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- PDB-7pv2: GA1 bacteriophage portal protein -

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Basic information

Entry
Database: PDB / ID: 7pv2
TitleGA1 bacteriophage portal protein
ComponentsHead-tail connector (Portal protein)
KeywordsVIRAL PROTEIN / Bacteriophage / Portal protein / cryoEM
Function / homologyPortal protein Gp10 / Phage Connector (GP10) / Portal protein Gp10 superfamily / Head-tail connector (Portal protein)
Function and homology information
Biological speciesBacillus phage GA-1 (virus)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.2 Å
AuthorsJaved, A. / Villanueva, H. / Orlova, E.V. / Savva, R.
Funding support United Kingdom, 4items
OrganizationGrant numberCountry
Biotechnology and Biological Sciences Research Council (BBSRC)BB/M009513 United Kingdom
Biotechnology and Biological Sciences Research Council (BBSRC)BB/R002622/1 United Kingdom
Wellcome Trust202679/Z/16/Z United Kingdom
Wellcome Trust206166/Z/17/Z United Kingdom
CitationJournal: Viruses / Year: 2021
Title: Cryo-EM Structures of Two Bacteriophage Portal Proteins Provide Insights for Antimicrobial Phage Engineering.
Authors: Abid Javed / Hugo Villanueva / Shadikejiang Shataer / Sara Vasciaveo / Renos Savva / Elena V Orlova /
Abstract: Widespread antibiotic resistance has returned attention to bacteriophages as a means of managing bacterial pathogenesis. Synthetic biology approaches to engineer phages have demonstrated genomic ...Widespread antibiotic resistance has returned attention to bacteriophages as a means of managing bacterial pathogenesis. Synthetic biology approaches to engineer phages have demonstrated genomic editing to broaden natural host ranges, or to optimise microbicidal action. Gram positive pathogens cause serious pastoral animal and human infections that are especially lethal in newborns. Such pathogens are targeted by the obligate lytic phages of the and families. These phages have relatively small ~20 kb linear protein-capped genomes and their compact organisation, relatively few structural elements, and broad host range, are appealing from a phage-engineering standpoint. In this study, we focus on portal proteins, which are core elements for the assembly of such tailed phages. The structures of dodecameric portal complexes from phage GA1, which targets , and phage phiCPV4 that infects , were determined at resolutions of 3.3 Å and 2.9 Å, respectively. Both are found to closely resemble the related phi29 portal protein fold. However, the portal protein of phiCPV4 exhibits interesting differences in the clip domain. These structures provide new insights on structural diversity in portal proteins and will be essential for considerations in phage structural engineering.
History
DepositionOct 1, 2021Deposition site: PDBE / Processing site: PDBE
Revision 1.0Oct 12, 2022Provider: repository / Type: Initial release
Revision 1.1Jun 7, 2023Group: Author supporting evidence / Refinement description / Source and taxonomy
Category: em_entity_assembly_naturalsource / em_virus_natural_host ...em_entity_assembly_naturalsource / em_virus_natural_host / entity_src_gen / pdbx_initial_refinement_model
Item: _em_entity_assembly_naturalsource.ncbi_tax_id / _em_entity_assembly_naturalsource.organism ..._em_entity_assembly_naturalsource.ncbi_tax_id / _em_entity_assembly_naturalsource.organism / _entity_src_gen.pdbx_gene_src_ncbi_taxonomy_id / _entity_src_gen.pdbx_gene_src_scientific_name
Revision 1.2Oct 25, 2023Group: Data collection / Database references / Refinement description
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / citation / citation_author / em_3d_fitting_list
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.journal_volume / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year / _em_3d_fitting_list.accession_code / _em_3d_fitting_list.initial_refinement_model_id / _em_3d_fitting_list.source_name / _em_3d_fitting_list.type

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Head-tail connector (Portal protein)
B: Head-tail connector (Portal protein)
C: Head-tail connector (Portal protein)
D: Head-tail connector (Portal protein)
E: Head-tail connector (Portal protein)
F: Head-tail connector (Portal protein)
G: Head-tail connector (Portal protein)
H: Head-tail connector (Portal protein)
I: Head-tail connector (Portal protein)
J: Head-tail connector (Portal protein)
K: Head-tail connector (Portal protein)
L: Head-tail connector (Portal protein)


Theoretical massNumber of molelcules
Total (without water)423,69412
Polymers423,69412
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein
Head-tail connector (Portal protein)


Mass: 35307.801 Da / Num. of mol.: 12
Source method: isolated from a genetically manipulated source
Details: In each subunit of the dodecamer, the DNA Tunnel loop residues (232 - 250) has not been modelled due to lack of EM density. Also missing in the pdb structure are 22 residues at the C- ...Details: In each subunit of the dodecamer, the DNA Tunnel loop residues (232 - 250) has not been modelled due to lack of EM density. Also missing in the pdb structure are 22 residues at the C-terminus and 7 residues at the N-terminus in each chain.
Source: (gene. exp.) Bacillus phage GA-1 (virus) / Gene: gene 10 / Production host: Escherichia coli (E. coli) / References: UniProt: Q9FZW5

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: GA1 bacteriophage portal protein / Type: COMPLEX / Details: A dodecamer complex made up of 12 subunits. / Entity ID: all / Source: RECOMBINANT
Source (natural)Organism: Bacillus phage GA-1 (virus)
Source (recombinant)Organism: Escherichia coli (E. coli)
Natural hostOrganism: Bacillus pumilus
Buffer solutionpH: 7.5
SpecimenConc.: 0.05 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: COPPER / Grid type: PELCO Ultrathin Carbon with Lacey Carbon
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 281 K / Details: 3ul of sample was applied on lacey carbon grids.

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 2000 nm / Nominal defocus min: 1200 nm / Cs: 2.7 mm / C2 aperture diameter: 70 µm / Alignment procedure: ZEMLIN TABLEAU
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Temperature (max): 98 K / Temperature (min): 95 K
Image recordingElectron dose: 1 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Num. of grids imaged: 1 / Num. of real images: 2249
EM imaging opticsEnergyfilter name: GIF Quantum LS / Energyfilter slit width: 20 eV
Image scansSampling size: 5 µm / Width: 3838 / Height: 3710 / Used frames/image: 1-12

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Processing

SoftwareName: PHENIX / Version: 1.15.1_3469: / Classification: refinement
EM software
IDNameVersionCategory
2EPUimage acquisition
4CTFFIND4.1CTF correction
7UCSF Chimeramodel fitting
8Cootmodel fitting
10PHENIXmodel refinement
11Cootmodel refinement
12ISOLDEmodel refinement
13RELION3initial Euler assignment
14RELION3final Euler assignment
15RELION3classification
16RELION3D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 50466
SymmetryPoint symmetry: C12 (12 fold cyclic)
3D reconstructionResolution: 3.2 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 22456 / Algorithm: FOURIER SPACE / Num. of class averages: 1 / Symmetry type: POINT
Atomic model buildingB value: 97.7 / Protocol: FLEXIBLE FIT / Space: REAL / Target criteria: Correlation coefficient
Atomic model buildingPDB-ID: 1FOU
Accession code: 1FOU / Source name: PDB / Type: experimental model

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