Peter from you description seems that big blob is not your ligand. In 3.0 resolution molecules with similar size, including ligands, tend to look similar in shape as well. I suggest you refine further your structure and check again later this N-terminal blob of density. It may tern out to be a bias from your molecular replacement model. If it is still there I will suggest to rum a MS and compare the MW with the one from your protein structure. That will give you an indication that probable you have something attached to N-terminal of your protein.
George _____ From: CCP4 bulletin board [mailto:[email protected]] On Behalf Of peter hudson Sent: Thursday, April 02, 2009 9:55 AM To: [email protected] Subject: [ccp4bb] ligand refinemnet Hello all I have refined and built model a dataset of 3.0A resolution dataset. This model is assocaied with a ligand. After final refinement and model building i found a big blob of Fo-Fc density of around 4sigma level at the N-terminal of the fianl model. My protein doesnot carry any tag at the N-terminal. But, the template i have used for the molecular replacement carrying a tag at the N-termianl, which can occupy only 1/4th of this positive density after superpostion. My crystallisation conditions component cannot occupy such higher level of positive density. Since my protein binds to a ligand, i looked carefully to the positive density and it seems very similar to the ligand density but its obscure. Refinement after fitting the ligand leads to very high B-factor of the ligan, which vary for various atoms from 50-90 and simultaneously positive density goes off. I also tried to change the occupancy of the ligand but as i reduce the occupancy the B factor comes down at the noraml expected average B factor value but again Fo-Fc density appears in the map over ligand. If i leave to refine the occupancy to the programme automatically, this lead to the zero occupancy of the few atoms in the ligand and avearge B factor remains normal. suggestions would be appreciated. Thanks in advance Peter
