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

Basic information

• Entry: Database: EMDB / ID: EMD-2816
• Title: Electron cryoEM structure of lactococcal siphophage 1358 virion
• Map data: Electron cryo-microscopy reconstruction of the capsid of phage 1358

Sample

• Sample: Capsid of phage 1358
• Virus: Lactococcus phage 1358 (virus)

• Keywords: Lactococcus lactis / Siphoviridae / electron microscopy / 1358 phage / capsid
• Biological species: Lactococcus phage 1358 (virus)
• Method: single particle reconstruction / cryo EM / Resolution: 16.0 Å
• Authors: Spinelli S / Bebeacua C / Orlov I / Tremblay D / Klaholz B / Moineau S / Cambillau C
• Citation: Journal: J Virol / Year: 2014; Title: Cryo-electron microscopy structure of lactococcal siphophage 1358 virion.; 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 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.

History

Deposition	Nov 14, 2014	-
Header (metadata) release	Dec 24, 2014	-
Map release	Feb 17, 2016	-
Update	Feb 17, 2016	-
Current status	Feb 17, 2016	Processing site: PDBe / Status: Released

Map

• File: Download / File: emd_2816.map.gz / Format: CCP4 / Size: 51.5 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
• Annotation: Electron cryo-microscopy reconstruction of the capsid of phage 1358
• Voxel size: X=Y=Z: 1.92 Å

Density

Contour Level	By AUTHOR: 0.004 / Movie #1: 0.004
Minimum - Maximum	-0.00000352 - 0.03249559
Average (Standard dev.)	0.00081323 (±0.00288972)

• Symmetry: Space group: 1

Details

EMDB XML:

Map geometry	Axis order X Y Z Origin 63 63 63 Dimensions 240 240 240 Spacing 240 240 240
Cell	A=B=C: 460.8 Å α=β=γ: 90.0 °

CCP4 map header:

mode	Image stored as Reals
Å/pix. X/Y/Z	1.92	1.92	1.92
M x/y/z	240	240	240
origin x/y/z	0.000	0.000	0.000
length x/y/z	460.800	460.800	460.800
α/β/γ	90.000	90.000	90.000
MAP C/R/S	1	2	3
start NC/NR/NS	63	63	63
NC/NR/NS	240	240	240
D min/max/mean	-0.000	0.032	0.001

Supplemental data

Sample components

Entire : Capsid of phage 1358

• Entire: Name: Capsid of phage 1358

Components

• Sample: Capsid of phage 1358
• Virus: Lactococcus phage 1358 (virus)

Supramolecule #1000: Capsid of phage 1358

• Supramolecule: Name: Capsid of phage 1358 / type: sample / ID: 1000 / Details: The sample contains the whole phage. / Oligomeric state: icosahedral / Number unique components: 1

Supramolecule #1: Lactococcus phage 1358

• Supramolecule: Name: Lactococcus phage 1358 / type: virus / ID: 1 / NCBI-ID: 741942 / Sci species name: Lactococcus phage 1358 / Virus type: OTHER / Virus isolate: SPECIES / Virus enveloped: No / Virus empty: No
• Host (natural): Organism: Lactoccocus lactis / synonym: BACTERIA(EUBACTERIA)
• Virus shell: Shell ID: 1 / Name: T7 / Diameter: 600 Å / T number (triangulation number): 7

Experimental details

Structure determination

• Method: cryo EM
• Processing: single particle reconstruction
• Aggregation state: particle

Sample preparation

• Buffer: pH: 7.4 / Details: Phage Buffer
• Vitrification: Cryogen name: ETHANE / Chamber humidity: 95 % / Instrument: FEI VITROBOT MARK I / Details: cryo plunge-freezing / Method: Blot for 3 seconds before plunging.

Electron microscopy

• Microscope: FEI TECNAI F30
• Electron beam: Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
• Electron optics: Illumination mode: SPOT SCAN / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.2 mm / Nominal defocus max: 3.5 µm / Nominal defocus min: 1.0 µm / Nominal magnification: 59000
• Specialist optics: Energy filter - Name: FEI
• Sample stage: Specimen holder model: GATAN LIQUID NITROGEN
• Alignment procedure: Legacy - Astigmatism: Objective lens astigmatism was corrected at 100,000 times magnification.
• Date: Sep 1, 2012
• Image recording: Category: CCD / Film or detector model: FEI EAGLE (4k x 4k) / Digitization - Sampling interval: 16 µm / Number real images: 200 / Average electron dose: 10 e/Ų / Bits/pixel: 16
• Experimental equipment: Model: Tecnai F30 / Image courtesy: FEI Company

Image processing

• CTF correction: Details: Images
• Final reconstruction: Applied symmetry - Point group: I (icosahedral) / Algorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 16.0 Å / Resolution method: OTHER / Software - Name: EMAN2, SPIDER, Xmipp / Number images used: 9211
• Details: capsid reconstructed using a Maximum Likelihood Approach.