Re: [ccp4bb] Trying to "digest" PISA results
On Wednesday, 31 August 2011, Jan Dohnalek wrote: > Wasn't the original question directed to our (growing) feeling that many > times PISA says No obvious oligomerization pattern but we already have > evidence of dimer formation etc.. > This should happen "occasionally" as the approach implied in the > calculations is statistical in a sense. We should not be getting such > contradictions on a regular basis. I think there are at least two possibilities 1) the interface seen in the crystal is a real dimer interface, but the PISA score fails to rate it as significant 2) the protein has crystallized as a monomer even though it [sometimes] exists in solution as a dimer. The interface seen in the crystal is not the "real" dimer interface and thus the PISA score is correct. I have no idea which, if either, of these might be the case for 1OYA. Ethan > > Possible I misunderstood the original point ... > > > Jan > > > On Thu, Sep 1, 2011 at 7:46 AM, Karthik S wrote: > > > http://www.ebi.ac.uk/msd-srv/prot_int/pi_ilist_css.html > > so it depends on how many 'stable assemblies' pisa can find i suppose. > > more interfaces and especially if stable enough will make your > > fraction go down. i would have been more surprised or worried if that > > conservative mutation showed radically different CSS scores say one > > close to zero and the other one or close to it. so the exclamation > > marks here are really pointless (since both values are close to zero). > > hence i would ignore the CSS in these two cases. CSS is a statistical > > measure and does not imply biological meaning. in making me (us) > > assume the latter through this one singular value leads to all > > misconceptions. > > > > -- > > Karthik > > > > On Wed, Aug 31, 2011 at 7:52 PM, Yuri Pompeu wrote: > > > I was playing around with PDBe PISA and came across the following: > > > For pdb entry 1OYA. The most promising interface has an area bury of > > around 720A^2 and DeltaG of -10.6Kcal/mol. sym_op(y,x,-z+1) and CSS of > > 0.039! Assembly analysis says it has no strong indications that point to > > stable quaternary structure. > > > This protein has been extensively studied and determined to be a dimer. > > > Entry 3RND is the same protein with one single conservative mutation deep > > in the active site. > > > They align with a RMSD of 0.3 A, 99.8% sequence identity. Superposition > > and inspection of the regions that contact > > > the adjacent monomer shows they are basically identical. > > > The interface here shows Area bury of 760 A^2 and DeltaG = -6.6Kcal/mol. > > sym_op (-y,-x,-z-1/2) CSS=0.00 ! > > > Assembly analysis basically says no stable oligomers form. This enzyme > > also is dimer according to gel filtration. > > > Could anyone ellaborate on this please, if they feel like they have the > > time... > > > Cheers > > > > > > > > >
Re: [ccp4bb] Trying to "digest" PISA results
Wasn't the original question directed to our (growing) feeling that many times PISA says No obvious oligomerization pattern but we already have evidence of dimer formation etc.. This should happen "occasionally" as the approach implied in the calculations is statistical in a sense. We should not be getting such contradictions on a regular basis. Possible I misunderstood the original point ... Jan On Thu, Sep 1, 2011 at 7:46 AM, Karthik S wrote: > http://www.ebi.ac.uk/msd-srv/prot_int/pi_ilist_css.html > so it depends on how many 'stable assemblies' pisa can find i suppose. > more interfaces and especially if stable enough will make your > fraction go down. i would have been more surprised or worried if that > conservative mutation showed radically different CSS scores say one > close to zero and the other one or close to it. so the exclamation > marks here are really pointless (since both values are close to zero). > hence i would ignore the CSS in these two cases. CSS is a statistical > measure and does not imply biological meaning. in making me (us) > assume the latter through this one singular value leads to all > misconceptions. > > -- > Karthik > > On Wed, Aug 31, 2011 at 7:52 PM, Yuri Pompeu wrote: > > I was playing around with PDBe PISA and came across the following: > > For pdb entry 1OYA. The most promising interface has an area bury of > around 720A^2 and DeltaG of -10.6Kcal/mol. sym_op(y,x,-z+1) and CSS of > 0.039! Assembly analysis says it has no strong indications that point to > stable quaternary structure. > > This protein has been extensively studied and determined to be a dimer. > > Entry 3RND is the same protein with one single conservative mutation deep > in the active site. > > They align with a RMSD of 0.3 A, 99.8% sequence identity. Superposition > and inspection of the regions that contact > > the adjacent monomer shows they are basically identical. > > The interface here shows Area bury of 760 A^2 and DeltaG = -6.6Kcal/mol. > sym_op (-y,-x,-z-1/2) CSS=0.00 ! > > Assembly analysis basically says no stable oligomers form. This enzyme > also is dimer according to gel filtration. > > Could anyone ellaborate on this please, if they feel like they have the > time... > > Cheers > > > -- Jan Dohnalek, Ph.D Institute of Macromolecular Chemistry Academy of Sciences of the Czech Republic Heyrovskeho nam. 2 16206 Praha 6 Czech Republic Tel: +420 296 809 390 Fax: +420 296 809 410
