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- EMDB-16624: HK97 Prohead II as part of a DNA packaging complex -

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Basic information

Entry
Database: EMDB / ID: EMD-16624
TitleHK97 Prohead II as part of a DNA packaging complex
Map dataHK97 bacteriophage prohead II i3 symmetry
Sample
  • Virus: Hendrixvirus
    • Protein or peptide: Major capsid subunit
Keywordsprohead / HK97 / bacteriophage / packaging / VIRAL PROTEIN
Function / homologyPhage capsid / Phage capsid family / Major capsid subunit
Function and homology information
Biological speciesHendrixvirus
Methodsingle particle reconstruction / cryo EM / Resolution: 3.06 Å
AuthorsHawkins DEDP / Antson AA
Funding support United Kingdom, 1 items
OrganizationGrant numberCountry
Biotechnology and Biological Sciences Research Council (BBSRC)AC014501 United Kingdom
CitationJournal: Nucleic Acids Res / Year: 2023
Title: Insights into a viral motor: the structure of the HK97 packaging termination assembly.
Authors: Dorothy E D P Hawkins / Oliver W Bayfield / Herman K H Fung / Daniel N Grba / Alexis Huet / James F Conway / Alfred A Antson /
Abstract: Double-stranded DNA viruses utilise machinery, made of terminase proteins, to package viral DNA into the capsid. For cos bacteriophage, a defined signal, recognised by small terminase, flanks each ...Double-stranded DNA viruses utilise machinery, made of terminase proteins, to package viral DNA into the capsid. For cos bacteriophage, a defined signal, recognised by small terminase, flanks each genome unit. Here we present the first structural data for a cos virus DNA packaging motor, assembled from the bacteriophage HK97 terminase proteins, procapsids encompassing the portal protein, and DNA containing a cos site. The cryo-EM structure is consistent with the packaging termination state adopted after DNA cleavage, with DNA density within the large terminase assembly ending abruptly at the portal protein entrance. Retention of the large terminase complex after cleavage of the short DNA substrate suggests that motor dissociation from the capsid requires headful pressure, in common with pac viruses. Interestingly, the clip domain of the 12-subunit portal protein does not adhere to C12 symmetry, indicating asymmetry induced by binding of the large terminase/DNA. The motor assembly is also highly asymmetric, showing a ring of 5 large terminase monomers, tilted against the portal. Variable degrees of extension between N- and C-terminal domains of individual subunits suggest a mechanism of DNA translocation driven by inter-domain contraction and relaxation.
History
DepositionFeb 3, 2023-
Header (metadata) releaseMay 24, 2023-
Map releaseMay 24, 2023-
UpdateAug 2, 2023-
Current statusAug 2, 2023Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_16624.png

Downloads & links

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Map

FileDownload / File: emd_16624.map.gz / Format: CCP4 / Size: 421.9 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationHK97 bacteriophage prohead II i3 symmetry
Voxel sizeX=Y=Z: 1.4014 Å
Density
Contour LevelBy AUTHOR: 0.03
Minimum - Maximum-0.07780039 - 0.13780151
Average (Standard dev.)0.0020531204 (±0.0072160102)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions480480480
Spacing480480480
CellA=B=C: 672.672 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_16624_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: HK97 bacteriophage prohead II i3 symmetry

Fileemd_16624_half_map_1.map
AnnotationHK97 bacteriophage prohead II i3 symmetry
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: HK97 bacteriophage prohead II i3 symmetry

Fileemd_16624_half_map_2.map
AnnotationHK97 bacteriophage prohead II i3 symmetry
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Hendrixvirus

EntireName: Hendrixvirus
Components
  • Virus: Hendrixvirus
    • Protein or peptide: Major capsid subunit

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Supramolecule #1: Hendrixvirus

SupramoleculeName: Hendrixvirus / type: virus / ID: 1 / Parent: 0 / Macromolecule list: all
Details: Proheads were produced by infection of E. coli 594 cells with HK97 amber mutant amC2, propagated using Escherichia LE392 cells. This map was derived from a prohead with portal and large terminase bound.
NCBI-ID: 2169654 / Sci species name: Hendrixvirus / Sci species strain: HK97 / Virus type: VIRION / Virus isolate: STRAIN / Virus enveloped: No / Virus empty: No
Host (natural)Organism: Escherichia phage EcSzw-2 (virus)
Molecular weightTheoretical: 560 KDa

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Macromolecule #1: Major capsid subunit

MacromoleculeName: Major capsid subunit / type: protein_or_peptide / ID: 1 / Number of copies: 7 / Enantiomer: LEVO
Source (natural)Organism: Hendrixvirus
Molecular weightTheoretical: 42.286453 KDa
Recombinant expressionOrganism: Escherichia phage EcSzw-2 (virus)
SequenceString: MSELALIQKA IEESQQKMTQ LFDAQKAEIE STGQVSKQLQ SDLMKVQEEL TKSGTRLFDL EQKLASGAEN PGEKKSFSER AAEELIKSW DGKQGTFGAK TFNKSLGSDA DSAGSLIQPM QIPGIIMPGL RRLTIRDLLA QGRTSSNALE YVREEVFTNN A DVVAEKAL ...String:
MSELALIQKA IEESQQKMTQ LFDAQKAEIE STGQVSKQLQ SDLMKVQEEL TKSGTRLFDL EQKLASGAEN PGEKKSFSER AAEELIKSW DGKQGTFGAK TFNKSLGSDA DSAGSLIQPM QIPGIIMPGL RRLTIRDLLA QGRTSSNALE YVREEVFTNN A DVVAEKAL KPESDITFSK QTANVKTIAH WVQASRQVMD DAPMLQSYIN NRLMYGLALK EEGQLLNGDG TGDNLEGLNK VA TAYDTSL NATGDTRADI IAHAIYQVTE SEFSASGIVL NPRDWHNIAL LKDNEGRYIF GGPQAFTSNI MWGLPVVPTK AQA AGTFTV GGFDMASQVW DRMDATVEVS REDRDNFVKN MLTILCEERL ALAHYRPTAI IKGTFSSGS

UniProtKB: Major capsid subunit

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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
Details: final buffer = 20 mM Tris pH 7.5, 10 mM MgCl2, 50 mM KGlu, 5 mM ATP-gamma-S
GridModel: Quantifoil / Material: COPPER / Mesh: 200 / Support film - Material: CARBON / Support film - topology: CONTINUOUS / Support film - Film thickness: 0.2 / Pretreatment - Type: PLASMA CLEANING
VitrificationCryogen name: ETHANE / Chamber humidity: 90 % / Instrument: FEI VITROBOT MARK IV
DetailsPackaging reactions 30 nM hk97 proheads, 2.5 micromolar large terminase, 5 micromolar small terminase and 30 nM DNA, 75 micromolar ATP were incubated for 2 mins then 5 micromolar ATP-gamma-S was added.

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Electron microscopy

MicroscopeTFS KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 2.5 µm / Nominal defocus min: 0.5 µm
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Number grids imaged: 1 / Average electron dose: 40.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 82279
Startup modelType of model: NONE
Initial angle assignmentType: NOT APPLICABLE
Final angle assignmentType: MAXIMUM LIKELIHOOD
Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: I (icosahedral) / Resolution.type: BY AUTHOR / Resolution: 3.06 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.1) / Number images used: 82275
FSC plot (resolution estimation)

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