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- EMDB-50359: Rhodobacter microvirus Ebor attached to B10 host cell reconstruct... -

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Basic information

Entry
Database: EMDB / ID: EMD-50359
TitleRhodobacter microvirus Ebor attached to B10 host cell reconstructed by single particle analysis with applied C5 symmetry
Map dataEbor attached to B10 host cell, class 1
Sample
  • Virus: Rhodobacter phage Ebor (virus)
    • Protein or peptide: Major capsid protein
KeywordsssDNA virus / Microviridae / Tainavirinae / Alphaproteobacteria / Rhodobacter capsulatus / aquatic virus / jelly-roll fold / VIRUS
Biological speciesRhodobacter capsulatus SB 1003 (bacteria) / Rhodobacter phage Ebor (virus)
Methodsingle particle reconstruction / cryo EM / Resolution: 7.84 Å
AuthorsBardy P / MacDonald CIW / Jenkins HT / Chechik M / Hart SJ / Turkenburg JP / Blaza JN / Fogg PCM / Antson AA
Funding support United Kingdom, 3 items
OrganizationGrant numberCountry
Wellcome Trust224067/Z/21/Z United Kingdom
Wellcome Trust206377 United Kingdom
Wellcome Trust109363/Z/15/A United Kingdom
CitationJournal: bioRxiv / Year: 2024
Title: A stargate mechanism of genome delivery unveiled by cryogenic electron tomography.
Authors: Pavol Bardy / Conor I W MacDonald / Paul C Kirchberger / Huw T Jenkins / Tibor Botka / Lewis Byrom / Nawshin T B Alim / Daouda A K Traore / Hannah C König / Tristan R Nicholas / Maria ...Authors: Pavol Bardy / Conor I W MacDonald / Paul C Kirchberger / Huw T Jenkins / Tibor Botka / Lewis Byrom / Nawshin T B Alim / Daouda A K Traore / Hannah C König / Tristan R Nicholas / Maria Chechik / Samuel J Hart / Johan P Turkenburg / James N Blaza / J Thomas Beatty / Paul C M Fogg / Alfred A Antson /
Abstract: Single-stranded DNA bacteriophages of the family are major components of the global virosphere. Microviruses are highly abundant in aquatic ecosystems and are prominent members of the mammalian gut ...Single-stranded DNA bacteriophages of the family are major components of the global virosphere. Microviruses are highly abundant in aquatic ecosystems and are prominent members of the mammalian gut microbiome, where their diversity has been linked to various chronic health disorders. Despite the clear importance of microviruses, little is known about the molecular mechanism of host infection. Here, we have characterized an exceptionally large microvirus, Ebor, and provide crucial insights into long-standing mechanistic questions. Cryogenic electron microscopy of Ebor revealed a capsid with trimeric protrusions that recognise lipopolysaccharides on the host surface. Cryogenic electron tomography of the host cell colonized with virus particles demonstrated that the virus initially attaches to the cell via five such protrusions, located at the corners of a single pentamer. This interaction triggers a stargate mechanism of capsid opening along the 5-fold symmetry axis, enabling delivery of the virus genome. Despite variations in specific virus-host interactions among different family viruses, structural data indicate that the stargate mechanism of infection is universally employed by all members of the family. Startlingly, our data reveal a mechanistic link for the opening of relatively small capsids made out of a single jelly-roll fold with the structurally unrelated giant viruses.
History
DepositionMay 23, 2024-
Header (metadata) releaseJun 12, 2024-
Map releaseJun 12, 2024-
UpdateJul 10, 2024-
Current statusJul 10, 2024Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_50359.png

Downloads & links

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Map

FileDownload / File: emd_50359.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationEbor attached to B10 host cell, class 1
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
2.14 Å/pix.
x 256 pix.
= 548.608 Å		2.14 Å/pix.
x 256 pix.
= 548.608 Å		2.14 Å/pix.
x 256 pix.
= 548.608 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 2.143 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.004
Minimum - Maximum-0.015209306 - 0.023703791
Average (Standard dev.)0.00012116539 (±0.0032055227)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 548.608 Å
α=β=γ: 90.0 °

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Supplemental data

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Additional map: Ebor attached to B10 host cell, class 2

Fileemd_50359_additional_1.map
AnnotationEbor attached to B10 host cell, class 2
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Additional map: Ebor attached to B10 host cell, class 3

Fileemd_50359_additional_2.map
AnnotationEbor attached to B10 host cell, class 3
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Additional map: mask used for 3D classification

Fileemd_50359_additional_3.map
Annotationmask used for 3D classification
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Additional map: half map 1 for class 2

Fileemd_50359_additional_4.map
Annotationhalf map 1 for class 2
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Additional map: half map 2 for class 2

Fileemd_50359_additional_5.map
Annotationhalf map 2 for class 2
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Additional map: half map 1 for class 3

