Re: [ccp4bb] Another paper structure retracted
As the macromolecular crystallography becomes more automated and user-friendly many biologists learn to solve structures and they can make mistakes. Besides, new data become available that can give new ideas etc. I don't think that's so horrible to make an honest mistake and retract papers. Even great scientists sometimes published papers with wrong analysis. Maia On 11/08/2011 1:05 PM, Judith Murray-Rust wrote: Just another point - the macromolecular community are not the only ones with a problem - I've just been shown http://retractionwatch.wordpress.com/ which sheds some light on retractions. And also maybe says something about why original data should be available/part of the review process. J From: CCP4 bulletin board [CCP4BB@JISCMAIL.AC.UK] On Behalf Of Gloria Borgstahl [gborgst...@gmail.com] Sent: 11 August 2011 19:32 To: CCP4BB@JISCMAIL.AC.UK Subject: Re: [ccp4bb] Another paper structure retracted Dale, This is exactly the conversation I just had with my student Jason, right on! The paper we are writing just now, this is figure 1. But I always get rejected by Nature, so go figure. On Thu, Aug 11, 2011 at 1:25 PM, Dale Tronruddet...@uoxray.uoregon.edumailto:det...@uoxray.uoregon.edu wrote: I agree with Prof. Tomchick: if the point of your paper is your crystal structure of the binding of a ligand to a protein you should include a figure with the omit map (displayed without a cover radius) that convinced you that binding took place. I prefer that map over some simulated, after-the-fact, omit map calculated just for publication. This is not simply a matter for reviewers to be gatekeepers, it is important for the readers to know what level of confidence to place in this result, and it is instructional for everyone to see what ligand binding density looks like. Apparently some people don't know what features to look for to distinguish between signal and noise. Dale Tronrud On 08/11/11 09:40, Diana Tomchick wrote: A quick glance at the header of the PDB file shows that there is one glaring discrepancy between it and the table in the paper that hasn't been mentioned yet in this forum. The data completeness (for data collection) reported in the paper is 95.7%, but in the header of the PDB file (actually, in both the 2QNS and the 3KJ5 depositions) the data completeness (for data collection) is reported as only 59.4%. The PDB header also contains an inconsistency, with the data completeness (for refinement) reported as 95.7%. Since the numbers of reflections reported for refinement versus data collection in the PDB header differ by less than 1%, it appears that there's been a bit of magical thinking that took place somewhere along the process from data processing to final model refinement. Small wonder that the refined geometry is so poor. Perhaps if these scientists had actually collected a complete dataset, we would not be having this conversation. Diana P.S. I have, on occasion, provided the coordinates and a map file to reviewers when they requested it. The last time it was requested was many years ago; I decided it was safer and easier if I provided as much information as possible in the manuscript (including better quality electron density figures than appear in this paper) to allow the reader to determine whether the work is valid or not. * * * * * * * * * * * * * * * * * * * * * * * * * * * * Diana R. Tomchick Associate Professor University of Texas Southwestern Medical Center Department of Biochemistry 5323 Harry Hines Blvd. Rm. ND10.214B Dallas, TX 75390-8816, U.S.A. Email: diana.tomch...@utsouthwestern.edu 214-645-6383tel:214-645-6383 (phone) 214-645-6353tel:214-645-6353 (fax) On Aug 10, 2011, at 5:45 PM, Dale Tronrud wrote: I've made a quick look at the model and the paper - and it doesn't need more than a quick look. The description of the model in the paper sounds great. The problems in the model are clear. My favorite is the quote Trp-477 of PTH1R makes several van der Waals contacts with Trp-339 and Lys-337 of G-beta-1 They are contacts all right. The distances between the 477:CH2 and 337:CE is 2.75 A and between 477:NE1 and 339:CH2 is 2.26 A. There are many more. In general the geometry of this entire model is terrible. In Table 1 the bond length rmsd is listed at 1.64 A and the bond angles are 0.0078 deg! Perhaps one is to presume the numbers should be swapped. In any case, the values I calculate for the model are 0.160 A and 4.46 deg! Absolutely dreadful. The PDB header lists the (swapped) values from the paper and then reports hundreds of outliers. The tools proposed by the Validation Task Force should cause a model like this to pop out clearly. Even the old tools show this model is quite unreliable. We just have to use them. Dale Tronrud On 08/10/11 14:35, Jacob Keller wrote: On the surface it doesn't seem as bad as others, i.e., it does not seem to
Re: [PyMOL] how to load two separate pdb files simultaneously
Also you can use load A.pdb B.pdb C.pdb or load *pdb leila karami wrote: Dear all very thanks for your time and attention. my problem was solved by load A.pdb load B.pdb -- All of the data generated in your IT infrastructure is seriously valuable. Why? It contains a definitive record of application performance, security threats, fraudulent activity, and more. Splunk takes this data and makes sense of it. IT sense. And common sense. http://p.sf.net/sfu/splunk-d2d-c2 ___ PyMOL-users mailing list (PyMOL-users@lists.sourceforge.net) Info Page: https://lists.sourceforge.net/lists/listinfo/pymol-users Archives: http://www.mail-archive.com/pymol-users@lists.sourceforge.net -- All of the data generated in your IT infrastructure is seriously valuable. Why? It contains a definitive record of application performance, security threats, fraudulent activity, and more. Splunk takes this data and makes sense of it. IT sense. And common sense. http://p.sf.net/sfu/splunk-d2d-c2 ___ PyMOL-users mailing list (PyMOL-users@lists.sourceforge.net) Info Page: https://lists.sourceforge.net/lists/listinfo/pymol-users Archives: http://www.mail-archive.com/pymol-users@lists.sourceforge.net
Re: [PyMOL] how to load two separate pdb files simultaneously
I made a mistake. I meant command-line pymol *pdb or pymol A.pdb B.pdb etc. Not load *pdb. Maia Cherney wrote: Also you can use load A.pdb B.pdb C.pdb or load *pdb leila karami wrote: Dear all very thanks for your time and attention. my problem was solved by load A.pdb load B.pdb -- All of the data generated in your IT infrastructure is seriously valuable. Why? It contains a definitive record of application performance, security threats, fraudulent activity, and more. Splunk takes this data and makes sense of it. IT sense. And common sense. http://p.sf.net/sfu/splunk-d2d-c2 ___ PyMOL-users mailing list (PyMOL-users@lists.sourceforge.net) Info Page: https://lists.sourceforge.net/lists/listinfo/pymol-users Archives: http://www.mail-archive.com/pymol-users@lists.sourceforge.net -- All of the data generated in your IT infrastructure is seriously valuable. Why? It contains a definitive record of application performance, security threats, fraudulent activity, and more. Splunk takes this data and makes sense of it. IT sense. And common sense. http://p.sf.net/sfu/splunk-d2d-c2 ___ PyMOL-users mailing list (PyMOL-users@lists.sourceforge.net) Info Page: https://lists.sourceforge.net/lists/listinfo/pymol-users Archives: http://www.mail-archive.com/pymol-users@lists.sourceforge.net
Re: [ccp4bb] Kd's in Crystals
Hi, We had a paper where we looked at Kd of arginine in the arginine repressor-DNA complex (p. 248-249). JMB,2010, *399*, pp.240-254. Maia Jacob Keller wrote: Yes, I think you are right--the somewhat counterintuitive case I was thinking of was, for example, when: Kd = 20nM [L] = 20uM [Po in crystal] = 20mM In this case, even though [L] = 20uM, since [L] is 1000 x Kd, the occupancy should be ~100%, and [PL] at equilibrium should be about 20mM, so in the crystal, the total [L] should be ~20mM. This explains, among other things, why bromophenol blue makes crystals bluer than the surrounding solution--the Kd is probably significantly lower than the BB concentration in the drop. Thanks for your clarifications! Jacob The question would remain, then, whether there is any utility in titrating ligands into crystals, and monitoring occupancies as a readout for binding. Although crystallization conditions are horribly non-physiological, perhaps there would be utility in the case where there are multiple known binding sites of various affinities, and other methods would have trouble resolving the binding events. One could start with: 1. totally saturated conditions, set occ=1 for all sites, refine B's, then 2. fix B's at this value, and refine the occ's in a subsequent series of dilutions. All of this is not totally theoretical--I am considering a set of experiments along these lines, where there really are multiple sites of varying affinity. *** Jacob Pearson Keller Northwestern University Medical Scientist Training Program cel: 773.608.9185 email: j-kell...@northwestern.edu ***
