Hi Shen, I agree with Eleanor that the split spots will cause worse statistics, but should not be a reason for molrep to fail. What I would do: Molecular replacement with a resolution cut of 3 or 3.5 Å. Process and run molecular replacement in P1. You may have some tricky pseudo-symmetry. With 8 molecules, molrep may take somewhat longer, but I think it is worth a try.
Best, Herman Von: CCP4 bulletin board <CCP4BB@JISCMAIL.AC.UK> Im Auftrag von Xu, Shenyuan Gesendet: Montag, 11. November 2019 16:51 An: CCP4BB@JISCMAIL.AC.UK Betreff: [EXTERNAL] Re: [ccp4bb] molecular replacement_protein-glycan complex EXTERNAL : Real sender is owner-ccp...@jiscmail.ac.uk<mailto:owner-ccp...@jiscmail.ac.uk> Hello Eleanor, I used the solved one as the input model. The space group of the solved crystals are: P1 21 1, and P 21. I will turn the anomalous processing off. Thanks, Shen On Mon, Nov 11, 2019 at 10:40 AM Eleanor Dodson <eleanor.dod...@york.ac.uk<mailto:eleanor.dod...@york.ac.uk>> wrote: Why dont you use one of your solved structures as a model? What space group are the solved ones in? The split spots are not good, but they are well separated and integration programs are not bad at such spacings? And your R merges etc look OK. Just a Q - why turn off the anomalous processing. You dont need to use it, but at some point it may help you find Ss or Ca or whatever.. Eleanor On Mon, 11 Nov 2019 at 15:24, Xu, Shenyuan <x...@miamioh.edu<mailto:x...@miamioh.edu>> wrote: Dear All, Thanks for all of your nice suggestions! (Sorry I did not respond to individual email to say Thanks.) I have used phaser to go through all possible space groups in the same point group(choose "all possible in same pointgroup" in the space group menu). I also try P43212 alone. The best result I get from phaser is (Top LLG: 26.490, Top TFZ: 6.7, Spacegroup: P41 2 2), and refinement ends in the R-work: ~0.54, R-free:~0.57. I go back to see the diffraction data, and it seems that spots in the high resolution bin are severely split (pictures are attached). I feel like the fail of replacement is caused by this. Is this caused by twinning? I am looking forward suggestions! Thanks, Shenyuan Xu On Wed, Nov 6, 2019 at 2:58 PM Kay Diederichs <kay.diederi...@uni-konstanz.de<mailto:kay.diederi...@uni-konstanz.de>> wrote: Dear Shenyuan, it seems you assumed P41 21 2 is correct but it isn't - did you try P43 21 2 in molecular replacement? The pointless log file mentions this alternative a few lines above the ones you've quoted. If that does not work, try the other P4x 2y 2 space groups. All of this can be done in a single phaser run (choose "all possible in same pointgroup" in the space group menu). Why did you cut the data at 2.29A? They are very strong in the high resolution shell! HTH, Kay On Wed, 6 Nov 2019 14:29:13 -0500, Xu, Shenyuan <x...@miamioh.edu<mailto:x...@miamioh.edu>> wrote: >Dear all, > >I am working on a protein-glycan complex trying to use molecular >replacement.This protein is a VP8* domain from rotavirus, adopting a >classical galectin-like fold. I already used MR to solve some other strains >(apo-form and complex form). However, When I want to use the same model to >solve this particular strain (sequence identity > 95%), MR seems to be >failed with R=0.54, Free R = 0.57, and the model does not agree with the >electron density map once viewed on coot. One difference of this >protein-glycan complex between others is that the glycan is longer, which >has six carbohydrate unit. > >I used imosflm trying to reindex and use pointless to check the space >group, it usually ends up with the current choice: >25_pointless.log > >WARNING! one or more zones have data systematically missing from the input >file >thus we cannot determine if reflections are truly systematically absent > >Best Solution: space group P 41 21 2 >Reindex operator: [h,k,l] >Laue group probability: 1.000 >Systematic absence probability: 0.995 >Total probability: 0.995 >Space group confidence: 0.993 >Laue group confidence 1.000 > >WARNING: You will have to resolve the enantiomorphic ambiguity later > >Unit cell: 106.78 106.78 138.59 90.00 90.00 90.00 > >84.58 to 2.43 - Resolution range used for Laue group search > >84.58 to 2.29 - Resolution range in file, used for systematic absence check > >Number of batches in file: 1 > >The data do not appear to be twinned, from the L-test >I also use Zanuda to validate the space group, it ends with the same choice: > >Step 3. > Refinement of the best model. > Candidate symmetry elements are added one by one. > > current time: Nov 06 16:17 GMT > expected end of job: Nov 06 