Re: [ccp4bb] Trying to "digest" PISA results
http://www.ebi.ac.uk/msd-srv/prot_int/pi_ilist_css.html so it depends on how many 'stable assemblies' pisa can find i suppose. more interfaces and especially if stable enough will make your fraction go down. i would have been more surprised or worried if that conservative mutation showed radically different CSS scores say one close to zero and the other one or close to it. so the exclamation marks here are really pointless (since both values are close to zero). hence i would ignore the CSS in these two cases. CSS is a statistical measure and does not imply biological meaning. in making me (us) assume the latter through this one singular value leads to all misconceptions. -- Karthik On Wed, Aug 31, 2011 at 7:52 PM, Yuri Pompeu wrote: > I was playing around with PDBe PISA and came across the following: > For pdb entry 1OYA. The most promising interface has an area bury of around > 720A^2 and DeltaG of -10.6Kcal/mol. sym_op(y,x,-z+1) and CSS of 0.039! > Assembly analysis says it has no strong indications that point to stable > quaternary structure. > This protein has been extensively studied and determined to be a dimer. > Entry 3RND is the same protein with one single conservative mutation deep in > the active site. > They align with a RMSD of 0.3 A, 99.8% sequence identity. Superposition and > inspection of the regions that contact > the adjacent monomer shows they are basically identical. > The interface here shows Area bury of 760 A^2 and DeltaG = -6.6Kcal/mol. > sym_op (-y,-x,-z-1/2) CSS=0.00 ! > Assembly analysis basically says no stable oligomers form. This enzyme also > is dimer according to gel filtration. > Could anyone ellaborate on this please, if they feel like they have the > time... > Cheers >
[ccp4bb] Trying to "digest" PISA results
I was playing around with PDBe PISA and came across the following: For pdb entry 1OYA. The most promising interface has an area bury of around 720A^2 and DeltaG of -10.6Kcal/mol. sym_op(y,x,-z+1) and CSS of 0.039! Assembly analysis says it has no strong indications that point to stable quaternary structure. This protein has been extensively studied and determined to be a dimer. Entry 3RND is the same protein with one single conservative mutation deep in the active site. They align with a RMSD of 0.3 A, 99.8% sequence identity. Superposition and inspection of the regions that contact the adjacent monomer shows they are basically identical. The interface here shows Area bury of 760 A^2 and DeltaG = -6.6Kcal/mol. sym_op (-y,-x,-z-1/2) CSS=0.00 ! Assembly analysis basically says no stable oligomers form. This enzyme also is dimer according to gel filtration. Could anyone ellaborate on this please, if they feel like they have the time... Cheers
Re: [ccp4bb] Temperature Factor statistics
On Aug 31, 2011, at 4:00 PM, Yuri Pompeu wrote: After i get my output file from baverage containing the average b- factor and rms by residues, How can I calculate and display the average (and or mean) B-factors? Is there a way of calculating it by protein, ligands and solvent separately? Chimera's Render by Attribute tool can depict atoms by their B-factor or residues by their average B-factor (assuming you have a PDB file with a B-factor column). It also shows a histogram of the B-factor values. You can use the Attribute Calculator tool to compute the average B-factor of any part of your structure and write it to a file. Some links: Chimera: www.cgl.ucsf.edu/chimera Render by Attribute: www.cgl.ucsf.edu/chimera/current/docs/ContributedSoftware/render/render.html Attribute Calculator: www.cgl.ucsf.edu/chimera/current/docs/ContributedSoftware/calculator/calculator.html a tutorial focused on coloring by B-factor: www.cgl.ucsf.edu/chimera/docs/UsersGuide/tutorials/bfactor.html --Eric Eric Pettersen UCSF Computer Graphics Lab http://www.cgl.ucsf.edu
Re: [ccp4bb] Windows 7 and Xtal Software
Dear Crystallographers, once again I am filled with graditude to this list--there were many helpful responses and even some geek humor. I have decided to go with a dual-boot windows7/linux, which seems easy enough. All the best, and thanks everyone for your quick and helpful advice. Jacob Keller
Re: [ccp4bb] Do manufacturers change their crystallogenesis screens?
