Hi Kyle, It's always worth questioning your data and your model, so yes I'd examine both carefully. I am guessing that this is molecular replacement solution. (It isn't as common to have a loop that disappears during model-building with experimental phasing because normally you would not build it in the first place unless the density was there. My suggestions would be similar in either case though.) So (still guessing) your starting MR model had some parts that are correct, some that are not, and some that are close. You carry our MR and place your model. You calculate a sigmaA-weighted map, probably after refinement. This map shows great density in many places, medium density in others, little density in some. You rebuild the parts that are clear to you, but some are not so clear, including the helix and loop in question. You refine some more, and now you want to improve the density in this unclear region. Then as you say below, you calculate an average kick map, and it doesn't look so good in this unclear region. You calculate an iterative build omit map and the density in this region is nearly gone. A comment or two first: 1. Yes, it is possible that the presence of a symmetry element very near the problematic area can make the density noisier. Normally this happens only right at the symmetry element however, not even a few A away. 2. The averaged kick maps and iterative build omit maps are ways to remove model bias. In general (with some exceptions) they are not great ways to "improve" maps. Generally your best maps for building are going to be your current 2Fo-Fc map (e.g., from phenix.refine) or a density modified map using model information (e.g., overall_best_denmod_map_coeffs.mtz from autobuild). The omit and kick maps are to help you determine whether you have overinterpreted your density. So if this is all about right, here is my suggestion: try to see if there is any evidence whatsoever for density in this region. A. Go back to the MR solution (very beginning of this structure solution, before any refinement except rigid-body). As this model has not been refined, you can delete any portion, refine the rest, and there will be no model bias in the map. B. So: delete the region in question completely from this unrefined starting model. C. Then refine the model, calculate a map, and look at the map in the region of your helix and linker. If there is density (even not great density) that clearly matches the structure of the helix and the linker, then you know for sure that this helix and linker belong there. If not...perhaps they are disordered or somewhere else. D. If no density yet, you can , iterative build the model, do whatever you want with this model to make it better except do not put the helix and linker back in...and then look at the map in the region of the helix and linker. Still you have not biased the map, so if you can find them they are really there. If you cannot see them...now it is time to call them "disordered" until you get further evidence. I hope that helps! All the best, Tom T On Aug 9, 2010, at 3:08 PM, Kyle Dolan wrote:
Hi all,
I attempted to improve the density around a helix and short linker region using an average kick map but it didn't make things look much better. I then tried creating a composite iterative-build omit map from phenix.autobuild, but in this case the electron density around the helix/linker in question largely disappeared! Is there another approach I can use that might improve my maps, or should I start questioning my data? The dataset has >95% completeness to 2.7 A so I am very curious as to why I am missing the backbone now, where before I was only missing the side chain densities. One thing I am wondering is if the problem could have something to do with symmetry; the part of the map I am looking at may be close to a two- fold crystallographic symmetry axis and I have been told that these regions tend to be very noisy and hard to model--comments?
Kyle
Kyle T. Dolan Department of Biochemistry and Molecular Biology The University of Chicago [email protected] _______________________________________________ phenixbb mailing list [email protected] http://phenix-online.org/mailman/listinfo/phenixbb
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