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- EMDB-5690: Cryo-electron microscopy of T7 tail complex formed by gp8, gp11, ... -

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Basic information

Entry
Database: EMDB / ID: EMD-5690
TitleCryo-electron microscopy of T7 tail complex formed by gp8, gp11, and gp12 proteins
Map dataReconstuction of T7 tail complex formed by proteins gp8, gp11, and gp12
Sample
  • Sample: T7 tail complex formed by proteins gp8, gp11, and gp12
  • Protein or peptide: gp8
  • Protein or peptide: gp11
  • Protein or peptide: gp12
KeywordsBacteriophage / DNA ejection / tail complex / connector / gatekeeper
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
Similarity search - Component
Biological speciesEnterobacteria phage T7 (virus)
Methodsingle particle reconstruction / cryo EM / Resolution: 12.0 Å
AuthorsCuervo A / Pulido-Cid M / Chagoyen M / Arranz R / Gonzalez-Garcia VA / Garcia-Doval C / Caston JR / Valpuesta JM / van Raaij MJ / Martin-Benito J / Carrascosa JL
CitationJournal: J Biol Chem / Year: 2013
Title: Structural characterization of the bacteriophage T7 tail machinery.
Authors: Ana Cuervo / Mar Pulido-Cid / Mónica Chagoyen / Rocío Arranz / Verónica A González-García / Carmela Garcia-Doval / José R Castón / José M Valpuesta / Mark J van Raaij / Jaime Martín- ...Authors: Ana Cuervo / Mar Pulido-Cid / Mónica Chagoyen / Rocío Arranz / Verónica A González-García / Carmela Garcia-Doval / José R Castón / José M Valpuesta / Mark J van Raaij / Jaime Martín-Benito / José L Carrascosa /
Abstract: Most bacterial viruses need a specialized machinery, called "tail," to inject their genomes inside the bacterial cytoplasm without disrupting the cellular integrity. Bacteriophage T7 is a well ...Most bacterial viruses need a specialized machinery, called "tail," to inject their genomes inside the bacterial cytoplasm without disrupting the cellular integrity. Bacteriophage T7 is a well characterized member of the Podoviridae family infecting Escherichia coli, and it has a short noncontractile tail that assembles sequentially on the viral head after DNA packaging. The T7 tail is a complex of around 2.7 MDa composed of at least four proteins as follows: the connector (gene product 8, gp8), the tail tubular proteins gp11 and gp12, and the fibers (gp17). Using cryo-electron microscopy and single particle image reconstruction techniques, we have determined the precise topology of the tail proteins by comparing the structure of the T7 tail extracted from viruses and a complex formed by recombinant gp8, gp11, and gp12 proteins. Furthermore, the order of assembly of the structural components within the complex was deduced from interaction assays with cloned and purified tail proteins. The existence of common folds among similar tail proteins allowed us to obtain pseudo-atomic threaded models of gp8 (connector) and gp11 (gatekeeper) proteins, which were docked into the corresponding cryo-EM volumes of the tail complex. This pseudo-atomic model of the connector-gatekeeper interaction revealed the existence of a common molecular architecture among viruses belonging to the three tailed bacteriophage families, strongly suggesting that a common molecular mechanism has been favored during evolution to coordinate the transition between DNA packaging and tail assembly.
History
DepositionJun 10, 2013-
Header (metadata) releaseJul 3, 2013-
Map releaseAug 7, 2013-
UpdateSep 18, 2013-
Current statusSep 18, 2013Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.01314
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 0.01314
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-3j4a, PDB-3j4b
  • Surface level: 0.01314
  • Imaged by UCSF Chimera
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  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-3j4a
  • Imaged by Jmol
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  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-3j4b
  • Imaged by Jmol
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_5690_1.jpg

Downloads & links

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Map

FileDownload / File: emd_5690.map.gz / Format: CCP4 / Size: 15.3 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationReconstuction of T7 tail complex formed by proteins gp8, gp11, and gp12
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
2.75 Å/pix.
x 160 pix.
= 440. Å		2.75 Å/pix.
x 160 pix.
= 440. Å		2.75 Å/pix.
x 160 pix.
= 440. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 2.75 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.01314 / Movie #1: 0.01314
Minimum - Maximum-0.04213833 - 0.04882427
Average (Standard dev.)0.00034643 (±0.00402175)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions160160160
Spacing160160160
CellA=B=C: 440.0 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z2.752.752.75
M x/y/z160160160
origin x/y/z0.0000.0000.000
length x/y/z440.000440.000440.000
α/β/γ90.00090.00090.000
start NX/NY/NZ-132-122-147
NX/NY/NZ250274261
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS160160160
D min/max/mean-0.0420.0490.000

