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- PDB-8c4h: CryoEM structure of the Hendra henipavirus nucleocapsid sauronoid... -

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Basic information

Entry
Database: PDB / ID: 8c4h
TitleCryoEM structure of the Hendra henipavirus nucleocapsid sauronoid assembly multimer
Components
  • Nucleocapsid
  • RNA (84-MER)
KeywordsVIRAL PROTEIN / Nucleoprotein / RNA-binding protein / Sauronoid
Function / homology
Function and homology information


helical viral capsid / viral nucleocapsid / host cell cytoplasm / ribonucleoprotein complex / structural molecule activity / RNA binding
Similarity search - Function
Paramyxovirus nucleocapsid protein / Paramyxovirus nucleocapsid protein
Similarity search - Domain/homology
RNA / RNA (> 10) / Nucleocapsid
Similarity search - Component
Biological speciesHendra henipavirus
Escherichia coli BL21 (bacteria)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.485 Å
AuthorsPasschier, T.C. / Maskell, D.P. / Edwards, T.A. / Barr, J.N.
Funding supportEuropean Union, 1items
OrganizationGrant numberCountry
H2020 Marie Curie Actions of the European Commission721367European Union
CitationJournal: Sci Rep / Year: 2024
Title: The cryoEM structure of the Hendra henipavirus nucleoprotein reveals insights into paramyxoviral nucleocapsid architectures.
Authors: Tim C Passchier / Joshua B R White / Daniel P Maskell / Matthew J Byrne / Neil A Ranson / Thomas A Edwards / John N Barr /
Abstract: We report the first cryoEM structure of the Hendra henipavirus nucleoprotein in complex with RNA, at 3.5 Å resolution, derived from single particle analysis of a double homotetradecameric RNA-bound ...We report the first cryoEM structure of the Hendra henipavirus nucleoprotein in complex with RNA, at 3.5 Å resolution, derived from single particle analysis of a double homotetradecameric RNA-bound N protein ring assembly exhibiting D14 symmetry. The structure of the HeV N protein adopts the common bi-lobed paramyxoviral N protein fold; the N-terminal and C-terminal globular domains are bisected by an RNA binding cleft containing six RNA nucleotides and are flanked by the N-terminal and C-terminal arms, respectively. In common with other paramyxoviral nucleocapsids, the lateral interface between adjacent N and N protomers involves electrostatic and hydrophobic interactions mediated primarily through the N-terminal arm and globular domains with minor contribution from the C-terminal arm. However, the HeV N multimeric assembly uniquely identifies an additional protomer-protomer contact between the N N-terminus and N C-terminal arm linker. The model presented here broadens the understanding of RNA-bound paramyxoviral nucleocapsid architectures and provides a platform for further insight into the molecular biology of HeV, as well as the development of antiviral interventions.
History
DepositionJan 4, 2023Deposition site: PDBE / Processing site: PDBE
Revision 1.0Jun 26, 2024Provider: repository / Type: Initial release

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
C: Nucleocapsid
D: Nucleocapsid
E: Nucleocapsid
F: Nucleocapsid
G: Nucleocapsid
H: Nucleocapsid
I: Nucleocapsid
J: Nucleocapsid
K: Nucleocapsid
L: Nucleocapsid
M: Nucleocapsid
N: Nucleocapsid
A: Nucleocapsid
1: RNA (84-MER)
B: Nucleocapsid
a: Nucleocapsid
2: RNA (84-MER)
b: Nucleocapsid
c: Nucleocapsid
d: Nucleocapsid
e: Nucleocapsid
f: Nucleocapsid
g: Nucleocapsid
h: Nucleocapsid
i: Nucleocapsid
j: Nucleocapsid
k: Nucleocapsid
l: Nucleocapsid
m: Nucleocapsid
n: Nucleocapsid


Theoretical massNumber of molelcules
Total (without water)1,690,50330
Polymers1,690,50330
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein ...
Nucleocapsid


Mass: 58541.320 Da / Num. of mol.: 28
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Hendra henipavirus / Plasmid: pET28a / Production host: Escherichia coli BL21(DE3) (bacteria) / Strain (production host): Rosetta2 / References: UniProt: A0A1L7B858
#2: RNA chain RNA (84-MER)


Mass: 25672.984 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli BL21(DE3) (bacteria) / Strain: Rosetta 2 / Production host: Escherichia coli BL21(DE3) (bacteria) / Strain (production host): Rosetta 2

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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: Sauronoid assembly of Hendra henipavirus nucleoprotein
Type: COMPLEX
Details: Recombinantly expressed Hendra henipavirus nucleoprotein forming sauronoid complexes.
Entity ID: all / Source: RECOMBINANT
Molecular weightValue: 1.691779 MDa / Experimental value: NO
Source (natural)Organism: Hendra henipavirus
Source (recombinant)Organism: Escherichia coli BL21(DE3) (bacteria) / Strain: Rosetta 2 / Plasmid: pET28a
Details of virusIsolate: SPECIES / Type: VIRION
Buffer solutionpH: 7.5
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: COPPER / Grid type: PELCO Ultrathin Carbon with Lacey Carbon
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 95 % / Chamber temperature: 281 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 magnification: 75000 X / Nominal defocus max: 2800 nm / Nominal defocus min: 700 nm
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingAverage exposure time: 1.5 sec. / Electron dose: 59.7 e/Å2 / Detector mode: INTEGRATING / Film or detector model: FEI FALCON III (4k x 4k) / Num. of grids imaged: 1 / Num. of real images: 3548

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Processing

EM software
IDNameVersionCategory
1RELION3.1particle selection
2EPUimage acquisition
4Gctf1.18CTF correction
10RELION3.1initial Euler assignment
11RELION3.1final Euler assignment
12RELION3.1classification
13RELION3.13D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: D14 (2x14 fold dihedral)
3D reconstructionResolution: 3.485 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 5683 / Num. of class averages: 1 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00289570
ELECTRON MICROSCOPYf_angle_d0.455121792
ELECTRON MICROSCOPYf_dihedral_angle_d6.03614022
ELECTRON MICROSCOPYf_chiral_restr0.03614112
ELECTRON MICROSCOPYf_plane_restr0.00415036

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