Dear Matthew, Robbie Hans and Oliver, Thanks for the advice, it seems that it is simply the geometry definitions in the dictionaries distributed with CCP4 6.4 are different to those calculated from the atomic coordinates. For example, the chiral volume definitions in the for the 4Fe4S cluster are the opposite sign to those in my protein. So swapping the atom names has sorted that, I am still working on the other clusters, but progress is being made.
Since I solved the structure by MR using coordinates downloaded from the pdb, I assumed the atom names would agree with the geometry definitions in Refmac (since the search model was also refined with refmac). Have the cif dictionaries been modified/updated recently? thanks again, Steve Dr Stephen Carr Research Complex at Harwell (RCaH) Rutherford Appleton Laboratory Harwell Oxford Didcot Oxon OX11 0FA United Kingdom Email stephen.c...@rc-harwell.ac.uk tel 01235 567717 ________________________________________ From: "Oliver Smart" [osm...@smartsci.uk] Sent: 13 October 2014 12:09 To: ccp4bb Subject: Re: [ccp4bb] Refinement of Iron-sulphur clusters Stephen, Robbie advice is 100% correct. Be careful about the naming of the sulphur atoms (is S1 opposite S4 or next to it). I recall that the distributed CCP4 dictionaries have different atom naming than the PDB chemical components dictionary. If this is the problem is that the naming is different this should result in large restraint violations that should be appear in the REFMAC output (others can tell you where). In general diffraction from Fe S clusters is so strong that the atom positions are pretty much determined from the electron density. But in case it helps I did some work data mining restraints for Fe2S2 and Fe4S4 from CSD structures (this has to be done manually). Please find attached the resulting FES.cif and SF4.cif restraint dictionaries (distributed with BUSTER). Good luck, Oliver ------------------- Dr Oliver Smart Director SmartSci Limited http://www.smartsci.uk/ & Consultant Global Phasing Ltd http://www.globalphasing.com/ on 10/10/14 5:33 PM, Robbie Joosten <robbie_joos...@hotmail.com> wrote: > Dear Stephen, > > The dictionary is very specific about atom names. If you have them swapped > (as many PDB entries do). The angle and chiral volume restraints will wreck > your cluster. You also need to make sure to provide LINK records to attach > the cluster to the surrounding cysteines. > > HTH, > Robbie > > Sent from my Windows Phone > ________________________________ > Van: <Stephen Carr> > Verzonden: 10-10-2014 18:07 > Aan: CCP4BB@JISCMAIL.AC.UK > Onderwerp: [ccp4bb] Refinement of Iron-sulphur clusters > > Dear CCP4 BBers, > > I am currently refining a model of a protein containing three iron sulphur > clusters using Refmac (v5.8.0078) and am getting some unusual results. One > of the clusters (3fe 4s) seems to behave resonably and the refined electron > density looks very nice. The atoms in the others clusters (4Fe4S and > 4Fe3S), however, migrate towards the centre of the cluster producing Fo-Fc > difference maps with strong negative density at the centre surrounded by > blobs of positive density where the atoms should be. > > I have checkedthe refmac dictionaries and there are entries for each of the > clusters, so I shouldn't have to explicitly include a cif file for the > clusters as an input right? In fact, when I did try to include a dictionary > file for one of the clusters refmac gave a warning message in the log about > duplicate monomers dictionaries and it made no difference to the > refinement/maps anyway. > > On checking the refmac log file there is no specific mention that there are > hetero-groups in the pdb file and no explicit mention that it is using any > dictionaries during the refinement. How can I tell if refmac is > reading/using the dictionaries? Also bonds within the clusters are flagged > up as outliers and deviating by more than 10 sigma from ideal suggesting > that they are not getting read? > > I realise that something chemically odd could be going on at the centres, > but if I omit them from the model and calculate maps very strong density > comes back around where the atoms sat before refinement, so I am pretty > confident that they are in the right place to start with. > > Any help with this problem would be greatly appreciated, I'm sure there is > something obvious that I am missing but can't see what that is at the > moment. It is particularly confusing since one cluster apparently behaves > while the others do not. I could, of course, try phenix or buster, but I > would like to get to the bottom of the problem with refmac if possible. > > Thanks very much in advance, > > Steve > > Dr Stephen Carr > Research Complex at Harwell (RCaH) > Rutherford Appleton Laboratory > Harwell Oxford > Didcot > Oxon OX11 0FA > United Kingdom > Email stephen.c...@rc-harwell.ac.uk > tel 01235 567717 > This email and any attachments may contain confidential, copyright and or > privileged material, and are for the use of the intended addressee only. 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