[phenixbb] Rfree refuse to come down

Francois Berenger berenger at riken.jp
Fri Mar 2 00:50:10 PST 2012

Maybe this tool I know from a previous colleague
may be of some help:

     "LAFIRE: software for automating the refinement process of protein 
structure analysis".
     YAO M., ZHOU Y., AND TANAKA I., Acta. Cryst., D62, 189-196
     "New algorithm for protein model building: extending partial model 
in map segment".
     ZHOU Y., YAO M., AND TANAKA I., J. Appl. Cryst., 39, 57-63

By the way, this tool shouldn't be in phenix? ;)


On 03/02/2012 04:36 PM, Vellieux Frederic wrote:
> Well, IMHO, one of the things to check is whether or not you are trying
> to refine far too many parameters for your data set (your observations),
> for example. You'd expect to see Rfree going up if you were to refine
> (at 2.4 A resolution) a structure with a full anisotropic temperature
> factor model (i.e. x, y, z and 6 Baniso values).
> However in your case I'd question first the molecular replacement
> solution. Make sure you have a valid solution... Did you see (in all 4
> protein molecules) density indicative of the sequence changes between
> your MR search model and your protein ? This is indicative of a real
> solution...
> And I'd certainly start refinement using the NCS present, to relax it
> (restraints) as the refinement proceeds (smoothly) and possibly not use
> any NCS in the final stages of the refinement. You may need to carry out
> annealing initially and so forth.
> Fred.
> #HEW KAI LI KELLY# wrote:
>> This may be a stupid question, but what are the things I should look
>> out for to change if my Rfree goes up?
>> Kelly
>> On 2 Mar, 2012, at 3:17 PM, "Vellieux Frederic"
>> <Frederic.Vellieux at ibs.fr> wrote:
>>> A couple of things:
>>> TFZ of 9 is on the lowish side therefore one could question the
>>> validity of the MR solution, are all protein molecules correctly
>>> positioned for example. Are there domains and subdomains that have
>>> moved in your structure relative to your search model?
>>> with 4-fold NCS you can use the redundancy to 1) improve the maps by
>>> iterative NCS averaging; 2- use NCS contraints (then restraints later
>>> on) during refinement.
>>> Fred.
>>> PS refinement of a model does not mean "bring R-free down", the
>>> R-free going up is a symptom that should be investigated and the
>>> causes of it then treated...
>>> #HEW KAI LI KELLY# wrote:
>>>> HI,
>>>> I have a few questions about a dataset that I am working on currently.
>>>> So I ran Xtraige and it tells me that other than pseudoNCS, there is
>>>> no other problems like twinning etc. And then I used autoMR and
>>>> Phaser from Phenix to run molecular replacement with a the structure
>>>> of the same protein (but that is with DNA bound). The TFZ is more
>>>> than 9 after from both the runs. Autobuild managed to build the 4
>>>> molecules in my ASU (Rfactor/Rfree = 28/32 for a resolution of
>>>> 2.4A). But because there are some clashes and outliers in the
>>>> Ramachandran Plot etc, I started to manually refine them in coot. In
>>>> addition, there is a loop that is unable to fit into the density, so
>>>> I chopped about 10 residues off each molecule in the ASU.
>>>> However, as I started to refine the model from AutoBuild, my Rfree
>>>> begins to climb. So currently, I am at Rfactor/Rfree=27/36. I don't
>>>> know if there is anything wrong with I had done. So many thanks in
>>>> advance for your advices on this!!
>>>> Warmest Regards,
>>>> Kelly Hew
>>>> Nanyang Technogical University
>>>> School of Biological Sciences
>>>> Division of Structural Biology & Biochemistry
>>>> Address : #07-01, IMCB, 61 Biopolis Drive, Proteos, Singapore 138673
>>>> Telephone (O): +65 65869673
>>>> Telephone (HP): +65 98713553
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