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Hi Appu, You have more than 4 reflections per atom. With restraints you should be able to refine individual B-factors in Refmac. Actually your reflections/atom is very high. Is that because your model is still rather incomplete or do you have a high solvent content? Anyway, your B-factors are not that scary but you can try a few things to make them more plausible: Reset them to, say, 30 at the start of refinement and set the B-factor restraint weight to a value greater than 1 (higher is tighter here). For your model PDB_REDO would try 2.5, 2.0, 1.5 and 1.2. See where the B-factors level out and wether the distribution is plausible (I.e. Lower B-factors in the core and higher at the surface of the protein; no large changes across atomic bonds). Once your model is in good shape you can try doing a Hamilton test to decide your B-factor model. For what it's worth, at more than 4 reflections per atom such a test would typically allow individual B-factors if your restraints are effective. Cheers, Robbie Http://xtal.nki.nl/PDB_REDO Sent with my Windows Phone ________________________________ Van: Appu kumar<mailto:[log in to unmask]> Verzonden: 22-7-2015 16:08 Aan: [log in to unmask]<mailto:[log in to unmask]> Onderwerp: [ccp4bb] High B-factor during refinement at 4A Dear All, I am facing troubles in refining structure (C2221 space group) at 4A resolution . The current Rwork/Rfree stalls at 35.8/37.7.Given low resolution, the total number of unique reflections accounts to *19356 while number of atoms in pdb corresponds to **3893. *At this very low resolution, the number of available observations are insufficient to support the refinement of four parameters (x, y Z, Biso) and therefore i think the modeled atomic displacement (B-factors) are excessively high ranging from 100-280 which is scary at this moment. Is there any illustrative parameters to work on which might be useful for putting control over the shooting up of B-factors during refinement. I tried incorporating the group B-factor refinement with TLS group but seems like is not much useful. I would appreciate any incoming suggestions and thoughts on adopting crucial refinement strategies pertaining to this resolution. Thank you Appu