Re: [PyMOL] [Fwd: Re: select residues from different chains]
Thanks, Zhijian It worked zjxu wrote: Dear Maia, Would you like to try this: select bridge, (c. a and i. 160+356+505+507+508) or (c. c and i. 275+355+128) Best Regards, Zhijian Xu Maia Cherney wrote: Hi Jason, I keep sending my e-mail from a wrong address that the pymol mailing list does nor recognize. I need a command that would allow me to select several residues from two chains, something like that: select bridge, /prot//A/160+356+505+507+508/+prot/C//275+355+128/ But I cannot find correct syntax. Maia -- Simplify data backup and recovery for your virtual environment with vRanger. Installation's a snap, and flexible recovery options mean your data is safe, secure and there when you need it. Discover what all the cheering's about. Get your free trial download today. http://p.sf.net/sfu/quest-dev2dev2 ___ PyMOL-users mailing list (PyMOL-users@lists.sourceforge.net) Info Page: https://lists.sourceforge.net/lists/listinfo/pymol-users Archives: http://www.mail-archive.com/pymol-users@lists.sourceforge.net -- Simplify data backup and recovery for your virtual environment with vRanger. Installation's a snap, and flexible recovery options mean your data is safe, secure and there when you need it. Discover what all the cheering's about. Get your free trial download today. http://p.sf.net/sfu/quest-dev2dev2 ___ PyMOL-users mailing list (PyMOL-users@lists.sourceforge.net) Info Page: https://lists.sourceforge.net/lists/listinfo/pymol-users Archives: http://www.mail-archive.com/pymol-users@lists.sourceforge.net
[PyMOL] [Fwd: Re: select residues from different chains]
Hi Jason, I keep sending my e-mail from a wrong address that the pymol mailing list does nor recognize. I need a command that would allow me to select several residues from two chains, something like that: select bridge, /prot//A/160+356+505+507+508/+prot/C//275+355+128/ But I cannot find correct syntax. Maia -- Simplify data backup and recovery for your virtual environment with vRanger. Installation's a snap, and flexible recovery options mean your data is safe, secure and there when you need it. Discover what all the cheering's about. Get your free trial download today. http://p.sf.net/sfu/quest-dev2dev2 ___ PyMOL-users mailing list (PyMOL-users@lists.sourceforge.net) Info Page: https://lists.sourceforge.net/lists/listinfo/pymol-users Archives: http://www.mail-archive.com/pymol-users@lists.sourceforge.net
Re: [ccp4bb] crystal bent once open cover slip
You probably use hanging drops. It's the surface tension effect. Check if sitting drops are better. Maia Sitting drops weikai wrote: Hi Folks, We have some membrane protein crystals that are grown in 30%PEG400, 0.1M Na Citrate pH 4.5, 0.1M LiCl. The protein is purified in DDM. The crystals are long rods and grown under room temperature in a hanging drop set up. But once we open the cover slip, we see the rods start to break and bend in a few seconds. Since it is in high PEG400, we just directly freeze the crystals. The diffraction only goes to 10 Ang at synchrotron. Have anybody had similar problem before and any suggestions? Thanks a lot, Weikai
Re: [ccp4bb] what to do with disordered side chains
I guess, most hydrophilic side chains on the surface are flexible, they don't keep the same conformation. If you cut those side chains off, the surface will be looking pretty hydrophobic and misleading (and very horrible). I prefer to see them intact. I know, most of them are flexible and don't have one exact position, but it's OK. I know they are there not far from the main chain. Usually, their exact position is irrelevant. Maia Jacob Keller wrote: Well, what about getting the default settings on the major molecular viewers to hide atoms with either occ=0 or bcutoff (novice mode?)? While the b cutoff is still be tricky, I assume we could eventually come to consensus on some reasonable cutoff (2 sigma from the mean?), and then this approach would allow each free-spirited crystallographer to keep his own preferred method of dealing with these troublesome sidechains and nary a novice would be led astray JPK On Sun, Apr 3, 2011 at 2:58 PM, Eric Bennett er...@pobox.com wrote: Most non-structural users are familiar with the sequence of the proteins they are studying, and most software does at least display residue identity if you select an atom in a residue, so usually it is not necessary to do any cross checking besides selecting an atom in the residue and seeing what its residue name is. The chance of somebody misinterpreting a truncated Lys as Ala is, in my experience, much much lower than the chance they will trust the xyz coordinates of atoms with zero occupancy or high B factors. What worries me the most is somebody designing a whole biological experiment around an over-interpretation of details that are implied by xyz coordinates of atoms, even if those atoms were not resolved in the maps. When this sort of error occurs it is a level of pain and wasted effort that makes the pain associated with having to build back in missing side chains look completely trivial. As long as the PDB file format is the way users get structural data, there is really no good way to communicate atom exists with no reliable coordinates to the user, given the diversity of software packages out there for reading PDB files and the historical lack of any standard way of dealing with this issue. Even if the file format is hacked there is no way to force all the existing software out there to understand the hack. A file format that isn't designed with this sort of feature from day one is not going to be fixable as a practical matter after so much legacy code has accumulated. -Eric On Apr 3, 2011, at 2:20 PM, Jacob Keller wrote: To the delete-the-atom-nik's: do you propose deleting the whole residue or just the side chain? I can understand deleting the whole residue, but deleting only the side chain seems to me to be placing a stumbling block also, and even possibly confusing for an experienced crystallographer: the .pdb says lys but it looks like an ala? Which is it? I could imagine a lot of frustration-hours arising from this practice, with people cross-checking sequences, looking in the methods sections for mutations... JPK
Re: [ccp4bb] Detergent/lipid crystal diffraction pattern?
Pius, Are you sure that you determined the correct cell. Which program did you use? Usually there are much less spots on an image when a crystal has so small unit size dimensions. To me the first crystal looks like protein. Send it to a synchrotron and process the data in XDS. They can process for you. Maia On 21/03/2011 4:33 PM, Maia Cherney wrote: Hi PS What is the unit cell dimensions in the first crystal? It looks like protein to me. Maia On 21/03/2011 2:03 PM, Pius Padayatti wrote: Hi all, We recently observed some diffraction from membrane protein crystallization drops diffraction that look like non-proteinaceous (please see attached files, from 4 different crystals grown in different conditions). Rains' question about about lipid and detergent diffraction is so relevant. This is most likely what lipids and detergent diffraction looks like? People with similar experience and know what could be these patterns might be from may have better suggestions and would like to hear all comments. first four images are from drops where detergent is DDM and and vapor diffusion while last image is from a crystal grown in mesophase (with monoolein). Padayatti PS On Sat, Mar 19, 2011 at 7:19 PM,Rain Field rainfiel...@163.com wrote: Hi All, I am wondering if the detergent or lipid crystal can have diffraction at low resolution. If they can, what does the diffraction pattern looks like? Are there any literatures describing these? Many thanks!
