[ccp4bb] Need suggestion
Dear expert, I am currently working on building my cryo-EM data using COOT. However, I have encountered an issue when trying to add monomers such as Glycerol or BCT using their letter code from the "Get monomer" option in COOT. Whenever I attempt to merge them into my coordinate, COOT crashes. I would greatly appreciate any suggestions or advice on how to overcome this issue. I am currently using version 0.9.8.5 of COOT on a Mac operating system. I am unsure if this is a bug or if there is a specific workaround for this situation. Thank you for your assistance. Best Regards ======= Afshan Begum To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=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/
[ccp4bb] Hello
Hi Experts, I have a problem to see the residue information on COOT if I click on residue info and place the curser their its just blink and disappear as I want to reduce some occupancy manually and it did not work but for water and the other stuff its work completely fine but not for the amino acid residue . Could you people help me to do so. I am working on MAC system Would be thankful to you. Best A.B To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=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/
[ccp4bb] Fixing Modified residue
Dear Expert, I have a trouble to fix the lysin modified residue in to the PDB coordinate. what i did,I get the KCX(Modified lysin) coordinate from monomer library in COOT (0.9) , merge in to the coordinate file , its listed into position 193 lysin then run rafmac 5 , its place the residue but as unconnected and produce its own number. In the second step I take the kcx coordinate in the text pdb file and place into between 192 and 194 and change reside number 193 . Save it. Now it can not be read for further refinement.Could you people suggest me how to fix it?Would be highly thankful to you. Best Regards === Afshan To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=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/
Re: [ccp4bb] Lysine coordinated ions
Hi Kaherine, It seems its a modified lysine in to kcx (Carbamylated lysine) but its only the case if you used during entire purification or crystallization BME in your buffer Best Regards AFSHAN On Wednesday, July 2, 2014 5:38 PM, Katherine Sippel katherine.sip...@gmail.com wrote: At various peoples' urgings (thanks for the contributions all) I went ahead and put in Mg based on the bond valence. If refines very nicely in the context of the density maps (see attached MgCl2_pH6point5_3.png). I ran it through the check my metal server and it clears the coordination geometry and bond distance parameters but is displeased with the coordinating ligands. I am also concerned about the rotamer (big ol' outlier) and the CE-NZ-Mg bond angle. Obviously the density looks good, but it feels weird from a basic chemical standpoint. The crystallization condition for the ion-coordinated lysine was 100 mM Bis Tris pH 6.5, 200 mM MgCl2, 19% PEG 3350. One of the other space groups has 100 mM Tris pH 8.5, 200 mM MgNO3, 27% PEG 1500 and there is no errant density (see attached MgNO3_pH8point5.png). The sodium I mentioned comes from NaCl in the protein storage buffer. Thank you for all of your help thus far. Katherine On Tue, Jul 1, 2014 at 9:35 PM, Nat Echols nathaniel.ech...@gmail.com wrote: On Tue, Jul 1, 2014 at 3:10 PM, Katherine Sippel katherine.sip...@gmail.com wrote: My google-fu has failed me once again so I am turning to the collective knowledge of the bb. I'm working on a blobology challenge at the moment and have hit a wall. Is anyone aware of an ion that coordinates to lysine and prefers octahedral geometry. The mystery ion seems to have perfect octahedral geometry with bond distances of ~2.1 angstrom, but the only direct side chain interaction is to a lysine NZ, the rest are waters. Lysine can coordinate a cation if the chemical environment is favorable - usually this means a high-pH buffer (what was the pH of your crystals?). The same is true for N-termini; I may be able to dig up a published example of this. (I think it is effectively impossible for Arg, however.) These interactions are certainly exceedingly rare (and I doubt they are ever present in vivo), but if the nitrogen loses a proton the lone pair will be able to coordinate a compatible ions. Since magnesium can be coordinated by the nitrogen histidine it seems like the most likely candidate - but I would still be very, very careful before assigning it, especially if the only other coordinating atoms are waters. -Nat -- Nil illegitimo carborundum- Didactylos
[ccp4bb] R factor geeting stuck