Dear Flip, I think with respect to the Formulatrix database, it would be useful to have the date of entry into the database for each screen input. I agree that there are discrepancies in the database, but they can generally be traced to a change from one catalog to the next. If you have the date of entry, then you can find the "correct" catalog to look at for your formulation information. It would be useful if the various crystallography suppliers would place their plate information in some format easy to incorporate into the Rock Maker database. Maybe this is something that can be discussed at the RAMC in France this fall. Kind regards, Bryan Prince -- Confidentiality Notice: This message is private and may contain confidential and proprietary information. If you have received this message in error, please notify us and remove it from your system and note that you must not copy, distribute or take any action in reliance on it. Any unauthorized use or disclosure of the contents of this message is not permitted and may be unlawful. -Original Message- From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of Flip Hoedemaeker Sent: Wednesday, August 31, 2011 5:01 AM To: CCP4BB@JISCMAIL.AC.UK Subject: Re: [ccp4bb] Do manufacturers change their crystallogenesis screens? Op 8/24/2011 17:21, Chris Morris schreef: Hi Chris, Yes, I have seen quite a few inconsistencies in screen formulations. Errors in listed conditions include recipe changes, but also typos both in the vendor description and in the database entries. At the moment I'm building a list of all discrepancies of the screens in the Formulatrix database together with the date found. This is a tedious work in progress but I'm happy to share the list sofar with people interested (offline). It will be published on the web site when complete. In the meantime, it is wise to double-check each hit condition found on the vendor web site, and in the cases where the exact composition is not published ask the vendor directly. Flip > HI, > > I've recently seen two examples where the description of a screen in a local database was different to the current one on the manufacturer's web site. This happened in two different labs, using different software, and with different screen manufacturers. > > This could potentially lead to an optimisation screen that finds no hits, because the wrong condition is being optimised. Does anyone have experience of this? Am I just looking at a few one-off errors, or is there a general problem here? > > The ideal solution is for screen manufacturers to give version numbers to their screens. Failing that, a good fix at the laboratory is to download the screen description every time a deep-well plate is received, and second best would be to download it every time a trial plate is set up. If there is a real concern here, we will implement one of these in xtalPiMS. > > Regards, > Chris > > > Chris Morris > chris.mor...@stfc.ac.uk > Tel: +44 1925 603689 Fax: +44 1925 603825 > Mobile: 07921-717915 > http://pims.instruct-fp7.eu/ > STFC, Daresbury Lab, Daresbury, Warrington, UK, WA4 4AD >
Re: [ccp4bb] Do manufacturers change their crystallogenesis screens?
