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- EMDB-2145: Electron microscopy of Influenza hemagglutinin (Malaysia/2506/200... -

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Basic information

Entry
Database: EMDB / ID: 2145
TitleElectron microscopy of Influenza hemagglutinin (Malaysia/2506/2004) in complex with neutralizing antibody (Fab CR8071
KeywordsInfluenza hemagglutinin (Malaysia/2506/2004) / neutralizing antibody (Fab CR8071) / negative stained / single particle analysis
SampleInfluenza hemagglutinin (Malaysia/2506/2004), neutralizing antibody (Fab CR8071), negative stained, single particle analysis
SourceInfluenza B virus (B/Malaysia/2506/2004) / virus / Influenza
Homo sapiens / human
Map dataInfluenza hemagglutinin (Malaysia/2506/2004), neutralizing antibody (Fab CR8071), negative stained, single particle analysis
Methodsingle particle reconstruction, at 19 A resolution
AuthorsKhayat R / Lee JH / Dreyfus C / Laursen NS / Wilson IA / Ward AB
CitationScience, 2012, 337, 1343-1348

Science, 2012, 337, 1343-1348 StrPapers
Highly conserved protective epitopes on influenza B viruses.
Cyrille Dreyfus / Nick S Laursen / Ted Kwaks / David Zuijdgeest / Reza Khayat / Damian C Ekiert / Jeong Hyun Lee / Zoltan Metlagel / Miriam V Bujny / Mandy Jongeneelen / Remko van der Vlugt / Mohammed Lamrani / Hans J W M Korse / Eric Geelen / Özcan Sahin / Martijn Sieuwerts / Just P J Brakenhoff / Ronald Vogels / Olive T W Li / Leo L M Poon / Malik Peiris / Wouter Koudstaal / Andrew B Ward / Ian A Wilson / Jaap Goudsmit / Robert H E Friesen

DateDeposition: Jun 26, 2012 / Header (metadata) release: Jul 9, 2012 / Map release: Aug 22, 2012 / Last update: Jun 26, 2012

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 4.8
  • Imaged by UCSF CHIMERA
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 4.8
  • Imaged by UCSF CHIMERA
  • Download
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Supplemental images

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Map

Fileemd_2145.map.gz (map file in CCP4 format, 16001 KB)
Projections & slices
Size
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Others
AxesZ (Sec.)Y (Row.)X (Col.)
160 pix
2.18 A/pix
= 348.8 A
160 pix
2.18 A/pix
= 348.8 A
160 pix
2.18 A/pix
= 348.8 A

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider package.

Voxel sizeX=Y=Z: 2.18 A
Density
Contour Level:4.8 (by author), 4.8 (movie #1):
Minimum - Maximum-6.27524042 - 17.14086151
Average (Standard dev.)0E-8 (1)
Details

EMDB XML:

Space Group Number1
Map Geometry
Axis orderXYZ
Dimensions160160160
Origin-80-80-80
Limit797979
Spacing160160160
CellA=B=C: 348.80002 A
Alpha=beta=gamma: 90 deg.

CCP4 map header:

modeImage stored as Reals
A/pix X/Y/Z2.182.182.18
M x/y/z160160160
origin x/y/z0.0000.0000.000
length x/y/z348.800348.800348.800
alpha/beta/gamma90.00090.00090.000
start NX/NY/NZ-32-32-32
NX/NY/NZ646464
MAP C/R/S123
start NC/NR/NS-80-80-80
NC/NR/NS160160160
D min/max/mean-6.27517.141-0.000

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Supplemental data

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Sample components

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Entire Influenza hemagglutinin (Malaysia/2506/2004), neutralizing antibo...

EntireName: Influenza hemagglutinin (Malaysia/2506/2004), neutralizing antibody (Fab CR8071), negative stained, single particle analysis
Details: The sample was monodisperse / Number of components: 2 / Oligomeric State: Trimer
MassTheoretical: 282 kDa / Experimental: 282 kDa / Measured by: Sequence of components

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Component #1: protein, Influenza hemagglutinin (Malaysia/2506/2004)

ProteinName: Influenza hemagglutinin (Malaysia/2506/2004) / Oligomeric Details: Trimer / Number of Copies: 3 / Recombinant expression: Yes
MassTheoretical: 49 kDa / Experimental: 49 kDa
SourceSpecies: Influenza B virus (B/Malaysia/2506/2004) / virus / Influenza
Source (engineered)Expression System: Sf9 / Vector: pDCE198
Source (natural)Cell: Sf9

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Component #2: protein, IgG Fab (CR8071)

ProteinName: IgG Fab (CR8071) / Oligomeric Details: 3 monomers / Recombinant expression: Yes / Number of Copies: 3
MassTheoretical: 45 kDa / Experimental: 45 kDa
SourceSpecies: Homo sapiens / human

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Experimental details

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Sample preparation

Specimen stateparticle
Sample solutionSpecimen conc.: 0.1 mg/ml / Buffer solution: 20 mM HEPES 8.0, 50mM NaCl / pH: 8
Support film400 mesh gold grid with thin carbon support, glow discharged
StainingGrids with adsorbed protein floated on Nano-W for 30 seconds.
VitrificationInstrument: NONE / Cryogen name: NONE

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Electron microscopy imaging

ImagingMicroscope: FEI TECNAI F20 / Date: Jan 12, 2012 / Details: Weak beam illumination
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 120 kV / Electron dose: 16 e/A2 / Illumination mode: FLOOD BEAM
LensMagnification: 100000 X (nominal), 100000 X (calibrated)
Astigmatism: Objective lens astigmatism was corrected at 100,000 times magnification
Cs: 2 mm / Imaging mode: BRIGHT FIELD / Defocus: 650 - 970 nm
Specimen HolderModel: SIDE ENTRY, EUCENTRIC / Tilt Angle: 0 - 55 deg.
CameraDetector: GATAN ULTRASCAN 4000 (4k x 4k)

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Image acquisition

Image acquisitionNumber of digital images: 200 / Scanner: OTHER / Sampling size: 10.9 microns / Bit depth: 16 / OD range: 1.4 / Details: Images were collected on a CCD

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Image processing

ProcessingMethod: single particle reconstruction / Number of projections: 7116
Details: Particles were selected using DoG Picker and processed using Sparx.
Applied symmetry: C3 (3 fold cyclic)
3D reconstructionAlgorithm: Cross-common lines / Software: EMAN2/Sparx / CTF correction: Each particle / Resolution: 19 A / Resolution method: FSC 0.5

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Atomic model buiding

Modeling #1Software: Molrep, Chimera / Refinement protocol: rigid body / Target criteria: Correlation coefficient / Refinement space: REAL
Details: Protocol: Rigid body. The hemagluttinin trimer was fit, then the Fabs were fit while imposing 3-fold non-cyrstallographic symmetry using Molrep.
Input PDB model: 4FQJ
Modeling #2Software: Molrep, Chimera / Refinement protocol: rigid body / Target criteria: Correlation coefficient / Refinement space: REAL
Details: Protocol: Rigid body. The hemagluttinin trimer was fit, then the Fabs were fit while imposing 3-fold non-cyrstallographic symmetry using Molrep.
Input PDB model: 4FQK

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