Dear Aleksandar, I did use SA-OMIT map feature in Phenix, implemented by Tom Terwilliger. I would strongly recommend you to use this feature. Remove the suspicious residues from the pdb and run first SA-omit map in phenix with default settings, and later with different starting temperatures. Make sure also to use harmonic restrains to prohibit neighboring residues to creep into the "omitted" region. Also, use MR solution and the raw mtz file with only FP and SIGFP that you have for the omit map. I have experienced that if you use refined model then it does not completely get rid of biases. At max, you can use rigid body refined one.
On the other side, if your molecule is not too big, then you can try the CNS. In that case, I would recommend remove the 'suspicious' resiudes and 1st use segmented rigid body refinement, i.e, consider each chain or subunit as rigid entity. Then, you can try group B factor with restrains along with DEN mode for simulated annealing. I strongly recommend the DEN mode which can help you to remove bias. Please note that it is necessary to try out several different temperatures and different temperature gradient as well for simulated annealing in order to reproduce the features. Otherwise, you never know if you are stuck at local minima or not. thanks shibom On Wed, Apr 29, 2015 at 4:16 AM, Aleksandar Bijelic < [email protected]> wrote: > Dear CCP4 users, > > I am currently solving a structure (2.8-2.9 A resolution) of a protein > complexed with a ligand using MR with the apo-form of this protein as model > (resolution of the model is 2.4). After MR-phasing I performed a regular > autobuild run giving me good outputs and thus I refined the best pdb > leading to good values according to R-values and geometry, however, the > denstiy doesn´t look well (but I think it´s due to the moderate > resolution). Now I want to get sure if the side chains which are involved > in the ligand binding are correctly positioned. However, the active site is > suspicously similar to the active site of the model (apo-form) and so I am > afraid that this could be due to model bias. My question is how to check > and to get rid of the bias (if present) at this stage (after several > refinements). I read the publication of Terwilliger about iterative-build > OMIT maps but since I am a bloody novice in this field I didn´t really > understand it. I originally thought iterative-build OMIT maps are performed > to compare the output map with one´s map in order to detect uncertainties, > but what to do next? Or should I start from the beginning but how to > proceed than, what should I do (I am using Phenix via GUI) ... Is it > possible and reasonable to run autobuild with iterative omit map option? Or > is it only reasonable if experimental phases are available? I didn´t run > iterative-build OMIT maps yet because I am not sure how to run it correctly > (what method is the best?) and at my institute the run will take more than > 1 day and I don´t want to block one computer until I am not sure if it is > reasonable. I hope you can give me some advice and help me. Thank you in > advance. > > Regards, > > Aleks > > -- > ------------------------------------------- > Aleksandar Bijelic, MSc. > > Institut für Biophysikalische Chemie > Universität Wien > Althanstrasse 14 > A-1090 Wien > > Tel: +43 1 4277 52536 > e-Mail: [email protected] > > -------------------------------------------- >
