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- EMDB-5594: Cryo-EM structure of short shafted adenovirus type 5 complexed wi... -

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Basic information

Entry
Database: EMDB / ID: EMD-5594
TitleCryo-EM structure of short shafted adenovirus type 5 complexed with factor VII
Map dataReconstruction of adenovirus type 5 modified to have a short shafted fiber with coagulation factor VII bound
Sample
  • Sample: Human zymogen coagulation factor VII bound to human adenovirus type 5 from species C. The virus has been modified to contain a short shafted fiber.
  • Virus: Human adenovirus 5
KeywordsAdenovirus / coagulation factor VII / innate immunity
Biological speciesHuman adenovirus 5
Methodsingle particle reconstruction / cryo EM / Resolution: 8.7 Å
AuthorsIrons EE / Flatt JW / Doronin K / Fox TL / Sumida JP / Acchione M / Stewart PL / Shayakhmetov DM
CitationJournal: J Virol / Year: 2013
Title: Coagulation factor binding orientation and dimerization may influence infectivity of adenovirus-coagulation factor complexes.
Authors: Eric E Irons / Justin W Flatt / Konstantin Doronin / Tara L Fox / Mauro Acchione / Phoebe L Stewart / Dmitry M Shayakhmetov /
Abstract: Adenoviruses (Ads) are promising vectors for therapeutic interventions in humans. When injected into the bloodstream, Ad vectors can bind several vitamin K-dependent blood coagulation factors, which ...Adenoviruses (Ads) are promising vectors for therapeutic interventions in humans. When injected into the bloodstream, Ad vectors can bind several vitamin K-dependent blood coagulation factors, which contributes to virus sequestration in the liver by facilitating transduction of hepatocytes. Although both coagulation factors FVII and FX bind the hexon protein of human Ad serotype 5 (HAdv5) with a very high affinity, only FX appears to play a role in mediating Ad-hepatocyte transduction in vivo. To understand the discrepancy between efficacy of FVII binding to hexon and its apparently poor capacity for supporting virus cell entry, we analyzed the HAdv5-FVII complex by using high-resolution cryo-electron microscopy (cryo-EM) followed by molecular dynamic flexible fitting (MDFF) simulations. The results indicate that although hexon amino acids T423, E424, and T425, identified earlier as critical for FX binding, are also involved in mediating binding of FVII, the FVII GLA domain sits within the surface-exposed hexon trimer depression in a different orientation from that found for FX. Furthermore, we found that when bound to hexon, two proximal FVII molecules interact via their serine protease (SP) domains and bury potential heparan sulfate proteoglycan (HSPG) receptor binding residues within the dimer interface. In contrast, earlier cryo-EM studies of the Ad-FX interaction showed no evidence of dimer formation. Dimerization of FVII bound to Ad may be a contributing mechanistic factor for the differential infectivity of Ad-FX and Ad-FVII complexes, despite high-affinity binding of both these coagulation factors to the virus.
History
DepositionMar 4, 2013-
Header (metadata) releaseJul 3, 2013-
Map releaseAug 28, 2013-
UpdateAug 28, 2013-
Current statusAug 28, 2013Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.992
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by radius
  • Surface level: 0.992
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_5594.tif

Downloads & links

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Map

FileDownload / File: emd_5594.map.gz / Format: CCP4 / Size: 976.6 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationReconstruction of adenovirus type 5 modified to have a short shafted fiber with coagulation factor VII bound
Voxel sizeX=Y=Z: 2.25 Å
Density
Contour LevelBy AUTHOR: 0.99 / Movie #1: 0.992
Minimum - Maximum0.8720004 - 1.17010355
Average (Standard dev.)0.99161088 (±0.02275172)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin-320-320-320
Dimensions640640640
Spacing640640640
CellA=B=C: 1440.0 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z2.252.252.25
M x/y/z640640640
origin x/y/z0.0000.0000.000
length x/y/z1440.0001440.0001440.000
α/β/γ90.00090.00090.000
start NX/NY/NZ-132-122-147
NX/NY/NZ250274261
MAP C/R/S123
start NC/NR/NS-320-320-320
NC/NR/NS640640640
D min/max/mean0.8721.1700.992

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Supplemental data

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Sample components

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Entire : Human zymogen coagulation factor VII bound to human adenovirus ty...

EntireName: Human zymogen coagulation factor VII bound to human adenovirus type 5 from species C. The virus has been modified to contain a short shafted fiber.
Components
  • Sample: Human zymogen coagulation factor VII bound to human adenovirus type 5 from species C. The virus has been modified to contain a short shafted fiber.
  • Virus: Human adenovirus 5

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Supramolecule #1000: Human zymogen coagulation factor VII bound to human adenovirus ty...

SupramoleculeName: Human zymogen coagulation factor VII bound to human adenovirus type 5 from species C. The virus has been modified to contain a short shafted fiber.
type: sample / ID: 1000
Oligomeric state: 240 factor VII molecules bind to one Ad virion
Number unique components: 2
Molecular weightTheoretical: 164 MDa

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Supramolecule #1: Human adenovirus 5

SupramoleculeName: Human adenovirus 5 / type: virus / ID: 1 / NCBI-ID: 28285 / Sci species name: Human adenovirus 5 / Database: NCBI / Virus type: VIRION / Virus isolate: OTHER / Virus enveloped: No / Virus empty: No
Host (natural)Organism: Homo sapiens (human) / synonym: VERTEBRATES
Virus shellShell ID: 2 / Name: hexon capsid / Diameter: 920 Å / T number (triangulation number): 25

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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

Concentration0.6 mg/mL
BufferpH: 7.6 / Details: 50mM Tris, 150mM NaCl, 2mM CaCl2, 2mM MgCl2
GridDetails: Quantifoil R2/4 holey carbon grids, glow discharged
VitrificationCryogen name: ETHANE / Chamber humidity: 30 % / Chamber temperature: 90 K / Instrument: HOMEMADE PLUNGER / Method: Blot for 6 seconds before plunging

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Electron microscopy

MicroscopeFEI POLARA 300
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsCalibrated magnification: 400000 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.26 mm / Nominal defocus max: 2.1 µm / Nominal defocus min: 0.5 µm / Nominal magnification: 310000
Sample stageSpecimen holder model: OTHER
Alignment procedureLegacy - Astigmatism: Objective lens astigmatism was corrected at 300,000 times magnification
Legacy - Electron beam tilt params: 0
DetailsLow dose
DateNov 14, 2010
Image recordingCategory: CCD / Film or detector model: GATAN ULTRASCAN 4000 (4k x 4k) / Digitization - Sampling interval: 15 µm / Number real images: 1503 / Average electron dose: 20 e/Å2 / Bits/pixel: 16
Experimental equipment
Model: Tecnai Polara / Image courtesy: FEI Company

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Image processing

CTF correctionDetails: Each particle
Final reconstructionAlgorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 8.7 Å / Resolution method: FSC 0.5 CUT-OFF / Software - Name: Frealign / Number images used: 837
DetailsThe particles were selected with an in-house script and processed using Frealign.

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