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- EMDB-2816: Electron cryoEM structure of lactococcal siphophage 1358 virion -

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Entry
Database: EMDB / Id: 2816
TitleElectron cryoEM structure of lactococcal siphophage 1358 virion
Map dataElectron cryo-microscopy reconstruction of the capsid of phage 1358
SampleCapsid of phage 1358:
virus
KeywordsLactococcus lactis / Siphoviridae / electron microscopy / 1358 phage / capsid
SourceLactococcus phage 1358 (bacteriophage)
Methodsingle particle reconstruction / cryo EM / Resolution: 16 Å
AuthorsSpinelli S / Bebeacua C / Orlov I / Tremblay D / Klaholz B / Moineau S / Cambillau C
CitationJournal: J. Virol. / Year: 2014
Title: Cryo-electron microscopy structure of lactococcal siphophage 1358 virion.
P-authors: Silvia Spinelli / Cecilia Bebeacua / Igor Orlov / Denise Tremblay / Bruno P Klaholz / Sylvain Moineau / Christian Cambillau /
Abstract: Lactococcus lactis, a Gram(+) lactic acid-producing bacterium used for the manufacture of several fermented dairy products, is subject to infection by diverse virulent tailed phages, leading to ...Lactococcus lactis, a Gram(+) lactic acid-producing bacterium used for the manufacture of several fermented dairy products, is subject to infection by diverse virulent tailed phages, leading to industrial fermentation failures. This constant viral risk has led to a sustained interest in the study of their biology, diversity, and evolution. Lactococcal phages now constitute a wide ensemble of at least 10 distinct genotypes within the Caudovirales order, many of them belonging to the Siphoviridae family. Lactococcal siphophage 1358, currently the only member of its group, displays a noticeably high genomic similarity to some Listeria phages as well as a host range limited to a few L. lactis strains. These genomic and functional characteristics stimulated our interest in this phage. Here, we report the cryo-electron microscopy structure of the complete 1358 virion. Phage 1358 exhibits noteworthy features, such as a capsid with dextro handedness and protruding decorations on its capsid and tail. Observations of the baseplate of virion particles revealed at least two conformations, a closed and an open, activated form. Functional assays uncovered that the adsorption of phage 1358 to its host is Ca(2+) independent, but this cation is necessary to complete its lytic cycle. Taken together, our results provide the complete structural picture of a unique lactococcal phage and expand our knowledge on the complex baseplate of phages of the Siphoviridae family.
Importance: Phages of Lactococcus lactis are investigated mainly because they are sources of milk fermentation failures in the dairy industry. Despite the availability of several antiphage measures, new phages keep emerging in this ecosystem. In this study, we provide the cryo-electron microscopy reconstruction of a unique lactococcal phage that possesses genomic similarity to particular Listeria phages and has a host range restricted to only a minority of L. lactis strains. The capsid of phage 1358 displays the almost unique characteristic of being dextro handed. Its capsid and tail exhibit decorations that we assigned to nonspecific sugar binding modules. We observed the baseplate of 1358 in two conformations, a closed and an open form. We also found that the adsorption to its host, but not infection, is Ca(2+) independent. Overall, this study advances our understanding of the adhesion mechanisms of siphophages.
DateDeposition: Nov 14, 2014 / Header (metadata) release: Dec 24, 2014 / Map release: Feb 17, 2016 / Last update: Feb 17, 2016

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Strvis

Movie
  • Surface view with section colored by density value
  • Surface level: 0.004
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by radius
  • Surface level: 0.004
  • Imaged by UCSF Chimera
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Structure viewerEM map:
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Supplemental images

Downlink

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Map

Fileemd_2816.map.gz (map file in CCP4 format, 54001 KB)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
240 pix
1.92 Å/pix.
= 460.8 Å
240 pix
1.92 Å/pix.
= 460.8 Å
240 pix
1.92 Å/pix.
= 460.8 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.92 Å
Density
Contour Level:0.004 (by author), 0.004 (movie #1):
Minimum - Maximum-0.00000352 - 0.03249559
Average (Standard dev.)0.00081323 (0.00288972)
Details

EMDB XML:

Space Group Number1
Map Geometry
Axis orderXYZ
Dimensions240240240
Origin636363
Limit302302302
Spacing240240240
CellA=B=C: 460.8 Å
α=β=γ: 90.0 deg.

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.921.921.92
M x/y/z240240240
origin x/y/z0.0000.0000.000
length x/y/z460.800460.800460.800
α/β/γ90.00090.00090.000
start NX/NY/NZ
NX/NY/NZ
MAP C/R/S123
start NC/NR/NS636363
NC/NR/NS240240240
D min/max/mean-0.0000.0320.001

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Supplemental data

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Sample components

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Entire Capsid of phage 1358

EntireName: Capsid of phage 1358 / Details: The sample contains the whole phage. / Number of components: 1 / Oligomeric State: icosahedral

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Component #1: virus, Lactococcus phage 1358

VirusName: Lactococcus phage 1358 / Class: OTHER / Empty: No / Enveloped: No / Isolate: SPECIES
SpeciesSpecies: Lactococcus phage 1358 (bacteriophage)
Source (natural)Host Species: Lactoccocus lactis / Host category: BACTERIA(EUBACTERIA)
Shell #1Name of element: T7 / Diameter: 600 Å / T number(triangulation number): 7

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Experimental details

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Sample preparation

SpecimenSpecimen state: particle / Method: cryo EM
Sample solutionBuffer solution: Phage Buffer / Ph: 7.4
VitrificationInstrument: FEI VITROBOT MARK I / Cryogen name: ETHANE / Humidity: 95 % / Method: Blot for 3 seconds before plunging. / Details: cryo plunge-freezing

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Electron microscopy imaging

Experimental equipment
Model: Tecnai F30 / Image courtesy: FEI Company
ImagingMicroscope: FEI TECNAI F30 / Date: Sep 1, 2012
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Electron dose: 10 e/Å2 / Illumination mode: SPOT SCAN
LensMagnification: 59000 X (nominal)
Astigmatism: Objective lens astigmatism was corrected at 100,000 times magnification.
Cs: 2.2 mm / Imaging mode: BRIGHT FIELD / Defocus: 1000 - 3500 nm / Energy filter: FEI
Specimen HolderModel: GATAN LIQUID NITROGEN
CameraDetector: FEI EAGLE (4k x 4k)

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Image acquisition

Image acquisitionNumber of digital images: 200 / Sampling size: 16 microns / Bit depth: 16

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Image processing

ProcessingMethod: single particle reconstruction / Applied symmetry: I (icosahedral) / Number of projections: 9211
Details: capsid reconstructed using a Maximum Likelihood Approach.
3D reconstructionAlgorithm: Maximum Likelihood / Software: EMAN2, SPIDER, Xmipp / CTF correction: Images / Resolution: 16 Å / Resolution method: FSC 0.5, gold-standard

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