Crystals with many copies of a single model are indeed notoriously difficult to solve by MR. There is often a confusion between crystal symmetry and non-crystallographic symmetry, but in your case that wont happen - since the SG is P1.
First thing i would check is the self-rotation function. Is there evidence of n-fold symmetry in the data - 2-folds? 3-folds? etc.. self rotations outputs are often confusing but there is useful information hidden in the MOLREP plots and log files (one confusingly called x.doc , which is a standard text file!) And if any model forms an assembly (PISA will document that) search with the assembly, and not a monomer. Luckily your spacegroup is P1 so you should find the whole assembly - if it exists - in the asymmetric unit.. Eleanor On Fri, 6 Sep 2019 at 08:01, Gerard Bricogne <g...@globalphasing.com> wrote: > Dear Napoleão, > > If you have such a large number of copies of the molecule in the > asymmetric unit, you should perhaps try and see whether there is some > orientational regularity in the way they are arranged by calculating a > self-rotation function. Any such indication might enable you to filter > solutions on that basis. > > > With best wishes, > > Gerard. > > -- > On Thu, Sep 05, 2019 at 11:03:42PM -0300, Napoleão wrote: > > Dear all, > > Thank you for your kind expert suggestions, and sorry for the late > > reply, I'm still trying all the suggested approaches. > > Some comments: > > - I can't use buccaneer yet, since I'm still working to resolve the > > structure. > > - Robyn Stanfield's suggestion of using Shelxe to try to verify the > > validity of the MR solutions was very helpful. I'm trying it a lot, > > however without success. Thank you Robyn and all Shelx developers! > > - The maps phased with partial molecular replacement solutions look > > reasonable, but do not show any sign of the rest of the molecules or of > > the missing side chains. > > - ContaMiner gave me green lights, so it does not appear to be a > > contaminant (thank you Ivan Shabalin). > > - Similarly, SAUC indicates there is no other PDB with cell parameters > > similar to this data set (thank you Jonathan Cooper). > > - I am currently using EPMR to try to resolve the structure by MR (thank > > you Roger Rowlett). > > - I am not sure if there is translational NCS, as Phil Jeffrey suggested > > despite phenix.xtriage's green lights. I'm not experienced with this > > issue, so I'm sending the log file to Randy Read as he kindly offered to > > help. > > > > I think one of my problems is that the AU is big (at least to me), > > apparently containing from 9 to 12 copies of my protein, so the MR > > solutions contain less than 10% of the scatters in the AU. > > Maps look good when I refine one chain of the MR solutions in Phenix > > (blue maps over most of the protein, with reasonable main chain > > continuity and very few green or red blobs), but the R values are 0.50 > > and I can't see any feature suggesting the solution is good (like a > > well-defined new side chain). > > I will keep trying. > > > > Thank you all again! And please let me know if any ideas cross your wise > > minds. > > Regards, > > Napo > > > > Em 2019-08-30 06:29, Randy Read escreveu: > > > > > Dear Napoleão, > > > Yes, I agree with Phil that it looks like a case where there *is* > translational NCS but it's not being used by Phaser, either because it > wasn't automatically detected (native Patterson peak just under the default > limit? more than one off-origin Patterson peak?) or because Phaser was told > to ignore tNCS. You should look in detail at the relevant section of the > logfile, and manually override Phaser's automated decision if you think > there's good evidence for tNCS. I would say that, as Phil noted, the fact > that two molecules are in basically the same orientation separated by a > translation is pretty strong evidence. In particular, the fact that > placing the second molecule in the same orientation gave such a large > increase in both LLG and TFZ is strong evidence: this tells us that having > two molecules in the same orientation (even if they're the wrong molecule > or in the wrong orientation) explains some feature of the data, i.e. the > modulation caused by tNCS. > > > > > > I'd be happy to look at the log file for you if you find it hard to > interpret. > > > > > > Best wishes, > > > > > > Randy Read > > > > > > On 29 Aug 2019, at 17:24, Phil Jeffrey <pjeff...