+Open data
-Basic information
Entry | Database: PDB / ID: 3j9b | ||||||
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Title | Electron cryo-microscopy of an RNA polymerase | ||||||
Components |
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Keywords | RNA BINDING PROTEIN/TRANSFERASE/RNA / Influenza RdRP / single particle reconstitution / replication / RNA BINDING PROTEIN-TRANSFERASE-RNA complex | ||||||
Function / homology | Function and homology information viral transcription / symbiont-mediated suppression of host mRNA transcription via inhibition of RNA polymerase II activity / cap snatching / 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 ...viral transcription / symbiont-mediated suppression of host mRNA transcription via inhibition of RNA polymerase II activity / cap snatching / 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 / identical protein binding / metal ion binding Similarity search - Function | ||||||
Biological species | Influenza A virus synthetic construct (others) | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4.3 Å | ||||||
Authors | Chang, S.H. / Sun, D.P. / Liang, H.H. / Wang, J. / Li, J. / Guo, L. / Wang, X.L. / Guan, C.C. / Boruah, B.M. / Yuan, L.M. ...Chang, S.H. / Sun, D.P. / Liang, H.H. / Wang, J. / Li, J. / Guo, L. / Wang, X.L. / Guan, C.C. / Boruah, B.M. / Yuan, L.M. / Feng, F. / Yang, M.R. / Wojdyla, J. / Wang, J.W. / Wang, M.T. / Wang, H.W. / Liu, Y.F. | ||||||
Citation | Journal: Mol Cell / Year: 2015 Title: Cryo-EM structure of influenza virus RNA polymerase complex at 4.3 Å resolution. Authors: Shenghai Chang / Dapeng Sun / Huanhuan Liang / Jia Wang / Jun Li / Lu Guo / Xiangli Wang / Chengcheng Guan / Bhargavi M Boruah / Lingmin Yuan / Feng Feng / Mingrui Yang / Lulan Wang / Yao ...Authors: Shenghai Chang / Dapeng Sun / Huanhuan Liang / Jia Wang / Jun Li / Lu Guo / Xiangli Wang / Chengcheng Guan / Bhargavi M Boruah / Lingmin Yuan / Feng Feng / Mingrui Yang / Lulan Wang / Yao Wang / Justyna Wojdyla / Lanjuan Li / Jiawei Wang / Meitian Wang / Genhong Cheng / Hong-Wei Wang / Yingfang Liu / Abstract: Replication and transcription of influenza virus genome mainly depend on its RNA-dependent RNA polymerase (RdRP), composed of the PA, PB1, and PB2 subunits. Although extensively studied, the ...Replication and transcription of influenza virus genome mainly depend on its RNA-dependent RNA polymerase (RdRP), composed of the PA, PB1, and PB2 subunits. Although extensively studied, the underlying mechanism of the RdRP complex is still unclear. Here we report the biochemical characterization of influenza RdRP subcomplex comprising PA, PB1, and N terminus of PB2, which exist as dimer in solution and can assemble into a tetramer state, regulated by vRNA promoter. Using single-particle cryo-electron microscopy, we have reconstructed the RdRP tetramer complex at 4.3 Å, highlighting the assembly and interfaces between monomers within the tetrameric structure. The individual RdRP subcomplex contains all the characterized motifs and appears as a cage-like structure. High-throughput mutagenesis profiling revealed that residues involved in the oligomer state formation are critical for viral life cycle. Our results lay a solid base for understanding the mechanism of replication of influenza and other negative-stranded RNA viruses. | ||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | Molecule: MolmilJmol/JSmol |
-Downloads & links
-Download
PDBx/mmCIF format | 3j9b.cif.gz | 311.5 KB | Display | PDBx/mmCIF format |
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PDB format | pdb3j9b.ent.gz | 245.5 KB | Display | PDB format |
PDBx/mmJSON format | 3j9b.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 3j9b_validation.pdf.gz | 810.4 KB | Display | wwPDB validaton report |
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Full document | 3j9b_full_validation.pdf.gz | 933.3 KB | Display | |
Data in XML | 3j9b_validation.xml.gz | 65.9 KB | Display | |
Data in CIF | 3j9b_validation.cif.gz | 101.3 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/j9/3j9b ftp://data.pdbj.org/pub/pdb/validation_reports/j9/3j9b | HTTPS FTP |
-Related structure data
Related structure data | 6202MC 6203C M: map data used to model this data C: citing same article (ref.) |
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Similar structure data |
-Links
-Assembly
Deposited unit |
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1 |
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-Components
-Protein , 3 types, 6 molecules AHBICJ
#1: Protein | Mass: 56930.676 Da / Num. of mol.: 2 / Fragment: UNP RESIDUES 203-716 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Influenza A virus / Strain: A/goose/Guangdong/3/1997(H5N1) / Gene: PA / Cell (production host): insect cell / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q91R78 #2: Protein | Mass: 38085.730 Da / Num. of mol.: 2 / Fragment: UNP RESIDUES 1-641 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Influenza A virus / Strain: A/Goose/Guangdong/1/1996 H5N1 genotype Gs/Gd / Gene: PB1 / Cell (production host): insect cell / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q9Q0V0, RNA-directed RNA polymerase #3: Protein | Mass: 5634.938 Da / Num. of mol.: 2 / Fragment: UNP RESIDUES 1-130 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Influenza A virus / Strain: A/Goose/Guangdong/1/1996 H5N1 genotype Gs/Gd / Gene: PB2 / Cell (production host): insect cell / Production host: Trichoplusia ni (cabbage looper) |
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-RNA chain , 3 types, 4 molecules DKEL
#4: RNA chain | Mass: 1930.277 Da / Num. of mol.: 2 / Source method: obtained synthetically / Source: (natural) synthetic construct (others) #5: RNA chain | | Mass: 1815.077 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (natural) synthetic construct (others) #6: RNA chain | | Mass: 1792.037 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (natural) synthetic construct (others) |
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-Details
Sequence details | THE AUTHOR KNOWS THE SEQUENCE OF CHAINS A/H, B/I AND C/J, BUT IS UNSURE OF THE COORDINATES ...THE AUTHOR KNOWS THE SEQUENCE OF CHAINS A/H, B/I AND C/J, BUT IS UNSURE OF THE COORDINATE |
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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: dimeric Influenza Virus RNA Polymerase / Type: COMPLEX |
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Specimen | Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Specimen support | Details: Quantifoil Cu R2/1 |
Vitrification | Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % Method: The grids were blotted for 6.5 s with 100% humidity and flash frozen in liquid ethane cooled at liquid nitrogen temperature using an FEI Vitrobot. |
-Electron microscopy imaging
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: FEI TITAN KRIOS / Date: Oct 2, 2014 |
Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM |
Electron lens | Mode: BRIGHT FIELD / Nominal magnification: 22500 X / Cs: 2.7 mm / Camera length: 0 mm |
Specimen holder | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Tilt angle max: 0 ° / Tilt angle min: 0 ° |
Image recording | Electron dose: 50 e/Å2 / Film or detector model: GATAN K2 (4k x 4k) |
-Processing
EM software |
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Symmetry | Point symmetry: C2 (2 fold cyclic) | ||||||||||||||||
3D reconstruction | Resolution: 4.3 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 67066 / Symmetry type: POINT | ||||||||||||||||
Atomic model building | Protocol: RIGID BODY FIT / Space: REAL / Details: REFINEMENT PROTOCOL--rigid body | ||||||||||||||||
Atomic model building | PDB-ID: 3CM8 Accession code: 3CM8 / Source name: PDB / Type: experimental model | ||||||||||||||||
Refinement step | Cycle: LAST
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