Re: [ccp4bb] Detergent/lipid crystal diffraction pattern?
Hi PS What is the unit cell dimensions in the first crystal? It looks like protein to me. Maia On 21/03/2011 2:03 PM, Pius Padayatti wrote: Hi all, We recently observed some diffraction from membrane protein crystallization drops diffraction that look like non-proteinaceous (please see attached files, from 4 different crystals grown in different conditions). Rains' question about about lipid and detergent diffraction is so relevant. This is most likely what lipids and detergent diffraction looks like? People with similar experience and know what could be these patterns might be from may have better suggestions and would like to hear all comments. first four images are from drops where detergent is DDM and and vapor diffusion while last image is from a crystal grown in mesophase (with monoolein). Padayatti PS On Sat, Mar 19, 2011 at 7:19 PM,Rain Field rainfiel...@163.com wrote: Hi All, I am wondering if the detergent or lipid crystal can have diffraction at low resolution. If they can, what does the diffraction pattern looks like? Are there any literatures describing these? Many thanks!
Re: [PyMOL] hbs
Hi, Can anybody advise me what is the right command for selecting or coloring many hbs? Instead of color black, hb1 color black, hb2 etc I want something like color black, hb1+hb2+hb3 etc. Maia -- Colocation vs. Managed Hosting A question and answer guide to determining the best fit for your organization - today and in the future. http://p.sf.net/sfu/internap-sfd2d ___ PyMOL-users mailing list (PyMOL-users@lists.sourceforge.net) Info Page: https://lists.sourceforge.net/lists/listinfo/pymol-users Archives: http://www.mail-archive.com/pymol-users@lists.sourceforge.net
Re: [PyMOL] delete an hb from a multiple hb object
Hi Jason, I generated hbs from mode=2 method. It made some wrong hbs. How can I delete those wrong hbs? Maia -- Colocation vs. Managed Hosting A question and answer guide to determining the best fit for your organization - today and in the future. http://p.sf.net/sfu/internap-sfd2d ___ PyMOL-users mailing list (PyMOL-users@lists.sourceforge.net) Info Page: https://lists.sourceforge.net/lists/listinfo/pymol-users Archives: http://www.mail-archive.com/pymol-users@lists.sourceforge.net
Re: [ccp4bb] [Fwd: Re: [ccp4bb] I/sigmaI of 3.0 rule]
Kay, Thank you for your explanation. The radiation damage was not the factor, but there was something strange about this crystal (actually two crystals had the same strange behavior). I could not process them in HKL2000, but it showed the problem (see pictures in the attachment). The processing in XDS was done at the CLS (Canadian Light Source). I know they always have the latest version of XDS. Maia Kay Diederichs wrote: Maia, provided radiation damage is not a major detrimental factor, your data are just fine, and useful also in the high resolution shell (which still has I/sigma of 2.84 so you could probably process a bit beyond 2.25A). There is nothing wrong with R_meas of 147.1% since, as others have said, R_meas is not limited to 59% (or similar) as a refinement R-factor is. Rather, R_meas is computed from a formula that has a denominator which in the asymptotic limit (noise) approaches zero - because there will be (almost) as many negative observations as positive ones! (The numerator however does not go to zero) Concerning radiation damage: First, take a look at your frames - but make sure you have the same crystal orientation, as anisotropy may mask radiation damage! Then, you can check (using CCP4's loggraph) the R_d plot provided by XDSSTAT (for a single dataset; works best for high-symmetry spacegroups), and you should also check ISa (printed in CORRECT.LP and XSCALE.LP). HTH, Kay P.S. I see one potential problem: XSCALE (VERSION December 6, 2007) when the calculation was done 28-Aug-2009. There were quite a number of improvements in XDS/XSCALE since that version. The reason may be that a licensed, non-expiring version was used - make sure you always rather use the latest version available! Original Message Subject: [Fwd: Re: [ccp4bb] I/sigmaI of 3.0 rule] Date: Thu, 3 Mar 2011 10:45:03 -0700 From: Maia Cherney ch...@ualberta.ca Original Message Subject: Re: [ccp4bb] I/sigmaI of 3.0 rule Date: Thu, 03 Mar 2011 10:43:23 -0700 From: Maia Cherney ch...@ualberta.ca To: Oganesyan, Vaheh oganesy...@medimmune.com References: 2ba9ce2f-c299-4ca9-a36a-99065d1b3...@unipd.it 4d6faed8.7040...@ualberta.ca 021001cbd9bc$f0ecc940$d2c65bc0$@gmail.com 4d6fcab6.3090...@ualberta.ca 4d6fcbff.2010...@ualberta.ca 73e543de77290c409c9bed6fa4ca34bb0173a...@md1ev002.medimmune.com Vaheh, The problem was with Rmerg. As you can see at I/sigma=2.84, the Rmerge (R-factor) was 143%. I am asking this question because B. Rupp wrote However, there is a simple relation between I/sigI and R-merge (provided no other indecency has been done to the data). It simply is (BMC) Rm=0.8/I/sigI. Maybe my data are indecent? This is the whole LP file. Maia MMC741_scale-2.25.LP ** XSCALE (VERSION December 6, 2007) 28-Aug-2009 ** Author: Wolfgang Kabsch Copy licensed until (unlimited) to Canadian Light Source, Saskatoon, Canada. No redistribution. inline: Cell1.gifinline: Distance1.gif
Re: [ccp4bb] weird crystals, I/sigma of 3.0 rule
Hi James, I remember that P1 did not help. That was like 2 years ago. That crystal was very important at that time, so I had to use it. There were many other crystals since then (native, mutants and complexes) in the same space group without problems. But also I had even a more weird crystal that Randy Read was interested to investigate fully. That crystal produced a nice diffraction to high resolution, scaled perfectly in any program, had no twinning. It had the same space group with slightly different cell dimensions . It gave huge LLG in phaser, but could never be refined. I sent those data to many people, and Randy figured out the type of disorder that crystal had. I guess, some crystals can be very weird. Maia
Re: [ccp4bb] I/sigmaI of 3.0 rule
