine |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 95 % / Chamber temperature: 77 K / Instrument: HOMEMADE PLUNGER / Method: Blot for ~2 sec before plunging |
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Electron microscopy
Microscope | FEI TECNAI F20 |
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Temperature | Min: 77 K / Max: 120 K / Average: 115 K |
Alignment procedure | Legacy - Astigmatism: Objective lens astigmatism was corrected at 125,000X magnification |
Date | Jul 30, 2008 |
Image recording | Category: CCD / Film or detector model: TVIPS TEMCAM-F415 (4k x 4k) / Number real images: 500 / Average electron dose: 10 e/Å2 / Bits/pixel: 8 |
Electron beam | Acceleration voltage: 120 kV / Electron source: ![]() |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.0 mm / Nominal defocus max: 3.8 µm / Nominal defocus min: 0.8 µm / Nominal magnification: 50000 |
Sample stage | Specimen holder model: GATAN LIQUID NITROGEN |
Experimental equipment | ![]() Model: Tecnai F20 / Image courtesy: FEI Company |
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Image processing
CTF correction | Details: Each CCD frame |
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Final reconstruction | Algorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 16.0 Å / Resolution method: FSC 0.5 CUT-OFF / Software - Name: Sparx,Spider / Number images used: 51000 |
-Atomic model buiding 1
Initial model | PDB ID: Chain - #0 - Chain ID: A / Chain - #1 - Chain ID: B / Chain - #2 - Chain ID: C / Chain - #3 - Chain ID: D / Chain - #4 - Chain ID: E / Chain - #5 - Chain ID: F / Chain - #6 - Chain ID: G / Chain - #7 - Chain ID: H / Chain - #8 - Chain ID: I / Chain - #9 - Chain ID: J / Chain - #10 - Chain ID: K / Chain - #11 - Chain ID: L |
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Software | Name: Monte Carlo Docking Analysis |
Details | Protocol: Rigid body fitting of individual RNA polymerase II structural modules. Fitted individual structural modules previously identified by X-ray crystallographic studies of RNA polymerase II under different crystallization conditions |
Refinement | Space: REAL / Protocol: RIGID BODY FIT |
Output model | ![]() PDB-3j1n: ![]() PDB-3j1o: |
-Atomic model buiding 2
Initial model | PDB ID: Chain - #0 - Chain ID: A / Chain - #1 - Chain ID: B / Chain - #2 - Chain ID: C / Chain - #3 - Chain ID: D / Chain - #4 - Chain ID: E / Chain - #5 - Chain ID: F / Chain - #6 - Chain ID: G |
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Software | Name: Monte Carlo Docking Analysis |
Details | Protocol: Rigid body fitting of individual Mediator Head structural modules. Fitted individual structural modules previously identified by EM and X-ray crystallographic studies of the Head module |
Refinement | Space: REAL / Protocol: RIGID BODY FIT |
Output model | ![]() PDB-3j1n: ![]() PDB-3j1o: |