Umm - this is tricky. First of all you need to reindex the C2221 data into the P21 cell - do you know the operator? then expand that data set to spacegroup P21. There is a cad option to do this.. Then add that FreeR to the re-processed P21 data. Eleanor
On 24 March 2013 14:37, Appu kumar <appu.kum...@gmail.com> wrote: > Sorry for the misconception. Yes i am expanding the space group from > merged mtz file. Actually i have enough number of images collected. when i > indexed, integrate, and scale the data in either C2221 or P 21, it fetches > the overall 98% completeness. But when i am trying to reindex the data > from C2221 to P21 keeping the Rfree flag of C2221, the completeness of data > reduced drastically to 40%. This is what i am not getting. I am a beginner > so i have to read a lot which i am doing also, but i had few practical > confusion which i shared and off course i am getting good response. Thank > you all for your kind response and educating me on the problem i faced. > Thank you all for your valuable response. > > On 24 March 2013 15:15, vellieux <frederic.velli...@ibs.fr> wrote: > >> Hello, >> >> Here we deal with symmetry and the unique part of reciprocal space (the >> reciprocal space "asymmetric unit" so to speak). >> >> C222(1) has eight asymmetric units (international tables, space group 20); >> >> P2(1) only has two. Assuming that Friedel's law does apply, then the >> minimum rotation range to collect a non-redundant data set (one observation >> per reflection) is 90 degrees, provided that the crystal is "correctly" and >> perfectly aligned. Normally with our current data collection methods where >> the crystal is randomly oriented, we would collect more than 90 degrees >> (180 degrees, or 360 degrees with the Pilatus detectors on an intense SR >> beamline where you cannot really check during data collection how well the >> crystal fares during exposure to the X-rays - "shoot first, think later". >> >> The "reciprocal space" asymmetric unit in C222(1) is smaller. >> >> I assume that what you are doing is to take the reduced data set file (an >> MTZ file probably) and reduce the symmetry from C222(1) to P2(1). You will >> not cover the monoclinic reciprocal space asymmetric unit by doing so. >> >> The way to do it is to take the file from processing, before >> (crystallographic symmetry) merging of the equivalents, and perform the >> scaling and merging in the P2(1) space group. Or reprocess the data frames >> in P2(1) if you have lost the unmerged data file. >> >> Now of course this will still give you a poor completeness if you have >> used a strategy to optimize data collection in the orthorhombic space group >> (you won't have collected enough data then for good completeness in the >> monoclinic space group). >> >> I hope this is clear ! >> >> HTH, >> >> Fred. >> >> >> On 24/03/13 11:20, Appu kumar wrote: >> >> I run the phenix.xtriage to evaluate the twining but it suggest no >> twining. When i reindex from C2221 to P21, the completeness of data reduced >> from 95 % to 35% whereas the map is very good and Rwork and Rfree are 26/31 >> for 2.2 resolution. I do not understand why the completeness of data >> reduced so much on reindexing. please Can anyone explain this phenomenon. >> Thank you >> >> On 24 March 2013 13:30, Matthias Zebisch < >> matthias.zebi...@bbz.uni-leipzig.de> wrote: >> >>> the p21 c2221 ambivalence can mean severe twinning (i had a similar >>> case just now - try several crystals from the same condition) ! >>> What do the twinning statistics suggest? >>> >>> cheers, Matthias >>> >>> ----------------------------------------- >>> Dr. Matthias Zebisch >>> Division of Structural Biology, >>> Wellcome Trust Centre for Human Genetics, >>> University of Oxford, >>> Roosevelt Drive, >>> Oxford OX3 7BN, UK >>> >>> Phone (+44) 1865 287549; >>> Fax (+44) 1865 287547 >>> Email matth...@strubi.ox.ac.uk >>> Website http://www.strubi.ox.ac.uk >>> ----------------------------------------- >>> >>> On 3/24/2013 7:46 AM, Appu kumar wrote: >>> >>> Thank you for the quick reply. After molecular replacement , i have done >>> only few cycle of refinement in refmac. I have not done any solvent >>> modification or NCS averaging. I have initially indexed the data in C2221 >>> but Rfree was not decreasing so i reindexed the data in data in P121 space >>> group keeping the Rfree flag of C2221. While analysing the symmetry mates , >>> i found large space but no density. structure of Ligand binding domain is >>> almost identical with 90% identity in sequence. I am stuck with this >>> problem and don't know how to process further. >>> Please give me your valuable suggestion. I will appreciate your effort. >>> Thank you >>> Appu >>> >>> On 24 March 2013 02:38, Raji Edayathumangalam <r...