Dear Bernhard I am wondering where I should cut my data off. Here is the statistics from XDS processing. Maia SUBSET OF INTENSITY DATA WITH SIGNAL/NOISE = -3.0 AS FUNCTION OF RESOLUTION RESOLUTION NUMBER OF REFLECTIONS COMPLET R-FACTOR R-FACTOR COMPARED I/SIGMA R-meas Rmrgd-F Anomal SigAno Nano LIMIT OBSERVED UNIQUE POSSIBLE OF DATA observed expected Corr 10.06 5509 304 364 83.5% 3.0% 4.4% 5509 63.83 3.1% 1.0% 11% 0.652 173 7.12 11785 595 595 100.0% 3.5% 4.8% 11785 59.14 3.6% 1.4% -10% 0.696 414 5.81 15168 736 736 100.0% 5.0% 5.6% 15168 51.88 5.1% 1.8% -9% 0.692 561 5.03 17803 854 854 100.0% 5.5% 5.7% 17803 50.02 5.6% 2.2% -10% 0.738 675 4.50 20258 964 964 100.0% 5.1% 5.4% 20258 52.61 5.3% 2.1% -16% 0.710 782 4.11 22333 1054 1054 100.0% 5.6% 5.7% 22333 50.89 5.8% 2.0% -16% 0.705 878 3.80 23312 1137 1137 100.0% 7.0% 6.6% 23312 42.95 7.1% 3.0% -13% 0.770 952 3.56 25374 1207 1208 99.9% 7.6% 7.3% 25374 40.56 7.8% 3.4% -18% 0.739 1033 3.35 27033 1291 1293 99.8% 9.7% 9.2% 27033 33.73 10.0% 4.1% -12% 0.765 1107 3.18 29488 1353 1353 100.0% 11.6% 11.6% 29488 28.16 11.9% 4.4% -7% 0.750 1176 3.03 31054 1419 1419 100.0% 15.7% 15.9% 31054 21.77 16.0% 6.9% -9% 0.741 1243 2.90 32288 1478 1478 100.0% 21.1% 21.6% 32288 16.99 21.6% 9.2% -6% 0.745 1296 2.79 33807 1542 1542 100.0% 28.1% 28.8% 33807 13.07 28.8% 12.9% -2% 0.783 1361 2.69 34983 1604 1604 100.0% 37.4% 38.7% 34983 9.95 38.3% 17.2% -2% 0.743 1422 2.60 35163 1653 1653 100.0% 48.8% 48.0% 35163 8.03 50.0% 21.9% -6% 0.754 1475 2.52 36690 1699 1699 100.0% 54.0% 56.0% 36690 6.98 55.3% 25.9% 0% 0.745 1517 2.44 37751 1757 1757 100.0% 67.9% 70.4% 37751 5.61 69.5% 32.5% -5% 0.733 1577 2.37 38484 1798 1799 99.9% 82.2% 84.5% 38484 4.72 84.2% 36.5% 2% 0.753 1620 2.31 39098 1842 1842 100.0% 91.4% 94.3% 39098 4.19 93.7% 43.7% -3% 0.744 1661 2.25 38809 1873 1923 97.4% 143.4% 139.3% 38809 2.84 147.1% 69.8% -2% 0.693 1696 total 556190 26160 26274 99.6% 11.9% 12.2% 556190 21.71 12.2% 9.7% -5% 0.739 22619 Bernhard Rupp (Hofkristallrat a.D.) wrote: I think this suppression of high resolution shells via I/sigI cutoffs is partially attributable to a conceptual misunderstanding of what these (darn) R-values mean in refinement versus data merging. In refinement, even a random atom structure follows the Wilson distribution, and therefore, even a completely wrong non-centrosymmetric structure will not - given proper scaling - give an Rf of more than 59%. There is no such limit for the basic linear merging R. However, there is a simple relation between I/sigI and R-merge (provided no other indecency has been done to the data). It simply is (BMC) Rm=0.8/I/sigI. I.e. for I/sigI -0.8 you get 100%, for 2 we obtain 40%, which, interpreted as Rf would be dreadful, but for I/sigI 3, we get Rm=0.27, and that looks acceptable for an Rf (or uninformed reviewer). Btw, I also wish to point out that the I/sig cutoffs are not exactly the cutoff criterion for anomalous phasing, a more direct measure is a signal cutoff such as delF/sig(delF); George I believe uses 1.3 for SAD. Interestingly, in almost all structures I played with, delF/sig(delF) for both, noise in anomalous data or no anomalous scatterer present, the anomalous signal was 0.8. I haven’t figured out yet or proved the statistics and whether this is generally true or just numerology... And, the usual biased rant - irrespective of Hamilton tests, nobody really needs these popular unweighted linear residuals which shall not be named, particularly on F. They only cause trouble. Best regards, BR - Bernhard Rupp 001 (925) 209-7429 +43 (676) 571-0536 b...@ruppweb.org hofkristall...@gmail.com http://www.ruppweb.org/ - Structural Biology is the practice of crystallography without a license. - -Original Message- From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of Bart Hazes Sent: Thursday, March 03, 2011 7:08 AM To: CCP4BB@JISCMAIL.AC.UK Subject: Re: [ccp4bb] I/sigmaI of 3.0 rule There seems to be an epidemic of papers with I/Sigma 3 (sometime much larger). In fact such cases have become so frequent that I fear some people start to believe that this is the proper procedure. I don't know where that has come from as the I/Sigma ~ 2 criterion has been established long ago and many consider that even a tad conservative. It simply pains me to see people going to the most advanced synchrotrons to boost their highest resolution data and then simply throw away much of it. I don't know what has caused this wave of high I/Sigma threshold use but here are some ideas - High I/Sigma cutoffs are normal for (S/M)AD data sets where a more strict focus on data quality is needed. Perhaps some people have started to think this is the norm. - For some dataset Rsym goes up strongly while I/SigI
Re: [ccp4bb] I/sigmaI of 3.0 rule
I have to resend my statistics. Maia Cherney wrote: Dear Bernhard I am wondering where I should cut my data off. Here is the statistics from XDS processing. Maia On 11-03-03 04:29 AM, Roberto Battistutta wrote: Dear all, I got a reviewer comment that indicate the need to refine the structures at an appropriate resolution (I/sigmaI of3.0), and re-submit the revised coordinate files to the PDB for validation.. In the manuscript I present some crystal structures determined by molecular replacement using the same protein in a different space group as search model. Does anyone know the origin or the theoretical basis of this I/sigmaI3.0 rule for an appropriate resolution? Thanks, Bye, Roberto. Roberto Battistutta Associate Professor Department of Chemistry University of Padua via Marzolo 1, 35131 Padova - ITALY tel. +39.049.8275265/67 fax. +39.049.8275239 roberto.battistu...@unipd.it www.chimica.unipd.it/roberto.battistutta/ VIMM (Venetian Institute of Molecular Medicine) via Orus 2, 35129 Padova - ITALY tel. +39.049.7923236 fax +39.049.7923250 www.vimm.it DETECTOR_SU SUBSET OF INTENSITY DATA WITH SIGNAL/NOISE = -3.0 AS FUNCTION OF RESOLUTION RESOLUTION NUMBER OF REFLECTIONSCOMPLETENESS R-FACTOR R-FACTOR COMPARED I/SIGMA R-meas Rmrgd-F Anomal SigAno Nano LIMIT OBSERVED UNIQUE POSSIBLE OF DATA observed expected Corr 10.065509 304 364 83.5% 3.0% 4.4% 5509 63.83 3.1% 1.0%11% 0.652 173 7.12 11785 595 595 100.0% 3.5% 4.8% 11785 59.14 3.6% 1.4% -10% 0.696 414 5.81 15168 736 736 100.0% 5.0% 5.6% 15168 51.88 5.1% 1.8%-9% 0.692 561 5.03 17803 854 854 100.0% 5.5% 5.7% 17803 50.02 5.6% 2.2% -10% 0.738 675 4.50 20258 964 964 100.0% 5.1% 5.4% 20258 52.61 5.3% 2.1% -16% 0.710 782 4.11 223331054 1054 100.0% 5.6% 5.7% 22333 50.89 5.8% 2.0% -16% 0.705 878 3.80 233121137 1137 100.0% 7.0% 6.6% 23312 42.95 7.1% 3.0% -13% 0.770 952 3.56 253741207 1208 99.9% 7.6% 7.3% 25374 40.56 7.8% 3.4% -18% 0.7391033 3.35 270331291 1293 99.8% 9.7% 9.2% 27033 33.7310.0% 4.1% -12% 0.7651107 3.18 294881353 1353 100.0% 11.6% 11.6% 29488 28.1611.9% 4.4%-7% 0.7501176 3.03 310541419 1419 100.0% 15.7% 15.9% 31054 21.7716.0% 6.9%-9% 0.7411243 2.90 322881478 1478 100.0% 21.1% 21.6% 32288 16.9921.6% 9.2%-6% 0.7451296 2.79 338071542 1542 100.0% 28.1% 28.8% 33807 13.0728.8%12.9%-2% 0.7831361 2.69 349831604 1604 100.0% 37.4% 38.7% 349839.9538.3%17.2%-2% 0.7431422 2.60 351631653 1653 100.0% 48.8% 48.0% 351638.0350.0%21.9%-6% 0.7541475 2.52 366901699 1699 100.0% 54.0% 56.0% 366906.9855.3%25.9% 0% 0.7451517 2.44 377511757 1757 100.0% 67.9% 70.4% 377515.6169.5%32.5%-5% 0.7331577 2.37 384841798 1799 99.9% 82.2% 84.5% 384844.7284.2%36.5% 2% 0.7531620 2.31 390981842 1842 100.0% 91.4% 94.3% 390984.1993.7%43.7%-3% 0.7441661 2.25 388091873 1923 97.4% 143.4%139.3% 388092.84 147.1%69.8%-2% 0.6931696 total 556190 26160 26274 99.6% 11.9% 12.2% 556190 21.7112.2% 9.7%-5% 0.739 22619
[ccp4bb] [Fwd: Re: [ccp4bb] I/sigmaI of 3.0 rule]
