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- PDB-8y9j: Structure of the Ebola virus nucleocapsid subunit -

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Basic information

Entry
Database: PDB / ID: 8y9j
TitleStructure of the Ebola virus nucleocapsid subunit
Components
  • Membrane-associated protein VP24
  • Nucleoprotein
  • RNA (12-MER)
KeywordsVIRAL PROTEIN/RNA / Complex / VIRAL PROTEIN / VIRAL PROTEIN-RNA complex
Function / homology
Function and homology information


host cell endomembrane system / viral RNA genome packaging / helical viral capsid / symbiont-mediated suppression of host JAK-STAT cascade via inhibition of STAT1 activity / viral budding from plasma membrane / viral nucleocapsid / host cell cytoplasm / symbiont-mediated suppression of host innate immune response / symbiont-mediated suppression of host type I interferon-mediated signaling pathway / ribonucleoprotein complex ...host cell endomembrane system / viral RNA genome packaging / helical viral capsid / symbiont-mediated suppression of host JAK-STAT cascade via inhibition of STAT1 activity / viral budding from plasma membrane / viral nucleocapsid / host cell cytoplasm / symbiont-mediated suppression of host innate immune response / symbiont-mediated suppression of host type I interferon-mediated signaling pathway / ribonucleoprotein complex / host cell plasma membrane / virion membrane / structural molecule activity / RNA binding / membrane
Similarity search - Function
Filovirus membrane-associated VP24 / Filovirus membrane-associated protein VP24 / Ebola nucleoprotein / Ebola nucleoprotein
Similarity search - Domain/homology
RNA / RNA (> 10) / Nucleoprotein / Membrane-associated protein VP24
Similarity search - Component
Biological speciesZaire ebolavirus
Homo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4.6 Å
AuthorsFujita-Fujiharu, Y. / Hu, S. / Hirabayashi, A. / Takamatsu, Y. / Ng, Y. / Houri, K. / Muramoto, Y. / Nakano, M. / Sugita, Y. / Noda, T.
Funding support Japan, 14items
OrganizationGrant numberCountry
Japan Society for the Promotion of Science (JSPS)21J12207 Japan
Japan Society for the Promotion of Science (JSPS)22KK0115 Japan
Japan Society for the Promotion of Science (JSPS)21K07059 Japan
Japan Society for the Promotion of Science (JSPS)22K19431 Japan
Japan Society for the Promotion of Science (JSPS)22KK0115 Japan
Japan Society for the Promotion of Science (JSPS)21K07052 Japan
Japan Science and TechnologyJPMJFR214S Japan
Japan Science and TechnologyJPMJCR20HA Japan
Japan Agency for Medical Research and Development (AMED)JP23fm0208101 Japan
Japan Agency for Medical Research and Development (AMED)JP22fm0208101 Japan
Japan Agency for Medical Research and Development (AMED)22wm0325023h9903 Japan
Japan Agency for Medical Research and Development (AMED)JP23fm0208101 Japan
Japan Agency for Medical Research and Development (AMED)22fk0108552h0001 Japan
Japan Agency for Medical Research and Development (AMED)JP23ama121001 Japan
CitationJournal: Nat Commun / Year: 2025
Title: Structural basis for Ebola virus nucleocapsid assembly and function regulated by VP24.
Authors: Yoko Fujita-Fujiharu / Shangfan Hu / Ai Hirabayashi / Yuki Takamatsu / Yen Ni Ng / Kazuya Houri / Yukiko Muramoto / Masahiro Nakano / Yukihiko Sugita / Takeshi Noda /
Abstract: The Ebola virus, a member of the Filoviridae family, causes severe hemorrhagic fever in humans. Filamentous virions contain a helical nucleocapsid responsible for genome transcription, replication, ...The Ebola virus, a member of the Filoviridae family, causes severe hemorrhagic fever in humans. Filamentous virions contain a helical nucleocapsid responsible for genome transcription, replication, and packaging into progeny virions. The nucleocapsid consists of a helical nucleoprotein (NP)-viral genomic RNA complex forming the core structure, to which VP24 and VP35 bind externally. Two NPs, each paired with a VP24 molecule, constitute a repeating unit. However, the detailed nucleocapsid structure remains unclear. Here, we determine the nucleocapsid-like structure within virus-like particles at 4.6 Å resolution using single-particle cryo-electron microscopy. Mutational analysis identifies specific interactions between the two NPs and two VP24s and demonstrates that each of the two VP24s in different orientations distinctively regulates nucleocapsid assembly, viral RNA synthesis, intracellular transport of the nucleocapsid, and infectious virion production. Our findings highlight the sophisticated mechanisms underlying the assembly and functional regulation of the nucleocapsid and provide insights into antiviral development.
History
DepositionFeb 6, 2024Deposition site: PDBJ / Processing site: PDBJ
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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Nucleoprotein
B: Nucleoprotein
C: Membrane-associated protein VP24
D: Membrane-associated protein VP24
K: RNA (12-MER)


