Missing density could point to incomplete data, how is the completeness at high and low resolution?
Sent from my iPhone On May 19, 2011, at 11:54 AM, Jon Schuermann <schue...@anl.gov> wrote: > Dear Joane, > > Bert's recommendations are very good, but I'd like to add a little > caution. If just part of the molecule has bad density then it is not unusual, > but if the whole chain does not look very good (missing side-chains or > backbone) you may have a couple of things going on. First, I would first > suspect NCS issues (as Bert has) where you might be trying to force too high > a space group since an NCS operator might be darn close to a crystallographic > operator. Second it could also be caused by twinning, but since your > R-factors are pretty respectable I would imagine that is probably not the > case although you may want to look into it. Third, it could be static > disorder where the crappy density might be two molecules on top of each other > but one is slightly shifted. A way to tell is to scale down in your electron > density maps to see if there are extra strands and helices next to the > strands and helices you have built. Hopefully this is not the case and it is > just NCS messing with you... > > Jon > > -- > Jonathan P. Schuermann, Ph. D. > Beamline Scientist > NE-CAT, Building 436E > Advanced Photon Source (APS) > Argonne National Laboratory > 9700 South Cass Avenue > Argonne, IL 60439 > > email: schue...@anl.gov > Tel: (630) 252-0682 > Fax: (630) 252-0687 > > > On 05/19/2011 10:06 AM, Van Den Berg, Bert wrote: >> >> One more thing though: have you refined with the NCS restraints off or on? >> Presumably on, seeing that you have a small gap between R and Rfree? It may >> be worth turning the restraints off or modify the NCS selections: if your 3 >> molecules are substantially different, applying NCS may make things worse >> rather than better (ie you’re averaging things that are different). >> >> Good luck, Bert >> >> >> On 5/19/11 9:02 AM, "Joane Kathelen Rustiguel" <jkrustig...@usp.br> wrote: >> >> Dear all >> >> >> I am refining a structure at 3.4 A resolution that contains 3 molecules in >> the >> a.u. The chain A sits on a 2-fold crystallographic axis forming the dimeric >> functional structure expected for this class of proteins. The other two >> chains >> B and C, which also form the functional dimer, seem to be, somehow, a lot >> more >> flexible than chain A. As a result, whereas the electron density map, >> b-factor >> and geometry for chain A is pretty reasonable for a 3.4 A resolution >> structure, the refinement for the other two chains (B and C) does not behave >> well. Even playing with different weights for geometry, analysing different >> levels of 2Fo-Fc/Fo-Fc maps, using NCS, TLS, etc..., nothing works. The map >> for the helical regions is ok, but the electron density map for strands and >> loops of chains B and C are broken along the main chain, B-factors are really >> high, and the geometry keeps being distorted. >> >> Right now, the R-factor and R-free are 24.2 and 28.6, respectively. >> >> Any suggestions in how to proceed the refinement? >> And even a more difficult question, how do we report this type of structure? >> How do we deposit those coordinates? We can certainly use chain A as a model >> to perform interesting studies of structure-function relationship, but we >> know >> that chain B and chain C have problems. >> >> Any help will be greatly appreciated. >> >> Regards >> >> Joane >> >> >> -- >> Joane Kathelen Rustiguel Bonalumi >> Faculdade de Ciências Farmacêuticas de Ribeirão Preto - USP >> Laboratório de Cristalografia de Proteínas >> Departamento de Física e Química >> Fone: +55.16.3602.4193 >> >> > >