Hi ccp4 experts, I have collected diffraction images to 0.96 Angstrom resolution to the edge of the detector. One data set give me the full completeness and below i have pasted my statistic values got from XDS. I have cut off data at 1.12 A which seems to be quite nice regarding all necessary parameter to consider. But the problem is the Rfree is 0.176 and Rwork is 0.151 but the maps look very good. Even If I cut off the data to 1.15 Angstrom the R factors not improved. The space group of my complex is P21212. I used anisotropic Bfactors, add Hydrogen on and also used TLS but unfortunately i can not reduce able to reduce R factor in a good way. so, could you people kindly give me some suggestion how can i improve my data quality. RESOLUTION NUMBER OF REFLECTIONS COMPLETENESS R-FACTOR R-FACTOR COMPARED I/SIGMA R-meas CC(1/2) Anomal SigAno Nano LIMIT OBSERVED UNIQUE POSSIBLE OF DATA observed expected Corr 1.12 104562 21143 21268 99.4% 37.0% 37.8% 103982 4.04 41.4% 91.8* 0 0.767 15016 1.04 86588 21777 23100 94.3% 98.4% 95.8% 84962 1.44 113.0% 74.1* -1 0.700 10431 I would be thankful for your valuable comments AFSHAN
[ccp4bb] need suggestions
Dear All, I have encountered one problem to optimization the crystallization condition manually. Actually i got the good shape and even size crystal in a screening plate. The condition are : 20% PEG 6000, 0.1M MES pH 6.0 whereas protein in 10mM Na-acetate pH 5.7 contain 50mM NaCl, 1mM EDTA 5mM BME. When I tried to optimize this on Linbro the drops are become heavily precipitate even a mints. I tried so many things to avoid this ppt such as start PEG conc from 7%, dilute protein at half conc., add more salt in a reservoir solution but no succeeds. The protein prep is same which was using to get the Hits. I would be very thankful if you people give me nice suggestions. Thanks Best Regards AFSHAN
[ccp4bb] having difficulties with ammonium ion
Hello all, I am refining my structures with REFMAC 5 and is failed. Actually i used ammonium dihydrogen phosphate buffer during crystallization and in the map file where i observed the density of ammonium ion (NH4). I get this ion from the coot library but when i run refmac its failed even i gave the cif file for this ion as a supplement but i could not get success so, if any body give me suggestion how to solve this problem. I would be thankful to all. this are the error message came from refmac log file ERROR : atom :H4_N NH4 1 FF is absent in the library ATTENTION: atom:N NH4 1 FF is missing in the structure ERROR : atom :H4_N NH4 2 Fa is absent in the library ATTENTION: atom:N NH4 2 Fa is missing in the structure ERROR : atom :H4_N NH4 3 Fb is absent in the library ATTENTION: atom:N NH4 3 Fb is missing in the structure ERROR : atom :H4_N NH4 4 Fc is absent in the library ATTENTION: atom:N NH4 4 Fc is missing in the structure ERROR : atom :H4_N NH4 5 Fd is absent in the library ATTENTION: atom:N NH4 5 Fd is missing in the structure ERROR : atom :H4_N NH4 6 Fe is absent in the library ATTENTION: atom:N NH4 6 Fe is missing in the structure ERROR : atom :H4_N NH4 7 Ff is absent in the library ATTENTION: atom:N NH4 7 Ff is missing in the structure Best Regards AFSHAN
[ccp4bb] Hi
Hope you get this on time, I made a trip to Aberdeen in Scotland and had my bag stolen from me with my passport,cash and credit cards in it. unfortunately for me,I have made contact with my bank but they are not providing a fast solution. I need you to lend me some money to sort my self out of this predicament,i will pay back as soon as i return. Western Union transfer is the best option to send money to me. Let me know if you need my details(Full names location) to make the transfer. You can reach me via email or the hotel's desk phone the numbers are,+447031804805 I await your response Thanks Afshan
[ccp4bb] chirality problem
Dear Users, I am facing difficulties to validate my structure according to PDB server. I have solved my structure and now want to submit in PDB but during validation process i have some chirality problem specially VAL and LEU amino acids there are total 18 amino acids which deviated from the chirality so how can i solve this problem. Any suggestion would be highly appreciated. Best Regards AFSHAN
[ccp4bb] how can merge two PDB
Hello CCP4 user I have collected a data set 2.1 for my complex. Actually after first run of Rafmac i can see the density for my inhibitor but the problem is my inhibitor is 6 KDa and i know the sequence of my inhibitor as well this inhibitor already crystallize with other protein molecule present in PDB data bank so how can i put in to that electron density i mean are there any ways to combine two Pdb in one molecule? I would be thankful for your help Best Regards AFSHAN
Re: [ccp4bb] how can merge two PDB