@brandeis.edu> wrote: >>> >>>> Dear Appu, >>>> >>>> I am not sure that I have a complete sense of the issue at hand since >>>> some of the information needed to think your issue through is missing in >>>> your email. For example, to what high resolution cut-off were the data >>>> measured? What resolution limits were used for the MR search? How do the >>>> unit cell dimensions and space group in the two cases compare? >>>> >>>> I am guessing the ligand binding domain in your protein has the >>>> identical sequence to that of the published ligand binding domain that you >>>> use as a template in your MR search. In any case, here are a couple of my >>>> thoughts: >>>> >>>> (1) It might be worth setting up different runs of MR with different >>>> numbers for expected copies (not just two copies but also one copy and >>>> three copies just in case you have one of the extreme cases of solvent >>>> content)? >>>> >>>> (2) If the MR solution is correct and there is physical room for a >>>> DNA binding domain in your lattice (check by displaying symmetry mates), >>>> perhaps the DNA binding domain is disordered. In that case (and if all >>>> attempts with current data fail), you may have to crystallize the protein >>>> in presence of DNA. >>>> >>>> >>>> Good luck! >>>> Raji >>>> >>>> >>>> >>>> >>>> On Sat, Mar 23, 2013 at 2:26 PM, Appu kumar <appu.kum...@gmail.com>wrote: >>>> >>>>> Dear members, >>>>> >>>>> I am doing a molecular replacement of a >>>>> transcription factor whose ligand binding structure(24000 Da) is available >>>>> in PDB but not for the DNA binding(13000 Da). When i am searching for the >>>>> two copies from ligand binding domain as a template model, i am getting >>>>> very good solution but i am not getting any density for the DNA binding >>>>> domain to build up in density. The space gorup is P 1 21 1 (4) and unit >>>>> cell parameters are Unit Cell: 57.43 69.36 105.99 90.00 90.00 >>>>> 90.00. Please guide me how to get the complete model structure. Table >>>>> below >>>>> show the matthews statistics >>>>> >>>>> For estimated molecular weight 37000. >>>>> Nmol/asym Matthews Coeff %solvent P(2.20) P(tot) >>>>> _____________________________________________________________ >>>>> 1 5.71 78.46 0.00 0.01 >>>>> 2 2.85 56.91 0.62 0.70 >>>>> 3 1.90 35.37 0.37 0.29 >>>>> 4 1.43 13.82 0.00 0.00 >>>>> _____________________________________________________________ >>>>> >>>>> >>>>> The phaser molecular replacement gives the following table. >>>>> istogram of relative frequencies of VM values >>>>> ---------------------------------------------- >>>>> Frequency of most common VM value normalized to 1 >>>>> VM values plotted in increments of 1/VM (0.02) >>>>> >>>>> <--- relative frequency ---> >>>>> 0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 >>>>> | | | | | | | | | | | >>>>> 10.00 - >>>>> 8.33 - >>>>> 7.14 - >>>>> 6.25 - >>>>> 5.56 - >>>>> 5.00 - >>>>> 4.55 - >>>>> 4.17 - >>>>> 3.85 -- >>>>> 3.57 --- >>>>> 3.33 ------ >>>>> 3.12 ---------- >>>>> 2.94 **************** (COMPOSITION*1) >>>>> 2.78 ----------------------- >>>>> 2.63 -------------------------------- >>>>> 2.50 ----------------------------------------- >>>>> 2.38 ------------------------------------------------ >>>>> 2.27 -------------------------------------------------- >>>>> 2.17 ----------------------------------------------- >>>>> 2.08 -------------------------------------- >>>>> 2.00 -------------------------- >>>>> 1.92 --------------- >>>>> 1.85 ------- >>>>> 1.79 --- >>>>> 1.72 - >>>>> 1.67 - >>>>> 1.61 - >>>>> 1.56 - >>>>> 1.52 - >>>>> 1.47 * (COMPOSITION*2) >>>>> 1.43 - >>>>> 1.39 - >>>>> 1.35 - >>>>> 1.32 - >>>>> 1.28 - >>>>> 1.25 - >>>>> >>>>> $TABLE : Cell Content Analysis: >>>>> $SCATTER >>>>> :N*Composition vs Probability:0|3x0|1:1,2: >>>>> $$ >>>>> N*Composition Probability >>>>> $$ loggraph $$ >>>>> 1 0.306066 >>>>> 2 0.00141804 >>>>> $$ >>>>> >>>>> Most probable VM for resolution = 2.27817 >>>>> Most probable MW of protein in asu for resolution = 92664.2 >>>>> >>>>> Thank a lot in advance >>>>> >>>>> >>>>> >>>>> >>>> >>>> -- >>>> Raji Edayathumangalam >>>> Instructor in Neurology, Harvard Medical School >>>> Research Associate, Brigham and Women's Hospital >>>> Visiting Research Scholar, Brandeis University >>>> >>>> >>> >>> >> >> >> -- >> Fred. Vellieux (B.Sc., Ph.D., hdr) >> ouvrier de la recherche >> IBS / ELMA >> 41 rue Jules Horowitz >> F-38027 Grenoble Cedex 01 >> Tel: +33 438789605 >> Fax: +33 438785494 >> >> >
