resolution assessment in cryoem
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Resolution Assessment in CryoEM. Kristin Shingler Friday, August 2 nd , 2013. What is Resolution?. In short, how close two objects can be without appearing as one. Atomic resolution is achieved in x-ray crystallography. - PowerPoint PPT PresentationTRANSCRIPT
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Resolution Assessment in CryoEM
Kristin ShinglerFriday, August 2nd, 2013
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What is Resolution?• In short, how close two objects can be without
appearing as one.• Atomic resolution is achieved in x-ray
crystallography.• Typically, cryoEM generates sub-nanometer
resolution, but recent reconstructions have approached atomic resolution.
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Semliki Forest Virus 1986
3.2 nanometer resolution Vogel et al., 1986 Nature
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Sub-nanometer Resolution
Hepatitis B Capsid, 9.1Å Resolution Conway et al., 1997 Science
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Near-Atomic Resolution
Epsilon 15 Bacteriophage, 4.5Å Resolution Jiang et al., 2008 Nature
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Atomic Resolution
Human Adenovirus 5, 3.6Å Resolution Liu et al., 2010 Science
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What does resolution really mean?
Zhou, Current Opinions in Structural Biology, 2008
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What does resolution really mean?
Baker et al., Methods in Enzymology, 2010
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Resolution Assessment in X-ray Crystallography
Modified Figure 5 from Wlodawer et al. 2007
• Determined by how closely the crystal lattice can be packed, while still maintaining diffraction
• Various statistical measurements correlate the resolution
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3D Reconstruction Work Flow
Baker, Olson, and Fuller 1999 Microbiology and Molecular Biology Reviews
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Fourier Shell Correlation
F1 – Structure Factor for Map #1F2 – Structure Factor for Map #2ri – individual voxel elementr – radius
“measures the normliazed cross-correlation coefficient between two 3D volumes as a function of spatial frequency” - van Heel
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Conventional Resolution Estimation versus Gold Standard
Conventional Gold Standard
Model
EvenMap
OddMap
FSC and Resolution Estimate
Model Model
½DataMap
½DataMap
FSC and Resolution Estimate
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Conventional Resolution Estimation versus Gold Standard
Scheres and Chene, 2012 Nature Methods
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Why this paper?
• Elife is an open access journal• It provides readers with both the reviewers
comments AND the author’s response to reviewers
• An excellent example of high quality work showing the disparity between conventional and gold standard resolution assessment
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Timeline of Events
• The authors first reported the resolution of their map to be 4.6Å by using conventional resolution assessment
• This method was challenged by reviewers • The authors reassessed resolution using the
gold standard method and found the map was 5.3Å resolution
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Reviewers Comment
Sun et al.,2013 Elife
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Author’s Response
Sun et al.,2013 Elife
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Chikungunya Virus Structure
Sun et al.,2013 Elife
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CryoEM Density Fitting
Sun et al.,2013 Elife