Original Message Subject:Re: [ccp4bb] I/sigmaI of 3.0 rule Date: Thu, 03 Mar 2011 10:43:23 -0700 From: Maia Cherney ch...@ualberta.ca To: Oganesyan, Vaheh oganesy...@medimmune.com References: 2ba9ce2f-c299-4ca9-a36a-99065d1b3...@unipd.it 4d6faed8.7040...@ualberta.ca 021001cbd9bc$f0ecc940$d2c65bc0$@gmail.com 4d6fcab6.3090...@ualberta.ca 4d6fcbff.2010...@ualberta.ca 73e543de77290c409c9bed6fa4ca34bb0173a...@md1ev002.medimmune.com Vaheh, The problem was with Rmerg. As you can see at I/sigma=2.84, the Rmerge (R-factor) was 143%. I am asking this question because B. Rupp wrote However, there is a simple relation between I/sigI and R-merge (provided no other indecency has been done to the data). It simply is (BMC) Rm=0.8/I/sigI. Maybe my data are indecent? This is the whole LP file. Maia ** XSCALE (VERSION December 6, 2007)28-Aug-2009 ** Author: Wolfgang Kabsch Copy licensed until (unlimited) to Canadian Light Source, Saskatoon, Canada. No redistribution. ** CONTROL CARDS ** MAXIMUM_NUMBER_OF_PROCESSORS=8 SPACE_GROUP_NUMBER=180 UNIT_CELL_CONSTANTS= 150.1 150.1 81.8 90.0 90.0 120.0 OUTPUT_FILE=XSCALE.HKL FRIEDEL'S_LAW=TRUE INPUT_FILE= XDS_ASCII.HKL XDS_ASCII INCLUDE_RESOLUTION_RANGE= 40 2.25 THE DATA COLLECTION STATISTICS REPORTED BELOW ASSUMES: SPACE_GROUP_NUMBER= 180 UNIT_CELL_CONSTANTS= 150.10 150.1081.80 90.000 90.000 120.000 * 12 EQUIVALENT POSITIONS IN SPACE GROUP #180 * If x',y',z' is an equivalent position to x,y,z, then x'=x*ML(1)+y*ML( 2)+z*ML( 3)+ML( 4)/12.0 y'=x*ML(5)+y*ML( 6)+z*ML( 7)+ML( 8)/12.0 z'=x*ML(9)+y*ML(10)+z*ML(11)+ML(12)/12.0 #1 2 3 45 6 7 89 10 11 12 11 0 0 00 1 0 00 0 1 0 20 -1 0 01 -1 0 00 0 1 8 3 -1 1 0 0 -1 0 0 00 0 1 4 4 -1 0 0 00 -1 0 00 0 1 0 50 1 0 0 -1 1 0 00 0 1 8 61 -1 0 01 0 0 00 0 1 4 70 1 0 01 0 0 00 0 -1 8 8 -1 0 0 0 -1 1 0 00 0 -1 4 91 -1 0 00 -1 0 00 0 -1 0 100 -1 0 0 -1 0 0 00 0 -1 8 111 0 0 01 -1 0 00 0 -1 4 12 -1 1 0 00 1 0 00 0 -1 0 ALL DATA SETS WILL BE SCALED TO XDS_ASCII.HKL ** READING INPUT REFLECTION DATA FILES ** DATAMEAN REFLECTIONSINPUT FILE NAME SET# INTENSITY ACCEPTED REJECTED 1 0.6203E+03 557303 0 XDS_ASCII.HKL ** CORRECTION FACTORS AS FUNCTION OF IMAGE NUMBER RESOLUTION ** RECIPROCAL CORRECTION FACTORS FOR INPUT DATA SETS MERGED TO OUTPUT FILE: XSCALE.HKL THE CALCULATIONS ASSUME FRIEDEL'S_LAW= TRUE TOTAL NUMBER OF CORRECTION FACTORS DEFINED 720 DEGREES OF FREEDOM OF CHI^2 FIT140494.9 CHI^2-VALUE OF FIT OF CORRECTION FACTORS 1.037 NUMBER OF CYCLES CARRIED OUT 3 CORRECTION FACTORS for visual inspection with VIEW DECAY_001.pck INPUT_FILE=XDS_ASCII.HKL XMIN= 0.1 XMAX= 179.9 NXBIN= 36 YMIN= 0.00257 YMAX= 0.19752 NYBIN= 20 NUMBER OF REFLECTIONS USED FOR DETERMINING CORRECTION FACTORS 238321 ** CORRECTION FACTORS AS FUNCTION OF X (fast) Y(slow) IN THE DETECTOR PLANE ** RECIPROCAL CORRECTION FACTORS FOR INPUT DATA SETS MERGED TO OUTPUT FILE: XSCALE.HKL THE CALCULATIONS ASSUME FRIEDEL'S_LAW= TRUE TOTAL NUMBER OF CORRECTION FACTORS DEFINED 4760 DEGREES OF FREEDOM OF CHI^2 FIT186486.8 CHI^2-VALUE OF FIT OF CORRECTION
Re: [ccp4bb] I/sigmaI of 3.0 rule
I see, there is no consensus about my data. Some people say 2.4A, other say all. Well, I chose 2.3 A. My rule was to be a little bit below Rmerg 100%. At 2.3A Rmerg was 98.7% Actually, I have published my paper in JMB. Yes, reviewers did not like that and even made me give Rrim and Rpim etc. Maia Bernhard Rupp (Hofkristallrat a.D.) wrote: First of all I would ask a XDS expert for that because I don't know exactly what stats the XDS program reports (shame on me, ok) nor what the quality of your error model is, or what you want to use the data for (I guess refinement - see Eleanor's response for that, and use all data). There is one point I'd like to make re cutoff: If one gets greedy and collects too much noise in high resolution shells (like way below I/sigI = 0.8 or so) the scaling/integration may suffer from an overabundance of nonsense data, and here I believe it makes sense to select a higher cutoff (like what exactly?) and reprocess the data. Maybe one of our data collection specialist should comment on that. BR -Original Message- From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of Maia Cherney Sent: Thursday, March 03, 2011 9:13 AM To: CCP4BB@JISCMAIL.AC.UK Subject: Re: [ccp4bb] I/sigmaI of 3.0 rule I have to resend my statistics. Maia Cherney wrote: Dear Bernhard I am wondering where I should cut my data off. Here is the statistics from XDS processing. Maia On 11-03-03 04:29 AM, Roberto Battistutta wrote: Dear all, I got a reviewer comment that indicate the need to refine the structures at an appropriate resolution (I/sigmaI of3.0), and re-submit the revised coordinate files to the PDB for validation.. In the manuscript I present some crystal structures determined by molecular replacement using the same protein in a different space group as search model. Does anyone know the origin or the theoretical basis of this I/sigmaI3.0 rule for an appropriate resolution? Thanks, Bye, Roberto. Roberto Battistutta Associate Professor Department of Chemistry University of Padua via Marzolo 1, 35131 Padova - ITALY tel. +39.049.8275265/67 fax. +39.049.8275239 roberto.battistu...@unipd.it www.chimica.unipd.it/roberto.battistutta/ VIMM (Venetian Institute of Molecular Medicine) via Orus 2, 35129 Padova - ITALY tel. +39.049.7923236 fax +39.049.7923250 www.vimm.it
Re: [PyMOL] Selecting ASP and GLU
For me, color red, resn asp+glu works. Maia Martin Hediger wrote: Dear all What is the selection syntax to select all GLU and ASP residues within an object? I'm trying it the way its written on the wiki: remove resn hoh# remove water h_add # add hydrogens as surface color grey90 color slate, resn lys # lysines in light blue color paleyellow, resn cys # cysteines in light yellow *color tv_red, (resn asp or(resn glu)) # aspartic and glutamic acid in light red* but, the selection kind of does not work for me (I'm assuming the operator for the logical AND is 'and'). What is it that I need to do differently? Kind regards Martin -- Free Software Download: Index, Search Analyze Logs and other IT data in Real-Time with Splunk. Collect, index and harness all the fast moving IT data generated by your applications, servers and devices whether physical, virtual or in the cloud. Deliver compliance at lower cost and gain new business insights. http://p.sf.net/sfu/splunk-dev2dev ___ PyMOL-users mailing list (PyMOL-users@lists.sourceforge.net) Info Page: https://lists.sourceforge.net/lists/listinfo/pymol-users Archives: http://www.mail-archive.com/pymol-users@lists.sourceforge.net -- Free Software Download: Index, Search Analyze Logs and other IT data in Real-Time with Splunk. Collect, index and harness all the fast moving IT data generated by your applications, servers and devices whether physical, virtual or in the cloud. Deliver compliance at lower cost and gain new business insights. http://p.sf.net/sfu/splunk-dev2dev ___ PyMOL-users mailing list (PyMOL-users@lists.sourceforge.net) Info Page: https://lists.sourceforge.net/lists/listinfo/pymol-users Archives: http://www.mail-archive.com/pymol-users@lists.sourceforge.net
