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Yorodumi- PDB-6oul: Cryo-EM structure of Escherichia coli RNAP polymerase bound to rp... -
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-Basic information
Entry | Database: PDB / ID: 6oul | ||||||
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Title | Cryo-EM structure of Escherichia coli RNAP polymerase bound to rpsTP2 promoter DNA | ||||||
Components |
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Keywords | TRANSCRIPTION / protein-DNA complex / transcription initiation / RNA polyermase / DNA promoter melting | ||||||
Function / homology | Function and homology information sigma factor antagonist complex / RNA polymerase complex / submerged biofilm formation / cellular response to cell envelope stress / cytosolic DNA-directed RNA polymerase complex / regulation of DNA-templated transcription initiation / sigma factor activity / bacterial-type flagellum assembly / bacterial-type flagellum-dependent cell motility / nitrate assimilation ...sigma factor antagonist complex / RNA polymerase complex / submerged biofilm formation / cellular response to cell envelope stress / cytosolic DNA-directed RNA polymerase complex / regulation of DNA-templated transcription initiation / sigma factor activity / bacterial-type flagellum assembly / bacterial-type flagellum-dependent cell motility / nitrate assimilation / DNA-directed RNA polymerase complex / transcription elongation factor complex / regulation of DNA-templated transcription elongation / transcription antitermination / DNA-templated transcription initiation / cell motility / ribonucleoside binding / DNA-directed 5'-3' RNA polymerase activity / DNA-directed RNA polymerase / response to heat / protein-containing complex assembly / intracellular iron ion homeostasis / protein dimerization activity / response to antibiotic / negative regulation of DNA-templated transcription / DNA-templated transcription / magnesium ion binding / DNA binding / zinc ion binding / membrane / metal ion binding / cytoplasm / cytosol Similarity search - Function | ||||||
Biological species | Escherichia coli (E. coli) Escherichia coli K-12 (bacteria) | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.4 Å | ||||||
Authors | Chen, J. / Chiu, C.E. / Campbell, E.A. / Darst, S.A. | ||||||
Funding support | United States, 1items
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Citation | Journal: Elife / Year: 2019 Title: TraR allosterically regulates transcription initiation by altering RNA polymerase conformation. Authors: James Chen / Saumya Gopalkrishnan / Courtney Chiu / Albert Y Chen / Elizabeth A Campbell / Richard L Gourse / Wilma Ross / Seth A Darst / Abstract: TraR and its homolog DksA are bacterial proteins that regulate transcription initiation by binding directly to RNA polymerase (RNAP) rather than to promoter DNA. Effects of TraR mimic the combined ...TraR and its homolog DksA are bacterial proteins that regulate transcription initiation by binding directly to RNA polymerase (RNAP) rather than to promoter DNA. Effects of TraR mimic the combined effects of DksA and its cofactor ppGpp, but the structural basis for regulation by these factors remains unclear. Here, we use cryo-electron microscopy to determine structures of RNAP, with or without TraR, and of an RNAP-promoter complex. TraR binding induced RNAP conformational changes not seen in previous crystallographic analyses, and a quantitative analysis revealed TraR-induced changes in RNAP conformational heterogeneity. These changes involve mobile regions of RNAP affecting promoter DNA interactions, including the βlobe, the clamp, the bridge helix, and several lineage-specific insertions. Using mutational approaches, we show that these structural changes, as well as effects on σ region 1.1, are critical for transcription activation or inhibition, depending on the kinetic features of regulated promoters. | ||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | Molecule: MolmilJmol/JSmol |
-Downloads & links
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PDBx/mmCIF format | 6oul.cif.gz | 737.5 KB | Display | PDBx/mmCIF format |
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PDB format | pdb6oul.ent.gz | 580.8 KB | Display | PDB format |
PDBx/mmJSON format | 6oul.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 6oul_validation.pdf.gz | 1.2 MB | Display | wwPDB validaton report |
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Full document | 6oul_full_validation.pdf.gz | 1.4 MB | Display | |
Data in XML | 6oul_validation.xml.gz | 129.6 KB | Display | |
Data in CIF | 6oul_validation.cif.gz | 194.2 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/ou/6oul ftp://data.pdbj.org/pub/pdb/validation_reports/ou/6oul | HTTPS FTP |
-Related structure data
Related structure data | 20203MC 0348C 0349C 6n57C 6n58C 6p1kC C: citing same article (ref.) M: map data used to model this data |
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Similar structure data |
-Links
-Assembly
Deposited unit |
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1 |
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-Components
-DNA-directed RNA polymerase subunit ... , 4 types, 6 molecules GHRIJK
#1: Protein | Mass: 36558.680 Da / Num. of mol.: 3 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Escherichia coli (E. coli) / Gene: rpoA / Production host: Escherichia coli BL21(DE3) (bacteria) References: UniProt: A0A073G207, UniProt: P0A7Z4*PLUS, DNA-directed RNA polymerase #2: Protein | | Mass: 150820.875 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Escherichia coli (E. coli) / Gene: rpoB, Z5560, ECs4910 / Production host: Escherichia coli BL21(DE3) (bacteria) References: UniProt: P0A8V4, UniProt: P0A8V2*PLUS, DNA-directed RNA polymerase #3: Protein | | Mass: 158073.609 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Escherichia coli (E. coli) / Gene: rpoC / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: U9YPW3, DNA-directed RNA polymerase #4: Protein | | Mass: 10249.547 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Escherichia coli (E. coli) / Gene: rpoZ, Z5075, ECs4524 / Production host: Escherichia coli BL21(DE3) (bacteria) References: UniProt: P0A802, UniProt: P0A800*PLUS, DNA-directed RNA polymerase |
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-Protein , 1 types, 1 molecules L
#5: Protein | Mass: 70715.609 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Escherichia coli (E. coli) / Gene: rpoD / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: Q0P6L9, UniProt: P00579*PLUS |
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-DNA chain , 2 types, 2 molecules PQ
#6: DNA chain | Mass: 26235.822 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Escherichia coli K-12 (bacteria) |
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#7: DNA chain | Mass: 26190.756 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Escherichia coli K-12 (bacteria) |
-Non-polymers , 3 types, 6 molecules
#8: Chemical | #9: Chemical | ChemComp-MG / | #10: Chemical | |
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-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
-Sample preparation
Component | Name: Cryo-EM structure of Escherichia coli RNAP polymerase bound to rpsTP2 promoter DNA Type: COMPLEX / Entity ID: #1-#7 / Source: RECOMBINANT |
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Source (natural) | Organism: Escherichia coli (E. coli) |
Source (recombinant) | Organism: Escherichia coli BL21(DE3) (bacteria) |
Buffer solution | pH: 8 |
Specimen | Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Vitrification | Cryogen name: ETHANE |
-Electron microscopy imaging
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: FEI TITAN KRIOS |
Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM |
Electron lens | Mode: BRIGHT FIELD |
Image recording | Electron dose: 1 e/Å2 / Film or detector model: GATAN K2 SUMMIT (4k x 4k) |
-Processing
CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION |
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Symmetry | Point symmetry: C1 (asymmetric) |
3D reconstruction | Resolution: 3.4 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 289670 / Symmetry type: POINT |