Dear Juergen Many thank for your response yes you have excatly understand my question we have a MR solution of the rest of our protein and just asking how to make my life easier to not built de novo 45-50 residues. so i could not find the option in coot find ligand so, from where i get it? Best Regards AFSHAN From: Afshan Begum afshan...@yahoo.com To: Bosch, Juergen jubo...@jhsph.edu Sent: Wednesday, October 19, 2011 4:58 PM Subject: Re: [ccp4bb] how can merge two PDB H.EDU To: CCP4BB@JISCMAIL.AC.UK Sent: Wednesday, October 19, 2011 4:29 PM Subject: Re: [ccp4bb] how can merge two PDB why don't you read in that chain in Coot and run the find ligand option with flexible ligand turned on and select that 6kDa ligand ? You should also choose Fo-Fc map to fit the ligand to maybe at 2.7 sigma. You might have to split up the ligand into pieces, not sure what the limitations in Coot/Find Ligand are. You already have a MR solution of the rest of your protein right ? So you are just asking how to make your life easier to not built de novo 45-50 residues ? Jürgen On Oct 19, 2011, at 10:00 AM, Ed Pozharski wrote: Why not do the molecular replacement - 6kDa is rather small but it most likely will work On Wed, 2011-10-19 at 06:13 -0700, Afshan Begum wrote: Hello CCP4 user I have collected a data set 2.1 for my complex. Actually after first run of Rafmac i can see the density for my inhibitor but the problem is my inhibitor is 6 KDa and i know the sequence of my inhibitor as well this inhibitor already crystallize with other protein molecule present in PDB data bank so how can i put in to that electron density i mean are there any ways to combine two Pdb in one molecule? I would be thankful for your help Best Regards AFSHAN -- Oh, suddenly throwing a giraffe into a volcano to make water is crazy? Julian, King of Lemurs .. Jürgen Bosch Johns Hopkins University Bloomberg School of Public Health Department of Biochemistry Molecular Biology Johns Hopkins Malaria Research Institute 615 North Wolfe Street, W8708 Baltimore, MD 21205 Office: +1-410-614-4742 Lab: +1-410-614-4894 Fax: +1-410-955-2926 http://web.mac.com/bosch_lab/
[ccp4bb] How can improve diffraction quality
Dear ccp4 user I am facing one crucial problem regarding diffraction. Actually the size of my crystal is good enough 0.5mm but it was diffracted only 4 A. The conditions of crystallization recipes are 4.5% PEG 1000, 0.1M Tris and 25mM Na citrate. I really need your suggestions regarding how can i improve my diffraction quality? Your support is highly appreciable. Best Regards AFSHAN
[ccp4bb] hello
Dear all, I have facing one problem during the refinement of my protein . Actually in my protein there are some modified amino acids are present like Cystein is modified into CME which i can get easily from monomer libraray in coot . but after refinement in Pdb text file indicated some gaps while in the structures there are no gap in between these amino acids so if any one suggest me what to do. I would appreciate your kind suggestions. LINKR GLU A 142 LEU A 144 gap LINKR SER A 328 GLY A 330 gap LINKR LEU A 138 GLU A 140 gap LINKR GLU A 126 ASP A 130 gap LINKR SER A 246 GLY A 248 gap Many thanks for your time Best regards Afshan
[ccp4bb] hello
Dear all, Could any one help me regarding my serious problem actually i have collected data at 3.0 and cut off 3.1 where the data statics showed the good values for the further processing. According to the methew coefficient there would be two molecule in the asymmetric unit but after running the molrep its provide only one monomer instead of two, for this reason R values is very high 50%. Actually homologous model having P6322 space group where as my one is F4132. I had tried to run phaser as well phenix but both were failed to process further. i really do not know how can i get the second chain in my structure. Please if you have some ideas i will appreciate and would be many thankful to you. Hope to hearing you soon Best Regards AFSHAN
[ccp4bb] need proper suggestion
Dear All, I have a severe prob lam to performed my ligand binding study with corresponding protein. I have taking the native diffraction data at 1.75 A and after that i have performed soaking as well co-crystallization experiment with my inhibitors. Problem is that at the active site phosphate ion always bind instead of inhibitors. I have used 1.6 M ammonium phosphate conc at the crystallization recipe which is a very weak inhibitor of my protein whereas the ligand is already clinically applicable but due to the very high conc. of phosphate i have not achieved my target. If some one can suggest me what else i can replace with ammonium phosphate or any other suggestions would be appreciated. I have tried to grown crystals some other condition but the crystal was not diffracted beyond 3.5 A. Best Regards AFSHAN