[ccp4bb] [Fwd: Re: [ccp4bb] Space group and R/Rfree value]
Original Message Subject:Re: [ccp4bb] Space group and R/Rfree value Date: Wed, 01 Dec 2010 09:44:06 -0700 From: Maia Cherney ch...@ualberta.ca To: Xiaopeng Hu huxp...@mail.sysu.edu.cn References: 643947201.129232.1291191478190.javamail.r...@zmbx0.sysu.edu.cn A dimer could be with a symmetry-related molecule in C2221, so there is no need to have a dimer in the asym. unit. Maia Xiaopeng Hu wrote: Dear all, I am working on a data-set (2.3A) and the space group problem bothers me a lot.The space group of the data-set could be C2221 or P212121, since our protein functions as a dimer, and P212121 gives two molecular in the asym-uint, I think P212121 is more reasonable than C2221.However with C2221, I can refine the R/Rfree to 20/24 or lower, while with P212121 only to 26/30. Also Phenix points out that the crystal is probably a twin with P212121 but is OK with C2221. I am not a real crystallographer, perhaps this problem is stupid, any help will be appreciated!! Best wishes, Xiaopeng Hu
Re: [ccp4bb] diverging Rcryst and Rfree
I found a practical solution to a similar problem. When I get large gap between Rf/R in refmac I repeat the refinement in PHENIX using the same model and the same mtz file, It has always worked for me. And I have no theory for that observation, but the tables in publications looked better. Maia Quoting Ian Tickle ianj...@gmail.com: Jackie I agree completely with Ed (for once!), not only for the reasons he gave, but also that it's valid to compare statistics such as likelihood and R factors ONLY if only the model is varied. Such a comparison is not valid if the data used are varied (in this case you are changing the data by deleting some of them). Cheers -- Ian On Tue, Oct 26, 2010 at 2:37 PM, Ed Pozharski epozh...@umaryland.edu wrote: Jackie, please note that (at least imho) the desire to obtain better R-factors does not justify excluding data from analysis. Weak reflections that you suggest should be rejected contain information, and excluding them will indeed artificially lower the R-factors while reducing the accuracy of your model. Cheers, Ed. On Mon, 2010-10-25 at 17:44 -0400, Jacqueline Vitali wrote: Also if your Rmerge is high and you include all reflections in refinement, Rfree is high. In my experience, by excluding F sigma reflections you drop Rfree a lot. -- I'd jump in myself, if I weren't so good at whistling. Julian, King of Lemurs
Re: [ccp4bb] diverging Rcryst and Rfree
I had a similar problem. It dissappeared when I switched the refinement to phenix. The R factors dropped and the difference between them became acceptable.
Re: [PyMOL] map lipophilic potential
Program Vasco can do it. Sebastian Kruggel wrote: dear all, i am looking for a possibility to map lipophilic potential on pymol generated surfaces. i didn't find anything in the documentation and in the mailing list the question came up once http://www.mail-archive.com/pymol-users@lists.sourceforge.net/msg03459.html but there was no reply. maybe anybody has an idea? thanks in advance, sebastian -- Start uncovering the many advantages of virtual appliances and start using them to simplify application deployment and accelerate your shift to cloud computing http://p.sf.net/sfu/novell-sfdev2dev ___ PyMOL-users mailing list (PyMOL-users@lists.sourceforge.net) Info Page: https://lists.sourceforge.net/lists/listinfo/pymol-users Archives: http://www.mail-archive.com/pymol-users@lists.sourceforge.net
Re: [ccp4bb] Molecular replacement question
Try balbes from G. Murshudov's website. It will find proper search model and use proper truncations automatically. In addition, it will put in your sequence. Paul Holland wrote: Hello fellow crystallographers, I am trying molecular replacement for a protein crystal dataset that has very high sequence similarity to the search model with several predicted flexible loop regions; however, all attempts at finding a solution have not produce very ideal starting solutions using Phaser and Molrep (CC = 0.3 and Z-score = 5). I am very confident that the unit cell parameters are C2 84.027 120.565 108.272 90.00 104.71 90.00, and there appears to be no evidence of twinning. The Matthews calculation predicts from anywhere from 2-4 monomers in the ASU, and calculation of the SRF in Molrep does not identify any peaks in higher order symmetry except for the expected crystallographic two-fold for C2. Below is the table from the calculated SRF in molrep. Any advice would be greatly appreciated. #thetaphichi alpha beta gamma Rf Rf/sigma Sol_RF 1 0.000.000.000.000.000.00 870.5 21.59 Sol_RF 258.61 -10.17 180.00 169.83 -117.23 10.17 162.5 4.03 Sol_RF 366.02 -0.00 180.00 180.00 -132.030.00 161.1 4.00 Sol_RF 458.42 -9.54 180.00 170.46 -116.859.54 159.8 3.96 Sol_RF 5 149.840.00 180.00 -180.00 60.320.00 156.0 3.87 Sol_RF 658.96 -5.52 180.00 174.48 -117.915.52 151.5 3.76 Sol_RF 765.59 20.95 180.00 20.95 131.18 159.05 143.9 3.57 Sol_RF 890.00 -98.96 180.000.00 180.00 17.92 142.9 3.55 Sol_RF 956.53 15.78 180.00 15.78 113.07 164.22 142.0 3.52 Sol_RF 1071.10 -19.94 180.00 160.06 -142.20 19.94 141.6 3.51 Sol_RF 1171.28 29.78 180.00 29.78 142.55 150.22 140.4 3.48 Sol_RF 1265.22 -15.88 180.00 164.12 -130.44 15.88 139.2 3.45 Sol_RF 1368.84 -0.00 180.00 180.00 -137.670.00 138.0 3.42 Sol_RF 1432.51 -180.00 180.00 -180.00 65.02 -0.00 137.9 3.42 Sol_RF 1575.02 -28.84 180.00 151.16 -150.04 28.84 134.7 3.34 Sol_RF 1671.69 -20.99 180.00 159.01 -143.37 20.99 133.0 3.30 Sol_RF 1792.13 101.46 179.93 102.35 -175.74 79.42 130.9 3.25 Sol_RF 18 107.89 144.73 179.79 145.06 -144.22 35.61 128.8 3.19 Sol_RF 1987.45 -78.19 180.00 101.81 -174.90 78.19 128.1 3.18 Sol_RF 2038.570.69 30.36 102.66 -18.79 -78.71 122.4 3.04 Sol_RF 2126.77 174.59 176.58 172.68 53.523.49 120.5 2.99 Sol_RF 22 116.66 178.08 175.143.49 126.48 187.32 120.5 2.99 Sol_RF 2375.56 -41.35 180.00 138.65 -151.12 41.35 119.8 2.97 Sol_RF 2466.12 36.35 180.00 36.35 132.24 143.65 116.6 2.89 Sol_RF 2583.87 71.62 180.00 71.62 167.74 108.38 114.7 2.85 Sol_RF 2669.24 -12.37 180.00 167.63 -138.48 12.37 112.3 2.79 Sol_RF 2759.75 15.26 172.297.64 119.07 157.12 112.2 2.78 Sol_RF 28 120.25 -164.74 172.29 22.88 119.07 172.36 112.2 2.78 Sol_RF 2996.68 -70.99 180.00 109.01 -166.63 70.99 110.9 2.75 Sol_RF 3063.23 -44.73 180.00 135.27 -126.47 44.73 108.9 2.70 Cheers, Paul Holland
Re: [ccp4bb] monomer-dimer
