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Yorodumi- PDB-7uwh: CryoEM Structure of E. coli Transcription-Coupled Ribonucleotide ... -
+Open data
-Basic information
Entry | Database: PDB / ID: 7uwh | |||||||||
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Title | CryoEM Structure of E. coli Transcription-Coupled Ribonucleotide Excision Repair (TC-RER) complex bound to ribonucleotide substrate | |||||||||
Components |
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Keywords | TRANSFERASE/HYDROLASE/DNA/RNA / Transcription-coupled RER / TRANSCRIPTION / TRANSFERASE-HYDROLASE-DNA-RNA complex | |||||||||
Function / homology | Function and homology information ribonuclease H2 complex / DNA replication, removal of RNA primer / ribonuclease H / submerged biofilm formation / cellular response to cell envelope stress / cytosolic DNA-directed RNA polymerase complex / regulation of DNA-templated transcription initiation / bacterial-type flagellum assembly / bacterial-type flagellum-dependent cell motility / mismatch repair ...ribonuclease H2 complex / DNA replication, removal of RNA primer / ribonuclease H / submerged biofilm formation / cellular response to cell envelope stress / cytosolic DNA-directed RNA polymerase complex / regulation of DNA-templated transcription initiation / bacterial-type flagellum assembly / bacterial-type flagellum-dependent cell motility / mismatch repair / 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 / RNA-DNA hybrid ribonuclease activity / manganese ion binding / response to heat / intracellular iron ion homeostasis / protein dimerization activity / response to antibiotic / DNA-templated transcription / magnesium ion binding / DNA binding / RNA binding / zinc ion binding / membrane / cytosol / cytoplasm Similarity search - Function | |||||||||
Biological species | Escherichia coli (E. coli) | |||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.1 Å | |||||||||
Authors | Hao, Z.T. / Grower, M. / Bharati, B. / Proshkin, S. / Epshtein, V. / Svetlov, V. / Nudler, E. / Shamovsky, I. | |||||||||
Funding support | United States, 2items
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Citation | Journal: Cell / Year: 2023 Title: RNA polymerase drives ribonucleotide excision DNA repair in E. coli. Authors: Zhitai Hao / Manjunath Gowder / Sergey Proshkin / Binod K Bharati / Vitaly Epshtein / Vladimir Svetlov / Ilya Shamovsky / Evgeny Nudler / Abstract: Ribonuclease HII (RNaseHII) is the principal enzyme that removes misincorporated ribonucleoside monophosphates (rNMPs) from genomic DNA. Here, we present structural, biochemical, and genetic evidence ...Ribonuclease HII (RNaseHII) is the principal enzyme that removes misincorporated ribonucleoside monophosphates (rNMPs) from genomic DNA. Here, we present structural, biochemical, and genetic evidence demonstrating that ribonucleotide excision repair (RER) is directly coupled to transcription. Affinity pull-downs and mass-spectrometry-assisted mapping of in cellulo inter-protein cross-linking reveal the majority of RNaseHII molecules interacting with RNA polymerase (RNAP) in E. coli. Cryoelectron microscopy structures of RNaseHII bound to RNAP during elongation, with and without the target rNMP substrate, show specific protein-protein interactions that define the transcription-coupled RER (TC-RER) complex in engaged and unengaged states. The weakening of RNAP-RNaseHII interactions compromises RER in vivo. The structure-functional data support a model where RNaseHII scans DNA in one dimension in search for rNMPs while "riding" the RNAP. We further demonstrate that TC-RER accounts for a significant fraction of repair events, thereby establishing RNAP as a surveillance "vehicle" for detecting the most frequently occurring replication errors. | |||||||||
History |
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-Structure visualization
Structure viewer | Molecule: MolmilJmol/JSmol |
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-Downloads & links
-Download
PDBx/mmCIF format | 7uwh.cif.gz | 633.4 KB | Display | PDBx/mmCIF format |
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PDB format | pdb7uwh.ent.gz | 501.6 KB | Display | PDB format |
PDBx/mmJSON format | 7uwh.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 7uwh_validation.pdf.gz | 1.2 MB | Display | wwPDB validaton report |
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Full document | 7uwh_full_validation.pdf.gz | 1.3 MB | Display | |
Data in XML | 7uwh_validation.xml.gz | 97.2 KB | Display | |
Data in CIF | 7uwh_validation.cif.gz | 152.9 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/uw/7uwh ftp://data.pdbj.org/pub/pdb/validation_reports/uw/7uwh | HTTPS FTP |
-Related structure data
Related structure data | 26832MC 7uweC C: citing same article (ref.) M: map data used to model this data |
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Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
-Assembly
Deposited unit |
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1 |
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-Components
-DNA-directed RNA polymerase subunit ... , 4 types, 5 molecules GHIJK
#4: Protein | Mass: 36558.680 Da / Num. of mol.: 2 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Escherichia coli (E. coli) / Gene: rpoA, pez, phs, sez, b3295, JW3257 Production host: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria) References: UniProt: P0A7Z4, DNA-directed RNA polymerase #5: 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-Gold(DE3)pLysS AG' (bacteria) References: UniProt: P0A8V4, DNA-directed RNA polymerase #6: Protein | | Mass: 155366.781 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Escherichia coli (E. coli) / Gene: rpoC, tabB, b3988, JW3951 Production host: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria) References: UniProt: P0A8T7, DNA-directed RNA polymerase #7: 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-Gold(DE3)pLysS AG' (bacteria) References: UniProt: P0A802, DNA-directed RNA polymerase |
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-DNA chain / DNA/RNA hybrid / Protein / RNA chain , 4 types, 4 molecules ABCR
#1: DNA chain | Mass: 18077.531 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Escherichia coli (E. coli) |
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#2: DNA/RNA hybrid | Mass: 18133.561 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Escherichia coli (E. coli) |
#3: Protein | Mass: 21499.986 Da / Num. of mol.: 1 / Mutation: E17A Source method: isolated from a genetically manipulated source Source: (gene. exp.) Escherichia coli (E. coli) Production host: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria) References: UniProt: W8T723, ribonuclease H |
#8: RNA chain | Mass: 5859.580 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Escherichia coli (E. coli) |
-Non-polymers , 2 types, 3 molecules
#9: Chemical | #10: Chemical | ChemComp-MG / | |
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-Details
Has ligand of interest | N |
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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: Elongation Complex(EC)-RNaseH2 complex bound to ribonucleotide substrate Type: COMPLEX / Entity ID: #1-#8 / Source: MULTIPLE SOURCES |
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Molecular weight | Experimental value: NO |
Source (natural) | Organism: Escherichia coli (E. coli) |
Source (recombinant) | Organism: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria) |
Buffer solution | pH: 8 |
Specimen | Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Vitrification | Cryogen name: OTHER |
-Electron microscopy imaging
Experimental equipment | Model: Talos Arctica / Image courtesy: FEI Company |
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Microscopy | Model: FEI TALOS ARCTICA |
Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 200 kV / Illumination mode: FLOOD BEAM |
Electron lens | Mode: BRIGHT FIELD / Nominal defocus max: 2000 nm / Nominal defocus min: 800 nm |
Image recording | Electron dose: 56 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) |
-Processing
CTF correction | Type: NONE |
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3D reconstruction | Resolution: 3.1 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 70705 / Symmetry type: POINT |