Theoretical massNumber of molelcules
Total (without water)226,9065
Polymers226,9065
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein Nucleoprotein / Ebola NP / eNP / Nucleocapsid protein / Protein N


Mass: 83387.500 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Zaire ebolavirus / Gene: NP / Plasmid: pCAGGS / Cell line (production host): HEK293T / Organ (production host): Kidney / Production host: Homo sapiens (human) / References: UniProt: P18272
#2: Protein Membrane-associated protein VP24 / Ebola VP24 / eVP24


Mass: 28250.811 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Zaire ebolavirus / Gene: VP24 / Cell line (production host): HEK293T / Organ (production host): Kidney / Production host: Homo sapiens (human) / References: UniProt: Q05322
#3: RNA chain RNA (12-MER)


Mass: 3629.032 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / Cell line: HEK293T / Organ: Kidney
Has protein modificationN

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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: Zaire ebolavirus / Type: VIRUS / Entity ID: all / Source: MULTIPLE SOURCES
Source (natural)Organism: Zaire ebolavirus
Source (recombinant)Organism: Homo sapiens (human) / Cell: HEK293T / Plasmid: pCAGGS
Details of virusEmpty: NO / Enveloped: YES / Isolate: STRAIN / Type: VIRUS-LIKE PARTICLE
Buffer solutionpH: 8
Buffer component
IDConc.NameFormulaBuffer-ID
110 mMtris hydroxymethyl aminomethane hydrochlorideTris-HCl1
2150 mMsodium chlorideNaCl1
31 mMethylenediaminetetraacetic acidEDTA1
SpecimenConc.: 1 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Details: The total 2.5 microlitre sample was applied to both sides of the glow-discharged grid.
Specimen supportGrid material: COPPER / Grid mesh size: 400 divisions/in. / Grid type: Quantifoil R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 281.15 K
Details: The sample was applied to both sides of the grid. The grids were blotted for 14 seconds.

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: TFS KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal magnification: 81000 X / Nominal defocus max: 2000 nm / Nominal defocus min: 600 nm
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingElectron dose: 60 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Num. of grids imaged: 1 / Num. of real images: 5897
EM imaging opticsEnergyfilter name: GIF Bioquantum / Energyfilter slit width: 20 eV
Spherical aberration corrector: Microscope was modified with a Cs corrector (CEOS GmbH)

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Processing

EM software
IDNameVersionCategory
1RELION4.0-betaparticle selection
2SerialEM4image acquisition
4CTFFIND4.1.14CTF correction
7UCSF ChimeraX1.15model fitting
9RELION4.0-betainitial Euler assignment
10cryoSPARC3.3.1final Euler assignment
12cryoSPARC3.3.13D reconstruction
13RosettaEMmodel refinement
20PHENIXmodel refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 263789
3D reconstructionResolution: 4.6 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 204348 / Symmetry type: POINT
Atomic model buildingProtocol: FLEXIBLE FIT
Atomic model building
IDPDB-ID 3D fitting-IDAccession codeInitial refinement model-IDSource nameType
16EHM

6ehm

16EHM1PDBexperimental model
25Z9W

5z9w

15Z9W2PDBexperimental model
34M0Q

4m0q

14M0Q3PDBexperimental model

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