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- PDB-6r21: Cryo-EM structure of T7 bacteriophage fiberless tail complex -

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Basic information

Entry
Database: PDB / ID: 6r21
TitleCryo-EM structure of T7 bacteriophage fiberless tail complex
Components
  • Portal protein
  • Tail tubular protein gp11
  • Tail tubular protein gp12
KeywordsVIRAL PROTEIN / viral complex / DNA ejection
Function / homology
Function and homology information


virus tail, tube / viral portal complex / symbiont genome ejection through host cell envelope, short tail mechanism / viral DNA genome packaging
Similarity search - Function
Tail tubular protein Gp11 / Tail tubular protein / Portal protein, Caudovirales / Head-to-tail connector protein, podovirus-type / Bacteriophage head to tail connecting protein
Similarity search - Domain/homology
Portal protein / Tail tubular protein gp11 / Tail tubular protein gp12
Similarity search - Component
Biological speciesEnterobacteria phage T7 (virus)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.33 Å
AuthorsCuervo, A. / Fabrega-Ferrer, M. / Machon, C. / Conesa, J.J. / Perez-Ruiz, M. / Coll, M. / Carrascosa, J.L.
Funding support Spain, 7items
OrganizationGrant numberCountry
Spanish Ministry of Science, Innovation, and UniversitiesBFU2014-54181 Spain
Spanish Ministry of Science, Innovation, and UniversitiesBFU2014-53550-P Spain
Spanish Ministry of Science, Innovation, and UniversitiesBFU2017-83720-P Spain
Spanish Ministry of Science, Innovation, and UniversitiesMDM-2014-0435 Spain
Spanish Ministry of Science, Innovation, and UniversitiesSEV-2013-0347 Spain
Spanish Ministry of Science, Innovation, and UniversitiesRYC-2011-09071 Spain
European Commission653706
CitationJournal: Nat Commun / Year: 2019
Title: Structures of T7 bacteriophage portal and tail suggest a viral DNA retention and ejection mechanism.
Authors: Ana Cuervo / Montserrat Fàbrega-Ferrer / Cristina Machón / José Javier Conesa / Francisco J Fernández / Rosa Pérez-Luque / Mar Pérez-Ruiz / Joan Pous / M Cristina Vega / José L Carrascosa / Miquel Coll /
Abstract: Double-stranded DNA bacteriophages package their genome at high pressure inside a procapsid through the portal, an oligomeric ring protein located at a unique capsid vertex. Once the DNA has been ...Double-stranded DNA bacteriophages package their genome at high pressure inside a procapsid through the portal, an oligomeric ring protein located at a unique capsid vertex. Once the DNA has been packaged, the tail components assemble on the portal to render the mature infective virion. The tail tightly seals the ejection conduit until infection, when its interaction with the host membrane triggers the opening of the channel and the viral genome is delivered to the host cell. Using high-resolution cryo-electron microscopy and X-ray crystallography, here we describe various structures of the T7 bacteriophage portal and fiber-less tail complex, which suggest a possible mechanism for DNA retention and ejection: a portal closed conformation temporarily retains the genome before the tail is assembled, whereas an open portal is found in the tail. Moreover, a fold including a seven-bladed β-propeller domain is described for the nozzle tail protein.
History
DepositionMar 15, 2019Deposition site: PDBE / Processing site: PDBE
Revision 1.0Sep 4, 2019Provider: repository / Type: Initial release

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Structure visualization

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Assembly

Deposited unit
A: Portal protein
B: Portal protein
C: Portal protein
D: Portal protein
E: Portal protein
F: Portal protein
G: Portal protein
H: Portal protein
I: Portal protein
J: Portal protein
K: Portal protein
L: Portal protein
M: Tail tubular protein gp11
N: Tail tubular protein gp11
O: Tail tubular protein gp11
P: Tail tubular protein gp11
Q: Tail tubular protein gp11
R: Tail tubular protein gp11
S: Tail tubular protein gp11
T: Tail tubular protein gp11
U: Tail tubular protein gp11
V: Tail tubular protein gp11
W: Tail tubular protein gp11
X: Tail tubular protein gp11
a: Tail tubular protein gp12
b: Tail tubular protein gp12
c: Tail tubular protein gp12
d: Tail tubular protein gp12
e: Tail tubular protein gp12
f: Tail tubular protein gp12


