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Yorodumi- PDB-6qcx: Crystal structure of influenza B polymerase initiation state with... -
+Open data
-Basic information
Entry | Database: PDB / ID: 6qcx | |||||||||
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Title | Crystal structure of influenza B polymerase initiation state with capped 15-mer RNA primer | |||||||||
Components |
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Keywords | VIRAL PROTEIN / RNA dependent RNA polymerase | |||||||||
Function / homology | Function and homology information viral transcription / symbiont-mediated suppression of host mRNA transcription via inhibition of RNA polymerase II activity / host cell mitochondrion / 7-methylguanosine mRNA capping / cap snatching / virion component / endonuclease activity / host cell cytoplasm / Hydrolases; Acting on ester bonds / RNA-directed RNA polymerase ...viral transcription / symbiont-mediated suppression of host mRNA transcription via inhibition of RNA polymerase II activity / host cell mitochondrion / 7-methylguanosine mRNA capping / cap snatching / virion component / endonuclease activity / host cell cytoplasm / Hydrolases; Acting on ester bonds / RNA-directed RNA polymerase / viral RNA genome replication / RNA-dependent RNA polymerase activity / DNA-templated transcription / nucleotide binding / host cell nucleus / RNA binding / metal ion binding Similarity search - Function | |||||||||
Biological species | Influenza B virus synthetic construct (others) | |||||||||
Method | X-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 3.08 Å | |||||||||
Authors | Cusack, S. / Drncova, P. | |||||||||
Funding support | European Union, France, 2items
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Citation | Journal: Nat Struct Mol Biol / Year: 2019 Title: Structural snapshots of actively transcribing influenza polymerase. Authors: Tomas Kouba / Petra Drncová / Stephen Cusack / Abstract: Influenza virus RNA-dependent RNA polymerase uses unique mechanisms to transcribe its single-stranded genomic viral RNA (vRNA) into messenger RNA. The polymerase is initially bound to a promoter ...Influenza virus RNA-dependent RNA polymerase uses unique mechanisms to transcribe its single-stranded genomic viral RNA (vRNA) into messenger RNA. The polymerase is initially bound to a promoter comprising the partially base-paired 3' and 5' extremities of the RNA. A short, capped primer, 'cap-snatched' from a nascent host polymerase II transcript, is directed towards the polymerase active site to initiate RNA synthesis. Here we present structural snapshots, as determined by X-ray crystallography and cryo-electron microscopy, of actively initiating influenza polymerase as it transitions towards processive elongation. Unexpected conformational changes unblock the active site cavity to allow establishment of a nine-base-pair template-product RNA duplex before the strands separate into distinct exit channels. Concomitantly, as the template translocates, the promoter base pairs are broken and the template entry region is remodeled. These structures reveal details of the influenza polymerase active site that will help optimize nucleoside analogs or other compounds that directly inhibit viral RNA synthesis. | |||||||||
History |
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-Structure visualization
Structure viewer | Molecule: MolmilJmol/JSmol |
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-Downloads & links
-Download
PDBx/mmCIF format | 6qcx.cif.gz | 961.8 KB | Display | PDBx/mmCIF format |
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PDB format | pdb6qcx.ent.gz | 795.7 KB | Display | PDB format |
PDBx/mmJSON format | 6qcx.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 6qcx_validation.pdf.gz | 518.1 KB | Display | wwPDB validaton report |
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Full document | 6qcx_full_validation.pdf.gz | 531.7 KB | Display | |
Data in XML | 6qcx_validation.xml.gz | 71 KB | Display | |
Data in CIF | 6qcx_validation.cif.gz | 98.7 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/qc/6qcx ftp://data.pdbj.org/pub/pdb/validation_reports/qc/6qcx | HTTPS FTP |
-Related structure data
Related structure data | 4511C 4512C 6qcsC 6qctC 6qcvC 6qcwC 5msgS C: citing same article (ref.) S: Starting model for refinement |
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Similar structure data |
-Links
-Assembly
Deposited unit |
