[phenixbb] Rfree and a low resolution data set

vincent Chaptal vincent.chaptal at ibcp.fr
Wed Apr 4 03:58:35 PDT 2018

Dear Toon,

if you have data up to 2.3A resolution, you should keep it, it is so 
valuable. The isotropic statistics that you are looking at and output by 
Aimless are I'm sure awful, because you simply don't have data (lack of 
completeness) in the high resolution shells, and the statistics are 
produced out of a mixture of real data (weak) and noise.

Staraniso has done a lot of work on outputting statistics more 
meaningful in term of Table 1 and anisotropy, so it could be worth a try 
to submit your data and see what comes out of it.
In any case, I would use the data to the resolution limit suggested by 
Aimless, and judge your electron density maps. I'm sure you will have a 
big improvement if you include the 2.3A limit, even though it is not 
going to look like an isotropic 2.3A.

If you use the data output by staraniso, please ask the server to also 
output your original data so you can deposit the original intensities in 
the PDB and investigators can look at it untouched in the future. There 
is a check box on the server to tick.

Best of luck with your data

On 04/04/2018 11:47, Toon Van Thillo wrote:
> Hi all,
> Currently I am refining a data set which showed anisotropic 
> diffraction. Aimless suggested cutoffs at 2.3, 2.6 and 3.6 angstrom 
> for the h,k and l axis.
> I chose a general 3.6 cutoff to obtain satisfactory statistics for 
> Rmeas, I/sd(I) and CC1/2. At this resolution the data set consists of 
> approximately 2800 reflections.
> Generally 5% of the set is set aside as the Rfree test set and I found 
> that a minimum of 500 reflections in total is used to produce a 
> reliable Rfree. However, 5% only amounts to 140 reflections in this 
> case. I am hesitant to include more reflections as I would have to go 
> up to 20% of the reflections to obtain more than 500 reflections for 
> the test set. In a discussion on the CCP4 message boards some time ago 
> it was suggested to do multiple refinements with different test sets:
> https://www.jiscmail.ac.uk/cgi-bin/webadmin?A2=ind1411&L=ccp4bb&F=&S=&P=125570
> In the thread it was also discussed that a least squares approach is 
> prefered when using a small test set. However, when using a LS target, 
> the resulting Rfree is very high (10% higher than when using the 
> automatic option) and /phenix.refine/​ produces awful geometry (24% 
> ramachandran outliers, 105 clashcore...). It seems that the refinement 
> is performed without restraints? Optimize X-ray/stereochemistry weight 
> does not result in improved stereochemistry. My question is if the LS 
> approach is still relevant and if so, is there an explanation (and 
> solution) for the bad statistics?
> Kind regards,
> Toon Van Thillo
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Vincent Chaptal, PhD

Institut de Biologie et Chimie des Protéines

Drug Resistance and Membrane Proteins Laboratory

7 passage du Vercors

69007 LYON


+33 4 37 65 29 01


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