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- EMDB-11226: Negative staining 3D reconstruction of p2 virion baseplate in act... -

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Basic information

Entry
Database: EMDB / ID: EMD-11226
TitleNegative staining 3D reconstruction of p2 virion baseplate in activated conformation
Map datanegative staining 3D reconstruction of p2 virion baseplate bound to VHH5
Sample
  • Virus: Lactococcus virus P2
    • Protein or peptide: Distal tail protein
    • Protein or peptide: Baseplate protein gp16Tripod (photography)
    • Protein or peptide: Receptor binding proteinReceptor (biochemistry)
Function / homology
Function and homology information


symbiont genome ejection through host cell envelope, long flexible tail mechanism / virus tail, baseplate / virus tail / entry receptor-mediated virion attachment to host cell / cell adhesion / symbiont entry into host cell / virion attachment to host cell
Similarity search - Function
: / : / : / Phage tail base-plate attachment protein C-terminal insertion domain / Phage tail base-plate attachment protein N0 domain / Phage tail base-plate attachment protein C-terminal barrel domain / Phage tail base-plate attachment protein ORF16 / Baseplate protein Gp15-like / Baseplate protein Gp16, domain D4 / Baseplate protein Gp16, domain 1 and 2 ...: / : / : / Phage tail base-plate attachment protein C-terminal insertion domain / Phage tail base-plate attachment protein N0 domain / Phage tail base-plate attachment protein C-terminal barrel domain / Phage tail base-plate attachment protein ORF16 / Baseplate protein Gp15-like / Baseplate protein Gp16, domain D4 / Baseplate protein Gp16, domain 1 and 2 / : / Distal tail protein, N-terminal domain / Phage tail base-plate attachment protein N-terminal barrel domain / Dit, C-terminal domain / : / : / Lactophage receptor-binding protein N-terminal shoulder domain / Lactococcus phage p2, Receptor binding protein, neck domain / Receptor-binding protein of phage tail base-plate Siphoviridae, head / Lactophage receptor-binding protein C-terminal head domain / Adenovirus pIV-like, attachment domain
Similarity search - Domain/homology
Baseplate protein gp16 / Distal tail protein / Receptor binding protein
Similarity search - Component
Biological speciesLactococcus phage p2 (virus) / Lactococcus virus P2
Methodsingle particle reconstruction / negative staining / Resolution: 28.7 Å
AuthorsSpinelli S / Cambillau C / Goulet A
CitationJournal: Viruses / Year: 2020
Title: Structural Insights into Lactococcal Siphophage p2 Baseplate Activation Mechanism.
Authors: Silvia Spinelli / Denise Tremblay / Sylvain Moineau / Christian Cambillau / Adeline Goulet /
Abstract: Virulent phages infecting , an industry-relevant bacterium, pose a significant risk to the quality of the fermented milk products. Phages of the Skunavirus genus are by far the most isolated ...Virulent phages infecting , an industry-relevant bacterium, pose a significant risk to the quality of the fermented milk products. Phages of the Skunavirus genus are by far the most isolated lactococcal phages in the cheese environments and phage p2 is the model siphophage for this viral genus. The baseplate of phage p2, which is used to recognize its host, was previously shown to display two conformations by X-ray crystallography, a rested state and an activated state ready to bind to the host. The baseplate became only activated and opened in the presence of Ca. However, such an activated state was not previously observed in the virion. Here, using nanobodies binding to the baseplate, we report on the negative staining electron microscopy structure of the activated form of the baseplate directly observed in the p2 virion, that is compatible with the activated baseplate crystal structure. Analyses of this new structure also established the presence of a second distal tail (Dit) hexamer as a component of the baseplate, the topology of which differs largely from the first one. We also observed an uncoupling between the baseplate activation and the tail tip protein (Tal) opening, suggesting an infection mechanism more complex than previously expected.
History
DepositionJun 26, 2020-
Header (metadata) releaseAug 26, 2020-
Map releaseAug 26, 2020-
UpdateAug 26, 2020-
Current statusAug 26, 2020Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 2.5
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 2.5
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6zih
  • Surface level: 2.5
  • Imaged by UCSF Chimera
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  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-6zih
  • Imaged by Jmol
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_11226.png

Downloads & links

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Map

FileDownload / File: emd_11226.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationnegative staining 3D reconstruction of p2 virion baseplate bound to VHH5
Voxel sizeX=Y=Z: 3.46 Å
Density
Contour LevelBy AUTHOR: 2.5 / Movie #1: 2.5
Minimum - Maximum-13.012129 - 18.571333
Average (Standard dev.)-0.00000000115 (±0.99999994)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 885.76 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z3.463.463.46
M x/y/z256256256
origin x/y/z0.0000.0000.000
length x/y/z885.760885.760885.760
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS256256256
D min/max/mean-13.01218.571-0.000

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Supplemental data

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Sample components

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Entire : Lactococcus virus P2

EntireName: Lactococcus virus P2
Components
  • Virus: Lactococcus virus P2
    • Protein or peptide: Distal tail protein
    • Protein or peptide: Baseplate protein gp16Tripod (photography)
    • Protein or peptide: Receptor binding proteinReceptor (biochemistry)

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Supramolecule #1: Lactococcus virus P2

SupramoleculeName: Lactococcus virus P2 / type: virus / ID: 1 / Parent: 0 / Macromolecule list: all / NCBI-ID: 254252 / Sci species name: Lactococcus virus P2 / Virus type: VIRION / Virus isolate: STRAIN / Virus enveloped: No / Virus empty: No
Host (natural)Organism: Lactococcus lactis (lactic acid bacteria)