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1 |
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Unit cell |
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-Components
-Protein , 3 types, 3 molecules ABC
#1: Protein | Mass: 85822.781 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Details: N-terminal His-tag C-terminal linker and TEV site / Source: (gene. exp.) Influenza B virus / Gene: PA / Production host: Trichoplusia ni (cabbage looper) References: UniProt: Q5V8Z9, Hydrolases; Acting on ester bonds |
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#2: Protein | Mass: 86207.016 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Details: N-terminal linker C-terminal linker and TEV site / Source: (gene. exp.) Influenza B virus / Gene: PB1 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q5V8Y6, RNA-directed RNA polymerase |
#3: Protein | Mass: 90844.109 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Details: N-terminal linker C-terminal STREP-tag and TEV site Source: (gene. exp.) Influenza B virus / Gene: PB2 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q5V8X3 |
-RNA chain , 3 types, 3 molecules MRV
#4: RNA chain | Mass: 5242.149 Da / Num. of mol.: 1 / Source method: obtained synthetically / Details: Capped 15-mer RNA / Source: (synth.) synthetic construct (others) |
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#5: RNA chain | Mass: 6525.820 Da / Num. of mol.: 1 / Source method: obtained synthetically / Details: 3' end vRNA promoter (extended) / Source: (synth.) synthetic construct (others) |
#6: RNA chain | Mass: 4557.820 Da / Num. of mol.: 1 / Source method: obtained synthetically / Details: 5' end vRNA promoter / Source: (synth.) synthetic construct (others) |
-Non-polymers , 3 types, 3 molecules
#7: Chemical | ChemComp-PO4 / |
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#8: Chemical | ChemComp-POP / |
#9: Chemical | ChemComp-MG / |
-Experimental details
-Experiment
Experiment | Method: X-RAY DIFFRACTION / Number of used crystals: 1 |
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-Sample preparation
Crystal | Density Matthews: 5.32 Å3/Da / Density % sol: 76.88 % |
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Crystal grow | Temperature: 277 K / Method: vapor diffusion, hanging drop / pH: 4.4 Details: Influenza B polymerase at 9 mg per ml in was mixed with 40 microM vRNA 5 prime end 14-mer, 40 microM vRNA 3 prime end 21-mer and 80 microM 15-mer capped RNA. The mother liquor contained 200 ...Details: Influenza B polymerase at 9 mg per ml in was mixed with 40 microM vRNA 5 prime end 14-mer, 40 microM vRNA 3 prime end 21-mer and 80 microM 15-mer capped RNA. The mother liquor contained 200 mM di-ammonium phosphate and 100 mM sodium acetate between pH 4.0 and 4.4. PH range: 4.0-4.4 |
-Data collection
Diffraction | Mean temperature: 100 K / Serial crystal experiment: N |
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Diffraction source | Source: SYNCHROTRON / Site: ESRF / Beamline: MASSIF-1 / Wavelength: 0.966 Å |
Detector | Type: DECTRIS PILATUS 6M-F / Detector: PIXEL / Date: May 29, 2018 |
Radiation | Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray |
Radiation wavelength | Wavelength: 0.966 Å / Relative weight: 1 |
Reflection | Resolution: 3.08→50 Å / Num. obs: 83143 / % possible obs: 75.2 % / Redundancy: 4.8 % / CC1/2: 0.999 / Rrim(I) all: 0.086 / Rsym value: 0.076 / Net I/σ(I): 14.3 |
Reflection shell | Resolution: 3.08→3.27 Å / Mean I/σ(I) obs: 1.7 / CC1/2: 0.54 / Rrim(I) all: 0.888 / Rsym value: 0.756 / % possible all: 61 |
-Processing
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Refinement | Method to determine structure: MOLECULAR REPLACEMENT Starting model: 5MSG Resolution: 3.08→173.56 Å / Cor.coef. Fo:Fc: 0.865 / Cor.coef. Fo:Fc free: 0.93 / SU B: 45.732 / SU ML: 0.324 / Cross valid method: THROUGHOUT / ESU R: 1.37 / ESU R Free: 0.376 / Stereochemistry target values: MAXIMUM LIKELIHOOD / Details: HYDROGENS HAVE BEEN ADDED IN THE RIDING POSITIONS
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Solvent computation | Ion probe radii: 0.8 Å / Shrinkage radii: 0.8 Å / VDW probe radii: 1.2 Å / Solvent model: MASK | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Displacement parameters | Biso mean: 114.342 Å2
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Refinement step | Cycle: 1 / Resolution: 3.08→173.56 Å
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