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Yorodumi- PDB-8f1k: SigN RNA polymerase early-melted intermediate bound to full duple... -
+Open data
-Basic information
Entry | Database: PDB / ID: 8f1k | ||||||
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Title | SigN RNA polymerase early-melted intermediate bound to full duplex DNA fragment dhsU36 (-12T) | ||||||
Components |
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Keywords | TRANSCRIPTION/DNA / promoter-bound / initiation / DNA melting / transcription bubble nucleation / TRANSCRIPTION / TRANSCRIPTION-DNA complex | ||||||
Function / homology | Function and homology information arginine metabolic process / DNA-binding transcription activator activity / 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 ...arginine metabolic process / DNA-binding transcription activator activity / 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 / nucleotidyltransferase activity / transcription elongation factor complex / regulation of DNA-templated transcription elongation / transcription antitermination / DNA-templated transcription initiation / cell motility / protein-DNA complex / ribonucleoside binding / DNA-directed 5'-3' RNA polymerase activity / DNA-directed RNA polymerase / response to heat / protein-containing complex assembly / intracellular iron ion homeostasis / transcription cis-regulatory region binding / protein dimerization activity / response to antibiotic / magnesium ion binding / DNA binding / zinc ion binding / membrane / cytoplasm / cytosol Similarity search - Function | ||||||
Biological species | Escherichia coli (E. coli) Aquifex aeolicus (bacteria) | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.8 Å | ||||||
Authors | Mueller, A.U. / Chen, J. / Darst, S.A. | ||||||
Funding support | United States, 1items
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Citation | Journal: Proc Natl Acad Sci U S A / Year: 2023 Title: A general mechanism for transcription bubble nucleation in bacteria. Authors: Andreas U Mueller / James Chen / Mengyu Wu / Courtney Chiu / B Tracy Nixon / Elizabeth A Campbell / Seth A Darst / Abstract: Bacterial transcription initiation requires σ factors for nucleation of the transcription bubble. The canonical housekeeping σ factor, σ, nucleates DNA melting via recognition of conserved bases ...Bacterial transcription initiation requires σ factors for nucleation of the transcription bubble. The canonical housekeeping σ factor, σ, nucleates DNA melting via recognition of conserved bases of the promoter -10 motif, which are unstacked and captured in pockets of σ. By contrast, the mechanism of transcription bubble nucleation and formation during the unrelated σ-mediated transcription initiation is poorly understood. Herein, we combine structural and biochemical approaches to establish that σ, like σ, captures a flipped, unstacked base in a pocket formed between its N-terminal region I (RI) and extra-long helix features. Strikingly, RI inserts into the nascent bubble to stabilize the nucleated bubble prior to engagement of the obligate ATPase activator. Our data suggest a general paradigm of transcription initiation that requires σ factors to nucleate an early melted intermediate prior to productive RNA synthesis. | ||||||
History |
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-Structure visualization
Structure viewer | Molecule: MolmilJmol/JSmol |
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-Downloads & links
-Download
PDBx/mmCIF format | 8f1k.cif.gz | 781.4 KB | Display | PDBx/mmCIF format |
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PDB format | pdb8f1k.ent.gz | 619.1 KB | Display | PDB format |
PDBx/mmJSON format | 8f1k.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 8f1k_validation.pdf.gz | 1.1 MB | Display | wwPDB validaton report |
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Full document | 8f1k_full_validation.pdf.gz | 1.2 MB | Display | |
Data in XML | 8f1k_validation.xml.gz | 98.8 KB | Display | |
Data in CIF | 8f1k_validation.cif.gz | 150.4 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/f1/8f1k ftp://data.pdbj.org/pub/pdb/validation_reports/f1/8f1k | HTTPS FTP |
-Related structure data
Related structure data | 28791MC 8f1iC 8f1jC 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 chain , 2 types, 4 molecules ACBD
#1: DNA chain | Mass: 11104.221 Da / Num. of mol.: 2 / Source method: obtained synthetically / Source: (synth.) Aquifex aeolicus (bacteria) / References: GenBank: AE000657.1 #7: DNA chain | Mass: 11045.092 Da / Num. of mol.: 2 / Source method: obtained synthetically / Source: (synth.) Aquifex aeolicus (bacteria) / References: GenBank: AE000657.1 |
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-DNA-directed RNA polymerase subunit ... , 4 types, 5 molecules GHIJK
#2: 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 (E. coli) / References: UniProt: P0A7Z4, DNA-directed RNA polymerase #3: 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, groN, nitB, rif, ron, stl, stv, tabD, b3987, JW3950 Production host: Escherichia coli (E. coli) / References: UniProt: P0A8V2, DNA-directed RNA polymerase #4: Protein | | Mass: 158105.672 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 (E. coli) / References: UniProt: P0A8T7, DNA-directed RNA polymerase #5: 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, b3649, JW3624 / Production host: Escherichia coli (E. coli) / References: UniProt: P0A800, DNA-directed RNA polymerase |
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-Protein , 1 types, 1 molecules M
#6: Protein | Mass: 54406.906 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Escherichia coli (E. coli) / Gene: rpoN, glnF, ntrA, b3202, JW3169 / Production host: Escherichia coli (E. coli) / References: UniProt: P24255 |
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-Non-polymers , 3 types, 13 molecules
#8: Chemical | ChemComp-MG / | ||
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#9: Chemical | #10: Water | ChemComp-HOH / | |
-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: EsN-dhsU36 / Type: COMPLEX / Entity ID: #1-#7 / Source: MULTIPLE SOURCES | |||||||||||||||||||||||||
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Molecular weight | Experimental value: NO | |||||||||||||||||||||||||
Buffer solution | pH: 8 | |||||||||||||||||||||||||
Buffer component |
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Specimen | Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES | |||||||||||||||||||||||||
Specimen support | Details: Self-built glow discharge unit was evacuated using a vacuum pump prior to application of voltage (no pressure and/or voltage readings available) Grid material: GOLD / Grid mesh size: 400 divisions/in. / Grid type: C-flat-1.2/1.3 | |||||||||||||||||||||||||
Vitrification | Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 295 K |
-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 / Nominal defocus max: 2500 nm / Nominal defocus min: 800 nm |
Image recording | Average exposure time: 2 sec. / Electron dose: 50.8 e/Å2 / Film or detector model: GATAN K3 (6k x 4k) / Num. of real images: 6088 Details: dose-fractionation, 0.05 s/frame (=40 frames total) |
EM imaging optics | Energyfilter name: GIF Bioquantum / Energyfilter slit width: 20 eV |
-Processing
EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||
3D reconstruction | Resolution: 2.8 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 892568 Details: Particle number and FSC resolution estimate of full map provided Symmetry type: POINT | ||||||||||||||||||||||||
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