17:06 GMT > > ^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^ > | >> 3 | C 1 2 1 | 0.3736 | 0.5334 | 0.5044 | 0.5586 | > --------------------------------------------------------------------- > | 1 | P 1 | 0.4058 | 0.5318 | 0.5020 | 0.5624 | > | 5 | P 1 21 1 | 0.4928 | -- | 0.4973 | 0.5701 | > | 9 | P 21 21 21 | 0.5470 | -- | 0.5011 | 0.5736 | > | 10 | P 41 21 2 | 0.5749 | -- | 0.5018 | 0.5778 | > --------------------------------------------------------------------- > | << 10 | P 41 21 2 | 0.5749 | -- | 0.5018 | 0.5778 | > --------------------------------------------------------------------- > > R-factor in the original subgroup is (almost) the best. > The original spacegroup assignment seems to be correct. > > The statistics of the dataset seems good: >Summary data for Project: Test Crystal: A Dataset: 1 > > Overall InnerShell OuterShell >Low resolution limit 84.58 84.58 2.41 >High resolution limit 2.29 7.23 2.29 > >Rmerge (within I+/I-) 0.086 0.036 0.564 >Rmerge (all I+ and I-) 0.088 0.036 0.581 >Rmeas (within I+/I-) 0.093 0.039 0.610 >Rmeas (all I+ & I-) 0.092 0.038 0.604 >Rpim (within I+/I-) 0.036 0.015 0.230 >Rpim (all I+ & I-) 0.026 0.011 0.165 >Rmerge in top intensity bin 0.045 - - >Total number of observations 466972 14511 70743 >Total number unique 37007 1338 5297 >Mean((I)/sd(I)) 20.4 44.5 5.0 >Mn(I) half-set correlation CC(1/2) 0.999 0.999 0.947 >Completeness 100.0 100.0 100.0 >Multiplicity 12.6 10.8 13.4 >Mean(Chi^2) 0.97 0.85 0.86 > >Anomalous completeness 100.0 100.0 100.0 >Anomalous multiplicity 6.5 6.6 6.7 >DelAnom correlation between half-sets -0.172 -0.180 -0.083 >Mid-Slope of Anom Normal Probability 0.857 - - > >Anomalous flag switched OFF in input, anomalous signal is weak > >Estimates of resolution limits: overall > from half-dataset correlation CC(1/2) > 0.30: limit = 2.29A == >maximum resolution > from Mn(I/sd) > 2.00: limit = 2.29A == >maximum resolution > from Mn(I/sd) > 2.00: limit = 2.29A == >maximum resolution > >Estimates of resolution limits in reciprocal lattice directions: > Along h k plane > from half-dataset correlation CC(1/2) > 0.30: limit = 2.29A == >maximum resolution > from Mn(I/sd) > 2.00: limit = 2.29A == >maximum resolution > Along l axis > from half-dataset correlation CC(1/2) > 0.30: limit = 2.29A == >maximum resolution > from Mn(I/sd) > 2.00: limit = 2.29A == >maximum resolution > >Anisotropic deltaB (i.e. range of principal components), A^2: 6.03 > >Average unit cell: 106.78 106.78 138.59 90.00 90.00 90.00 >Space group: P 41 21 2 >Average mosaicity: 0.27 > >Minimum and maximum SD correction factors: Fulls 1.00 1.35 Partials >0.00 0.00 > >Any suggestions are welcomed! > >Thanks, > >Shenyuan Xu > >######################################################################## > >To unsubscribe from the CCP4BB list, click the following link: >https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB&A=1<https://urldefense.proofpoint.com/v2/url?u=https-3A__www.jiscmail.ac.uk_cgi-2Dbin_webadmin-3FSUBED1-3DCCP4BB-26A-3D1&d=DwMFaQ&c=Dbf9zoswcQ-CRvvI7VX5j3HvibIuT3ZiarcKl5qtMPo&r=HK-CY_tL8CLLA93vdywyu3qI70R4H8oHzZyRHMQu1AQ&m=DivhmpKd5Pwuefz41rsUIvaEj3_QOwMPpIv44-xpi1k&s=tfgHlzCP0-vbR3kmX7qNza4dK5ot2hqFhwDLegNgWq8&e=> > ######################################################################## To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB&A=1<https://urldefense.proofpoint.com/v2/url?u=https-3A__www.jiscmail.ac.uk_cgi-2Dbin_webadmin-3FSUBED1-3DCCP4BB-26A-3D1&d=DwMFaQ&c=Dbf9zoswcQ-CRvvI7VX5j3HvibIuT3ZiarcKl5qtMPo&r=HK-CY_tL8CLLA93vdywyu3qI70R4H8oHzZyRHMQu1AQ&m=DivhmpKd5Pwuefz41rsUIvaEj3_QOwMPpIv44-xpi1k&s=tfgHlzCP0-vbR3kmX7qNza4dK5ot2hqFhwDLegNgWq8&e=> ________________________________ To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB&A=1<https://urldefense.proofpoint.com/v2/url?u=https-3A__www.jiscmail.ac.uk_cgi-2Dbin_webadmin-3FSUBED1-3DCCP4BB-26A-3D1&d=DwMFaQ&c=Dbf9zoswcQ-CRvvI7VX5j3HvibIuT3ZiarcKl5qtMPo&r=HK-CY_tL8CLLA93vdywyu3qI70R4H8oHzZyRHMQu1AQ&m=DivhmpKd5Pwuefz41rsUIvaEj3_QOwMPpIv44-xpi1k&s=tfgHlzCP0-vbR3kmX7qNza4dK5ot2hqFhwDLegNgWq8&e=> ________________________________ To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB&A=1<https://urldefense.proofpoint.com/v2/url?u=https-3A__www.jiscmail.ac.uk_cgi-2Dbin_webadmin-3FSUBED1-3DCCP4BB-26A-3D1&d=DwMFaQ&c=Dbf9zoswcQ-CRvvI7VX5j3HvibIuT3ZiarcKl5qtMPo&r=HK-CY_tL8CLLA93vdywyu3qI70R4H8oHzZyRHMQu1AQ&m=DivhmpKd5Pwuefz41rsUIvaEj3_QOwMPpIv44-xpi1k&s=tfgHlzCP0-vbR3kmX7qNza4dK5ot2hqFhwDLegNgWq8&e=> ######################################################################## To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB&A=1
