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- EMDB-57884: PLD-fold vaccinia virus endonuclease K4 with DNA -

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Basic information

Entry
Database: EMDB / ID: EMD-57884
TitlePLD-fold vaccinia virus endonuclease K4 with DNA
Map data
Sample
  • Complex: Octameric complex of K4 containing mutated histidines in the HKD motif, with DNA bound in chains A and D
    • Protein or peptide: Virion nicking-joining enzyme
    • DNA: 19mer-ssDNA-primer
KeywordsNuclease / PLD / Phospholipase D / vaccinia virus / poxvirus / DNase / ssDNA / VIRAL PROTEIN
Function / homology
Function and homology information


virion component / endonuclease activity / hydrolase activity
Similarity search - Function
Phospholipase D / PLD-like domain / PLD3/4, PLD-like domain / : / Phospholipase D. Active site motifs. / Phospholipase D/Transphosphatidylase / Phospholipase D phosphodiesterase active site profile.
Similarity search - Domain/homology
Virion nicking-joining enzyme
Similarity search - Component
Biological speciesVaccinia virus Copenhagen / Orthopoxvirus vaccinia / synthetic construct (others)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.6 Å
AuthorsGroger H / Burmeister WP / Tarbouriech N
Funding support France, 1 items
OrganizationGrant numberCountry
Agence Nationale de la Recherche (ANR)ANR-22-CE11-0007-01 France
CitationJournal: J Mol Biol / Year: 2026
Title: High-resolution Structure of the Vaccinia Virus Phospholipase D-fold Endonuclease K4.
Authors: Henri Gröger / Candice Trouba / Jade Barbaste / Guillaume Lacroix / Guy Schoehn / Wim P Burmeister / Nicolas Tarbouriech /
Abstract: Vaccinia virus (VACV) is an orthopoxvirus closely related to mpox virus, which started global outbreaks in 2022. Poxvirus genomes are flanked by short, inverted complementary hairpin telomeres that ...Vaccinia virus (VACV) is an orthopoxvirus closely related to mpox virus, which started global outbreaks in 2022. Poxvirus genomes are flanked by short, inverted complementary hairpin telomeres that feature mismatched bases and insertions essential for viral replication. In this context, a role of the late protein K4 has been proposed. K4 is present in the virion, is apparently non-essential and has a phospholipase D (PLD)-fold as has VACV F13 protein. It also shares fold and nuclease activity with its closest homologue, mammalian PLD3. We established an endonuclease activity against ssDNA and hairpin loops and bubbles in a dsDNA context while RNA is resistant to cleavage. The 2.4 Å cryo-EM structure of K4 shows an unusual octameric assembly, also present in solution. At low concentration, tetramers and dimers similar to the one of hPLD3 are also present. Despite its nuclease activity, in K4 a C-terminal extension blocks the DNA binding pockets. Using an inactive mutant, fortuitously, a DNA 19mer bound simultaneously to 2 sites of the octamer where it displaced the C-termini. DNA binding uses similar residues as the hPLD3 5'-exonuclease, despite different activities and orientations of the DNA. The role of K4 and the control of its activity by the observed auto-inhibition remain enigmatic.
History
DepositionMay 4, 2026-
Header (metadata) releaseSep 30, 2026-
Map releaseSep 30, 2026-
UpdateSep 30, 2026-
Current statusSep 30, 2026Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_57884.png

Downloads & links

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Map

FileDownload / File: emd_57884.map.gz / Format: CCP4 / Size: 244.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.84 Å/pix.
x 400 pix.
= 336. Å		0.84 Å/pix.
x 400 pix.
= 336. Å		0.84 Å/pix.
x 400 pix.
= 336. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.84 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.29
Minimum - Maximum-0.62889564 - 1.3408339
Average (Standard dev.)0.0024704484 (±0.045485012)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions400400400
Spacing400400400
CellA=B=C: 336.0 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_57884_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #2

Fileemd_57884_half_map_1.map
Projections & Slices
AxesZYX

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Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_57884_half_map_2.map
Projections & Slices
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Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Octameric complex of K4 containing mutated histidines in the HKD ...

EntireName: Octameric complex of K4 containing mutated histidines in the HKD motif, with DNA bound in chains A and D
Components
  • Complex: Octameric complex of K4 containing mutated histidines in the HKD motif, with DNA bound in chains A and D
    • Protein or peptide: Virion nicking-joining enzyme
    • DNA: 19mer-ssDNA-primer

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Supramolecule #1: Octameric complex of K4 containing mutated histidines in the HKD ...

