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- EMDB-17410: Vaccinia Virus palisade layer A10 trimer -

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Basic information

Entry
Database: EMDB / ID: EMD-17410
TitleVaccinia Virus palisade layer A10 trimer
Map dataFinal map after Phenix density modification of RELION-refined Vaccinia virus A10 trimer cryo-EM data.
Sample
  • Virus: Vaccinia virus Western Reserve
    • Protein or peptide: Core protein OPG136
KeywordsPoxvirus / Vaccinia Virus / core / cryo-electron tomography / AlphaFold / VIRAL PROTEIN
Function / homologyPoxvirus P4A / Poxvirus P4A protein / virion component / structural molecule activity / Major core protein OPG136 precursor
Function and homology information
Biological speciesVaccinia virus Western Reserve
Methodsingle particle reconstruction / cryo EM / Resolution: 3.8 Å
AuthorsDatler J / Hansen JM / Thader A / Schloegl A / Hodirnau VV / Schur FKM
Funding support1 items
OrganizationGrant numberCountry
Not funded
CitationJournal: Nat Struct Mol Biol / Year: 2024
Title: Multi-modal cryo-EM reveals trimers of protein A10 to form the palisade layer in poxvirus cores.
Authors: Julia Datler / Jesse M Hansen / Andreas Thader / Alois Schlögl / Lukas W Bauer / Victor-Valentin Hodirnau / Florian K M Schur /
Abstract: Poxviruses are among the largest double-stranded DNA viruses, with members such as variola virus, monkeypox virus and the vaccination strain vaccinia virus (VACV). Knowledge about the structural ...Poxviruses are among the largest double-stranded DNA viruses, with members such as variola virus, monkeypox virus and the vaccination strain vaccinia virus (VACV). Knowledge about the structural proteins that form the viral core has remained sparse. While major core proteins have been annotated via indirect experimental evidence, their structures have remained elusive and they could not be assigned to individual core features. Hence, which proteins constitute which layers of the core, such as the palisade layer and the inner core wall, has remained enigmatic. Here we show, using a multi-modal cryo-electron microscopy (cryo-EM) approach in combination with AlphaFold molecular modeling, that trimers formed by the cleavage product of VACV protein A10 are the key component of the palisade layer. This allows us to place previously obtained descriptions of protein interactions within the core wall into perspective and to provide a detailed model of poxvirus core architecture. Importantly, we show that interactions within A10 trimers are likely generalizable over members of orthopox- and parapoxviruses.
History
DepositionMay 22, 2023-
Header (metadata) releaseJan 17, 2024-
Map releaseJan 17, 2024-
UpdateFeb 21, 2024-
Current statusFeb 21, 2024Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

Downloads & links

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Map

FileDownload / File: emd_17410.map.gz / Format: CCP4 / Size: 125 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationFinal map after Phenix density modification of RELION-refined Vaccinia virus A10 trimer cryo-EM data.
Voxel sizeX=Y=Z: 1.06 Å
Density
Contour LevelBy AUTHOR: 0.9
Minimum - Maximum-12.96467 - 17.902683
Average (Standard dev.)0.0000007090825 (±0.20514202)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions320320320
Spacing320320320
CellA=B=C: 339.19998 Å
α=β=γ: 90.0 °

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Supplemental data

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Additional map: Final map after Phenix density modification of RELION-refined...

Fileemd_17410_additional_1.map
AnnotationFinal map after Phenix density modification of RELION-refined Vaccinia virus A10 trimer cryo-EM data.
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Half map: Final map after Phenix density modification of RELION-refined...

Fileemd_17410_half_map_1.map
AnnotationFinal map after Phenix density modification of RELION-refined Vaccinia virus A10 trimer cryo-EM data.
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Half map: 10A low-pass filtered map after Phenix density modification...

