[phenixbb] simulated annealing omit map

Pavel Afonine pafonine at lbl.gov
Thu Nov 13 23:59:39 PST 2014

Hello Ivan,

>      I want to see a ligand in the structure I have. I am using Phenix
> for refinement.
>      When I do a molecular replacement with the model or part of the
> model, it looks like there is continues density of the ligand on 2Fo-Fc
> and Fo-Fc map at lower counter level.

I presume at this point you have not placed the ligand yet, and 
therefore the ligand map you look at is a "discovery map" in Dale 
Tronrud's terms (see my previous email where I mention this).

>      1st QUESTION: Once we got the phases from the model which serves for
> MR, what is the counter level limit for a 2Fo-Fc and Fo-Fc maps we can
> look at ??

It is a good question with no universal answer. First, it depends how 
you scale the map; for recent sort of review see:

Acta Cryst. (2014). D70, 2593-2606
Metrics for comparison of crystallographic maps
A. Urzhumtsev, P. V. Afonine, V. Y. Lunin, T. C. Terwilliger and P. D. Adams

If map is "sigma scaled" then the rule of thumb is 1sigma for 2mFo-DFc 
and 3sigma for mFo-DFc. Now, this applies for fully occupied typical 
atoms. For mobile partially partially occupied atoms (like those of 
ligands) you can go well below these 1 or 3 "sigma".

"1sigma" rule applies to composite residual OMIT map computed with 
phenix.composite_omit_map (see previous email).

If you use FEM map:
then contouring doesn't really matter, and anything below or equal to 
"1sigma" is suitable.

>      Further going on the structure refinement, at some point I add the
> compound I poorly see and I do 3 different refinements !! One with
> compound occupancy 0.8, another one with 0.7 and another one with 0.6.

Well, I presume you do not just set occupancy to these numbers, but you 
actually refine group occupancy starting from these values. If this is 
correct, then I hope you are getting the same result. If you just set 
these occupancies but you do not refine them, then I don't think it is a 
good idea.

> Then the density for the compound after refinement appears. At that
> point I am on the second option in Pavel's suggestions below - I get
> probably a biased positive density in the map. BUT, I would like to
> notice that my Fwork and Ffree go significantly down.
>      2nd QUESTION: Does it mean the compound is there if statistics do
> down??

Usually, a ligand generates a small contribution to the total scattering 
so it is next to "invisible" by global metrics such as Rwork and Rfree. 
Therefore using local map and model quality metrics is a better way to 
do assessments. However, if adding it improves both Rs then I would say 
it is a good sign.

All the best,

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