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Hi Garib, This is a side-question. What you just said writing to Abhinav explains why I have problem of some repulsion between identical ligands with different tail conformations when I try to refine them at 25% and 25% occupancy. In my case the overall occupancy of the ligand is not expected to be 100% (the ligand is naturally partially lost) so in order to have the well behaving refinement I will have to change these occupancies to 50-50, right? And just pay the price with the higher B factors? Or is there any other way to deal with this situation? Aleks On 30 Jul 2008, at 16:15, Garib Murshudov wrote: If sum of occupancies of atoms is less than or equal to one and atoms are not in the same residue with the same alt code then they do not see each other. Otherwise they see each other and there is vdw repulsion between them. this has not changed substantially since the first version. Waters are not good way of modelling unknown ligands.  If you want to model unknown model then you can use DUM atoms with DUM residue name (just like the arp/warp uses it). Then atoms can come much closer to each other (up to 1.2A or so. this value can be controlled) Garib On 30 Jul 2008, at 16:05, Kumar, Abhinav wrote: Arial0000,0000,FFFFI have had a similar experience. Arial0000,0000,FFFF  Arial0000,0000,FFFFI was trying to model a bunch of waters to simulate an unknown ligand (UNL) in an unmodeled density. The waters were all in different alternate conformations of the same residue number. When the occupancies of these waters were set to 0.5, the vdw repulsion became absent; waters stayed within density at close distances. But, as soon as the occupancies were changed to anything but 0.5, waters got pushed out of density due to vdw repulsion. Arial0000,0000,FFFF  Arial0000,0000,FFFFSo my feeling is that at 0.5 occupancy the atom does not see vdw repulsion (?). Change it to something else and atom should crawl back into its density. Arial0000,0000,FFFF  Arial0000,0000,FFFFI was using refmac5 version 5.2.0019. Arial0000,0000,FFFF  Arial0000,0000,FFFFThanks  Arial0000,0000,FFFFAbhinav Arial8080,0000,0000Stanford Synchrotron Radiation Laboratory  Arial8080,0000,0000Joint Center for Structural Genomics  Arial8080,0000,0000Mail Stop 99  Arial8080,0000,0000Phone: (650) 926-2992  Arial8080,0000,0000Fax: (650) 926-3292 Times New Roman0000,0000,FFFF  TahomaFrom:Tahoma CCP4 bulletin board [0000,0000,EEEEmailto:[log in to unmask]] On Behalf Of Garib Murshudov TahomaSent:Tahoma Wednesday, July 30, 2008 5:18 AM TahomaTo:Tahoma 0000,0000,EEEE[log in to unmask] TahomaSubject:Tahoma Re: [ccp4bb] Preventing close contact between protein and ligand Times New Roman  Times New RomanDear Snageetha Times New Roman  Times New Roman1) Could you check please if specified atoms have zero occupancy. Atoms with zero occupancy are considered as absent and there are not restraints on them Times New Roman2) symm y at the end of instructions means that the program check all possible symmetry operators and finds minimal distance. Most probably 5.024 is the distance between symmetry related atoms Times New Roman3) to remove antibumping between different chains there is an undocumented keyword. It can be used. the keyword is (as an example) Times New Roman  Times New Romanvdwrestraints exclude between chains A B  Times New Roman  Times New Roman  Times New RomanPlease let me know if this instruction does not work. Times New RomanNB: This option should not be used unless you know what you are doing (that is the reason why it has not been documented). If there are clashes between chains then there are reasons for that. For example Times New Romanif ligand has half occupancy then it is very likely that surrounding atoms also have multiple conformation and you should model them.  Times New Roman  Times New Roman  Times New Romanregards Times New RomanGarib Times New Roman  Times New Roman  Times New RomanOn 29 Jul 2008, at 18:24, Sangeetha Vedula wrote: Times New RomanDear bb users, Times New RomanI am refining a protein-ligand complex (at 1.68 A resolution) in which the ligand lies on a 2-fold crystallographic symmetry axis. The ligand occupancy is, therefore, 0.5 in each asymmetric unit. Times New RomanI am almost at the end of the refinement but one problem has me stumped. Refmac keeps moving a carbon in the ligand too close to a serine OG and an oxygen too close to an arginine CD. Given that the ligand is at the interface, the density is not perfect. However, I rebuild the ligand to eliminate close contacts and still be within density and refmac pulls it right back close to the protein. The refined position does not even look better than the rebuilt one! It almost always looks worse! Would refmac put less weight on close contacts with the ligand because it is only partially occupied? Times New RomanI tried to use external restraints between the ligand and the residues so that they are kept further away.  Times New RomanUpon searching the net, I found this command line: Times New Romanexternal distance first chain [ch] residue [res] insertion [ins] - Times New Romanatom [n] [altcode [a]] second chain [ch] residue [res] insertion [ins]- Times New Romanatom [n] [altcode [a] ] value [v] sigma [s] [symm y/n] Times New RomanI thought (hoped) that the distance herein is the minimum distance of approach between the specified atoms, I added these lines from within "Developer options" in refmac interface: Times New Romanexte dist first chain A resi 59 atom CD seco chain X resi 2001 atom O1 valu 3.2 sigm 0.02 symm Y Times New Romanexte dist first chain A resi 27 atom OG seco chain X resi 2001 atom C10 valu 3.2 sigm 0.02 symm Y Times New RomanIt didn't recognize these restraints at all. Times New RomanHowever, when I change these lines to: Times New Romanexte dist first chain A resi 59 atom CA seco chain X resi 2001 atom O1 valu 3.2 sigm 0.02 symm Y Times New Romanexte dist first chain A resi 27 atom OG seco chain X resi 2001 atom C10 valu 3.2 sigm 0.02 symm Y Times New RomanRefmac recognizes the first line but not the second - lines from log file: Times New RomanBond distance deviations from the ideal >10.000Sigma will be monitored Times New Roman  Times New RomanA     59 ARG CA  . - X   2001 DIE O1  . mod.= 5.024 id.= 3.200 dev= -1.824 sig.= 0.020 Times New RomanThis raises two concerns: Times New RomanConcern 1: From the first line of output: the restraints here don't seem to be minimizing close contact at all; it seems to think they are bonded somehow (the distance between these atoms is not 5.024; it is 6.26 A; I don't know what 5.024 A is!).  Times New RomanI am missing something here. It'd be great if someone can tell me what that is! Times New RomanConcern 2: This command only works when the first atom specified is a C-alpha atom (or maybe a main chain atom; I didn't try using other main chain atoms). Why is that? Times New RomanAND ULTIMATELY, Times New Romanis there some way I can tell refmac not to make the ligand and protein clash? Times New RomanI'd really appreciate any help! Times New RomanThanks, Times New RomanSangeetha. Times New Roman  -------------------------------------------------------------------------------------------------------------------------- Aleksander W. Roszak, PhD E-mail: [log in to unmask] Protein Crystallography Web: www.chem.gla.ac.uk/~aleks University of Glasgow Fax: +44-(0)141-330 3779 Level 3 Room B 317 Tel (office): +44-(0)141-330 4476 Glasgow Biomedical Research Centre Tel (X-ray lab): +44-(0)141-330 3589 120 University Place Mobile: +44-(0)780 9559996 Glasgow G12 8TA Scotland, UK