HI, Mark: Thank you for your suggestions.
>try all possible spacegroups with PHASER Yes, this is done with all the modeling. >make sure you are using a monomer as search model I also tried with monomer as search model. When run with refmac5, the R-value is above 0.5. I exam the model in coot. Some parts of model match with density. But some parts of model do not matched to the density. >At what resolution are the templates in the PDB? 3.0 - 3.9 A At this stage, I would like to compare the structure at backbone level, not the side chain. May have to adjust model piece by piece to fit into the map. Thanks Uma On Wed, Jul 30, 2014 at 12:43 PM, Mark J van Raaij <mjvanra...@cnb.csic.es> wrote: > try all possible spacegroups with PHASER, make sure you are using a > monomer as search model, not a crystallographic multimer. > In any case 3.9 A is not going to give you much information and you will > be hard-pressed to see the mutation differences - so perhaps forget about > this data and improve the crystals first to get higher-resolution data to > 2.5A or preferably even better. At what resolution are the templates in the > PDB? You should be able to get similar resolution to those. > > > Mark J van Raaij > Lab 20B > Dpto de Estructura de Macromoleculas > Centro Nacional de Biotecnologia - CSIC > c/Darwin 3 > E-28049 Madrid, Spain > tel. (+34) 91 585 4616 > http://www.cnb.csic.es/~mjvanraaij > Associate Editor of Virology Journal > Academic Editor of PLoS One > > > > > > > On 30 Jul 2014, at 18:38, Uma Ratu wrote: > > > Dear All: > > > > I try to solve a structure by using Phaser. > > > > The data set was collected at 3.9 A with 98.1% (95%) completeness. Based > on pointless, the best space group for the data set is P21212. And the data > can be processed with P21, P212121 as well. The I/sigma value for the data > is 13.3 (1.9). > > > > There are some structure models available from PDB with space groups at > P21, P212121 or P21212. I used these structures as templates to get initial > models. I then use Refmac5 to refine the models. But the R-factors/R-free > are around 0.5 /0.5. When I exam the .pdb and .mtz in coot, the models are > not aligned with density. > > > > I then use the templates (PDB entries) against the processed .mtz files > using Refmac5. Again, the R-factors are higher than 0.5. > > > > I notice that my data set has different cell units with templates. When > the space group match to one template, the cell dimensions is differ from > the template. When the cell dimension matches to one template, the space > group is different. Is this the problem for modeling? > > > > Sequence identity between my protein and templates is 99%. Two residues > were mutated in my protein. > > > > I use Phaser for molecular replacement, and Refmac5 for refinement. Data > was processed using xds or mosfilm. > > > > Thank you for advice > > > > Uma > > > > > > > > > > On Wed, Aug 1, 2012 at 2:27 PM, Uma Ratu <rosiso2...@gmail.com> wrote: > > Dear All: > > > > I try to use Phaser to solve the structure by Molecular Replacement. > > > > The data set is collected @180 degree. I process the data using HKL, and > have resonable good score: rejection (0.05), Linear R-factor (0.038), > completeness (98.3), resolution (50-1.5). > > > > I then use Phaser to do MR. The parameter setting are: > > automated search > > components in asymmetric unit; number of residue 1332; number in > asymmetric unit 1 > > perform search search using "ensemble1" number of copies to search for 4 > > > > The protein is in tetramer form. I define this by using the residue > number (1332) which is 4 x monomer. > > > > After run, Phaser only gave 9 partial solutions, and no solution with > all components. The resulted PDB contains only dimer form of the protein, > not the tetramer. And the first TFZ score is around 2.5, which is too low > for MR. > > > > I have the report file of data processing and the summary of Phaser > attached. > > > > Could you please advice which part is wrong, why can I get the tetramer > form of the protein? > > > > Thank you > > > > Uma > > > >
