Hi Mike, Often, I generate independent freeR set (especially in cases where "soak" dataset is of different resolution (usually worse) compared to the native dataset and do following two things to get rid-off the bias: 1. add a noise to the coordinates (this can be done using PDBSET). 2. set the Bvalues to the wilson B of the "soak" dataset (within Refmac5 before rigidbody refinement).
HTH, Partha On Mon, Nov 21, 2011 at 5:47 PM, Michael Thompson <mi...@chem.ucla.edu>wrote: > ----- Forwarded Message ----- > From: "Michael Thompson" <mi...@chem.ucla.edu> > To: "e dodson" <e.dod...@ysbl.york.ac.uk> > Sent: Monday, November 21, 2011 11:30:17 AM GMT -08:00 US/Canada Pacific > Subject: Re: [ccp4bb] LESS MR pleae.. 1.95A, different phase > > A question regarding the plea for less MR (which I support): > > There have been several recent instances in which I have used the solution > of an isomorphous structure to do rigid body refinement for a new crystal > (as described by Eleanor). It has always produced good results. My question > is about how to best handle the free set of reflections when doing this? I > have heard a number of differing opinions about whether or not it is > important to carry the freeR flags from the original structure over to the > new data set. I have heard equally convincing arguments from both sides, so > my young and impressionable mind does not know who to believe. I was hoping > I could get an opinion from the "advocates for less MR." > > Sorry for hijacking this thread, but hopefully it will provide some > insight that is relevant to the original post. > > Thanks! > > Mike > > > > > ----- Original Message ----- > From: "Eleanor Dodson" <c...@ysbl.york.ac.uk> > To: CCP4BB@JISCMAIL.AC.UK > Sent: Monday, November 21, 2011 2:23:21 AM GMT -08:00 US/Canada Pacific > Subject: Re: [ccp4bb] LESS MR pleae.. 1.95A, different phase > > Just a plea for less molecular replacement. > > If you get a new crystal of a known protein with the same cell > dimension as youur old crystal, the most likely scenario is that it has > the same group, and you really should not try MR - use the previous > solution as input to do rigid body refinement, and then > a) the R factor will tell you if this is a reasonable hypothesis (it > usually is..) and > b) you dont have this awful problem of not being able to compare the > solutions.. > > Eleanor > > On 11/20/2011 03:57 PM, Napoleão Valadares wrote: > > Thank you all for the replies. Felix Frolow, Dan Leahy, Hans > > Brandstetter, Boaz Shaanan and Tim Gruene you really helped a lot. > > > > I think I understand it now, I always thought the "one ring to rule them > > all" translated in the crystallography realms to "one origin to rule > > them all". That probably means I have a long road in front of me. > > > > I'm still half confused, I definitely need to read more, as much as I > > read about symmetry and space groups I never seem to improve or get a > > better understanding, but I'll keep trying. > > > > About the same origin: > > The pdbs of both Solution-1 and Solution-2 present the same space group > > and cell, as observed opening the pdbs as text files or in pymol. When I > > open both maps on coot they are not superposed but present the same cell > > and origin. > > > > If I open both solutions on pymol they clash. If I generate the symmetry > > mates of both solutions none of them are superposed, instead they clash. > > But I think they are related as you all pointed, I'll check it out. > > > > Thank you all for your kind answers and your patience with a beginner. > > Regards from a sunny Brazil, > > Napo > > > > > > On 11/20/2011 2:58 AM, Felix Frolow wrote: > >> Napoleao, > >> It is so called alternative origins play a game with you. You do not > >> change your structure by shifting 1/2 translation (or even combination > >> of these translations) > >> into directions of the main axes of your unit cell. Structure factors > >> after this operation stay the same, however phases change > >> systematically, producing however the same > >> map features. > >> Would I be a begin crystallographer now, I would read a bit more old > >> fashioned books > >> on crystallography such as probably Jensen and Stout… > >> FF > >> Dr Felix Frolow > >> Professor of Structural Biology and Biotechnology > >> Department of Molecular Microbiology > >> and Biotechnology > >> Tel Aviv University 69978, Israel > >> > >> Acta Crystallographica F, co-editor > >> > >> e-mail: mbfro...@post.tau.ac.il <mailto:mbfro...@post.tau.ac.il> > >> Tel: ++972-3640-8723 > >> Fax: ++972-3640-9407 > >> Cellular: 0547 459 608 > >> > >> On Nov 20, 2011, at 07:42 , Napoleão Valadares wrote: > >> > >>> Hello, > >>> I'm observing a very strange phenomena (at least to me, I'm a > >>> beginner). It is related to symmetry (I think). > >>> > >>> I got a data set at 1.95A (I/Sigma 3.5, R-Factor and R-meas < 35% in > >>> the last shell) and a partially refined solution with R/Rfree 22/24, > >>> 166 aminoacids observed and around 30 solvent molecules. I'll call > >>> this Solution-1. The refinement was smooth, the densities were very > >>> clearly "asking" for the correct missing side chains and the map > >>> looks good. > >>> > >>> The space group I'm using is P212121, pointless and XDS agree with > >>> that (but me and pointless both have a long history of being wrong > >>> about space groups). Phenix.xtriage says there's no twinning. > >>> > >>> I took Solution-1 and used it as a template in a molecular > >>> replacement in the same space group (P212121) using the same mtz as > >>> the one used to refine the template. I got a different (not > >>> superposed in space) solution (called Solution-2, scores by Phaser > >>> RFZ=24.2 TFZ=33.0 PAK=0 LLG=1413 LLG=1899) that was readily refined > >>> in Phenix to R/Rfree 24/26 without any solvent molecule. > >>> > >>> - The solutions are not superposed in space, although they are > >>> near-identical and can be superposed yielding a C-alpha rmd =0.001. > >>> - Both structures present VERY similar density maps. The maps are not > >>> superposed in space, but when you "run the chain" in one map in Coot > >>> and do the same in the other it they the present exactly the same > >>> features. It is impossible to ignore their similarities. > >>> - Both structures and maps present the same origin and unit cell. > >>> - If I add to Solution-2 the equivalent solvent molecules of > >>> Solution-1 (I did this by superposing Solution-1 to Solution-2 then > >>> copy/pasting the solvent molecules), the R/Rfree become 22/24. This > >>> is a clear indication that the solutions are related. > >>> - I can't find any MR solutions using the same template and space > >>> groups P222, P2221 and P21212. > >>> > >>> How two different sets of phases can yield maps with the same > >>> features? What is happening, wrong space group? I have a feeling my > >>> lack of experience is the problem. > >>> Thank you. > >>> Regards, > >>> Napo > >> > > > > > > -- > Michael C. Thompson > > Graduate Student > > Biochemistry & Molecular Biology Division > > Department of Chemistry & Biochemistry > > University of California, Los Angeles > > mi...@chem.ucla.edu > > -- > Michael C. Thompson > > Graduate Student > > Biochemistry & Molecular Biology Division > > Department of Chemistry & Biochemistry > > University of California, Los Angeles > > mi...@chem.ucla.edu >
