Dear all I have a structure at 1.4 A currently with an Rfactor of 21/26 %. I notice from the plot by cycle that everytime the RMS bonds/angles reduce, the Rfactor increases a few percent. Is there any way of telling phenix.refine to not 'restrain' the model too much? I have already used automatic weighting. Also, RINGER identified a few alternate conformations, so I'm thinking that modeling this might account for a better model. However, as I have an NCS of 4, this can be tedious to do by hand, and AFAIK, phenix.refine does not create new conformations. Is there an alternative? Final question, some residues were flagged as having high B factors but not in all the copies. How do I deal with this? Some background: I initially had problems with refinement being stuck at 30-35 % in SG P 3 2 1 (as suggested by Pointless and Xtriage). However, the beamline scientist suggested that there could've been lattice translocation and that it is actually P 3. Reintegrating seems to have solved the problem. With integrated data up to 1.2 A, I tried cut-off at 1.4, 1.3 and 1.2 A. Together with CC1/2 and refinement results, I chose 1.4 A, but should I cut it a bit lower? Thanks.
Hi Mohamed,
I have a structure at 1.4 A currently with an Rfactor of 21/26 %.
R-factors are a bit high... At this resolution one would expect Rwork ~15-17 and Rfree ~16-21 (lookig at statistics for similar resolution structures in PDB). Did you add water, refine anisotropic ADPs, add H, model alternative conformations?
I notice from the plot by cycle that everytime the RMS bonds/angles reduce, the Rfactor increases a few percent. Is there any way of telling phenix.refine to not 'restrain' the model too much?
Try running refinement with weights optimization - usually this addresses this issue.
I have already used automatic weighting.
Oh I see you did.
However, as I have an NCS of 4, this can be tedious to do by hand, and AFAIK, phenix.refine does not create new conformations. Is there an alternative?
I'm not aware of software that builds alternative conformations fast and reliably. Make sure you do not use NCS restraints - at this resolution using them may be counterproductive.
Final question, some residues were flagged as having high B factors but not in all the copies. How do I deal with this?
At this resolution it's expected to observe differences
Some background: I initially had problems with refinement being stuck at 30-35 % in SG P 3 2 1 (as suggested by Pointless and Xtriage). However, the beamline scientist suggested that there could've been lattice translocation and that it is actually P 3. Reintegrating seems to have solved the problem. With integrated data up to 1.2 A, I tried cut-off at 1.4, 1.3 and 1.2 A. Together with CC1/2 and refinement results, I chose 1.4 A, but should I cut it a bit lower?
I see.. Crystal oddity may contribute to higher (than anticipated) R-factors. If you modeled everything you can model, model to map is ok, and model passes all other validation then perhaps there isn't much else you can do. More data you cut lower R factors you may get but a) R-factors computed using different sets of reflections are not comparable, and b) is lowering R really ultimate goal? Pavel
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mohamed noor -
Pavel Afonine