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- EMDB-43629: Cryo-EM structure of phage DEV ejection proteins gp72:gp73 -

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Basic information

Entry
Database: EMDB / ID: EMD-43629
TitleCryo-EM structure of phage DEV ejection proteins gp72:gp73
Map data
Sample
  • Virus: Pseudomonas phage vB_PaeP_DEV (virus)
    • Protein or peptide: gp72
    • Protein or peptide: N4 gp52-like protein
Keywordsphage / bacteriophage / STRUCTURAL PROTEIN / VIRAL PROTEIN / outer membrane protein / gp73 / gp74 / DEV
Function / homologyUncharacterized protein / N4 gp52-like protein
Function and homology information
Biological speciesPseudomonas phage vB_PaeP_DEV (virus)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.1 Å
AuthorsIglesias SM / Cingolani G
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Cancer Institute (NIH/NCI)GM140733 United States
CitationJournal: Res Sq / Year: 2024
Title: Integrative structural analysis of phage DEV reveals a genome ejection motor.
Authors: Gino Cingolani / Ravi Lokareddy / Chun-Feng Hou / Francesca Forti / Stephano Iglesias / Fenglin Li / Mikhail Pavlenok / Michael Niederweis / Federica Briani
Abstract: DEV is an obligatory lytic phage of the N4-like genus, recently reclassified as . The DEV genome encodes 91 ORFs, including a 3,398 amino acid virion-associated RNA polymerase. Here, we describe the ...DEV is an obligatory lytic phage of the N4-like genus, recently reclassified as . The DEV genome encodes 91 ORFs, including a 3,398 amino acid virion-associated RNA polymerase. Here, we describe the complete architecture of DEV, determined using a combination of cryo-electron microscopy localized reconstruction, biochemical methods, and genetic knockouts. We built de structures of all capsid factors and tail components involved in host attachment. We demonstrate that DEV long tail fibers are essential for infection of and dispensable for infecting mutants with a truncated lipopolysaccharide devoid of the O-antigen. We identified DEV ejection proteins and, unexpectedly, found that the giant DEV RNA polymerase, the hallmark of the family, is an ejection protein. We propose that DEV ejection proteins form a genome ejection motor across the host cell envelope and that these structural principles are conserved in all .
History
DepositionFeb 5, 2024-
Header (metadata) releaseApr 3, 2024-
Map releaseApr 3, 2024-
UpdateOct 16, 2024-
Current statusOct 16, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_43629.png

Downloads & links

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Map

FileDownload / File: emd_43629.map.gz / Format: CCP4 / Size: 244.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

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AxesX (Sec.)Y (Row.)Z (Col.)
1.21 Å/pix.
x 400 pix.
= 484. Å		1.21 Å/pix.
x 400 pix.
= 484. Å		1.21 Å/pix.
x 400 pix.
= 484. Å

Surface

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Images are generated by Spider.

Voxel sizeX=Y=Z: 1.21 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.435
Minimum - Maximum-6.182148 - 10.354398
Average (Standard dev.)-0.0007386073 (±0.15847953)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderZYX
Origin000
Dimensions400400400
Spacing400400400
CellA=B=C: 484.0 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_43629_msk_1.map
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Half map: #2

Fileemd_43629_half_map_1.map
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Half map: #1

Fileemd_43629_half_map_2.map
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Sample components

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Entire : Pseudomonas phage vB_PaeP_DEV

EntireName: Pseudomonas phage vB_PaeP_DEV (virus)
Components
  • Virus: Pseudomonas phage vB_PaeP_DEV (virus)
    • Protein or peptide: gp72
    • Protein or peptide: N4 gp52-like protein

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Supramolecule #1: Pseudomonas phage vB_PaeP_DEV

SupramoleculeName: Pseudomonas phage vB_PaeP_DEV / type: virus / ID: 1 / Parent: 0 / Macromolecule list: all / NCBI-ID: 2034344 / Sci species name: Pseudomonas phage vB_PaeP_DEV / Virus type: VIRION / Virus isolate: OTHER / Virus enveloped: No / Virus empty: No
Host (natural)Organism: Pseudomonas aeruginosa (bacteria)

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Macromolecule #1: gp72

MacromoleculeName: gp72 / type: protein_or_peptide / ID: 1 / Number of copies: 9 / Enantiomer: LEVO
Source (natural)Organism: Pseudomonas phage vB_PaeP_DEV (virus)
Molecular weightTheoretical: 57.183238 KDa
SequenceString: MAQEITWRNI GATVSPGSAS SMSAGTTGVQ QALGALGDII SRQQEMNVNN AKLQREANTQ SYLDQVAAST LEQLSNADYR SGLEAQRDA MGMNLDRAAT RDAITKQISA QQNQAAATQK FDDMQAEVGQ RGIVDQLRTL SAEGRAGEVN QILAEQQLIN E GEIRKELT ...String:
MAQEITWRNI GATVSPGSAS SMSAGTTGVQ QALGALGDII SRQQEMNVNN AKLQREANTQ SYLDQVAAST LEQLSNADYR SGLEAQRDA MGMNLDRAAT RDAITKQISA QQNQAAATQK FDDMQAEVGQ RGIVDQLRTL SAEGRAGEVN QILAEQQLIN E GEIRKELT GVQDAIQNRQ YRAAGEQRAQ AAANRAAEAH SLSMAAGREN LAFTREQRDE LRRDRDEAKL VSGTIATTFQ DY DESRQAQ SEIMRIVGKE VGMPTDDQGM PDMSRASQDQ LDAFSNALNE AGVQANTSPT ERRNAVLKSL VDAGVSSKGI AQA KQEMEL RESLEGLAPQ DRTKVEATIG AVNAELDTLQ RTATEDYERE VARNPFVEPD KDPLGSVNKI VDKAVKSGFG WEGD RQDLN NMLVDFATNG IKLPDGRTAV VPSKLLEQAF NTTNTWLFKN AGDVEKRIIE LMTTDGMTQM REDAPTIREN FLKTV SDIA NQKRSNAVKV TRSAEREKGV TMDPTDDLTF ALRGRKR

UniProtKB: Uncharacterized protein

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Macromolecule #2: N4 gp52-like protein

MacromoleculeName: N4 gp52-like protein / type: protein_or_peptide / ID: 2 / Number of copies: 9 / Enantiomer: LEVO
Source (natural)Organism: Pseudomonas phage vB_PaeP_DEV (virus)
Molecular weightTheoretical: 16.639414 KDa
SequenceString:
MAYPYSDMPF GVELDTSTLG SFGLGGPQTQ LQMQMPAVDV NAAASGSGGF MAGFSNIFSR DSMFGGVAPS GAQTGGWVLP ALGIGQAVF GAIGANRQQR AARDQLAESR RQFDMNYGAQ RQSINTNLED RQRARVASNP TAYESVDSYM ERNRIR

UniProtKB: N4 gp52-like protein

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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: 7.5
GridModel: Quantifoil R2/1 / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 15 sec. / Pretreatment - Atmosphere: OTHER
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277.15 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: FEI FALCON IV (4k x 4k) / Average electron dose: 1.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 100.0 µm / Illumination mode: OTHER / Imaging mode: OTHER / Cs: 2.7 mm / Nominal defocus max: 2.2 µm / Nominal defocus min: 0.8 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

Startup modelType of model: NONE
Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: C9 (9 fold cyclic) / Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 3.1 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 61482
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
FSC plot (resolution estimation)

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