SupramoleculeName: Octameric complex of K4 containing mutated histidines in the HKD motif, with DNA bound in chains A and D
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Vaccinia virus Copenhagen
Molecular weightTheoretical: 392.61 KDa

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Macromolecule #1: Virion nicking-joining enzyme

MacromoleculeName: Virion nicking-joining enzyme / type: protein_or_peptide / ID: 1 / Number of copies: 4 / Enantiomer: LEVO
Source (natural)Organism: Orthopoxvirus vaccinia / Strain: Copenhagen
Molecular weightTheoretical: 52.079234 KDa
Recombinant expressionOrganism: Trichoplusia ni (cabbage looper)
SequenceString: MSYYHHHHHH DYDIPTTENL YFQGAMANPD NTIAVITETI PIGMQFDKVY LSTFNMWREI LSNTTKTLDI SSFYWSLSDE VGTNFGTII LNEIVQLPKR GVRVRVAVNK SNKPLKDVER LQMAGVEVRY IDITNILGGV LQTKFWISDN THIYLGSANM D WRSLTQVK ...String:
MSYYHHHHHH DYDIPTTENL YFQGAMANPD NTIAVITETI PIGMQFDKVY LSTFNMWREI LSNTTKTLDI SSFYWSLSDE VGTNFGTII LNEIVQLPKR GVRVRVAVNK SNKPLKDVER LQMAGVEVRY IDITNILGGV LQTKFWISDN THIYLGSANM D WRSLTQVK ELGIAIFNNR NLAADLTQIF EVYWYLGVNN LPYNWKNFYP SYYNTDHPLS INVSGVPHSV FIASAPQQLC TM ERTNDLT ALLSCIRNAS KFVYVSVMNF IPIIYSKAGK ILFWPYIEDE LRRSAIDRQV SVKLLISCWQ RSSFIMRNFL RSI AMLKSK NIDIEVKLFI VPDADPPIPY SRVNQAKYMV TDKTAYIGTS NWTGNYFTDT CGASINITPD DGLGLRQQLE DIFM RDWNS KYSYELYDTS PTKRCKLLKN MKQCTNDIYC DEIQPEKEIP EYSLE

UniProtKB: Virion nicking-joining enzyme

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Macromolecule #2: 19mer-ssDNA-primer

MacromoleculeName: 19mer-ssDNA-primer / type: dna / ID: 2 / Number of copies: 2 / Classification: DNA
Source (natural)Organism: synthetic construct (others)
Molecular weightTheoretical: 6.005881 KDa
SequenceString:
(DC)(DT)(DT)(DA)(DG)(DG)(DT)(DA)(DG)(DG) (DG)(DG)(DA)(DG)(DG)(DA)(DT)(DG)(DG)

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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

Concentration1 mg/mL
BufferpH: 8.5 / Details: 20 mM Tris-HCl (pH 8.5), 100 mM NaCl, 2 mM DTT
GridModel: UltrAuFoil R1.2/1.3 / Material: GOLD / Mesh: 300 / Support film - Material: GOLD / Support film - topology: HOLEY ARRAY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 50 sec. / Pretreatment - Atmosphere: AIR / Details: EMS instruments
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 293 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeTFS KRIOS
Specialist opticsEnergy filter - Name: GIF Quantum LS / Energy filter - Slit width: 20 eV
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Detector mode: COUNTING / Number grids imaged: 1 / Number real images: 6075 / Average electron dose: 42.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.4 µm / Nominal defocus min: 0.7000000000000001 µm / Nominal magnification: 105000
Sample stageCooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

CTF correctionSoftware - Name: cryoSPARC (ver. 4) / Type: PHASE FLIPPING AND AMPLITUDE CORRECTION
Startup modelType of model: NONE
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 2.6 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 4) / Number images used: 23730
Initial angle assignmentType: RANDOM ASSIGNMENT / Software - Name: cryoSPARC (ver. 4)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4)
Final 3D classificationSoftware - Name: cryoSPARC (ver. 4)
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelPDB ID:

30ie


Chain - Source name: PDB / Chain - Initial model type: experimental model
RefinementSpace: REAL / Protocol: FLEXIBLE FIT / Overall B value: 133.56 / Target criteria: cross-correlation coefficient
Output model

PDB-30mn:
PLD-fold vaccinia virus endonuclease K4 with DNA

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