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
Vincent

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
_______________________________________________
phenixbb mailing list
[email protected]
http://phenix-online.org/mailman/listinfo/phenixbb
Unsubscribe: [email protected]

--

Vincent Chaptal, PhD

Institut de Biologie et Chimie des Protéines

Drug Resistance and Membrane Proteins Laboratory

7 passage du Vercors

69007 LYON

FRANCE

+33 4 37 65 29 01

http://www.ibcp.fr