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Title:Difference imaging of adenovirus: bridging the resolution gap between X-ray crystallography and electron microscopy.
Authors:Stewart PL, Fuller SD, Burnett RM
Sample:adenovirus type 2
Aggregation state:Icosahedral (25 angstroms resolution)
Red flagLatest update:2011-05-26
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Sample
Sample name: adenovirus type 2
Oligomeric state: T=25 particle
Components:
ID Type Name Exp. MW (MDa) Oligomeric details Recombinant expression Synthetic Organism GO identifier InterPro identifier Virus identifier Details
1virusAdenovvirus type 2Adenovvirus type 201.0.1.0.001
Experiment
Sample preparation:
pHSample conc.DetailsStainingSample support details
7.410 mg/mLSample stored in CsCl was diluted in Tris (10mM) NaCl (100 mM) pH 7.4
Vitrification:
Cryogen nameHumidityTemp.Instr.MethodTime resolvedDetails
ETHANE60%23 KHOMEMADE PLUNGERBlot for 2 sec msVitrification instrument: EMBL plunger. plunging at ambient temperature and humidity
Imaging:
MicroscopeVoltageIllumination modeImaging modeCs:Defocus min.Defocus max.Nominal mag.Calibrated mag.Electron sourceDetectorDetector distanceAstigmatism
FEI/PHILIPS EM40080 kVFLOOD BEAMBRIGHT FIELD2 mm1000 nm8800 nm34000TUNGSTEN HAIRPINSO 163 film mm

Specimen holderHolder modelTilt min.Tilt max.Energy filterEnergy windowTemp.Temp. min.Temp. max.Beam tiltElectron doseOther detailsDate
eucentric - Philips EM400GATAN LIQUID NITROGEN°12° eV105 K K K mrad8 e/Å2Details of the microscopy are in Stewart, P. L., Burnett, R. M., Cyrklaff, M. and Fuller, S. D. (1991). Image reconstruction reveals the complex molecular organization of adenovirus. Cell 67, 145-54. Images were taken in triples at 1, 2.6 and 8.8 microns underfocus. Some images were taken at 12 degrees tilt to compensate for the preference for the virus to orient with the fibres away from the water layer. For image reconstruction see: Stewart, P. L., Burnett, R. M.,Cyrklaff, M. and Fuller, S. D. (1991). Image reconstruction reveals the complex molecular organization of adenovirus. Cell 67, 145-54. For reconstruction details see: P.L. Stewart, S.D. Fuller, R. M. Burnett, (1993). Difference imaging of adenovirus: bridging the resolution gap between X- ray crystallography and electron microscopy EMBO J.Stewart PL, Burnett RM. Adenovirus structure by X-ray crystallography and electron microscopy. Curr Top Microbiol Immunol. 1995, 199 (1):25-38. The PDB code for the Ad2 hexon model is 1DHX, and the reference Athappilly, F. K., Murali, R., Rux, J. J., Cai, Z., Burnett, R. M.: The refined crystal structure of hexon, the major coat protein of adenovirus type 2, at 2.9 A resolution. J Mol Biol 242 pp. 430 (1994) 1990-01-01
Processing
Software:EMBL-ICOS
CTF correction:initially different defocuses were combined by summing in resolution bands
Resolution by author:25 Å
Resolution method:comparison with atomic structure of hexon at various resolutions
Unit cell:
Scanned images:
Num. imagesSampling sizeOD rangeQuant. bit numberOther detailsScanner
5025 μm/pixel18linkOPTRONICS
Fitting:
PDBProtocolTarget crit.SoftwareB valueFitting spacePDB chainDetails
rigid bodymax cross correlationmaximizing cross correlation3000REAL
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