Re: [ccp4bb] software or server to validate ligand density
Dear Ansuman, The PDB_REDO server has extensive ligand validation, but it won't tell you explicitly whether the ligand is there or not. You have to figure that out from the validation scores. High real-space R values or low real-space correlation coefficients are a hint that your ligand isn't there. High energy of formation indicates ligand strain which may also be the result of building a ligand that is either non-existent or just misidentified. Poor ligand-binding site interactions are also informative, but are sensitive to the chemical properties of the ligand. Cheers, Robbie Sent from my Windows Phone Van: ansuman biswas Verzonden: 20-9-2014 0:08 Aan: CCP4BB@JISCMAIL.AC.UK Onderwerp: [ccp4bb] software or server to validate ligand density Dear All, I have collected a diffraction dataset from a crystal soaked in a solution containing the ligand of interest. After refining a few cycles, I can see some density in the active site pocket, but not so clear to model the ligand unambiguously. Is any tool available to validate whether the ligand is actually there or not ? The Twilight server appears to be for PDB files that have already been deposited. thanks and regards, Ansuman Biswas, dept. of Physics, Indian Institute of science
Re: [ccp4bb] Phaser MR problem
Just to add to what Herman said: The statistics are good for placing the domain represented by ensemble 1 (TFZ=14.3) and the first copy of the domain represented by ensemble 2 (TFZ=11.9), but not for the two possible solutions for placing the second copy of ensemble 2 (TFZs of 4.5 and 4.7). So it’s possible that only the placement of the first two components is correct. If you turn on symmetry in coot, do those two domains come together to form a sensible whole protein? If so, you could try using that composite model to look for more copies of the whole protein (keeping in mind Herman’s point about the possible numbers of copies). That said, it’s going to be a challenge to finish up the structure rebuilding and refinement when you have limited resolution and relatively low sequence identity starting models. With 3-fold or greater NCS, averaging would help a great deal (2-fold helps somewhat but is less powerful); otherwise, you’re likely to need some additional phase information. Best wishes, Randy Read - Randy J. Read Department of Haematology, University of Cambridge Cambridge Institute for Medical ResearchTel: +44 1223 336500 Wellcome Trust/MRC Building Fax: +44 1223 336827 Hills RoadE-mail: rj...@cam.ac.uk Cambridge CB2 0XY, U.K. www-structmed.cimr.cam.ac.uk On 19 Sep 2014, at 10:33, Veerendra KUMAR (IMCB) veerend...@imcb.a-star.edu.sg wrote: Dear CCP4 members, Recently I have collected native data at 3.3 A resolution. The structure of the protein should have two domains. The structure of c terminal domain from homologous (30 seq similarity) is known. I took n terminal domain from another homologous protein. I ran the phaser using these two ensembles and got the following solutions. # [No title given] SOLU SET RFZ=3.9 TFZ=14.3 PAK=4 LLG=144 TFZ==10.5 RFZ=2.4 TFZ=11.9 PAK=9 LLG=259 TFZ==7.0 RFZ=4.0 TFZ=4.5 PAK=24 LLG=207 TFZ==7.4 SOLU SPAC I 41 3 2 SOLU 6DIM ENSE ensemble1 EULER 43.364 14.638 271.356 FRAC 0.81509 0.22245 0.92422 BFAC -7.23070 SOLU 6DIM ENSE ensemble2 EULER 173.495 55.758 234.557 FRAC 1.25797 -0.71934 0.10135 BFAC 1.81823 SOLU 6DIM ENSE ensemble2 EULER 351.235 68.606 254.802 FRAC 0.64931 0.81795 0.27592 BFAC 5.82534 SOLU ENSE ensemble1 VRMS 1.157 SOLU ENSE ensemble2 VRMS 1.540 SOLU SET RFZ=3.9 TFZ=14.3 PAK=4 LLG=144 TFZ==10.5 RFZ=2.4 TFZ=11.9 PAK=9 LLG=259 TFZ==7.0 RFZ=2.7 TFZ=4.7 PAK=19 LLG=202 SOLU SPAC I 41 3 2 SOLU 6DIM ENSE ensemble1 EULER 43.364 14.638 271.356 FRAC 0.81509 0.22245 0.92422 BFAC -7.23070 SOLU 6DIM ENSE ensemble2 EULER 173.495 55.758 234.557 FRAC 1.25797 -0.71934 0.10135 BFAC 1.81823 SOLU 6DIM ENSE ensemble2 EULER 41.452 118.850 315.630 FRAC 0.61025 0.05816 1.60902 BFAC 5.47988 SOLU ENSE ensemble1 VRMS 1.157 SOLU ENSE ensemble2 VRMS 1.540 The TFZ score 14.3 suggests a solution. Then I did refinement using Ploy Ala phaser model. The R values are 0.590/0.62. I use the map to build the model in Buccaneer but it did not build anything. Is the pahser solution correct? Why are the R values so high despite the good TFZ score? Any suggestions are greatly appreciated. Thank you Veerendra Note: This message may contain confidential information. If this Email/Fax has been sent to you by mistake, please notify the sender and delete it immediately. Thank you.
