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- EMDB-28010: Pseudomonas phage phiKZ PhuZ Filament -

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Basic information

Entry
Database: EMDB / ID: EMD-28010
TitlePseudomonas phage phiKZ PhuZ Filament
Map dataunsharpened and filtered to the FSC0.143 cutoff
Sample
  • Complex: Pseudomonas phage phiKZ PhuZ Filament
Keywordsbacteriophage / structural protein / viral tubulin / VIRAL PROTEIN
Biological speciesPseudomonas phage phiKZ (virus)
Methodsubtomogram averaging / cryo EM / Resolution: 22.0 Å
AuthorsLaughlin TG / Villa E
Funding support United States, 3 items
OrganizationGrant numberCountry
Howard Hughes Medical Institute (HHMI) United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R01GM129245 United States
National Science Foundation (NSF, United States)DBI 1920374 United States
CitationJournal: bioRxiv / Year: 2023
Title: Identifying the core genome of the nucleus-forming bacteriophage family and characterization of phage RAY.
Abstract: We recently discovered that some bacteriophages establish a nucleus-like replication compartment (phage nucleus), but the core genes that define nucleus-based phage replication and their phylogenetic ...We recently discovered that some bacteriophages establish a nucleus-like replication compartment (phage nucleus), but the core genes that define nucleus-based phage replication and their phylogenetic distribution were unknown. By studying phages that encode the major phage nucleus protein chimallin, including previously sequenced yet uncharacterized phages, we discovered that chimallin-encoding phages share a set of 72 highly conserved genes encoded within seven distinct gene blocks. Of these, 21 core genes are unique to this group, and all but one of these unique genes encode proteins of unknown function. We propose that phages with this core genome comprise a novel viral family we term Chimalliviridae. Fluorescence microscopy and cryo-electron tomography studies of phage vB_EamM_RAY confirm that many of the key steps of nucleus-based replication encoded in the core genome are conserved among diverse chimalliviruses, and reveal that non-core components can confer intriguing variations on this replication mechanism. For instance, unlike previously studied nucleus-forming phages, RAY doesn't degrade the host genome, and its PhuZ homolog appears to form a five-stranded filament with a lumen. This work expands our understanding of phage nucleus and PhuZ spindle diversity and function, providing a roadmap for identifying key mechanisms underlying nucleus-based phage replication.
History
DepositionAug 31, 2022-
Header (metadata) releaseMar 15, 2023-
Map releaseMar 15, 2023-
UpdateJan 17, 2024-
Current statusJan 17, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_28010.png

Downloads & links

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Map

FileDownload / File: emd_28010.map.gz / Format: CCP4 / Size: 333 KB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationunsharpened and filtered to the FSC0.143 cutoff
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
3.46 Å/pix.
x 44 pix.
= 152.108 Å		3.46 Å/pix.
x 44 pix.
= 152.108 Å		3.46 Å/pix.
x 44 pix.
= 152.108 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 3.457 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.05
Minimum - Maximum-0.18022408 - 0.40515885
Average (Standard dev.)0.008424343 (±0.11442117)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions444444
Spacing444444
CellA=B=C: 152.108 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_28010_msk_1.map
Projections & Slices
AxesZYX

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Half map: #1

Fileemd_28010_half_map_1.map
Projections & Slices
AxesZYX

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Density Histograms

Histogram

Histogram (log scale)

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Half map: #2

Fileemd_28010_half_map_2.map
Projections & Slices
AxesZYX

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Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Pseudomonas phage phiKZ PhuZ Filament

EntireName: Pseudomonas phage phiKZ PhuZ Filament
Components
  • Complex: Pseudomonas phage phiKZ PhuZ Filament

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Supramolecule #1: Pseudomonas phage phiKZ PhuZ Filament

SupramoleculeName: Pseudomonas phage phiKZ PhuZ Filament / type: complex / ID: 1 / Parent: 0
Source (natural)Organism: Pseudomonas phage phiKZ (virus)

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Experimental details

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Structure determination

Methodcryo EM
Processingsubtomogram averaging
Aggregation statecell

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Sample preparation

BufferpH: 7.5
GridModel: Quantifoil R2/1 / Material: COPPER / Mesh: 200 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 60 sec. / Pretreatment - Atmosphere: OTHER / Pretreatment - Pressure: 0.019 kPa / Details: 20 mA in a PELCO easiGLO
VitrificationCryogen name: ETHANE-PROPANE / Instrument: HOMEMADE PLUNGER
Detailscell suspension

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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 K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Number grids imaged: 2 / Average electron dose: 2.5 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 70.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 6.0 µm / Nominal defocus min: 4.0 µm / Nominal magnification: 42000
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 - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 22.0 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.1.3) / Number subtomograms used: 1487
ExtractionNumber tomograms: 14 / Number images used: 4776 / Method: manual picking and over-sampling
Software: (Name: IMOD, Dynamo (ver. 1.1514), MATLAB (ver. 2019b))
Details: Manually picked the start and end points of the filaments in IMOD and used coordinates to generate over-sampled cropping models in Dynamo.
Final 3D classificationSoftware - Name: RELION
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.1.3)
FSC plot (resolution estimation)

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