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 xray crystallography ID: 557534
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Slide1
Resolution Assessment in CryoEM
Kristin Shingler
Friday, August 2
nd
, 2013Slide2
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.Slide3
Semliki Forest Virus 1986
3.2 nanometer
r
esolution Vogel
et al.,
1986
NatureSlide4
Sub-nanometer Resolution
Hepatitis B Capsid, 9.1
Å
Resolution Conway
et al.,
1997
ScienceSlide5
Near-Atomic Resolution
Epsilon 15 Bacteriophage, 4.5
Å
Resolution Jiang
et al.,
2008
NatureSlide6
Atomic Resolution
Human Adenovirus 5, 3.6
Å
Resolution Liu
et al.,
2010
ScienceSlide7
What does resolution really mean?
Zhou,
Current Opinions in Structural Biology
, 2008Slide8
What does resolution really mean?
Baker
et al.
,
Methods in Enzymology
, 2010Slide9
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 resolutionSlide10
3D Reconstruction Work Flow
Baker, Olson, and Fuller 1999
Microbiology and Molecular Biology ReviewsSlide11
Fourier Shell Correlation
F
1
– Structure Factor for Map #1
F
2
– Structure Factor for Map #2
r
i
– individual voxel element
r – radius
“measures the
normliazed
cross-correlation coefficient between two 3D volumes as a function of spatial frequency” - van HeelSlide12
Conventional Resolution Estimation versus Gold Standard
Conventional
Gold Standard
Model
Even
Map
Odd
Map
FSC and Resolution Estimate
Model
Model
½
Data
Map
½
Data
Map
FSC and Resolution EstimateSlide13
Conventional Resolution Estimation versus Gold Standard
Scheres
and
Chen
e
,
2012
Nature MethodsSlide14Slide15
Why this paper?
Elife
is an open access journal
It provides readers with both the reviewers comments AND the author’s response to reviewersAn excellent example of high quality work showing the disparity between conventional and gold standard resolution assessmentSlide16
Timeline of Events
The authors first reported the resolution of their map to be 4.6
Å 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 Slide17
Reviewers Comment
Sun
et al.,
2013 ElifeSlide18
Author’s Response
Sun
et al.,
2013
ElifeSlide19
Chikungunya
Virus Structure
Sun
et al.,
2013
ElifeSlide20
CryoEM Density Fitting
Sun
et al.,
2013
Elife