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- EMDB-1231: Structure of the connector of bacteriophage T7 at 8A resolution: ... -

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Basic information

Entry
Database: EMDB / ID: EMD-1231
TitleStructure of the connector of bacteriophage T7 at 8A resolution: structural homologies of a basic component of a DNA translocating machinery.
Map data3D map of T7-connector
Sample
  • Sample: Connector of Bacteriophage T7
  • Protein or peptide: x 12 types
Biological speciesEnterobacteria phage T7 (virus)
Methodsingle particle reconstruction / cryo EM / negative staining / Resolution: 8.0 Å
AuthorsAgirrezabala X / Martin-Benito J / Valle M / Gonzalez JM / Valencia A / Valpuesta JM / Carrascosa JL
CitationJournal: J Mol Biol / Year: 2005
Title: Structure of the connector of bacteriophage T7 at 8A resolution: structural homologies of a basic component of a DNA translocating machinery.
Authors: Xabier Agirrezabala / Jaime Martín-Benito / Mikel Valle / José M González / Alfonso Valencia / José María Valpuesta / José L Carrascosa /
Abstract: The three-dimensional structure of the bacteriophage T7 head-to-tail connector has been obtained at 8A resolution using cryo-electron microscopy and single-particle analysis from purified recombinant ...The three-dimensional structure of the bacteriophage T7 head-to-tail connector has been obtained at 8A resolution using cryo-electron microscopy and single-particle analysis from purified recombinant connectors. The general morphology of the T7 connector is that of a 12-folded toroidal homopolymer with a channel that runs along the longitudinal axis of the particle. The structure of the T7 connector reveals many structural similarities with the connectors from other bacteriophages. Docking of the atomic structure of the varphi29 connector into the three-dimensional reconstruction of T7 connector reveals that the narrow, distal region of the two oligomers are almost identical. This region of the varphi29 connector has been suggested to be involved in DNA translocation, and is composed of an alpha-beta-alpha-beta-beta-alpha motif. A search for alpha-helices in the same region of the T7 three-dimensional map has located three alpha-helices in approximately the same position as those of the varphi29 connector. A comparison of the predicted secondary structure of several bacteriophage connectors, including among others T7, varphi29, P22 and SPP1, reveals that, despite the lack of sequence homology, they seem to contain the same alpha-beta-alpha-beta-beta-alpha motif as that present in the varphi29 connector. These results allow us to suggest a common architecture related to a basic component of the DNA translocating machinery for several viruses.
History
DepositionMay 25, 2006-
Header (metadata) releaseMay 25, 2006-
Map releaseMay 25, 2007-
UpdateMay 26, 2011-
Current statusMay 26, 2011Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.48780768
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 0.48780768
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

1231.gif

Downloads & links

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Map

FileDownload / File: emd_1231.map.gz / Format: CCP4 / Size: 10.2 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotation3D map of T7-connector
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
2.18 Å/pix.
x 140 pix.
= 305.2 Å		2.18 Å/pix.
x 140 pix.
= 305.2 Å		2.18 Å/pix.
x 140 pix.
= 305.2 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 2.18 Å
Density

Histogram

Histogram (log scale)
Contour Level1: 0.371 / Movie #1: 0.4878077
Minimum - Maximum-0.535083 - 5.68371
Average (Standard dev.)0.0686053 (±0.332779)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions140140140
Spacing140140140
CellA=B=C: 305.2 Å
α=β=γ: 90 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z2.182.182.18
M x/y/z140140140
origin x/y/z0.0000.0000.000
length x/y/z305.200305.200305.200
α/β/γ90.00090.00090.000
start NX/NY/NZ-64-64-64
NX/NY/NZ128128128
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS140140140
D min/max/mean-0.5355.6840.069

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Supplemental data

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Sample components

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Entire : Connector of Bacteriophage T7

