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- PDB-8snx: Cryo-EM structure of the respiratory syncytial virus polymerase (... -

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Basic information

Entry
Database: PDB / ID: 8snx
TitleCryo-EM structure of the respiratory syncytial virus polymerase (L:P) bound to the leader promoter
Components
  • Phosphoprotein
  • RNA (5'-R(*UP*UP*UP*UP*UP*CP*GP*CP*GP*U)-3')
  • RNA-directed RNA polymerase L
KeywordsVIRAL PROTEIN/RNA / Respiratory syncytial virus / RNA-dependent RNA polymerase / VIRAL PROTEIN-RNA complex
Function / homology
Function and homology information


NNS virus cap methyltransferase / Respiratory syncytial virus genome transcription / GDP polyribonucleotidyltransferase / Translation of respiratory syncytial virus mRNAs / negative stranded viral RNA replication / Respiratory syncytial virus genome replication / RSV-host interactions / Assembly and release of respiratory syncytial virus (RSV) virions / Maturation of hRSV A proteins / Hydrolases; Acting on acid anhydrides; In phosphorus-containing anhydrides ...NNS virus cap methyltransferase / Respiratory syncytial virus genome transcription / GDP polyribonucleotidyltransferase / Translation of respiratory syncytial virus mRNAs / negative stranded viral RNA replication / Respiratory syncytial virus genome replication / RSV-host interactions / Assembly and release of respiratory syncytial virus (RSV) virions / Maturation of hRSV A proteins / Hydrolases; Acting on acid anhydrides; In phosphorus-containing anhydrides / Respiratory syncytial virus (RSV) attachment and entry / virion component / mRNA 5'-cap (guanine-N7-)-methyltransferase activity / host cell cytoplasm / RNA-directed RNA polymerase / RNA-dependent RNA polymerase activity / GTPase activity / ATP binding / metal ion binding
Similarity search - Function
Phosphoprotein, pneumoviral / Pneumovirus phosphoprotein / RNA-directed RNA polymerase L methyltransferase domain, rhabdovirus / Virus-capping methyltransferase, MT domain / RNA-directed RNA polymerase, paramyxovirus / Mononegavirales RNA-directed RNA polymerase catalytic domain / Mononegavirus L protein 2-O-ribose methyltransferase / Mononegavirales mRNA-capping domain V / RNA-directed RNA polymerase L, C-terminal / Mononegavirales RNA dependent RNA polymerase ...Phosphoprotein, pneumoviral / Pneumovirus phosphoprotein / RNA-directed RNA polymerase L methyltransferase domain, rhabdovirus / Virus-capping methyltransferase, MT domain / RNA-directed RNA polymerase, paramyxovirus / Mononegavirales RNA-directed RNA polymerase catalytic domain / Mononegavirus L protein 2-O-ribose methyltransferase / Mononegavirales mRNA-capping domain V / RNA-directed RNA polymerase L, C-terminal / Mononegavirales RNA dependent RNA polymerase / Mononegavirales mRNA-capping region V / RdRp of negative ssRNA viruses with non-segmented genomes catalytic domain profile. / Mononegavirus L protein 2'-O-ribose methyltransferase domain profile.
Similarity search - Domain/homology
RNA / Phosphoprotein / RNA-directed RNA polymerase L
Similarity search - Component
Biological speciesRespiratory syncytial virus A2
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.4 Å
AuthorsCao, D. / Gao, Y. / Chen, Z. / Gooneratne, I. / Roesler, C. / Mera, C. / Liang, B.
Funding support United States, 5items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R01GM130950 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)U24GM116788 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)U24GM116789 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)U24GM129539 United States
Simons FoundationSF349247 United States
CitationJournal: Nature / Year: 2024
Title: Structures of the promoter-bound respiratory syncytial virus polymerase.
Authors: Dongdong Cao / Yunrong Gao / Zhenhang Chen / Inesh Gooneratne / Claire Roesler / Cristopher Mera / Paul D'Cunha / Anna Antonova / Deepak Katta / Sarah Romanelli / Qi Wang / Samantha Rice / ...Authors: Dongdong Cao / Yunrong Gao / Zhenhang Chen / Inesh Gooneratne / Claire Roesler / Cristopher Mera / Paul D'Cunha / Anna Antonova / Deepak Katta / Sarah Romanelli / Qi Wang / Samantha Rice / Wesley Lemons / Anita Ramanathan / Bo Liang /
Abstract: The respiratory syncytial virus (RSV) polymerase is a multifunctional RNA-dependent RNA polymerase composed of the large (L) protein and the phosphoprotein (P). It transcribes the RNA genome into ten ...The respiratory syncytial virus (RSV) polymerase is a multifunctional RNA-dependent RNA polymerase composed of the large (L) protein and the phosphoprotein (P). It transcribes the RNA genome into ten viral mRNAs and replicates full-length viral genomic and antigenomic RNAs. The RSV polymerase initiates RNA synthesis by binding to the conserved 3'-terminal RNA promoters of the genome or antigenome. However, the lack of a structure of the RSV polymerase bound to the RNA promoter has impeded the mechanistic understanding of RSV RNA synthesis. Here we report cryogenic electron microscopy structures of the RSV polymerase bound to its genomic and antigenomic viral RNA promoters, representing two of the first structures of an RNA-dependent RNA polymerase in complex with its RNA promoters in non-segmented negative-sense RNA viruses. The overall structures of the promoter-bound RSV polymerases are similar to that of the unbound (apo) polymerase. Our structures illustrate the interactions between the RSV polymerase and the RNA promoters and provide the structural basis for the initiation of RNA synthesis at positions 1 and 3 of the RSV promoters. These structures offer a deeper understanding of the pre-initiation state of the RSV polymerase and could aid in antiviral research against RSV.
History
DepositionApr 28, 2023Deposition site: RCSB / Processing site: RCSB
Revision 1.0Dec 20, 2023Provider: repository / Type: Initial release
Revision 1.1Jan 3, 2024Group: Database references / Category: citation / citation_author / Item: _citation.pdbx_database_id_PubMed / _citation.title
Revision 1.2Jan 31, 2024Group: Database references / Category: citation
Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation.year