Thank you. Now I understand the difference. I thought there was separation. Maia Xuewu Zhang wrote: Hi Maia, I have seen your post regarding this before and I just want to point out that you may have confused AUC (analytical ultracentrifugation) with gradient-based ultra-centrifugation methods for separating macromolecules. AUC does not involve separation of different species in the sample. There are two types of AUCs: sedimentation velocity and sedimentation equilibrium. In sedimentation equilibrium experiments, the system reaches the equilibrium at the end, and the monomer/dimer ratio, Kd, etc parameters can be worked out by fitting the data to a model globally. The shape of the molecule does not matter. For starters: http://en.wikipedia.org/wiki/Ultracentrifuge Xuewu Zhang On Wed, Aug 11, 2010 at 10:37 AM, chern ch...@ualberta.ca mailto:ch...@ualberta.ca wrote: Hi Anastassis, We are back to the same argument that AUC is not a good method. As everyone knows, it's a dynamic equilibrium between monomers and dimers that exists before separation. Once you started separation in any method, the equilibrium is disturbed now in each separated band. That will cause re-equilibration and constant migration of newly formed dimers from the monomer band and newly formed monomers from the dimer band. The t(eq) is the re-equilibration time. Your method of separation of monomers and dimers should be quick enough before any re-equilibration occurs (t(sep)t(eq)). Otherwise, you get a mess and smearing of bands. Also, most conventional methods depend on shape etc. I find SEC is most convenient. Maia - Original Message - *From:* Anastassis Perrakis mailto:a.perra...@nki.nl *To:* chern mailto:ch...@ualberta.ca *Sent:* Monday, July 05, 2010 2:38 PM *Subject:* Re: [ccp4bb] monomeric coiled coil--updated On 5 Jul 2010, at 22:04, chern wrote: Hi, Anastassis If you had just a monomer at the start time then t(eq) is the time to get to equilibrium with the dimer and vice versa. sorry to say but the definition of that time in a biophysical sense, is in my opinion equal to infinity and cannot be defined. I am being a bit pedantic here, but I am just saying that t(eq) cannot be defined, it can be approximated, and thus t(eq) is wrong to define. Why not talk about kD and kON and kOFF that have robust definitions based on kinetic properties and a physical meaning? When you separated the two bands (monomers and dimers) in AUC, and then the equilibrium is quickly established in each band again what's the point? So, to be successful in this method, you need to have t(eq) much lower than the separation run. Ideally, if you could separate monomers and dimers instantly and freeze them in the separated state, then you can have good estimate of the both fractions. I think this is clear. But, I disagree and I think what you say is wrong. The equilibrium is dynamic. Why do you insist there is a point in 'separation'? The monomer changes to a dimer and vise versa in a continuous fashion. All you can say is that in a given concentration the equilibrium is shifted towards one or the other form. But its a dynamic one. Even at a concentration which is 50-50 between two states, the molecules that are in one state or another are changing according to kinetic parameters that are characteristic for the complex. Even at 100% - lets say of a dimer - by your definition, (100% cannot exist since its reached asymptotically by any derivation about equilibriums) molecules will fall to monomer and will reassemble to a dimer rapidly. To be honest I think that talking about t(eq) is largely wrong in biophysical terms, since it does not exist. A. That's what I meant. Maia - Original Message - *From:* Anastassis Perrakis mailto:a.perra...@nki.nl *To:* chern mailto:ch...@ualberta.ca *Sent:* Monday, July 05, 2010 11:45 AM *Subject:* Re: [ccp4bb] monomeric coiled coil--updated On 5 Jul 2010, at 19:30, chern wrote: Thank you for reply. 1.It will be nice to have mass-spec method for non-covalent complexes. Carol Robinson is doing these
Re: [ccp4bb] monomer-dimer
Hi ccp4bb Could you please send me some references with the sedimentation equilibrium calculations of Kd, monomer/dimer ratio etc. Maia Maia Cherney wrote: Thank you. Now I understand the difference. I thought there was separation. Maia Xuewu Zhang wrote: Hi Maia, I have seen your post regarding this before and I just want to point out that you may have confused AUC (analytical ultracentrifugation) with gradient-based ultra-centrifugation methods for separating macromolecules. AUC does not involve separation of different species in the sample. There are two types of AUCs: sedimentation velocity and sedimentation equilibrium. In sedimentation equilibrium experiments, the system reaches the equilibrium at the end, and the monomer/dimer ratio, Kd, etc parameters can be worked out by fitting the data to a model globally. The shape of the molecule does not matter. For starters: http://en.wikipedia.org/wiki/Ultracentrifuge Xuewu Zhang On Wed, Aug 11, 2010 at 10:37 AM, chern ch...@ualberta.ca mailto:ch...@ualberta.ca wrote: Hi Anastassis, We are back to the same argument that AUC is not a good method. As everyone knows, it's a dynamic equilibrium between monomers and dimers that exists before separation. Once you started separation in any method, the equilibrium is disturbed now in each separated band. That will cause re-equilibration and constant migration of newly formed dimers from the monomer band and newly formed monomers from the dimer band. The t(eq) is the re-equilibration time. Your method of separation of monomers and dimers should be quick enough before any re-equilibration occurs (t(sep)t(eq)). Otherwise, you get a mess and smearing of bands. Also, most conventional methods depend on shape etc. I find SEC is most convenient. Maia - Original Message - *From:* Anastassis Perrakis mailto:a.perra...@nki.nl *To:* chern mailto:ch...@ualberta.ca *Sent:* Monday, July 05, 2010 2:38 PM *Subject:* Re: [ccp4bb] monomeric coiled coil--updated On 5 Jul 2010, at 22:04, chern wrote: Hi, Anastassis If you had just a monomer at the start time then t(eq) is the time to get to equilibrium with the dimer and vice versa. sorry to say but the definition of that time in a biophysical sense, is in my opinion equal to infinity and cannot be defined. I am being a bit pedantic here, but I am just saying that t(eq) cannot be defined, it can be approximated, and thus t(eq) is wrong to define. Why not talk about kD and kON and kOFF that have robust definitions based on kinetic properties and a physical meaning? When you separated the two bands (monomers and dimers) in AUC, and then the equilibrium is quickly established in each band again what's the point? So, to be successful in this method, you need to have t(eq) much lower than the separation run. Ideally, if you could separate monomers and dimers instantly and freeze them in the separated state, then you can have good estimate of the both fractions. I think this is clear. But, I disagree and I think what you say is wrong. The equilibrium is dynamic. Why do you insist there is a point in 'separation'? The monomer changes to a dimer and vise versa in a continuous fashion. All you can say is that in a given concentration the equilibrium is shifted towards one or the other form. But its a dynamic one. Even at a concentration which is 50-50 between two states, the molecules that are in one state or another are changing according to kinetic parameters that are characteristic for the complex. Even at 100% - lets say of a dimer - by your definition, (100% cannot exist since its reached asymptotically by any derivation about equilibriums) molecules will fall to monomer and will reassemble to a dimer rapidly. To be honest I think that talking about t(eq) is largely wrong in biophysical terms, since it does not exist. A. That's what I meant. Maia - Original Message - *From:* Anastassis Perrakis mailto:a.perra...