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Macromolecule #1: Distal tail protein

MacromoleculeName: Distal tail protein / type: protein_or_peptide / ID: 1 / Number of copies: 12 / Enantiomer: LEVO
Source (natural)Organism: Lactococcus phage p2 (virus)
Molecular weightTheoretical: 34.511992 KDa
SequenceString: MVRQYKIHTN LDGTDDKVWD VTNGKVRFYQ PSNLGLQSTN NIWQSNGIGV MGTRSITQPQ IEFKLETFGE SLEENYQLMK DFVNDILSK KFVTLEYQTE IFQVYADLAL ADVTKTEGYG KNGTFSEKIT FDIITKWYTY ENLTFDKIQN GKVIAGMSKI Y GGTAPGNY ...String:
MVRQYKIHTN LDGTDDKVWD VTNGKVRFYQ PSNLGLQSTN NIWQSNGIGV MGTRSITQPQ IEFKLETFGE SLEENYQLMK DFVNDILSK KFVTLEYQTE IFQVYADLAL ADVTKTEGYG KNGTFSEKIT FDIITKWYTY ENLTFDKIQN GKVIAGMSKI Y GGTAPGNY KYIKGTSYTY YGESDIDRLS RWDIKEEIFS FMGILYPKLP KTPAGVRFLD DIGNEYTAIV FKTEQVQDYI LI NTDVNDE TYQGWKGTTA LNLFPVMDFE RYRTRIIEKG QMELINLSKA EFKIKRKADF V

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Macromolecule #2: Baseplate protein gp16

MacromoleculeName: Baseplate protein gp16 / type: protein_or_peptide / ID: 2 / Number of copies: 3 / Enantiomer: LEVO
Source (natural)Organism: Lactococcus phage p2 (virus)
Molecular weightTheoretical: 42.883266 KDa
SequenceString: MLEANVYDNF NPNYYNISDF SMPNGKKEKR GLPIPKARCQ VINYELWETG YLYTSSATLT VSVEVGDIVQ ILFPEVVPIE EALGKKKKL NLDMVYLVTD VDESNKATLK NYFWAMIESL DVPNAITKTT NFAIIDYLID PNKNNLMSYG YFFNSSIFAG K ATINRKAE ...String:
MLEANVYDNF NPNYYNISDF SMPNGKKEKR GLPIPKARCQ VINYELWETG YLYTSSATLT VSVEVGDIVQ ILFPEVVPIE EALGKKKKL NLDMVYLVTD VDESNKATLK NYFWAMIESL DVPNAITKTT NFAIIDYLID PNKNNLMSYG YFFNSSIFAG K ATINRKAE TSSAHDVAKR IFSKVQFQPT TTIQHAPSET DPRNLLFINF ASRNWNRKRI TTRVDIKQSV TMDTETIVER SA YNFAVVF VKNKATDDYT DPPKMYIAKN NGDVIDYSTY HGDGTDLPDV RTAKTLFYDR DDHGNPPELS TIKVEISPST IVT RLIFNQ NELLPLYVND LVDIWYEGKL YSGYIADRVK TEFNDRLIFV ESGDKPNVI

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Macromolecule #3: Receptor binding protein

MacromoleculeName: Receptor binding protein / type: protein_or_peptide / ID: 3 / Number of copies: 18 / Enantiomer: LEVO
Source (natural)Organism: Lactococcus phage p2 (virus)
Molecular weightTheoretical: 28.668057 KDa
SequenceString: MTIKNFTFFS PNSTEFPVGS NNDGKLYMML TGMDYRTIRR KDWSSPLNTA LNVQYTNTSI IAGGRYFELL NETVALKGDS VNYIHANID LTQTANPVSL SAETANNSNG VDINNGSGVL KVCFDIVTTS GTGVTSTKPI VQTSTLDSIS VNDMTVSGSI D VPVQTLTV ...String:
MTIKNFTFFS PNSTEFPVGS NNDGKLYMML TGMDYRTIRR KDWSSPLNTA LNVQYTNTSI IAGGRYFELL NETVALKGDS VNYIHANID LTQTANPVSL SAETANNSNG VDINNGSGVL KVCFDIVTTS GTGVTSTKPI VQTSTLDSIS VNDMTVSGSI D VPVQTLTV EAGNGLQLQL TKKNNDLVIV RFFGSVSNIQ KGWNMSGTWV DRPFRPAAVQ SLVGHFAGRD TSFHIDINPN GS ITWWGAN IDKTPIATRG NGSYFIK

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Experimental details

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Structure determination

Methodnegative staining
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

BufferpH: 7.5
StainingType: NEGATIVE / Material: uranyl acetate

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Electron microscopy

MicroscopeFEI TECNAI SPIRIT
Electron beamAcceleration voltage: 120 kV / Electron source: LAB6
Electron opticsIllumination mode: SPOT SCAN / Imaging mode: BRIGHT FIELDBright-field microscopy
Image recordingFilm or detector model: OTHER / Average electron dose: 25.0 e/Å2
Experimental equipment
Model: Tecnai Spirit / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 293
CTF correctionSoftware - Name: CTFFIND
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.0)
Final 3D classificationSoftware - Name: RELION (ver. 3.0)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.0)
Final reconstructionApplied symmetry - Point group: D6 (2x6 fold dihedral) / Resolution.type: BY AUTHOR / Resolution: 28.7 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.0) / Number images used: 261

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Atomic model buiding 1

Initial modelPDB ID:

2x53

RefinementProtocol: RIGID BODY FIT
Output model

PDB-6zih:
Topological model of p2 virion baseplate in activated conformation

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