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- EMDB-5538: CryoEM Structure of human defensin 5 bound to a neutralization se... -

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Basic information

Entry
Database: EMDB / ID: EMD-5538
TitleCryoEM Structure of human defensin 5 bound to a neutralization sensitive adenovirus chimera
Map data3D reconstruction of defensin complexed with a neutralization sensitive adenovirus chimera
Sample
  • Sample: Complex of an adenovirus chimera Ad5.F35 with human alpha defensin 5
  • Virus: Human adenovirus 5
Keywordsadenovirus / human defensin 5 / neutralization / cryoEM / MDFF / innate immunity
Biological speciesHuman adenovirus 5
Methodsingle particle reconstruction / cryo EM / Resolution: 9.6 Å
AuthorsFlatt JW / Smith JG / Nemerow GR / Stewart PL
CitationJournal: PLoS One / Year: 2013
Title: An intrinsically disordered region of the adenovirus capsid is implicated in neutralization by human alpha defensin 5.
Authors: Justin W Flatt / Robert Kim / Jason G Smith / Glen R Nemerow / Phoebe L Stewart /
Abstract: Human α-defensins are proteins of the innate immune system that suppress viral and bacterial infections by multiple mechanisms including membrane disruption. For viruses that lack envelopes, such as ...Human α-defensins are proteins of the innate immune system that suppress viral and bacterial infections by multiple mechanisms including membrane disruption. For viruses that lack envelopes, such as human adenovirus (HAdV), other, less well defined, mechanisms must be involved. A previous structural study on the interaction of an α-defensin, human α-defensin 5 (HD5), with HAdV led to a proposed mechanism in which HD5 stabilizes the vertex region of the capsid and blocks uncoating steps required for infectivity. Studies with virus chimeras comprised of capsid proteins from sensitive and resistant serotypes supported this model. To further characterize the critical binding site, we determined subnanometer resolution cryo-electron microscopy (cryoEM) structures of HD5 complexed with both neutralization-sensitive and -resistant HAdV chimeras. Models were built for the vertex regions of these chimeras with monomeric and dimeric forms of HD5 in various initial orientations. CryoEM guided molecular dynamics flexible fitting (MDFF) was used to restrain the majority of the vertex model in well-defined cryoEM density. The RGD-containing penton base loops of both the sensitive and resistant virus chimeras are predicted to be intrinsically disordered, and little cryoEM density is observed for them. In simulations these loops from the sensitive virus chimera, interact with HD5, bridge the penton base and fiber proteins, and provides significant stabilization with a three-fold increase in the intermolecular nonbonded interactions of the vertex complex. In the case of the resistant virus chimera, simulations revealed fewer bridging interactions and reduced stabilization by HD5. This study implicates a key dynamic region in mediating a stabilizing interaction between a viral capsid and a protein of the innate immune system with potent anti-viral activity.
History
DepositionDec 7, 2012-
Header (metadata) releaseJul 3, 2013-
Map releaseJul 3, 2013-
UpdateJul 3, 2013-
Current statusJul 3, 2013Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.022
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by radius
  • Surface level: 0.022
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_5538.jpg

Downloads & links

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Map

FileDownload / File: emd_5538.map.gz / Format: CCP4 / Size: 3.2 GB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotation3D reconstruction of defensin complexed with a neutralization sensitive adenovirus chimera
Voxel sizeX=Y=Z: 1.5 Å
Density
Contour LevelBy AUTHOR: 0.022 / Movie #1: 0.022
Minimum - Maximum-0.12764664 - 0.23956536
Average (Standard dev.)-0.00216073 (±0.02941307)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin-480-480-480
Dimensions960960960
Spacing960960960
CellA=B=C: 1440.0 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.51.51.5
M x/y/z960960960
origin x/y/z0.0000.0000.000
length x/y/z1440.0001440.0001440.000
α/β/γ90.00090.00090.000
start NX/NY/NZ-5029166
NX/NY/NZ106122134
MAP C/R/S123
start NC/NR/NS-480-480-480
NC/NR/NS960960960
D min/max/mean-0.1280.240-0.002

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Supplemental data

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Sample components

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Entire : Complex of an adenovirus chimera Ad5.F35 with human alpha defensin 5

EntireName: Complex of an adenovirus chimera Ad5.F35 with human alpha defensin 5
Components
  • Sample: Complex of an adenovirus chimera Ad5.F35 with human alpha defensin 5
  • Virus: Human adenovirus 5

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Supramolecule #1000: Complex of an adenovirus chimera Ad5.F35 with human alpha defensin 5

SupramoleculeName: Complex of an adenovirus chimera Ad5.F35 with human alpha defensin 5
type: sample / ID: 1000
Oligomeric state: ~3000 molecules bind each adenovirus particle
Number unique components: 2
Molecular weightTheoretical: 150 MDa

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Supramolecule #1: Human adenovirus 5

SupramoleculeName: Human adenovirus 5 / type: virus / ID: 1 / Name.synonym: Human adenovirus type 5 with short fiber / NCBI-ID: 28285 / Sci species name: Human adenovirus 5 / Database: NCBI / Virus type: VIRION / Virus isolate: OTHER / Virus enveloped: No / Virus empty: No / Syn species name: Human adenovirus type 5 with short fiber
Host (natural)Organism: Homo sapiens (human) / synonym: VERTEBRATES
Molecular weightTheoretical: 150 MDa
Virus shellShell ID: 1 / Name: Capsid / Diameter: 1170 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

Concentration0.16 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 4 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
DateJul 12, 2010
Image recordingCategory: FILM / Film or detector model: GATAN ULTRASCAN 4000 (4k x 4k) / Digitization - Scanner: OTHER / Digitization - Sampling interval: 15 µm / Number real images: 3515 / 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: 9.6 Å / Resolution method: FSC 0.5 CUT-OFF / Software - Name: Frealign / Number images used: 1014
DetailsThe particles were selected with an in-house script and processed using Frealign

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