Theoretical massNumber of molelcules
Total (without water)1,561,40530
Polymers1,561,40530
Non-polymers00
Water0
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_5551
Buried area264030 Å2
ΔGint-981 kcal/mol
Surface area433530 Å2
MethodPISA

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Components

#1: Protein
Portal protein / Gene product 8 / Gp8 / Head-to-tail connector


Mass: 59173.984 Da / Num. of mol.: 12
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Enterobacteria phage T7 (virus) / Production host: Escherichia coli (E. coli) / References: UniProt: P03728
#2: Protein
Tail tubular protein gp11 / Gene product 11 / Gp11


Mass: 26202.820 Da / Num. of mol.: 12
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Enterobacteria phage T7 (virus) / Gene: 11 / Production host: Escherichia coli (E. coli) / References: UniProt: P03746
#3: Protein
Tail tubular protein gp12 / Gene product 12 / Gp12


Mass: 89480.594 Da / Num. of mol.: 6
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Enterobacteria phage T7 (virus) / Gene: 12 / Production host: Escherichia coli (E. coli) / References: UniProt: P03747

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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

Component
IDNameTypeEntity IDParent-IDSource
1Bacteriophage tail complexCOMPLEXall0RECOMBINANT
2Portal proteinCOMPLEX#11RECOMBINANT
3Adaptor proteinCOMPLEX#21RECOMBINANT
4Nozzle proteinCOMPLEX#31RECOMBINANT
Molecular weight
IDEntity assembly-IDValue (°)Experimental value
111.5 MDaYES
210.706 MDaNO
310.3 MDaNO
410.534 MDaNO
Source (natural)
IDEntity assembly-IDOrganismNcbi tax-ID
21Enterobacteria phage T7 (virus)10760
32Enterobacteria phage T7 (virus)10760
43Enterobacteria phage T7 (virus)10760
54Enterobacteria phage T7 (virus)10760
Source (recombinant)
IDEntity assembly-IDOrganismNcbi tax-ID
21Escherichia coli (E. coli)562
32Escherichia coli (E. coli)562
43Escherichia coli (E. coli)562
54Escherichia coli (E. coli)562
Details of virusEmpty: NO / Enveloped: NO / Isolate: STRAIN / Type: VIRION
Buffer solutionpH: 7.8 / Details: 50 mM Tris-HCl pH 7.8, 100 mM NaCl, 10 mM MgCl2
Buffer component
IDConc.FormulaBuffer-ID
1100 mMNaClSodium chloride1
210 mMMgCl21
350 mMTris-HClTris1
SpecimenConc.: 0.8 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportDetails: acetone and treated with 0.1% w/v poly-L-Lysine (Sigma) for 1 min in water
Grid material: COPPER/RHODIUM / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R2/2
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 95 % / Chamber temperature: 295.15 K

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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 FIELDBright-field microscopy / Alignment procedure: COMA FREE
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingElectron dose: 0.84 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k)

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Processing

SoftwareName: PHENIX / Version: (1.12_2829:phenix.real_space_refine) / Classification: refinement
EM software
IDNameCategoryFitting-IDDetails
1Gautomatchparticle selection
2EPUimage acquisition
4GctfCTF correction
7Cootmodel fitting1
9RELIONinitial Euler assignment
10RELIONfinal Euler assignment
11RELIONclassification
12RELION3D reconstruction
14Scipionimage acquisitionOn the fly processing
26REFMACmodel refinement1
27PHENIXmodel refinement1
42Cootmodel fitting2
47REFMACmodel refinement2
48PHENIXmodel refinement2
67Cootmodel fitting3
68REFMACmodel refinement3
69PHENIXmodel refinement3
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: C6 (6 fold cyclic)
3D reconstructionResolution: 3.33 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 92382 / Symmetry type: POINT
Atomic model building
IDProtocolSpace
1RIGID BODY FITREAL
2RIGID BODY FIT
3AB INITIO MODEL
Atomic model building
IDPDB-ID 3D fitting-ID
16QWP

6qwp

1
25MU4

5mu4

2
33
RefinementHighest resolution: 3.33 Å / Cross valid method: THROUGHOUT
Displacement parametersBiso max: 184.86 Å2 / Biso mean: 90.971 Å2 / Biso min: 49.88 Å2
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.008103008
ELECTRON MICROSCOPYf_angle_d1.157139758
ELECTRON MICROSCOPYf_chiral_restr0.06515390
ELECTRON MICROSCOPYf_plane_restr0.00718408
ELECTRON MICROSCOPYf_dihedral_angle_d11.98662052

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