Dear Jeorge just as a curiosity: 1- do you see a continuous electron density for the ligand, before and after performing the refinement? 2- do you have only one copy of the molecule(s), and if not, do you see the same behaviour for all the ligands? 3- what are the B-factor values for the ligand? Any hint that the phosphate could be very flexible and actually missing? 4- do you see any red (/ green) FoFc densities where the phosphate should (/ not) be? If so, do you see anything else by using 'less data', i.e. data that would be less prone to radiation damage? 5- what is the resolution of your data? 6- is it a known ligand? If you do have the same ligand, even only once, present in the PDB, why don't you take the coordinates and libraries from this already refined structure and try to fit it in your coordinates? 7- do you have chemical data showing that the ligand can stay for very long times in your protein without being shortened? 8- do you know how pure is your ligand solution? i.e. is your ligand stable at room temperature (or whatever temperature you did the soaking / complex formation). 9- similarly, if you did co-crystallisation instead of soaking, it could be that your protein somehow cleaved the molecule. It will all depend on how stable is the ligand in high concentrations of the protein, pH dependency and more.
Those are not real answers to your original question, just points of curiosity... Cheers, Leo > On Nov 22, 2014, at 4:26 AM, jeorgemarley thomas <kirtswab...@gmail.com> > wrote: > > Dear all, > > I am working on my molecule, but after running > Refmac5 restrained refinement the bonding distances and angles are broken. > I used Phenix to create the ligand and used Coot to place the ligand into the > correct location in the molecule, merged the ligand and molecule coordinates, > and saved the file. Then I went to run restrained refinement on > the merged coordinates using the cif file provided from the phenix to the > refmac library,the rest of the settings were set at the defaults. During > fitting the model in the 2Fo-Fc map of the ligand there were no issue, but it > fitted well and after refinement of the same, the bond breaks and > distorted as observed in coot. The resolution is 1.6 Ang. > phosphate linkage bond is broken. What could be the best possible reason for > this and what can be solution for this ? Could you please suggest what should > I do for a complete model for the ligand ? I would like to run a round of > restrained refinement without breaking any geometric restraints that should > be imposed on my ligand. > > Thank you for your help in advance, > > Jeorge >