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: RNA-directed RNA polymerase L
B: Phosphoprotein
C: Phosphoprotein
D: Phosphoprotein
E: Phosphoprotein
T: RNA (5'-R(*UP*UP*UP*UP*UP*CP*GP*CP*GP*U)-3')


Theoretical massNumber of molelcules
Total (without water)362,4616
Polymers362,4616
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author&software
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1

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Components

#1: Protein RNA-directed RNA polymerase L / Protein L / Large structural protein / Replicase / Transcriptase


Mass: 250704.484 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Respiratory syncytial virus A2 / Production host: Spodoptera frugiperda (fall armyworm)
References: UniProt: P28887, RNA-directed RNA polymerase, Hydrolases; Acting on acid anhydrides; In phosphorus-containing anhydrides, GDP polyribonucleotidyltransferase, NNS virus cap methyltransferase
#2: Protein
Phosphoprotein


Mass: 27165.838 Da / Num. of mol.: 4
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Respiratory syncytial virus A2 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: G3C7Q7
#3: RNA chain RNA (5'-R(*UP*UP*UP*UP*UP*CP*GP*CP*GP*U)-3')


Mass: 3092.812 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Respiratory syncytial virus A2

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: The respiratory syncytial virus polymerase (L:P) in complex with the leader promoter
Type: COMPLEX / Entity ID: all / Source: MULTIPLE SOURCES
Molecular weightExperimental value: NO
Source (natural)Organism: Respiratory syncytial virus A2
Source (recombinant)Organism: Spodoptera frugiperda (fall armyworm)
Buffer solutionpH: 7.4
SpecimenConc.: 0.35 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid type: UltrAuFoil R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 293.15 K

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 2500 nm / Nominal defocus min: 800 nm
Image recordingElectron dose: 51.11 e/Å2 / Film or detector model: GATAN K3 (6k x 4k)

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Processing

CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 3.4 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 358385 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00314093
ELECTRON MICROSCOPYf_angle_d0.56919087
ELECTRON MICROSCOPYf_dihedral_angle_d14.6475373
ELECTRON MICROSCOPYf_chiral_restr0.042216
ELECTRON MICROSCOPYf_plane_restr0.0042374

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