[Chimera-users] calculate FSC curve between experimental cryo EM map and model

[Tom Goddard]

Hi Tofayel,

  Your procedure from computing a Fourier shell correlation between a cryoEM map (at about 4 Angstroms resolution) and a predicted map for an atomic model sounds reasonable.  But keep in mind that the predicted map from the atomic model is just a sum of Gaussians at the atom centers that all have the same width (standard deviation) (molmap command in Chimera), and this may be a completely unrealistic approximation to the expected cryoEM map, especially at relatively high 4 Angstroms resolution.  The electrons in EM are scattered primarily by the electrons of the atoms in the sample, and the distribution of those electrons is not just the sum of Gaussians of equal width centered on the atoms.  I don’t know how bad that approximation is at 4 Angstroms.  I don’t know if the EMAN2 program e2pdb2mrc.py uses the same sum of Gaussians.  The following message says that you need to take care that your predicted map aligns with the experimental map with zero MRC header offsets when using e2proc3d.py to compute the FSC since it apparently ignores the offsets.

	https://groups.google.com/forum/#!topic/eman2/fTGriONtY0A <https://groups.google.com/forum/#!topic/eman2/fTGriONtY0A>

I think you have that taken care of with the Chimera “vop resample” operation but that resampling does a trilinear interpolation that changes the map somewhat and will effect your result somewhat — how much is discused in the the Chimera mailing list post you cited.

  Another thing to consider is that for purposes of Fourier shell correlation it shouldn’t matter what resolution you use for the simulated map when using Chimera molmap, since different resolutions are related by just convolution with a Gaussian in real space, or pointwise multiplication by a Gaussian in Fourier space, so that shells each get scaled by the same factor and scaling every shell will not change the correlations (ignoring shell thickness) with the experimental map Fourier shells.

	Tom


> On Apr 29, 2017, at 4:40 AM, #AHMED TOFAYEL# <TOFAYEL001 at e.ntu.edu.sg> wrote:
> 
> Hi Elaine,
> 
> Thanks for your reply. Yes, I have ardently gone through http://www.cgl.ucsf.edu/pipermail/chimera-users/2016-June/012366.html <http://www.cgl.ucsf.edu/pipermail/chimera-users/2016-June/012366.html>  but ended up being confused as it seems no conclusion had been reached there, or I failed altogether to get the take home message from the discussion.
> 
> Also, the "LocalFSC" thingy is not what I am looking for...
> 
> But nevermind, lets wait for our "map expert" to be back in action and meanwhile I am waiting for others to bring in some appropriate answers.
> 
> Thanks a lot for taking time.
> 
> Best regards,
> Tofayel
> 
>    
> From: Elaine Meng <meng at cgl.ucsf.edu>
> Sent: Saturday, April 29, 2017 5:59:22 AM
> To: #AHMED TOFAYEL#
> Cc: UCSF Chimera Mailing List
> Subject: Re: [Chimera-users] calculate FSC curve between experimental cryo EM map and model
>  
> Dear Tofayel,
> Our map expert is away this week and next week.  All I can suggest is to take a look at this previous post (which you may have seen already):
> <http://www.cgl.ucsf.edu/pipermail/chimera-users/2016-June/012366.html <http://www.cgl.ucsf.edu/pipermail/chimera-users/2016-June/012366.html>>
> 
> There is also a “localFSC” plugin to Chimera that you could get separately, see:
> <http://www.rbvi.ucsf.edu/chimera/plugins/plugins.html <http://www.rbvi.ucsf.edu/chimera/plugins/plugins.html>>
> … however, I don’t know if it is relevant to your question.
> 
> Sorry I couldn’t provide a more confident answer,  Maybe other EM folks on this list will have some thoughts. 
> Best,
> Elaine
> -----
> Elaine C. Meng, Ph.D.                       
> UCSF Computer Graphics Lab (Chimera team) and Babbitt Lab
> Department of Pharmaceutical Chemistry
> University of California, San Francisco
> 
> > On Apr 28, 2017, at 6:09 AM, #AHMED TOFAYEL# <TOFAYEL001 at e.ntu.edu.sg> wrote:
> > 
> > Dear all,
> > If I want to calculate FSC curve between experimental cryo EM map and model, what is the correct procedure to do so?
> > 
> > I will explain what I have tried so far. Although I have variable local resolution in my map, according to FSC=0.143 criterion I have a 4 Ang map (say 1.mrc), I have used 'molmap' command in Chimera to generate a simulated map at 4 Ang. Then I used "vop resample" to take care of the pixel and box values ( then saved as 2.mrc). Thereafter, I have used "e2proc3d.py 1.mrc fsc.txt --calcfsc=2.mrc and plotted the fsc.txt file to get the curve. 
> > 
> > I am not sure if I am on the right track. My pixel size is 1.28.
> > Best regards,
> > Tofayel
> 
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