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- EMDB-10911: Cryo-EM structure of T7 bacteriophage DNA translocation gp15 core... -

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Basic information

Entry
Database: EMDB / ID: EMD-10911
TitleCryo-EM structure of T7 bacteriophage DNA translocation gp15 core protein intermediate assembly
Map dataT7 DNA ejection complex
Sample
  • Complex: gp15 tubular core protein
    • Protein or peptide: Internal virion protein gp15
KeywordsVIRAL PROTEIN / DNA TRANSLOCATION / VIRAL INFECTION / PERIPLASMIC SPACE COMPLEX.
Function / homologyInternal virion protein Gp15 / host cell periplasmic space / symbiont genome ejection through host cell envelope, short tail mechanism / virion component / Internal virion protein gp15
Function and homology information
Biological speciesEscherichia phage T7 (virus)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.6 Å
AuthorsPerez-Ruiz M / Pulido-Cid M
Funding support Spain, 3 items
OrganizationGrant numberCountry
Spanish Ministry of Science, Innovation, and UniversitiesBFU 2014-54181 Spain
Spanish Ministry of Science, Innovation, and UniversitiesSEV-2013-0347 Spain
Spanish Ministry of Science, Innovation, and UniversitiesBES-2015-073615 Spain
CitationJournal: Proc Natl Acad Sci U S A / Year: 2021
Title: Assisted assembly of bacteriophage T7 core components for genome translocation across the bacterial envelope.
Authors: Mar Pérez-Ruiz / Mar Pulido-Cid / Juan Román Luque-Ortega / José María Valpuesta / Ana Cuervo / José L Carrascosa /
Abstract: In most bacteriophages, genome transport across bacterial envelopes is carried out by the tail machinery. In viruses of the family, in which the tail is not long enough to traverse the bacterial ...In most bacteriophages, genome transport across bacterial envelopes is carried out by the tail machinery. In viruses of the family, in which the tail is not long enough to traverse the bacterial wall, it has been postulated that viral core proteins assembled inside the viral head are translocated and reassembled into a tube within the periplasm that extends the tail channel. Bacteriophage T7 infects , and despite extensive studies, the precise mechanism by which its genome is translocated remains unknown. Using cryo-electron microscopy, we have resolved the structure of two different assemblies of the T7 DNA translocation complex composed of the core proteins gp15 and gp16. Gp15 alone forms a partially folded hexamer, which is further assembled upon interaction with gp16 into a tubular structure, forming a channel that could allow DNA passage. The structure of the gp15-gp16 complex also shows the location within gp16 of a canonical transglycosylase motif involved in the degradation of the bacterial peptidoglycan layer. This complex docks well in the tail extension structure found in the periplasm of T7-infected bacteria and matches the sixfold symmetry of the phage tail. In such cases, gp15 and gp16 that are initially present in the T7 capsid eightfold-symmetric core would change their oligomeric state upon reassembly in the periplasm. Altogether, these results allow us to propose a model for the assembly of the core translocation complex in the periplasm, which furthers understanding of the molecular mechanism involved in the release of T7 viral DNA into the bacterial cytoplasm.
History
DepositionApr 23, 2020-
Header (metadata) releaseSep 8, 2021-
Map releaseSep 8, 2021-
UpdateJul 10, 2024-
Current statusJul 10, 2024Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.0114
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 0.0114
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6ysz
  • Surface level: 0.02
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_10911.png

Downloads & links

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Map

FileDownload / File: emd_10911.map.gz / Format: CCP4 / Size: 52.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationT7 DNA ejection complex
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.85 Å/pix.
x 240 pix.
= 204. Å		0.85 Å/pix.
x 240 pix.
= 204. Å		0.85 Å/pix.
x 240 pix.
= 204. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.85 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.0114 / Movie #1: 0.0114
Minimum - Maximum-0.003546688 - 0.22533487
Average (Standard dev.)0.00015627204 (±0.0071606752)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions240240240
Spacing240240240
CellA=B=C: 204.0 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z0.850.850.85
M x/y/z240240240
origin x/y/z0.0000.0000.000
length x/y/z204.000204.000204.000
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS240240240
D min/max/mean-0.0040.2250.000

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Supplemental data

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Sample components

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Entire : gp15 tubular core protein

