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- EMDB-52708: Enterobacteriaphage PRD1 - P12 protein filament in complex with p... -

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Basic information

Entry
Database: EMDB / ID: EMD-52708
TitleEnterobacteriaphage PRD1 - P12 protein filament in complex with poly(dT) ssDNA
Map dataPrimary map
Sample
  • Complex: P12 filament bound to poly(dT) ssDNA
    • Protein or peptide: Single-stranded DNA-binding protein
    • DNA: ssDNA poly(dT), 80mer
KeywordsSSB / Protein primed replication / Protein filament / Homooligomer / ssDNA-binding / REPLICATION
Function / homologynucleotide binding / DNA binding / Single-stranded DNA-binding protein
Function and homology information
Biological speciesEnterobacteria phage PRD1 (virus) / synthetic construct (others)
Methodhelical reconstruction / cryo EM / Resolution: 2.75 Å
AuthorsDegen M / Traeger KL / Hiller S
Funding support Switzerland, 1 items
OrganizationGrant numberCountry
Swiss Nanoscience Institute Switzerland
CitationJournal: Nucleic Acids Res / Year: 2025
Title: Structural basis for cooperative ssDNA binding by bacteriophage protein filament P12.
Authors: Lena K Träger / Morris Degen / Joana Pereira / Janani Durairaj / Raphael Dias Teixeira / Sebastian Hiller / Nicolas Huguenin-Dezot /
Abstract: Protein-primed DNA replication is a unique mechanism, bioorthogonal to other known DNA replication modes. It relies on specialised single-stranded DNA (ssDNA)-binding proteins (SSBs) to stabilise ...Protein-primed DNA replication is a unique mechanism, bioorthogonal to other known DNA replication modes. It relies on specialised single-stranded DNA (ssDNA)-binding proteins (SSBs) to stabilise ssDNA intermediates by unknown mechanisms. Here, we present the structural and biochemical characterisation of P12, an SSB from bacteriophage PRD1. High-resolution cryo-electron microscopy reveals that P12 forms a unique, cooperative filament along ssDNA. Each protomer binds the phosphate backbone of 6 nucleotides in a sequence-independent manner, protecting ssDNA from nuclease degradation. Filament formation is driven by an intrinsically disordered C-terminal tail, facilitating cooperative binding. We identify residues essential for ssDNA interaction and link the ssDNA-binding ability of P12 to toxicity in host cells. Bioinformatic analyses place the P12 fold as a distinct branch within the OB-like fold family. This work offers new insights into protein-primed DNA replication and lays a foundation for biotechnological applications.
History
DepositionFeb 4, 2025-
Header (metadata) releaseMar 19, 2025-
Map releaseMar 19, 2025-
UpdateMar 19, 2025-
Current statusMar 19, 2025Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_52708.png

Downloads & links

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Map

FileDownload / File: emd_52708.map.gz / Format: CCP4 / Size: 178 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationPrimary map
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.82 Å/pix.
x 360 pix.
= 295.2 Å		0.82 Å/pix.
x 360 pix.
= 295.2 Å		0.82 Å/pix.
x 360 pix.
= 295.2 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.82 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.0595
Minimum - Maximum-0.44134465 - 0.5889039
Average (Standard dev.)0.00003586459 (±0.012084629)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions360360360
Spacing360360360
CellA=B=C: 295.2 Å
α=β=γ: 90.0 °

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Supplemental data

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Additional map: DeepEMhancer output of primary Map

Fileemd_52708_additional_1.map
AnnotationDeepEMhancer output of primary Map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half Map B

Fileemd_52708_half_map_1.map
AnnotationHalf_Map_B
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half Map A

Fileemd_52708_half_map_2.map
AnnotationHalf_Map_A
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : P12 filament bound to poly(dT) ssDNA

EntireName: P12 filament bound to poly(dT) ssDNA
Components
  • Complex: P12 filament bound to poly(dT) ssDNA
    • Protein or peptide: Single-stranded DNA-binding protein
    • DNA: ssDNA poly(dT), 80mer

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Supramolecule #1: P12 filament bound to poly(dT) ssDNA

SupramoleculeName: P12 filament bound to poly(dT) ssDNA / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Enterobacteria phage PRD1 (virus)

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Macromolecule #1: Single-stranded DNA-binding protein

MacromoleculeName: Single-stranded DNA-binding protein / type: protein_or_peptide / ID: 1 / Number of copies: 6 / Enantiomer: LEVO
Source (natural)Organism: Enterobacteria phage PRD1 (virus)
Molecular weightTheoretical: 16.671012 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString:
MEIVSKLTLK TIGAQPKPHS VKENTALASI YGRVRGKKVG QSTFGDFIKF EGEFEGVNIA TGEVFRSGAL ILPKVLESLL AGAVDGENT VDFAVEIWAK PSEKGNTGYE YGVKPLIEPA ASDELAALRN QVKAALPAPA AAGEAAAEAK PAAKAKAKAE A

UniProtKB: Single-stranded DNA-binding protein

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Macromolecule #2: ssDNA poly(dT), 80mer

MacromoleculeName: ssDNA poly(dT), 80mer / type: dna / ID: 2 / Number of copies: 2 / Classification: DNA
Source (natural)Organism: synthetic construct (others)
Molecular weightTheoretical: 24.290438 KDa
SequenceString: (DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT) (DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT) (DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT) (DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT) (DT) (DT)(DT)(DT)(DT)(DT)(DT) ...String:
(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT) (DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT) (DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT) (DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT) (DT) (DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT) (DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT) (DT)(DT) (DT)(DT)(DT)(DT)(DT)(DT)(DT) (DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT) (DT)(DT)(DT)

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Experimental details

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Structure determination

Methodcryo EM
Processinghelical reconstruction
Aggregation statefilament

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Sample preparation

BufferpH: 8.5
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeTFS KRIOS
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Average electron dose: 48.7 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: 4.2 µm / Nominal defocus min: 0.39 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Final reconstructionApplied symmetry - Helical parameters - Δz: 27.461 Å
Applied symmetry - Helical parameters - Δ&Phi: 19.184 °
Applied symmetry - Helical parameters - Axial symmetry: C1 (asymmetric)
Resolution.type: BY AUTHOR / Resolution: 2.75 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 4.4.1) / Number images used: 463309
Startup modelType of model: PDB ENTRY
PDB model - PDB ID:

9gfq

Final angle assignmentType: NOT APPLICABLE / Software - Name: cryoSPARC (ver. 4.4.1)
FSC plot (resolution estimation)

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