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- EMDB-19599: Structural characterization of Thogoto Virus nucleoprotein provid... -

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Basic information

Entry
Database: EMDB / ID: EMD-19599
TitleStructural characterization of Thogoto Virus nucleoprotein provides insights into RNA encapsidation and assembly
Map datafiltered to 18A, non-sharpened
Sample
  • Complex: Ribonucleoprotein (RNP) complexes of Thogoto virus
    • Other: THOV RNP
KeywordsViral replication / nucleoprotein / RNA binding / oligomerization / orthomyxovirus / VIRAL PROTEIN
Function / homologyhelical viral capsid / viral penetration into host nucleus / host cell / viral nucleocapsid / ribonucleoprotein complex / symbiont entry into host cell / host cell nucleus / RNA binding / Nucleoprotein
Function and homology information
Biological speciesThogotovirus
Methodsubtomogram averaging / cryo EM / Resolution: 18.0 Å
AuthorsRoske Y / Mikirtumov V / Daumke O / Kudryashev M / Dick A
Funding support Germany, 1 items
OrganizationGrant numberCountry
German Research Foundation (DFG)DA 1127/1-2 and SFB958/A12 Germany
CitationJournal: Structure / Year: 2024
Title: Structural characterization of Thogoto Virus nucleoprotein provides insights into viral RNA encapsidation and RNP assembly.
Authors: Alexej Dick / Vasilii Mikirtumov / Jonas Fuchs / Ferdinand Krupp / Daniel Olal / Elias Bendl / Thiemo Sprink / Christoph Diebolder / Mikhail Kudryashev / Georg Kochs / Yvette Roske / Oliver Daumke /
Abstract: Orthomyxoviruses, such as influenza and thogotoviruses, are important human and animal pathogens. Their segmented viral RNA genomes are wrapped by viral nucleoproteins (NPs) into helical ...Orthomyxoviruses, such as influenza and thogotoviruses, are important human and animal pathogens. Their segmented viral RNA genomes are wrapped by viral nucleoproteins (NPs) into helical ribonucleoprotein complexes (RNPs). NP structures of several influenza viruses have been reported. However, there are still contradictory models of how orthomyxovirus RNPs are assembled. Here, we characterize the crystal structure of Thogoto virus (THOV) NP and found striking similarities to structures of influenza viral NPs, including a two-lobed domain architecture, a positively charged RNA-binding cleft, and a tail loop important for trimerization and viral transcription. A low-resolution cryo-electron tomography reconstruction of THOV RNPs elucidates a left-handed double helical assembly. By providing a model for RNP assembly of THOV, our study suggests conserved NP assembly and RNA encapsidation modes for thogoto- and influenza viruses.
History
DepositionFeb 9, 2024-
Header (metadata) releaseJun 19, 2024-
Map releaseJun 19, 2024-
UpdateJun 19, 2024-
Current statusJun 19, 2024Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_19599.png

Downloads & links

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Map

FileDownload / File: emd_19599.map.gz / Format: CCP4 / Size: 27 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationfiltered to 18A, non-sharpened
Voxel sizeX=Y=Z: 2.6 Å
Density
Contour LevelBy AUTHOR: 0.00732
Minimum - Maximum-0.0021238325 - 0.013839871
Average (Standard dev.)0.00031548986 (±0.0014727503)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions192192192
Spacing192192192
CellA=B=C: 499.19998 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_19599_msk_1.map
Projections & Slices
AxesZYX

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Half map: #1

Fileemd_19599_half_map_1.map
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Half map: #2

Fileemd_19599_half_map_2.map
Projections & Slices
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Sample components

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Entire : Ribonucleoprotein (RNP) complexes of Thogoto virus

EntireName: Ribonucleoprotein (RNP) complexes of Thogoto virus
Components
  • Complex: Ribonucleoprotein (RNP) complexes of Thogoto virus
    • Other: THOV RNP

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Supramolecule #1: Ribonucleoprotein (RNP) complexes of Thogoto virus

SupramoleculeName: Ribonucleoprotein (RNP) complexes of Thogoto virus / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Thogotovirus

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Macromolecule #1: THOV RNP

MacromoleculeName: THOV RNP / type: other / ID: 1 / Details: Nucleoprotein / Classification: other
Source (natural)Organism: Thogotovirus / Strain: THOV strain SiAr 126
SequenceString: MATDQMDISG PPPKKQHVDT ESQIPKMYEM IRDQMRTLAS THKIPLNIDH NCEVIGSIIM AACTNNRDLR PVDKYWFLMG PAGAEVMTEV EIDIQPQLQW AKGAVHDPKY KGQWYPFLAL LQISNKTKDT ILWQKYPVTQ ELEISNSLEI YANGHGIKDR LKNSRPRSVG ...String:
MATDQMDISG PPPKKQHVDT ESQIPKMYEM IRDQMRTLAS THKIPLNIDH NCEVIGSIIM AACTNNRDLR PVDKYWFLMG PAGAEVMTEV EIDIQPQLQW AKGAVHDPKY KGQWYPFLAL LQISNKTKDT ILWQKYPVTQ ELEISNSLEI YANGHGIKDR LKNSRPRSVG PLVHLLHLKR LQENPPKSPA VNGIRKSIVG HLKRQCIGET QKAMINQFEM GRWESLSTFA ASLLAIKPRI ENHFVLTYPL IANCEDFAGA TLSDEWVFKA MEKISNKKTL RVCGPDEKWI SFMNQIYIHS VFQTTGEDLG VLEWVFGGRF CQRKEFGRYC KKSQTKVIGL FTFQYEYWSK PLKSAPRSIE GSKRGQISCR PSFKGKRPSY NNFTSIDALQ SASGSQTVSF YDQVREECQK YMDLKVEGTT CFYRKGGHVE VEFPGSAHCN TYLFG
Recombinant expressionOrganism: Mesocricetus auratus (golden hamster)

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Experimental details

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Structure determination

Methodcryo EM
Processingsubtomogram averaging
Aggregation statefilament

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Sample preparation

BufferpH: 7
GridModel: Quantifoil R1.2/1.3 / Material: GOLD / Mesh: 300 / Support film - Material: GOLD / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE
VitrificationCryogen name: ETHANE / Chamber humidity: 95 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Digitization - Dimensions - Width: 5760 pixel / Digitization - Dimensions - Height: 4092 pixel / Number real images: 1 / Average exposure time: 1.2 sec. / Average electron dose: 3.5 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsCalibrated defocus max: 5.0 µm / Calibrated defocus min: 3.0 µm / Calibrated magnification: 33000 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 5.0 µm / Nominal defocus min: 3.0 µm / Nominal magnification: 33000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Final reconstructionNumber classes used: 1
Applied symmetry - Helical parameters - Δz: 24.6 Å
Applied symmetry - Helical parameters - Δ&Phi: -55.5 °
Applied symmetry - Helical parameters - Axial symmetry: C1 (asymmetric)
Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 18.0 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 4) / Number subtomograms used: 3128
ExtractionNumber tomograms: 33 / Number images used: 16101
Method: template matching with manual annotation on untilted images
Software: (Name: RELION (ver. 4), Dynamo)
Final 3D classificationNumber classes: 8 / Avg.num./class: 2000 / Software - Name: RELION (ver. 4)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 4)
FSC plot (resolution estimation)

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