Op 8/24/2011 17:21, Chris Morris schreef: Hi Chris, Yes, I have seen quite a few inconsistencies in screen formulations. Errors in listed conditions include recipe changes, but also typos both in the vendor description and in the database entries. At the moment I'm building a list of all discrepancies of the screens in the Formulatrix database together with the date found. This is a tedious work in progress but I'm happy to share the list sofar with people interested (offline). It will be published on the web site when complete. In the meantime, it is wise to double-check each hit condition found on the vendor web site, and in the cases where the exact composition is not published ask the vendor directly. Flip HI, I've recently seen two examples where the description of a screen in a local database was different to the current one on the manufacturer's web site. This happened in two different labs, using different software, and with different screen manufacturers. This could potentially lead to an optimisation screen that finds no hits, because the wrong condition is being optimised. Does anyone have experience of this? Am I just looking at a few one-off errors, or is there a general problem here? The ideal solution is for screen manufacturers to give version numbers to their screens. Failing that, a good fix at the laboratory is to download the screen description every time a deep-well plate is received, and second best would be to download it every time a trial plate is set up. If there is a real concern here, we will implement one of these in xtalPiMS. Regards, Chris Chris Morris chris.mor...@stfc.ac.uk Tel: +44 1925 603689 Fax: +44 1925 603825 Mobile: 07921-717915 http://pims.instruct-fp7.eu/ STFC, Daresbury Lab, Daresbury, Warrington, UK, WA4 4AD
Re: [ccp4bb] twinning in hexagonal system
Dear John, It is not the sequence identity/similarity that counts, but how similar the protein folds are. For many protein families, the fold is identical although the sequence identity is very low. With 25% sequence identity and presumably a protein from the same family, I give you a good chance that e.g. Phaser will find the solution. However, if there is a hinge movement, or some other deformation, you may not find a solution despite very high sequence identity. Concerning your twinning: with 25% twinning fraction, you could try to detwin using Yates algorithm, but I would just try Phaser first with the data as is. I have done this with data sets with a twinning fraction close to 50% and what happened was that phaser would find two solutions, one for each twin orientation. The "raw" electron density map using this twinned data was amazingly good, presumably because the data of the twin fraction with the same orientation as the molecule used for phasing would produce very nice electron density, while the data of the the twin fraction with the other orientation would just produce noise, because for this data phases would be more or less random. So the bottom line: I would just run Phaser with the P6x data as is (with SGALTERNATIVE ALL) and only if that does not work try more elaborate methods. Best, Herman -Original Message- From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of john peter Sent: Tuesday, August 30, 2011 9:32 PM To: CCP4BB@JISCMAIL.AC.UK Subject: [ccp4bb] twinning in hexagonal system Hello All, This is regarding twinning in a data set. I collected a native data set to resolution, 1.8 A. I used XDS suite to process and scale the data set. It scaled well in P622 and I found systematic absence (l=6n present). Hence thought the space group may be P6122/P6522. SFCHECK did not show any twinning and also it did not detect pseudo-translation. Twin test in http://nihserver.mbi.ucla.edu/pystats/ ( Merohedral Twin Detector: Padilla-Yeates Algorithm ) showed perfect twinning. Scaled in P61/65 and SFCHECK reported twinning fraction 0.272 & no pseudo-translation. http://nihserver.mbi.ucla.edu/pystats/ ( Merohedral Twin Detector: Padilla-Yeates Algorithm ) showed perfect twinning. Another server from ucla http://nihserver.mbi.ucla.edu/Twinning/ showed partial twinning with twin fraction 0.23 as follows. 2 along a, b, a*, b* No. Twin Law Related Reflections = 18033 (pairs) No. Twin Law Pairs Considered = 9016 = 0.266149 = 0.095303 Twin Fraction = 0.233249 +/- 0.000602 (SHELXL Commands: TWIN 1 0 0 -1 -1 0 0 0 -1 and BASF 0.233249) In P61/65, I got the following matthews-coeffs mol/asym Matthews Coeff %solvent P(1.73) P(tot) _ 19.7587.39 .00 .00 24.8874.79 .00 .01 33.2562.18 .06 .14 4 2.4449.57 .57 .63 5 1.9536.97 .36 .21 6 1.6324.36 .00 .00 7 1.3911.75 .00 .00 _ May I ask what could be the real twin fraction and what is the likelihood of solving the structure by molecular replacement by models with 25 % sequence identity and 30 % sequence similarity. Thank you so much for reading this mail during your busy hours and all suggestions, comments would be gratefully welcome & appreciated. thank you ccp4 mailing list. John
