Satvik - if you attach your data processing logs I can tell you what to look out for - too abstract to do without a concrete example to discuss. Eleanor
On 20 September 2017 at 14:00, <herman.schreu...@sanofi.com> wrote: > Dear Satvik, > > > > An R/Rfree of 0.29/0.35 after one round of automatic model building > indicates that your solution is correct. You can proceed with refinement > and rebuilding. You can take either the monomer-based or dimer-based > solution, it does not matter. Personally I would take the monomer-based > solution since it allowed for some small difference in orientation of the > monomers within the dimer. > > > > Best, > > Herman > > > > *Von:* CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] *Im Auftrag von > *Satvik Kumar > *Gesendet:* Mittwoch, 20. September 2017 14:40 > *An:* CCP4BB@JISCMAIL.AC.UK > *Betreff:* Re: [ccp4bb] doubt regarding MR search model > > > > Hello, > > Thanks all for your valuable suggestions. Two things that stuck me were "one > gets to know if the space group is correct only after solving the > structure" and "to try all possible space groups until one lands up with > the correct solution". > > I ran Phaser with "all alternative space groups". The solution is still in > the same space group P212121. This holds true with both the monomer as well > as dimer search models. I hope I have checked all orthorhombic space groups > by doing this as I am unsure what "all alternative space groups" means. > > I also ran Zanuda which tells me that the space group P212121 is correct > in both cases. > > Pointless output indicates that "Systematic absence probability: 0.818". > Can I take this to be convincing or do I need to run other tests? > > > > I dont know how to check for non-crystallographic symmetry? Please tell me > how I can do so. > > > > The model is 60 percent identical to my protein. Also, one round of > automated model building has brought down the R values. The Rwork,Rfree > stand at 0.29,0.35 (monomer search model) and 0.30, 0.35 (dimer search > model). There is good scope for building residues in both cases. But which > one do I go ahead with is my question. > > > > Thanks, > > Satvik > > > > > > On Tue, Sep 19, 2017 at 11:40 PM, Eleanor Dodson < > eleanor.dod...@york.ac.uk> wrote: > > Well - you haven't said what the sequence identity between model and your > protein is, nor if you have a non-crystallographic translation. > > > > With low homology that R factor drop is acceptable and rebuilding can fix > it, However if there is high homology you might expect better. > > > > But this sort of conjecture is pretty pointless - check in all > orthorhombic space groups as Mark suggests. > > > > Eleanor > > > > On 19 September 2017 at 15:16, Mark J van Raaij <mjvanra...@cnb.csic.es> > wrote: > > With Rs of 43/48% I don't think you can be sure that your spacegroup is > right. > > You should always try all the spacegroup possibilities until you get a > solution you are sure is right, i.e. that refines to Rs of around 35% or > preferably even lower. > > More so in the case of screw axes, so try P222, P2122, P2212, P2221, > P21212, P21221, P22121 and P212121. Phaser can do this automatically for > you by clicking the right box. > > If necessary, then try lower symmetry like P21 and perhaps P1. > > Programs like Xanuda can help. > > > > > > Mark J van Raaij > Dpto de Estructura de Macromoleculas > Centro Nacional de Biotecnologia - CSIC > calle Darwin 3 > <https://urldefense.proofpoint.com/v2/url?u=https-3A__maps.google.com_-3Fq-3Dcalle-2BDarwin-2B3-26entry-3Dgmail-26source-3Dg&d=DwMFaQ&c=Dbf9zoswcQ-CRvvI7VX5j3HvibIuT3ZiarcKl5qtMPo&r=HK-CY_tL8CLLA93vdywyu3qI70R4H8oHzZyRHMQu1AQ&m=WuFY958mWwJOc-_oZdihE6B4h0tlRsfBpLyZ0agnz9E&s=wsbi7Hs3zKGXbwDzjt4Up9v2OvmNaexqb0vzGpBG7j8&e=> > E-28049 Madrid, Spain > tel. (+34) 91 585 4616 <+34%20915%2085%2046%2016> > http://wwwuser.cnb.csic.es/~mjvanraaij > <https://urldefense.proofpoint.com/v2/url?u=http-3A__wwwuser.cnb.csic.es_-7Emjvanraaij&d=DwMFaQ&c=Dbf9zoswcQ-CRvvI7VX5j3HvibIuT3ZiarcKl5qtMPo&r=HK-CY_tL8CLLA93vdywyu3qI70R4H8oHzZyRHMQu1AQ&m=WuFY958mWwJOc-_oZdihE6B4h0tlRsfBpLyZ0agnz9E&s=xcDWCasKH3VKSzXrKFpceN-EhF-q4nC9GwjZ338cYrA&e=> > > Editor of Acta Crystallographica F, Structural Biology Communications > > http://journals.iucr.org/f/ > <https://urldefense.proofpoint.com/v2/url?u=http-3A__journals.iucr.org_f_&d=DwMFaQ&c=Dbf9zoswcQ-CRvvI7VX5j3HvibIuT3ZiarcKl5qtMPo&r=HK-CY_tL8CLLA93vdywyu3qI70R4H8oHzZyRHMQu1AQ&m=WuFY958mWwJOc-_oZdihE6B4h0tlRsfBpLyZ0agnz9E&s=AGltbWDVK377w5vwK7tqER8JFzGUMWMblni0CyuiDUs&e=> > > > > On 19 Sep 2017, at 16:01, Satvik Kumar <kumarsatvi...