Fileemd_50359_additional_6.map
Annotationhalf map 1 for class 3
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Additional map: half map 2 for class 3

Fileemd_50359_additional_7.map
Annotationhalf map 2 for class 3
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Half map: half map 2 for class 1

Fileemd_50359_half_map_1.map
Annotationhalf map 2 for class 1
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Half map: half map 1 for class 1

Fileemd_50359_half_map_2.map
Annotationhalf map 1 for class 1
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Sample components

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Entire : Rhodobacter phage Ebor

EntireName: Rhodobacter phage Ebor (virus)
Components
  • Virus: Rhodobacter phage Ebor (virus)
    • Protein or peptide: Major capsid protein

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Supramolecule #1: Rhodobacter phage Ebor

SupramoleculeName: Rhodobacter phage Ebor / type: virus / ID: 1 / Parent: 0 / Macromolecule list: all
Details: Phage, purified from Rhodobacter capsulatus strain SB1003 by ion-exchange chromatography, attached to host cell (B10 strain)
NCBI-ID: 3144506 / Sci species name: Rhodobacter phage Ebor / Sci species strain: variant R120 / Virus type: VIRION / Virus isolate: OTHER / Virus enveloped: No / Virus empty: No
Host (natural)Organism: Rhodobacter capsulatus (bacteria) / Strain: B10
Molecular weightTheoretical: 5.7 MDa
Virus shellShell ID: 1 / Name: Genome-releasing capsid / Diameter: 460.0 Å / T number (triangulation number): 1

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Macromolecule #1: Major capsid protein

MacromoleculeName: Major capsid protein / type: protein_or_peptide / ID: 1 / Enantiomer: LEVO
Source (natural)Organism: Rhodobacter capsulatus SB 1003 (bacteria) / Strain: SB1003
SequenceString: MSYQTKRNVR REARTLMGRF KAGKLAPVMA VPVKGSEGGM LSQSVSFELD PIAGRMATPI TAEMCAVFVP VQACDALKNP EADYAGMTEI VREKLLSGNP LFVLEPETDV SKRCGVNPRR NNGLMRVNEI VRLAHNCAVN FLRRRRYVDA VQLTAANHST TPAILSQTVL ...String:
MSYQTKRNVR REARTLMGRF KAGKLAPVMA VPVKGSEGGM LSQSVSFELD PIAGRMATPI TAEMCAVFVP VQACDALKNP EADYAGMTEI VREKLLSGNP LFVLEPETDV SKRCGVNPRR NNGLMRVNEI VRLAHNCAVN FLRRRRYVDA VQLTAANHST TPAILSQTVL DRFNGALDPD PNVNGAVQLS MPDMRLPVAS DAEKAVTAPL TVKRGNENRR LDAGISRLAA AEVAAATDAA LYAVFGGREA GNVSLTDFYN AQKMDELTRV MRKICDDNPE YGEEMVLRWA HGLSVDPGRV PFLLAEKSVV LGRQIIGATD TAGVEDGVKR SDMAAQLSFT VPIPTTELGG IIVTFACIKP DETLSSQPHP ILADHWRLDN FVADELALDP QPVMARELDY KVAQANETTV VFYTGLNELK KTYVSYGLCR ALDPNTVESK NAVWQLEVPL SVTPETVLYP ADLPQYPFAD QQAEVCTYVV QSTAVMPTPM IFGPSPVEQL AVIETEDLFE D

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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: 7.75
Component:
ConcentrationNameFormula
20.0 mMTris-HCl
100.0 mMsodium chlorideNaCl
5.0 mMcalcium chlorideCaCl2
GridModel: Quantifoil R2/1 / Material: COPPER / Mesh: 200 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 30 sec. / Pretreatment - Atmosphere: AIR
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 295.15 K / Instrument: FEI VITROBOT MARK IV / Details: blot time 2 sec, blot force 0, wait time 8 sec.
Detailsconcentration of 7x10^10 PFU/ml

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Electron microscopy

MicroscopeTFS KRIOS
Specialist opticsEnergy filter - Name: GIF Bioquantum
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Number grids imaged: 1 / Number real images: 2750 / Average electron dose: 27.9 e/Å2 / Details: acquired using lacey carbon setup in EPU
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 3.0 µm / Nominal defocus min: 0.75 µm
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

Particle selectionNumber selected: 75291
Startup modelType of model: OTHER / Details: map of the native virus
Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: C5 (5 fold cyclic) / Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 7.84 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 5)
Details: RELION skip align run, low passed filtered to the final estimated resolution using relion_image_handler
Number images used: 18179
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 5)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 5)
Details: icosahedral symmetry followed by symmetry expansion and classification in C5
Final 3D classificationNumber classes: 3 / Software - Name: RELION (ver. 5)
Details: class 1 = 18179 particles, class 2 = 2741 particles, class 3 = 2175 particles

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