Fileemd_17410_half_map_2.map
Annotation10A low-pass filtered map after Phenix density modification of RELION-refined Vaccinia virus A10 trimer cryo-EM data.
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Sample components

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Entire : Vaccinia virus Western Reserve

EntireName: Vaccinia virus Western Reserve
Components
  • Virus: Vaccinia virus Western Reserve
    • Protein or peptide: Core protein OPG136

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Supramolecule #1: Vaccinia virus Western Reserve

SupramoleculeName: Vaccinia virus Western Reserve / type: virus / ID: 1 / Parent: 0 / Macromolecule list: all
Details: Purified from HeLa cells infected with Vaccinia Virus Western Reserve.
NCBI-ID: 696871 / Sci species name: Vaccinia virus Western Reserve / Virus type: VIRION / Virus isolate: STRAIN / Virus enveloped: No / Virus empty: No

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Macromolecule #1: Core protein OPG136

MacromoleculeName: Core protein OPG136 / type: protein_or_peptide / ID: 1 / Number of copies: 3 / Enantiomer: LEVO
Source (natural)Organism: Vaccinia virus Western Reserve
Molecular weightTheoretical: 69.068242 KDa
SequenceString: MMPIKSIVTL DQLEDSEYLF RIVSTVLPHL CLDYKVCDQL KTTFVHPFDI LLNNSLGSVT KQDELQAAIS KLGINYLIDT TSRELKLFN VTLNAGNIDI INTPINISSE TNPIINTHSF YDLPPFTQHL LNIRLTDTEY RARFIGGYIK PDGSDSMDVL A EKKYPDLN ...String:
MMPIKSIVTL DQLEDSEYLF RIVSTVLPHL CLDYKVCDQL KTTFVHPFDI LLNNSLGSVT KQDELQAAIS KLGINYLIDT TSRELKLFN VTLNAGNIDI INTPINISSE TNPIINTHSF YDLPPFTQHL LNIRLTDTEY RARFIGGYIK PDGSDSMDVL A EKKYPDLN FDNTYLFNIL YKDVINAPIK EFKAKIVNGV LSRQDFDNLI GVRQYITIQD RPRFDDAYNI ADAARHYGVN LN TLPLPNV DLTTMPTYKH LIMFEQYFIY TYDRVDIYYN GNKMLFDDEI INFTISMRYQ SLIPRLVDFF PDIPVNNNIV LHT RDPQNA AVNVTVALPN VQFVDINRNN KFFINFFNLL AKEQRSTAIK VTKSMFWDGM DYEEYKSKNL QDMMFINSTC YVFG LYNHN NTTYCSILSD IISAEKTPIR VCLLPRVVGG KTVTNLISET LKSISSMTIR EFPRKDKSIM HIGLSETGFM RFFQL LRLM ADKPHETAIK EVVMAYVGIK LGDKGSPYYI RKESYQDFIY LLFASMGFKV TTRRSIMGSN NISIISIRPR VTKQYI VAT LMKTSCSKNE AEKLITSAFD LLNFMVSVSD FRDYQS

UniProtKB: Major core protein OPG136 precursor

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

BufferpH: 9
Component:
ConcentrationFormulaName
210.0 mMKClPotassium chloride
1.0 mMTris-HClTrisTris hydrochloride

Details: diluted 1:1 with 0.25% Trypsin (final concentration 0.125%) and 1:1 4% paraformaldehyde (final concentration 2%) in 1 mM Tris-HCl.
GridModel: Quantifoil R2/2 / Material: COPPER / Mesh: 200 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 180 sec. / Pretreatment - Atmosphere: AIR
VitrificationCryogen name: ETHANE / Chamber humidity: 80 % / Chamber temperature: 277 K / Instrument: LEICA EM GP / Details: Leica GP2.
DetailsIsolated viral cores with some detached soluble monodispersed particles.

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 50.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 3.0 µm / Nominal defocus min: 1.25 µm / Nominal magnification: 81000
Specialist opticsEnergy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Digitization - Dimensions - Width: 5760 pixel / Digitization - Dimensions - Height: 4092 pixel / Number real images: 9264 / Average electron dose: 53.04 e/Å2 / Details: 34 frames total.
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: NONE / Details: ab initio without symmetry
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4.0)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 4.0)
Final reconstructionApplied symmetry - Point group: C3 (3 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 3.8 Å / Resolution method: OTHER / Software - Name: RELION (ver. 4.0) / Details: Phenix FSCref following density modification / Number images used: 24943
Detailsdose weighted and motion corrected

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Atomic model buiding 1

Initial modelChain - Source name: AlphaFold / Chain - Initial model type: in silico model / Details: version 2.3
DetailsRosetta relaxation
RefinementProtocol: OTHER
Output model

PDB-8p4k:
Vaccinia Virus palisade layer A10 trimer

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