@gmail.com> wrote: > > > > Hello, > > Thanks everyone for your explanations. > > I have pasted the pointless output to provide more information. > > Best Solution: space group P 21 21 21 > > Laue group probability: 0.959 > > Systematic absence probability: 0.818 > > Total probability: 0.785 > > Space group confidence: 0.751 > > Laue group confidence 0.951 > > > > Unit cell: 82.10 100.51 157.11 90.00 90.00 90.00 > > Also based on L-test, pointless says data does not suggest twinning. > > Yes, the R values go down when I refine in both cases. After 20 rounds of > restrained refinement using the coordinates generated by monomer as search > model, the Rwork and Rfree are 0.43 and 0.48 > > respectively. Refinement using the coordinates generated by using dimer as > search model also results in similar R values. I have attached the plots to > show that the R values indeed reduce in both cases. > > Is my space group correct? Do I need to reexamine the space group even > though the probability is high? > > If my space group is indeed correct, how do I decide whether to go ahead > with the results generated by the monomer search model or the dimer? > > Please share your thoughts. > > Thanks, > > Satvik > > > > On Mon, Sep 18, 2017 at 7:36 PM, Eleanor Dodson <eleanor.dod...@york.ac.uk> > wrote: > > You need to provide a bit more information. > > First of all about the data processing.. > > Is the space group correct? > > ways of being misled are: > > Non-crystallographic translations with a shift of ~0.5 along an axis - say > a. This will generate absences in the odd h 0 0 reflections and can make > the space group appear to be P 21 21 21 whilst it is really P 2 21 21.. > > Perfect twinning can have the same effect. In an orthorhombix space group > this can usually only occur if two axes have approximately the same length, > but the data processing stats can indicate if that is the case. > > Then - re PHASER. The packing rejection criteria may be set too severely - > that seems the case for your solution. > > Best check on any MR solution is: does it refine - give it 20 cycles of > mindless refinement and see if the R and FreeR go down. > > Then look at the maps and see if there are obvious corrections to be made.. > > Eleanor > > > > On 18 September 2017 at 14:59, Satvik Kumar <kumarsatvi...@gmail.com> > wrote: > > Dear Crystallographers, > > I am trying to solve a structure in the space group P212121. Based on > Matthews coefficient, there are 4 molecules in the asymmetric unit. > > Based on my limited reading about using of Phaser, I understand that a > single chain should be used as search model even though many copies are > present in asymmetric unit. Am I correct? > > So when I use a single chain as search model and ask Phaser to search for > 4 molecules, Phaser identifies a single solution with a warning "The top > solution from a TF rescoring did not pack" and a warning "Search request > requires more scattering than defined in composition. Composition increased > to accommodate search components". But the final values reported "PAK=2 > LLG=1065 TFZ==22.6" indicate that phaser has solved the problem. > > Can anyone please explain the meaning of the warning. > > When I inspect the arrangement of the chains (attachment), I observe > minimal contact between the chains and a large cavity in the center. Can a > crystal form this way? > > I have also tried using the dimer as search model and asking phaser to > search for 2 molecules. Even in this case, Phaser finds a single solution > but the warning and the advisory still appear as before. The numbers > reported reduce a bit to "PAK=1 LLG=722 TFZ==29.2". > > Please help me in understanding these results. > > Thanks, > > Satvik > > > > > > > > <Monomer_searchmodel.pdf><Dimer_searchmodel.pdf> > > > > > > >