EntireName: Connector of Bacteriophage T7
Components
  • Sample: Connector of Bacteriophage T7
  • Protein or peptide: gp8
  • Protein or peptide: gp8
  • Protein or peptide: gp8
  • Protein or peptide: gp8
  • Protein or peptide: gp8
  • Protein or peptide: gp8
  • Protein or peptide: gp8
  • Protein or peptide: gp8
  • Protein or peptide: gp8
  • Protein or peptide: gp8
  • Protein or peptide: gp8
  • Protein or peptide: gp8

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Supramolecule #1000: Connector of Bacteriophage T7

SupramoleculeName: Connector of Bacteriophage T7 / type: sample / ID: 1000 / Oligomeric state: dodecamer / Number unique components: 12
Molecular weightExperimental: 708 KDa / Theoretical: 708 KDa

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Macromolecule #1: gp8

MacromoleculeName: gp8 / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Oligomeric state: monomer / Recombinant expression: Yes
Source (natural)Organism: Enterobacteria phage T7 (virus) / synonym: phage T7
Molecular weightExperimental: 57 MDa / Theoretical: 57 MDa
Recombinant expressionOrganism: Escherichia coli BL21 (bacteria) / Recombinant plasmid: pAR2545

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Macromolecule #2: gp8

MacromoleculeName: gp8 / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Oligomeric state: monomer / Recombinant expression: Yes
Source (natural)Organism: Enterobacteria phage T7 (virus) / synonym: phage T7
Molecular weightExperimental: 57 MDa / Theoretical: 57 MDa
Recombinant expressionOrganism: Escherichia coli BL21 (bacteria) / Recombinant plasmid: pAR2545

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Macromolecule #3: gp8

MacromoleculeName: gp8 / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Oligomeric state: monomer / Recombinant expression: Yes
Source (natural)Organism: Enterobacteria phage T7 (virus) / synonym: phage T7
Molecular weightExperimental: 57 MDa / Theoretical: 57 MDa
Recombinant expressionOrganism: Escherichia coli BL21 (bacteria) / Recombinant plasmid: pAR2545

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Macromolecule #4: gp8

MacromoleculeName: gp8 / type: protein_or_peptide / ID: 4 / Number of copies: 1 / Oligomeric state: monomer / Recombinant expression: Yes
Source (natural)Organism: Enterobacteria phage T7 (virus) / synonym: phage T7
Molecular weightExperimental: 57 MDa / Theoretical: 57 MDa
Recombinant expressionOrganism: Escherichia coli BL21 (bacteria) / Recombinant plasmid: pAR2545

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Macromolecule #5: gp8

MacromoleculeName: gp8 / type: protein_or_peptide / ID: 5 / Number of copies: 1 / Oligomeric state: monomer / Recombinant expression: Yes
Source (natural)Organism: Enterobacteria phage T7 (virus) / synonym: phage T7
Molecular weightExperimental: 57 MDa / Theoretical: 57 MDa
Recombinant expressionOrganism: Escherichia coli BL21 (bacteria) / Recombinant plasmid: pAR2545

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Macromolecule #6: gp8

MacromoleculeName: gp8 / type: protein_or_peptide / ID: 6 / Number of copies: 1 / Oligomeric state: monomer / Recombinant expression: Yes
Source (natural)Organism: Enterobacteria phage T7 (virus) / synonym: phage T7
Molecular weightExperimental: 57 MDa / Theoretical: 57 MDa
Recombinant expressionOrganism: Escherichia coli BL21 (bacteria) / Recombinant plasmid: pAR2545

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Macromolecule #7: gp8

MacromoleculeName: gp8 / type: protein_or_peptide / ID: 7 / Number of copies: 1 / Oligomeric state: monomer / Recombinant expression: Yes
Source (natural)Organism: Enterobacteria phage T7 (virus) / synonym: phage T7
Molecular weightExperimental: 57 MDa / Theoretical: 57 MDa
Recombinant expressionOrganism: Escherichia coli BL21 (bacteria) / Recombinant plasmid: pAR2545