Re: [ccp4bb] Phaser MR problem
Or perhaps there is only 1 copy and more solvent than you expect? This is not that uncommon. On 20 Sep 2014 12:09, Randy Read rj...@cam.ac.uk wrote: Just to add to what Herman said: The statistics are good for placing the domain represented by ensemble 1 (TFZ=14.3) and the first copy of the domain represented by ensemble 2 (TFZ=11.9), but not for the two possible solutions for placing the second copy of ensemble 2 (TFZs of 4.5 and 4.7). So it’s possible that only the placement of the first two components is correct. If you turn on symmetry in coot, do those two domains come together to form a sensible whole protein? If so, you could try using that composite model to look for more copies of the whole protein (keeping in mind Herman’s point about the possible numbers of copies). That said, it’s going to be a challenge to finish up the structure rebuilding and refinement when you have limited resolution and relatively low sequence identity starting models. With 3-fold or greater NCS, averaging would help a great deal (2-fold helps somewhat but is less powerful); otherwise, you’re likely to need some additional phase information. Best wishes, Randy Read - Randy J. Read Department of Haematology, University of Cambridge Cambridge Institute for Medical Research Tel: +44 1223 336500 Wellcome Trust/MRC Building Fax: +44 1223 336827 Hills Road E-mail: rj...@cam.ac.uk Cambridge CB2 0XY, U.K. www-structmed.cimr.cam.ac.uk On 19 Sep 2014, at 10:33, Veerendra KUMAR (IMCB) veerend...@imcb.a-star.edu.sg wrote: Dear CCP4 members, Recently I have collected native data at 3.3 A resolution. The structure of the protein should have two domains. The structure of c terminal domain from homologous (30 seq similarity) is known. I took n terminal domain from another homologous protein. I ran the phaser using these two ensembles and got the following solutions. # [No title given] SOLU SET RFZ=3.9 TFZ=14.3 PAK=4 LLG=144 TFZ==10.5 RFZ=2.4 TFZ=11.9 PAK=9 LLG=259 TFZ==7.0 RFZ=4.0 TFZ=4.5 PAK=24 LLG=207 TFZ==7.4 SOLU SPAC I 41 3 2 SOLU 6DIM ENSE ensemble1 EULER 43.364 14.638 271.356 FRAC 0.81509 0.22245 0.92422 BFAC -7.23070 SOLU 6DIM ENSE ensemble2 EULER 173.495 55.758 234.557 FRAC 1.25797 -0.71934 0.10135 BFAC 1.81823 SOLU 6DIM ENSE ensemble2 EULER 351.235 68.606 254.802 FRAC 0.64931 0.81795 0.27592 BFAC 5.82534 SOLU ENSE ensemble1 VRMS 1.157 SOLU ENSE ensemble2 VRMS 1.540 SOLU SET RFZ=3.9 TFZ=14.3 PAK=4 LLG=144 TFZ==10.5 RFZ=2.4 TFZ=11.9 PAK=9 LLG=259 TFZ==7.0 RFZ=2.7 TFZ=4.7 PAK=19 LLG=202 SOLU SPAC I 41 3 2 SOLU 6DIM ENSE ensemble1 EULER 43.364 14.638 271.356 FRAC 0.81509 0.22245 0.92422 BFAC -7.23070 SOLU 6DIM ENSE ensemble2 EULER 173.495 55.758 234.557 FRAC 1.25797 -0.71934 0.10135 BFAC 1.81823 SOLU 6DIM ENSE ensemble2 EULER 41.452 118.850 315.630 FRAC 0.61025 0.05816 1.60902 BFAC 5.47988 SOLU ENSE ensemble1 VRMS 1.157 SOLU ENSE ensemble2 VRMS 1.540 The TFZ score 14.3 suggests a solution. Then I did refinement using Ploy Ala phaser model. The R values are 0.590/0.62. I use the map to build the model in Buccaneer but it did not build anything. Is the pahser solution correct? Why are the R values so high despite the good TFZ score? Any suggestions are greatly appreciated. Thank you Veerendra Note: This message may contain confidential information. If this Email/Fax has been sent to you by mistake, please notify the sender and delete it immediately. Thank you.
