Hi All, perhaps rank scaling of maps as described here
Acta Cryst. (2014). D70, 2593-2606 instead of "sigma" scaling is a way to eliminate the ambiguity inherent to "sigma" scaling. Pavel On Wed, May 27, 2015 at 1:31 PM, Gerard Bricogne <[email protected]> wrote: > Dear Sagar and Pavel, > > Yes, it may be model bias, but you also have to bear in mind that > the rmsd that is used as a unit in the choice of contouring level > should not be thought of as the standard deviation of a Gaussian > random variable and therefore implicitly associated to a "significance > level" relative to a noise level. This is unfortunately a widespread > misconception. > > We have seen a case where an entire copy of a molecule had been > missed at first because its density was visible only at a contour > level of 0.35 "sigma", which at first sight would make anybody shrink > back in horror. When it was nevertheless modelled and refined, there > was no doubt whatsoever that it was there, making perfectly sensible > contacts with its nearest neighbour molecules, obeying good NCS with > the already placed copies and markedly improving the fit to the data. > The peculiarity of that extra copy was that a clash around a symmetry > element with its symmetry mate caused it to have a mean B-factor that > was twice that of the other, well-ordered, copies of the molecule. > > Now, if you look at the expression for the Debye-Waller factor in > real space, you will find the B-factor raised to the power 3/2 in the > denominator. That means that the blurring effect of that D-W factor in > real space results in a lowering of the maximum value of an atomic > electron density to which it is applied by B^(3/2). > > The consequence is that electron density for a molecule whose > mean B-factor is double that of another molecule will look similar at > a contour level of 1.0/(2^(3/2)) = 0.35 to what the other molecule > looks like when examined at a contour level of 1.0. > > It is therefore a good idea to remember that the "sigma" level is > mainly determined by the better-ordered region(s) of the molecule(s) > present in the asymmetric units, and that less-well ordered regions > will automatically have a contouring level handicap that is not to be > automatically interpreted as a lower level of significance. Omit maps > are a good idea of course, but do bear in mind the (B/B0)^(-3/2) > effect, where B0 is the mean B in the well-ordered region, and B is > the same quantity in a less well ordered one. Above all, don't confuse > "sigma" with a noise level! > > > With best wishes, > > Gerard. > > -- > On Wed, May 27, 2015 at 12:18:25PM +0100, Pavel Afonine wrote: > > Sagar, > > > > what you see may be model bias unless you calculated these maps without > > that region of molecule. > > > > Pavel > > > > On Wed, May 27, 2015 at 12:03 PM, Sagar De'Biomimic < > > [email protected]> wrote: > > > > > Dear all, > > > > > > We have solved a structure of Protein-DNA complex. In an ASU we have > two > > > protein homodimers and two DNA duplexes. Additionally we were able to > model > > > N-terminal region (NTR) missing in the previously reported structure. > > > > > > I had modelled poly-alanine chains in the positive density and refined. > > > These chains were then joined to form NTR. Finally the entire NTR was > > > refined for occupancies. > > > > > > I am doubtful about the validity of modelling of NTR as i had dropped > down > > > sigma levels to as low as 0.8 (as i could not see much of the noise) of > > > FO-FC and modelled in positive density. After refinment the region > shows > > > the density at 0.5 sigma level of 2FO-FC. I have attached a pdf file > > > showing snapshots of NTR at 0.4, 0.5, 0.6, 0.7 sigma level of 2FO-FC > map. > > > > > > I would like to know crystallography community's opinion on validity of > > > such modelling. > > > > > > This NTR region is highly flexible. Main intention of modelling it in > such > > > a weak signal was to complete the model for our Molecular dynamics > > > studies. > > > > > > Additionally we have confirmed with SAXS DATA using Ensemble optimised > > > modelling (EOM 2.0), the high flexibility and multiple conformations of > > > this NTR region. > > > > > > I would like to know if there is a possibility of having a tool > similar to > > > EOM to model highly flexible regions of a protein in electron Density. > As > > > far as I know xMDFF does better at lower resolutions but fails to model > > > highly flexible regions such as ours. > > > > > > Thank you. > > > > > > Regards > > > > > > Sagar > > > > > > > > > > > > -- > > > Sagar Khavnekar > > > Project student, > > > Structural Biology and Molecular Biophysics lab, > > > UM-DAE Centre for Excellence in Basic Sciences > > > University of Mumbai, Vidyanagari Campus, Kalina, Santacruz (East) > > > Mumbai 400098, India. > > > > > -- > > =============================================================== > * * > * Gerard Bricogne [email protected] * > * * > * Global Phasing Ltd. * > * Sheraton House, Castle Park Tel: +44-(0)1223-353033 * > * Cambridge CB3 0AX, UK Fax: +44-(0)1223-366889 * > * * > =============================================================== >