@princeton.edu> > wrote: > > > > > > Are you *sure* there's no translational NCS ? > > > > > > For example your first molecular replacement solution out of Phenix > shows > > > > > > EULER 293.6 27.7 288.7 > > > FRAC -0.02 0.02 0.02 > > > (that's "first molecule at origin in P1") > > > > > > and > > > > > > EULER 294.0 27.9 288.8 > > > FRAC -0.37 0.02 0.02 > > > > > > which is essentially the same orientation, and a translation down one > crystallographic axis (a*) > > > > > > And this suggests to me that either Xtriage or Phaser is missing > something here. Does Phaser find translational NCS in its initial data > analysis ? Unmodeled translational NCS could cause significant problems > with the molecular replacement search. > > > > > > Phil Jeffrey > > > Princeton > > > > > > On 8/29/19 11:28 AM, Napoleão wrote: > > > Deal all, > > > Sorry for the long post. > > > I have a data set obtained from a crystal produced after incubating a > protease with a protein which is mostly composed by an antiparallel beta > sheet. I have tried numerous approaches to solve it, and failed. Molecular > replacement using Phaser, and the protease or the protein as a template > yields no solution. However, molecular replacement using only part of the > beta sheet yields LLG=320 TFZ==28.0 (see below). > > > The apparently good data extends to 1.9 A, as processed by XDS, and > the space group is P1 (pointless agree). XDS info below: > > > SPACE_GROUP_NUMBER= 1 > > > UNIT_CELL_CONSTANTS= 44.43 72.29 77.30 97.802 89.939 101.576 > > > a b ISa > > > 9.647E-01 3.176E-03 18.07 > > > RESOLUTION NUMBER OF REFLECTIONS COMPLETENESS R-FACTOR > R-FACTOR COMPARED I/SIGMA R-meas CC(1/2) Anomal SigAno Nano > > > LIMIT OBSERVED UNIQUE POSSIBLE OF DATA observed expected > Corr > > > 1.90 24890 19149 23814 80.4% 58.1% 63.7% > 11482 0.77 82.2% 63.8* 3 0.694 492 > > > total 163756 125884 146938 85.7% 10.6% 10.8% > 75744 3.78 15.0% 99.0* -3 0.761 5834 > > > Xtriage in Phenix 1.16-3549 gives me all green lights (print below), > suggesting the data presents no twinning, no translational NCS, no ice > rings and is not anisotropic. > > > http://fullonline.org/science/phenix_xtriage_green.png > > > Molecular replacement in Phaser yields single solutions like: > > > Solution annotation (history): > > > SOLU SET RFZ=3.0 TFZ=* PAK=0 LLG=29 RFZ=2.8 TFZ=8.8 PAK=1 LLG=310 > TFZ==27.6 > > > LLG=320 TFZ==28.0 > > > SOLU SPAC P 1 > > > SOLU 6DIM ENSE ensemble1 EULER 293.6 27.7 288.7 FRAC -0.02 0.02 > 0.02 BFAC > > > -6.03 > > > SOLU 6DIM ENSE ensemble1 EULER 294.0 27.9 288.8 FRAC -0.37 0.02 > 0.02 BFAC > > > -6.52 > > > SOLU ENSEMBLE ensemble1 VRMS DELTA -0.1983 RMSD 0.49 #VRMS 0.21 > > > or partial solutions like: > > > Partial Solution #1 annotation (history): > > > SOLU SET RFZ=3.7 TFZ=* PAK=0 LLG=32 RFZ=2.8 TFZ=13.0 PAK=0 LLG=317 > TFZ==30.2 > > > LLG=331 TFZ==30.5 RFZ=2.4 TFZ=7.2 PAK=0 LLG=464 TFZ==18.5 RFZ=2.7 > TFZ=5.7 PAK=1 > > > LLG=501 TFZ==6.8 LLG=509 TFZ==6.6 > > > SOLU SPAC P 1 > > > SOLU 6DIM ENSE ensemble1 EULER 85.4 153.0 138.5 FRAC -0.01 -0.00 > -0.00 BFAC > > > -12.30 > > > SOLU 6DIM ENSE ensemble1 EULER 86.2 153.2 139.5 FRAC -0.36 -0.01 > -0.01 BFAC > > > -9.16 > > > SOLU 6DIM ENSE ensemble1 EULER 83.8 152.3 135.9 FRAC -0.00 0.00 > -0.25 BFAC > > > 1.52 > > > SOLU 6DIM ENSE ensemble1 EULER 191.2 109.1 39.3 FRAC -0.27 -0.01 > 0.22 BFAC > > > 10.18 > > > SOLU ENSEMBLE ensemble1 VRMS DELTA -0.0447 RMSD 0.49 #VRMS 0.44 > > > However, after 1 refinement round in Phenix_Refine (Final: r_work = > 0.4881 r_free = 0.5009) I got densities that are part good and part bad, > and if I delete the bad parts and refine again, the good parts become bad. > Please check the prints: > > > http://fullonline.org/science/good_part_of_density.png > > > http://fullonline.org/science/bad_part_of_density.png > > > What is the explanation for these molecular replacement results? > > > What else should I try? Arcimboldo takes 2 days+ to run and yields no > good solution. > > > Thank you! > > > Regards, > > > Napo > > > > ------------------------------------------------------------------------ > > > To unsubscribe from the CCP4BB list, click the following link: > > > https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB&A=1 > > > > > > > ######################################################################## > > > > > > To unsubscribe from the CCP4BB list, click the following link: > > > https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB&A=1 > > > > ------ > > Randy J. Read > > Department of Haematology, University of Cambridge > > Cambridge Institute for Medical Research Tel: + 44 1223 336500 > > The Keith Peters Building Fax: + 44 1223 > > 336827 > > Hills Road E-mail: > > rj...@cam.ac.uk > > Cambridge CB2 0XY, U.K. > > www-structmed.cimr.cam.ac.uk [1] > > > > ------------------------- > > > > To unsubscribe from the CCP4BB list, click the following link: > > https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB&A=1 > > > > Links: > > ------ > > [1] http://www-structmed.cimr.cam.ac.uk > > > > ######################################################################## > > > > To unsubscribe from the CCP4BB list, click the following link: > > https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB&A=1 > > ######################################################################## > > To unsubscribe from the CCP4BB list, click the following link: > https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB&A=1 > ######################################################################## To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB&A=1
