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- EMDB-49121: In situ cryoET of an EPI vesicle in a Goslar infected chmA KD E. ... -

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Basic information

Entry
Database: EMDB / ID: EMD-49121
TitleIn situ cryoET of an EPI vesicle in a Goslar infected chmA KD E. coli cell 90 mpi
Map data
Sample
  • Cell: Goslar infected chmA KD E. coli cell 90 mpi
Keywordsvesicle / phage / genome / VIRUS
Biological speciesEscherichia coli (E. coli)
Methodelectron tomography / cryo EM / Resolution: 15.0 Å
AuthorsHutchings J / Rodriguez ZK / Klusch N / Villa E
Funding support United States, 1 items
OrganizationGrant numberCountry
Howard Hughes Medical Institute (HHMI) United States
CitationJournal: Cell Host Microbe / Year: 2025
Title: Sequential membrane- and protein-bound organelles compartmentalize genomes during phage infection.
Authors: Emily G Armbruster / Phoolwanti Rani / Jina Lee / Niklas Klusch / Joshua Hutchings / Lizbeth Y Hoffman / Hannah Buschkaemper / Eray Enustun / Benjamin A Adler / Koe Inlow / Arica R VanderWal ...Authors: Emily G Armbruster / Phoolwanti Rani / Jina Lee / Niklas Klusch / Joshua Hutchings / Lizbeth Y Hoffman / Hannah Buschkaemper / Eray Enustun / Benjamin A Adler / Koe Inlow / Arica R VanderWal / Madelynn Y Hoffman / Daksh Daksh / Ann Aindow / Amar Deep / Zaida K Rodriguez / Chase J Morgan / Majid Ghassemian / Thomas G Laughlin / Emeric Charles / Brady F Cress / David F Savage / Jennifer A Doudna / Kit Pogliano / Kevin D Corbett / Elizabeth Villa / Joe Pogliano /
Abstract: Many eukaryotic viruses require membrane-bound compartments for replication, but no such organelles are known to be formed by prokaryotic viruses. Bacteriophages of the Chimalliviridae family ...Many eukaryotic viruses require membrane-bound compartments for replication, but no such organelles are known to be formed by prokaryotic viruses. Bacteriophages of the Chimalliviridae family sequester their genomes within a phage-generated organelle, the phage nucleus, which is enclosed by a lattice of the viral protein ChmA. We show that inhibiting phage nucleus formation arrests infections at an early stage in which the injected phage genome is enclosed within a membrane-bound early phage infection (EPI) vesicle. Early phage genes are expressed from the EPI vesicle, demonstrating its functionality as a prokaryotic, transcriptionally active, membrane-bound organelle. We also show that the phage nucleus is essential, with genome replication beginning after the injected DNA is transferred from the EPI vesicle to the phage nucleus. Our results show that Chimalliviridae require two sophisticated subcellular compartments of distinct compositions and functions that facilitate successive stages of the viral life cycle.
History
DepositionFeb 7, 2025-
Header (metadata) releaseApr 9, 2025-
Map releaseApr 9, 2025-
UpdateApr 23, 2025-
Current statusApr 23, 2025Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_49121.png

Downloads & links

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Map

FileDownload / File: emd_49121.map.gz / Format: CCP4 / Size: 344.3 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
15 Å/pix.
x 288 pix.
= 4320. Å		15 Å/pix.
x 664 pix.
= 9960. Å		15 Å/pix.
x 472 pix.
= 7080. Å

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

generated in cubic-lattice coordinate

Voxel sizeX=Y=Z: 15 Å
Density

Histogram

Histogram (log scale)
Minimum - Maximum-0.6622478 - 1.4708278
Average (Standard dev.)-0.000000000001093 (±0.08077936)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions664472288
Spacing472664288
CellA: 7080.0 Å / B: 9960.0 Å / C: 4320.0 Å
α=β=γ: 90.0 °

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Supplemental data

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Sample components

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Entire : Goslar infected chmA KD E. coli cell 90 mpi

EntireName: Goslar infected chmA KD E. coli cell 90 mpi
Components
  • Cell: Goslar infected chmA KD E. coli cell 90 mpi

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Supramolecule #1: Goslar infected chmA KD E. coli cell 90 mpi

SupramoleculeName: Goslar infected chmA KD E. coli cell 90 mpi / type: cell / ID: 1 / Parent: 0
Source (natural)Organism: Escherichia coli (E. coli)

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Experimental details

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Structure determination

Methodcryo EM
Processingelectron tomography
Aggregation statecell

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Sample preparation

BufferpH: 7.5
GridModel: Quantifoil R2/1 / Material: COPPER / Mesh: 200 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 60 sec.
VitrificationCryogen name: ETHANE-PROPANE
SectioningFocused ion beam - Instrument: OTHER / Focused ion beam - Ion: OTHER / Focused ion beam - Voltage: 30 / Focused ion beam - Current: 0.03 / Focused ion beam - Duration: 600 / Focused ion beam - Temperature: 78 K / Focused ion beam - Initial thickness: 200 / Focused ion beam - Final thickness: 140
Focused ion beam - Details: The value given for _em_focused_ion_beam.instrument is Aquilos 2 dual beam microscope. This is not in a list of allowed values {'OTHER', 'DB235'} so OTHER is written into the XML file.

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Electron microscopy

MicroscopeTFS KRIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 3.8 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: 5.0 µm / Nominal defocus min: 4.0 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Final reconstructionResolution.type: BY AUTHOR / Resolution: 15.0 Å / Software - Name: Warp (ver. 1.1.0) / Number images used: 37

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