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Macromolecule #8: gp8

MacromoleculeName: gp8 / type: protein_or_peptide / ID: 8 / Number of copies: 1 / Oligomeric state: monomer / Recombinant expression: Yes
Source (natural)Organism: Enterobacteria phage T7 (virus) / synonym: phage T7
Molecular weightExperimental: 57 MDa / Theoretical: 57 MDa
Recombinant expressionOrganism: Escherichia coli BL21 (bacteria) / Recombinant plasmid: pAR2545

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Macromolecule #9: gp8

MacromoleculeName: gp8 / type: protein_or_peptide / ID: 9 / Number of copies: 1 / Oligomeric state: monomer / Recombinant expression: Yes
Source (natural)Organism: Enterobacteria phage T7 (virus) / synonym: phage T7
Molecular weightExperimental: 57 MDa / Theoretical: 57 MDa
Recombinant expressionOrganism: Escherichia coli BL21 (bacteria) / Recombinant plasmid: pAR2545

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Macromolecule #10: gp8

MacromoleculeName: gp8 / type: protein_or_peptide / ID: 10 / Number of copies: 1 / Oligomeric state: monomer / Recombinant expression: Yes
Source (natural)Organism: Enterobacteria phage T7 (virus) / synonym: phage T7
Molecular weightExperimental: 57 MDa / Theoretical: 57 MDa
Recombinant expressionOrganism: Escherichia coli BL21 (bacteria) / Recombinant plasmid: pAR2545

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Macromolecule #11: gp8

MacromoleculeName: gp8 / type: protein_or_peptide / ID: 11 / Number of copies: 1 / Oligomeric state: monomer / Recombinant expression: Yes
Source (natural)Organism: Enterobacteria phage T7 (virus) / synonym: phage T7
Molecular weightExperimental: 57 MDa / Theoretical: 57 MDa
Recombinant expressionOrganism: Escherichia coli BL21 (bacteria) / Recombinant plasmid: pAR2545

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Macromolecule #12: gp8

MacromoleculeName: gp8 / type: protein_or_peptide / ID: 12 / Number of copies: 1 / Oligomeric state: monomer / Recombinant expression: Yes
Source (natural)Organism: Enterobacteria phage T7 (virus) / synonym: phage T7
Molecular weightExperimental: 57 MDa / Theoretical: 57 MDa
Recombinant expressionOrganism: Escherichia coli BL21 (bacteria) / Recombinant plasmid: pAR2545

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Experimental details

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Structure determination

Methodnegative staining, cryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration0.4 mg/mL
BufferpH: 7.8
Details: 20mM Tris HCl pH:7.8, 3mM B-mercaptoethanol, 4%(v/v) glicerol, 2mM Na3EDTA, 100 mM NaCl
StainingType: NEGATIVE / Details: cryoEM
GridDetails: Quantifoil grids with 2 um hole size
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TECNAI F20
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsCalibrated magnification: 64220 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.26 mm / Nominal defocus max: 2.3 µm / Nominal defocus min: 0.9 µm / Nominal magnification: 62000
Sample stageSpecimen holder: Side entry liquid nitrogen-cooled cryo specimen holder, GATAN. Eucentric
Specimen holder model: GATAN LIQUID NITROGEN
Alignment procedureLegacy - Astigmatism: FFT with ssCCD
Image recordingCategory: FILM / Film or detector model: KODAK SO-163 FILM / Digitization - Scanner: ZEISS SCAI / Digitization - Sampling interval: 14 µm / Average electron dose: 10 e/Å2
Tilt angle min0
Tilt angle max0
Experimental equipment
Model: Tecnai F20 / Image courtesy: FEI Company

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Image processing

CTF correctionDetails: Wiener filter, defocus groups
Final angle assignmentDetails: SPIDER
Final reconstructionApplied symmetry - Point group: C12 (12 fold cyclic) / Algorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 8.0 Å / Resolution method: FSC 0.5 CUT-OFF / Software - Name: SPIDER / Number images used: 26492

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