Hmm - I cant quite understand your map - that density is not for the lysine ? It looks like a well ordered PHE contoured at quite a high level? As Edward suggests - if you omit a well ordered feature the resultant difference maps often show high positive density and a surrounding complementary "sag" of negative density - not really something to worry about.. But I wonder why you don't just do a bog standard difference map? Set the occupancies of the loop you want to omit to 0.00 - do a few cycles of refinement and see what comes back? A prediction - the well ordered features will show up loud and clear and your surface LYS wont! High resolution structures show they often are in multiple conformations, which would be hard to model at 2.1A.
And just a thought about negative features in difference maps. I often see random "holes" which don't seem to have any logical explanation.. I sloppily put them down to "data defects - missing data? poorly measured data? etc - but does anyone have a more satisfactory explanation? Or dont other people see them?? Eleanor On Thu, 25 Jul 2024 at 01:48, Edward A. Berry <[email protected]> wrote: > Does your difference map have mean value ~zero (over 1 ASU or cell)? If > maps are constructed by > Fourier transform without the 0 0 0 reflection, they have mean of zero > (because the mean of a > sinusoid over one period is zero). That means that any time you add > positive (difference) density, > which raises the mean value of the absolute map, the Fourier map has to > sink down a little to bring > that new mean value to zero. > Now if your data were complete except for the 0 0 0 reflection, this would > move the floor down > everywhere by a small amount to balance the large increase at the local > peak, and it would not go > below the negative contour limit. But without the ultra-low resolution > amplitudes the FT cannot make > a constant offset, and instead you get sagging around the peak, like a > water-bed sagging around the > spot where a child (the peak) is standing. This could lead to the sagging > part going below the > negative contour and giving you the red density. > However I would not have expected the negative density to be quite so > localized around the peak - > depending on your low-res cutoff. As John Bacik suggested you should check > for a mis-modeled part of > a symm-related molecule, but if that were the case, you should see the red > density even before > omitting the loop, and you should see the red density somewhere else on > your model. If it just > appeared after omitting the loop, it could be due to the map sagging under > the weight of the peak, > and should go away as soon as the residue is replaced. > eab > > Renuka Kadirvelraj wrote: > > Hi CCP4bb, > > We would greatly appreciate your advice regarding an odd problem that > has cropped up with one of our > > crystal structures. We have a protein structure in space group P6(3)22 > at 2.1 A resolution and in > > the final refinement stages with Rwork of 0.20 and Rfree of 0.22. When > we ran simulated annealing to > > rebuild an outlier Lysine residue in a loop (using Phenix), the Sim Omit > map showed the positive > > density expected from the omission of the loop (colored green in > attached pics). However, flanking > > the positive density, there is a complimentary negative density (in > red). We are puzzled as to the > > cause of the appearance of the negative density. Can you help us? > > Many thanks, > > Renu > > > > --------------------------------------------- > > Renu Kadirvelraj, Ph.D. > > Research Scientist > > A428, Biochemistry and Molecular Biology > > 120, East Green Street > > University of Georgia > > Athens, GA 30602 > > Tel: (706) 583 0303 > > > > > ---------------------------------------------------------------------------------------------------- > > > > To unsubscribe from the CCP4BB list, click the following link: > > https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 > > > > ######################################################################## > > To unsubscribe from the CCP4BB list, click the following link: > https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 > > This message was issued to members of www.jiscmail.ac.uk/CCP4BB, a > mailing list hosted by www.jiscmail.ac.uk, terms & conditions are > available at https://www.jiscmail.ac.uk/policyandsecurity/ > ######################################################################## To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 This message was issued to members of www.jiscmail.ac.uk/CCP4BB, a mailing list hosted by www.jiscmail.ac.uk, terms & conditions are available at https://www.jiscmail.ac.uk/policyandsecurity/
