[English] 日本語
Yorodumi- PDB-5nsr: Cryo-EM structure of RNA polymerase-sigma54 holo enzyme with prom... -
+Open data
-Basic information
Entry | Database: PDB / ID: 5nsr | |||||||||
---|---|---|---|---|---|---|---|---|---|---|
Title | Cryo-EM structure of RNA polymerase-sigma54 holo enzyme with promoter DNA closed complex | |||||||||
Components |
| |||||||||
Keywords | TRANSCRIPTION / transcription initiation / RNA polymerase / sigma54 | |||||||||
Function / homology | Function and homology information nitrogen fixation / 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 ...nitrogen fixation / 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 / DNA-directed RNA polymerase complex / nucleotidyltransferase activity / 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 / magnesium ion binding / DNA binding / zinc ion binding / membrane / cytoplasm / cytosol Similarity search - Function | |||||||||
Biological species | Escherichia coli K-12 (bacteria) Klebsiella pneumoniae (bacteria) | |||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.8 Å | |||||||||
Authors | Glyde, R. / Ye, F.Z. / Darbari, V.C. / Zhang, N. / Buck, M. / Zhang, X.D. | |||||||||
Funding support | United Kingdom, 2items
| |||||||||
Citation | Journal: Mol Cell / Year: 2017 Title: Structures of RNA Polymerase Closed and Intermediate Complexes Reveal Mechanisms of DNA Opening and Transcription Initiation. Authors: Robert Glyde / Fuzhou Ye / Vidya Chandran Darbari / Nan Zhang / Martin Buck / Xiaodong Zhang / Abstract: Gene transcription is carried out by RNA polymerases (RNAPs). For transcription to occur, the closed promoter complex (RPc), where DNA is double stranded, must isomerize into an open promoter complex ...Gene transcription is carried out by RNA polymerases (RNAPs). For transcription to occur, the closed promoter complex (RPc), where DNA is double stranded, must isomerize into an open promoter complex (RPo), where the DNA is melted out into a transcription bubble and the single-stranded template DNA is delivered to the RNAP active site. Using a bacterial RNAP containing the alternative σ factor and cryoelectron microscopy, we determined structures of RPc and the activator-bound intermediate complex en route to RPo at 3.8 and 5.8 Å. Our structures show how RNAP-σ interacts with promoter DNA to initiate the DNA distortions required for transcription bubble formation, and how the activator interacts with RPc, leading to significant conformational changes in RNAP and σ that promote RPo formation. We propose that DNA melting is an active process initiated in RPc and that the RNAP conformations of intermediates are significantly different from that of RPc and RPo. | |||||||||
History |
|
-Structure visualization
Movie |
Movie viewer |
---|---|
Structure viewer | Molecule: MolmilJmol/JSmol |
-Downloads & links
-Download
PDBx/mmCIF format | 5nsr.cif.gz | 684.5 KB | Display | PDBx/mmCIF format |
---|---|---|---|---|
PDB format | pdb5nsr.ent.gz | 532.5 KB | Display | PDB format |
PDBx/mmJSON format | 5nsr.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 5nsr_validation.pdf.gz | 791.8 KB | Display | wwPDB validaton report |
---|---|---|---|---|
Full document | 5nsr_full_validation.pdf.gz | 855.8 KB | Display | |
Data in XML | 5nsr_validation.xml.gz | 94 KB | Display | |
Data in CIF | 5nsr_validation.cif.gz | 145.2 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/ns/5nsr ftp://data.pdbj.org/pub/pdb/validation_reports/ns/5nsr | HTTPS FTP |
-Related structure data
Related structure data | 3695MC 3696C 3697C 5nssC M: map data used to model this data C: citing same article (ref.) |
---|---|
Similar structure data |
-Links
-Assembly
Deposited unit |
|
---|---|
1 |
|
-Components
-DNA-directed RNA polymerase subunit ... , 4 types, 5 molecules ABCDE
#1: Protein | Mass: 36558.680 Da / Num. of mol.: 2 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Escherichia coli K-12 (bacteria) / Gene: rpoA, pez, phs, sez, b3295, JW3257 / Plasmid: pGEM / Production host: Escherichia coli BL21(DE3) (bacteria) / Variant (production host): Gold pLysS AG / References: UniProt: P0A7Z4, 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 K-12 (bacteria) Gene: rpoB, groN, nitB, rif, ron, stl, stv, tabD, b3987, JW3950 Plasmid: pGEM / Production host: Escherichia coli BL21(DE3) (bacteria) / Variant (production host): Gold pLysS AG / References: UniProt: P0A8V2, DNA-directed RNA polymerase #3: Protein | | Mass: 155366.781 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Escherichia coli K-12 (bacteria) / Gene: rpoC, tabB, b3988, JW3951 / Plasmid: pGEM / Production host: Escherichia coli BL21(DE3) (bacteria) / Variant (production host): Gold pLysS AG / References: UniProt: P0A8T7, 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 K-12 (bacteria) / Gene: rpoZ, b3649, JW3624 / Plasmid: paCYCduet1-omega / Production host: Escherichia coli BL21(DE3) (bacteria) / Variant (production host): Gold pLysS AG / References: UniProt: P0A800, DNA-directed RNA polymerase |
---|
-Protein , 1 types, 1 molecules M
#5: Protein | Mass: 62158.797 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Klebsiella pneumoniae (bacteria) / Gene: rpoN, SAMEA3531867_02981, SM87_03359 / Plasmid: pET28b-sigma54 / Production host: Escherichia coli BL21(DE3) (bacteria) / Variant (production host): Gold pLysS AG / References: UniProt: A0A0J4U551, UniProt: P06223*PLUS |
---|
-DNA chain , 2 types, 2 molecules FG
#6: DNA chain | Mass: 19404.410 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Klebsiella pneumoniae (bacteria) / Production host: Escherichia coli BL21(DE3) (bacteria) |
---|---|
#7: DNA chain | Mass: 19439.387 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Klebsiella pneumoniae (bacteria) / Gene: nifH / Production host: Escherichia coli BL21(DE3) (bacteria) |
-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
---|---|
EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
-Sample preparation
Component |
| ||||||||||||||||||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Source (natural) |
| ||||||||||||||||||||||||||||||
Source (recombinant) |
| ||||||||||||||||||||||||||||||
Buffer solution | pH: 8 | ||||||||||||||||||||||||||||||
Specimen | Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES | ||||||||||||||||||||||||||||||
Vitrification | Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 K |
-Electron microscopy imaging
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
---|---|
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 |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Image recording | Electron dose: 1.4 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k) |
-Processing
Software | Name: PHENIX / Version: 1.10.1_2155: / Classification: refinement | ||||||||||||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
EM software |
| ||||||||||||||||||||||||
CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||
Symmetry | Point symmetry: C1 (asymmetric) | ||||||||||||||||||||||||
3D reconstruction | Resolution: 3.8 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 80810 / Symmetry type: POINT | ||||||||||||||||||||||||
Atomic model building | Protocol: RIGID BODY FIT / Space: REAL | ||||||||||||||||||||||||
Refine LS restraints |
|