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Supplemental data

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Sample components

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Entire : T7 tail complex formed by proteins gp8, gp11, and gp12

EntireName: T7 tail complex formed by proteins gp8, gp11, and gp12
Components
  • Sample: T7 tail complex formed by proteins gp8, gp11, and gp12
  • Protein or peptide: gp8
  • Protein or peptide: gp11
  • Protein or peptide: gp12

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Supramolecule #1000: T7 tail complex formed by proteins gp8, gp11, and gp12

SupramoleculeName: T7 tail complex formed by proteins gp8, gp11, and gp12
type: sample / ID: 1000 / Oligomeric state: gp8 (12mer), gp11 (12mer), gp12 (6mer) / Number unique components: 3
Molecular weightTheoretical: 1.5 MDa

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Macromolecule #1: gp8

MacromoleculeName: gp8 / type: protein_or_peptide / ID: 1 / Name.synonym: Connector / Number of copies: 12 / Oligomeric state: 12mer / Recombinant expression: Yes
Source (natural)Organism: Enterobacteria phage T7 (virus) / synonym: T7 Bacteriophage
Molecular weightTheoretical: 59 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli) / Recombinant strain: C41 / Recombinant plasmid: pRSET-B

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Macromolecule #2: gp11

MacromoleculeName: gp11 / type: protein_or_peptide / ID: 2 / Name.synonym: gatekeeper / Details: This protein was cloned with and His-tag / Number of copies: 12 / Oligomeric state: 12mer / Recombinant expression: Yes
Source (natural)Organism: Enterobacteria phage T7 (virus) / synonym: T7 bacteriophage
Molecular weightTheoretical: 25 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli) / Recombinant strain: C41 / Recombinant plasmid: pRSET-B

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Macromolecule #3: gp12

MacromoleculeName: gp12 / type: protein_or_peptide / ID: 3 / Number of copies: 6 / Oligomeric state: 6mer / Recombinant expression: No / Database: NCBI
Source (natural)Organism: Enterobacteria phage T7 (virus) / synonym: T7 bacteriophage
Molecular weightTheoretical: 90 KDa

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration0.1 mg/mL
BufferpH: 7.8 / Details: 50mM Tris-HCl, 10mM MgCl2, 100 mM NaCl
GridDetails: R2/2 Quantifoil coated with a thin carbon layer
VitrificationCryogen name: ETHANE / Instrument: LEICA EM CPC
Method: Samples were applied to grids for 1 minute, blotted, and plunged into liquid ethane.

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Electron microscopy

MicroscopeFEI TECNAI F20
TemperatureMin: 91 K / Max: 113 K
DateNov 8, 2012
Image recordingCategory: CCD / Film or detector model: FEI EAGLE (4k x 4k) / Number real images: 264 / Average electron dose: 10 e/Å2
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsCalibrated magnification: 108696 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.26 mm / Nominal defocus max: 3.5 µm / Nominal defocus min: 1.5 µm / Nominal magnification: 108696
Sample stageSpecimen holder model: GATAN LIQUID NITROGEN
Experimental equipment
Model: Tecnai F20 / Image courtesy: FEI Company

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Image processing

CTF correctionDetails: each micrograph
Final reconstructionAlgorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 12.0 Å / Resolution method: FSC 0.33 CUT-OFF / Software - Name: XMIPP, SPIDER, EMAN / Number images used: 1820

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Atomic model buiding 1

Initial modelPDB ID:

1vt0
PDB Unreleased entry

SoftwareName: Chimera
DetailsOne monomer was manually fitted and then the oligomer was generated using SITUS program.
RefinementSpace: REAL / Protocol: RIGID BODY FIT / Target criteria: Volumetric correlation
Output model

PDB-3j4a:
Structure of gp8 connector protein

PDB-3j4b:
Structure of T7 gatekeeper protein (gp11)

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Atomic model buiding 2

Initial modelPDB ID:

3lj5


Chain - Chain ID: L
SoftwareName: Chimera
DetailsOne monomer was manually fitted and then the oligomer was generated using SITUS program.
RefinementSpace: REAL / Protocol: RIGID BODY FIT / Target criteria: Volumetric correlation
Output model

PDB-3j4a:
Structure of gp8 connector protein

PDB-3j4b:
Structure of T7 gatekeeper protein (gp11)

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