EntireName: gp15 tubular core protein
Components
  • Complex: gp15 tubular core protein
    • Protein or peptide: Internal virion protein gp15

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Supramolecule #1: gp15 tubular core protein

SupramoleculeName: gp15 tubular core protein / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Escherichia phage T7 (virus)
Molecular weightTheoretical: 543 KDa

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Macromolecule #1: Internal virion protein gp15

MacromoleculeName: Internal virion protein gp15 / type: protein_or_peptide / ID: 1 / Number of copies: 6 / Enantiomer: LEVO
Source (natural)Organism: Escherichia phage T7 (virus)
Molecular weightTheoretical: 88.377219 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MRGSHHHHHH GMASMTGGQQ MGRDLYDDDD KDPSSMSKIE SALQAAQPGL SRLRGGAGGM GYRAATTQAE QPRSSLLDTI GRFAKAGAD MYTAKEQRAR DLADERSNEI IRKLTPEQRR EALNNGTLLY QDDPYAMEAL RVKTGRNAAY LVDDDVMQKI K EGVFRTRE ...String:
MRGSHHHHHH GMASMTGGQQ MGRDLYDDDD KDPSSMSKIE SALQAAQPGL SRLRGGAGGM GYRAATTQAE QPRSSLLDTI GRFAKAGAD MYTAKEQRAR DLADERSNEI IRKLTPEQRR EALNNGTLLY QDDPYAMEAL RVKTGRNAAY LVDDDVMQKI K EGVFRTRE EMEEYRHSRL QEGAKVYAEQ FGIDPEDVDY QRGFNGDITE RNISLYGAHD NFLSQQAQKG AIMNSRVELN GV LQDPDML RRPDSADFFE KYIDNGLVTG AIPSDAQATQ LISQAFSDAS SRAGGADFLM RVGDKKVTLN GATTTYRELI GEE QWNALM VTAQRSQFET DAKLNEQYRL KINSALNQED PRTAWEMLQG IKAELDKVQP DEQMTPQREW LISAQEQVQN QMNA WTKAQ AKALDDSMKS MNKLDVIDKQ FQKRINGEWV STDFKDMPVN ENTGEFKHSD MVNYANKKLA EIDSMDIPDG AKDAM KLKY LQADSKDGAF RTAIGTMVTD AGQEWSAAVI NGKLPERTPA MDALRRIRNA DPQLIAALYP DQAELFLTMD MMDKQG IDP QVILDADRLT VKRSKEQRFE DDKAFESALN ASKAPEIARM PASLRESARK IYDSVKYRSG NESMAMEQMT KFLKEST YT FTGDDVDGDT VGVIPKNMMQ VNSDPKSWEQ GRDILEEARK GIIASNPWIT NKQLTMYSQG DSIYLMDTTG QVRVRYDK E LLSKVWSENQ KKLEEKAREK ALADVNKRAP IVAATKAREA AAKRVREKRK QTPKFIYGRK E

UniProtKB: Internal virion protein gp15

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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

Concentration1 mg/mL
BufferpH: 8
Component:
ConcentrationFormula
100.0 mMTris
100.0 mMNaCitrate
10.0 mMMgCl2
GridModel: Quantifoil R2/2 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 60 sec. / Pretreatment - Atmosphere: AIR
VitrificationCryogen name: ETHANE / Chamber humidity: 95 % / Chamber temperature: 293.15 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TALOS ARCTICA
Image recordingFilm or detector model: FEI FALCON III (4k x 4k) / Detector mode: COUNTING / Number grids imaged: 1 / Number real images: 2470 / Average exposure time: 35.0 sec. / Average electron dose: 30.0 e/Å2
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsCalibrated magnification: 120000 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Talos Arctica / Image courtesy: FEI Company

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Image processing

Startup modelType of model: INSILICO MODEL
Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: C6 (6 fold cyclic) / Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 3.6 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION / Number images used: 50980
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION
Final 3D classificationSoftware - Name: RELION
FSC plot (resolution estimation)

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Atomic model buiding 1

RefinementSpace: REAL / Protocol: AB INITIO MODEL
Output model

PDB-6ysz:
Cryo-EM structure of T7 bacteriophage DNA translocation gp15 core protein intermediate assembly

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