Dear Professor, Thank you for your valuable suggestions.
I have attempted to investigate the NCS operator, but it seems to be a monomer. Nevertheless, I will reconsider your suggestion regarding this matter. I employed the concept of using high-resolution data (2 Angstroms) without cutoff for model building and refinement cycling, along with phases from MR (2.7 Angstroms). I will revisit this process. I will follow your advice for the peak search. If I cannot find any leads, I hope to seek your guidance. Thank you again for your support. *Warm Regards-* *Devbrat Kumar* On Wed, Nov 15, 2023 at 4:38 PM Eleanor Dodson < [email protected]> wrote: > You seem pretty near to having solved your structure! > Ignoring the data problems.. > > Extra steps I might have used. > 1) Self rotation function. (in CCP4I2 the task is under data analysis ..) > Does it suggest a NCS operator? > If so is this a two fold? which might mean you have a dimer.. > > > 2) Now you have a reasonable R factor try extending the resolution. the > refinement program will weight down the high resolution less reliable data > but the extra information might marginally improve the maps. > > 3) Use the calculated phases to run an anomalous map - REFMAC will produce > the appropriate coefficients and you can display the map in COOT. > Do a peaksearch and you should of course see the sites SHELX found.. > But sometimes you get peaks over S atoms and that makes you pretty > confident that the sequence there is correct. > > > > On Wed, 15 Nov 2023 at 10:42, Kay Diederichs < > [email protected]> wrote: > >> Hello Devbrat, >> >> your project is difficult and there is no magic bullet to solve its >> problems. Your approach is good because it always pays off to process >> the data carefully. >> In this respect, let me make a few comments. >> 1) you don't say why you call the diffraction patterns "multi-lattice". >> What exactly do you mean by that? Non-merohedral >> twinning? How many lattices superimposed and visible on all frames? Can >> they be separately indexed by XDS >> (see https://wiki.uni-konstanz.de/xds/index.php/Indexing)? >> 2) "XDS processing" _is_ integrating; what AIMLESS does is called scaling. >> 3) what do you mean by "monomer NCS"? NCS implies two or more copies of >> the same molecule in the asymmetric unit. >> These copies often form dimers, trimers, tetramers, ... by making >> more-or-less strong and specific interactions. >> 4) you've advanced amazingly far and it sounds to me that with a >> combination of your refined MR model with the SAD data you >> should be able to improve your solution. Look up the MR-SAD pipeline (for >> SAD after MR and for model rebuilding using anomalous >> data) of Crank2. >> >> If you want me to take a look at your raw data, upload the best datasets >> (native and SeMet) to a cloud service and send me the link. >> >> Good luck, >> Kay >> >> On Tue, 14 Nov 2023 21:25:30 +0530, Devbrat Kumar <[email protected]> >> wrote: >> >> >Hello everyone, >> > >> >The issue with the crystal is its multi-lattice nature; even the >> truncated >> >protein, which has been crystallized, exhibits multi-lattice >> >characteristics (detectable only after XRD). >> > >> >I have multiple native and selenium datasets with similar unit cell >> >parameters. (One axis is excessively long.) The XRD images were processed >> >using XDS in the P2 spacegroup, with unit cell parameters as follows: a = >> >27.75 Å, b = 293.9 Å, c = 34.6 Å, and β = 113°. The XDS-processed data >> were >> >integrated with the data reduction tool AIMLESS in the CCP4i2 suite. In >> >CRANK2, the Estimation of Matthews coefficient (Program used: GCX) >> >suggested the presence of monomer NCS with a solvent content of 63.6%. >> The >> >FA estimation and substructure detection were performed by SHELXC, which >> >detected a very weak signal below 3.4 Å. Substructure determination was >> >carried out using SHELXD, yielding a maximum figure of merit of 27.8 >> after >> >640 trials and suggesting 11 atoms in the substructure with an occupancy >> of >> >at least 25%. Phasing and substructure refinement were conducted using >> the >> >BP3 program, resulting in an FOM of 0.2. During hand determination, the >> >programs suggested combined DM (density modification) FOM and phasing CLD >> >score for hand one as 6.0 and for hand two as 4.783375. The tool didn't >> >choose the hand because the value is less than the threshold. Density >> >modification with Fourier recycling suggests that the final FOM for hand >> >one and hand two is 0.428 and 0.482, respectively, while REFMAC5 gives >> the >> >R factor and Rfree factor as 0.4262 and 0.4912. >> > >> >One of the MR templates (model with balbes) works(For MR, Identity with >> the >> >PDB template is 21%), but R & Rfree are stuck at 33 & 37 for the 2.7 >> >Angstrom cut-off (the total resolution in the dataset is 2 Angstrom). >> The R >> >& Rfree is not decreasing for the dataset. I have played with detector >> >distances for spot resolution, but at one pHi the spots have merged as a >> >single spot, while at 90 degrees will give us the streak of spots. >> > >> >Looking forward to hearing from you regarding dataset processing ideas >> for >> >multi-lattice crystals(Native & Se dataset) and structure solution >> strategy. >> > >> >Thank you. >> >Regards >> >Devbrat >> > >> >######################################################################## >> > >> >To unsubscribe from the CCP4BB list, click the following link: >> >https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 >> > >> >This message was issued to members of www.jiscmail.ac.uk/CCP4BB, a >> mailing list hosted by www.jiscmail.ac.uk, terms & conditions are >> available at https://www.jiscmail.ac.uk/policyandsecurity/ >> > >> >> ######################################################################## >> >> To unsubscribe from the CCP4BB list, click the following link: >> https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 >> >> This message was issued to members of www.jiscmail.ac.uk/CCP4BB, a >> mailing list hosted by www.jiscmail.ac.uk, terms & conditions are >> available at https://www.jiscmail.ac.uk/policyandsecurity/ >> > > ------------------------------ > > To unsubscribe from the CCP4BB list, click the following link: > https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 > ######################################################################## To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 This message was issued to members of www.jiscmail.ac.uk/CCP4BB, a mailing list hosted by www.jiscmail.ac.uk, terms & conditions are available at https://www.jiscmail.ac.uk/policyandsecurity/
