Download - Resolution Assessment in CryoEM
Resolution Assessment in CryoEM
Kristin ShinglerFriday, August 2nd, 2013
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.
Semliki Forest Virus 1986
3.2 nanometer resolution Vogel et al., 1986 Nature
Sub-nanometer Resolution
Hepatitis B Capsid, 9.1Å Resolution Conway et al., 1997 Science
Near-Atomic Resolution
Epsilon 15 Bacteriophage, 4.5Å Resolution Jiang et al., 2008 Nature
Atomic Resolution
Human Adenovirus 5, 3.6Å Resolution Liu et al., 2010 Science
What does resolution really mean?
Zhou, Current Opinions in Structural Biology, 2008
What does resolution really mean?
Baker et al., Methods in Enzymology, 2010
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
3D Reconstruction Work Flow
Baker, Olson, and Fuller 1999 Microbiology and Molecular Biology Reviews
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
Conventional Resolution Estimation versus Gold Standard
Conventional Gold Standard
Model
EvenMap
OddMap
FSC and Resolution Estimate
Model Model
½DataMap
½DataMap
FSC and Resolution Estimate
Conventional Resolution Estimation versus Gold Standard
Scheres and Chene, 2012 Nature Methods
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
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
Reviewers Comment
Sun et al.,2013 Elife
Author’s Response
Sun et al.,2013 Elife
Chikungunya Virus Structure
Sun et al.,2013 Elife
CryoEM Density Fitting
Sun et al.,2013 Elife