@nki.nl *To:* chern mailto:ch...@ualberta.ca *Sent:* Monday, July 05, 2010 11:45 AM *Subject:* Re: [ccp4bb] monomeric coiled coil--updated On 5 Jul 2010, at 19:30, chern wrote: Thank you for reply
Re: [ccp4bb] monomer-dimer
To determine the oligomeric state of a protein (monomer or dimer in your case), it's useful to use the PISA server. You upload your pdb file from the crystal structure.The server calculates the areas of interfaces (buried area) and deltaG (change in Gibbs energy) upon oligomer dissociation. (E. Krissinel and K. Henrick (2007). /Inference of macromolecular assemblies from crystalline state/. J. Mol. Biol. *372*, 774--797 . E. Krissinel and K. Henrick (2005). /Detection of Protein Assemblies in Crystals/. In: M.R. Berthold /et.al./ (Eds.): CompLife 2005, LNBI 3695, pp. 163--174 http://dx.doi.org/10.1007/11560500_15. E. Krissinel (2009). /Crystal contacts as nature's docking solutions/. J. Comp. Chem., in press; published on-line 6 May 2009; DOI 10.1002/jcc.21303} If the interface area (divided by 2 per one protomer) is greater than 1000 A2 and delta G is more than 5kcal/mol (the higher the better), it's a dimer. However, don't forget that most dimers can dissociate into monomers upon dilution. There is a dynamic equilibrium between dimers (oligomers) and monomers that depends on their concentration and the Kdiss. Separating them in any method will disturb this equilibrium. If the re-equilibration time is greater than the separation time, you can see both monomers and dimers. You can even roughly calculate the dissociation constant: Kdiss=[monomer]2/[dimer] where brackets mean concentrations. To give you an estimate, at Kdiss=10(-3)M, you have roughly equal concentration of dimers and monomers at 10-3 M and only 10% dimers at 10-4 M. Sometimes, protein needs to dissociate easily for the biological function. Maia intekhab alam wrote: Hi everyone Sorry for some non specific query! i am working with a protein that shows a dimer in the crystal structure but when i tried to figure out that with standard molecular markers in gel filteration (superdex-200, 24ml column) it turned out to be a monnomer. Native gel analysis after incubating the protein at 20 degree, 37 degree showed more dimer at 20 degree celcius as compared to 37. I tried similar strategy in gel filteration by incubating my protein at various temperature,where a lot of precipitation was observed at 37 degree celcius and after removing the precipitates i run the gel filteration that has 0.5 ml higher elution volume as compared to samples incubated at 20 degree celcius and 4 degree celcius.( Is this significant) Furthermore i have done some experiments in cold room (4 degree) where the elution volume is stuck at a point irrespective of the conditions (as Flow rate, concentration of protein etc) and that is higher than that of the room temperature by 1 ml. Standard moleculr weight markers also show higher elution volume in cold room in comparison to the room temperature by 1 ml. I will be highly obliged if someone suggest some literature or any otherway to do gel filtrtaion so that i can clearly resolve this issue. Also let me know if there is some literature available on effect of temperature on the elution volume of proteins. Thanks in advance -- INTEKHAB ALAM LABORATORY OF STRUCTURAL BIOINFORMATICS KOREA UNIVERSITY, SEOUL
Re: [PyMOL] pymol dies when minimizing
Hi Jason, I had my problem in ubuntu, not windows. I could open pymol only from the education menu (not from a terminal). I wrote several e-mails to pymol list, and nothing helped me. Then incidentally I upgraded my ubuntu from 8.04 to 9.04 and several things started working automatically, such as pymol started to open from a terminal window. Of course, some other programs stopped working etc. Now everything is ok. The pymol version that I have has come with ubuntu package, it's Version 1.1r2pre. Works fine. Maia Jason Vertrees wrote: Maia and Robin, What version of Windows and PyMOL are you using? Cheers, -- Jason On Wed, Jun 2, 2010 at 10:52 AM, Maia Cherney ch...@ualberta.ca wrote: I had once a problem with pymol that itr would not open from a terminal window. After a long battle with it, the problem was solved after I upgraded my OS. Robin Emig wrote: Does anyone else have the problem where pymol dies, or wont update the display after minimizing on windows? Is there a way to fix this? -Robin -- ___ PyMOL-users mailing list (PyMOL-users@lists.sourceforge.net) Info Page: https://lists.sourceforge.net/lists/listinfo/pymol-users Archives: http://www.mail-archive.com/pymol-users@lists.sourceforge.net -- ___ PyMOL-users mailing list (PyMOL-users@lists.sourceforge.net) Info Page: https://lists.sourceforge.net/lists/listinfo/pymol-users Archives: http://www.mail-archive.com/pymol-users@lists.sourceforge.net -- ThinkGeek and WIRED's GeekDad team up for the Ultimate GeekDad Father's Day Giveaway. ONE MASSIVE PRIZE to the lucky parental unit. See the prize list and enter to win: http://p.sf.net/sfu/thinkgeek-promo ___ PyMOL-users mailing list (PyMOL-users@lists.sourceforge.net) Info Page: https://lists.sourceforge.net/lists/listinfo/pymol-users Archives: http://www.mail-archive.com/pymol-users@lists.sourceforge.net
Re: [PyMOL] pymol dies when minimizing
I had once a problem with pymol that itr would not open from a terminal window. After a long battle with it, the problem was solved after I upgraded my OS. Robin Emig wrote: Does anyone else have the problem where pymol dies, or wont update the display after minimizing on windows? Is there a way to fix this? -Robin -- ___ PyMOL-users mailing list (PyMOL-users@lists.sourceforge.net) Info Page: https://lists.sourceforge.net/lists/listinfo/pymol-users Archives: http://www.mail-archive.com/pymol-users@lists.sourceforge.net -- ___ PyMOL-users mailing list (PyMOL-users@lists.sourceforge.net) Info Page: https://lists.sourceforge.net/lists/listinfo/pymol-users Archives: http://www.mail-archive.com/pymol-users@lists.sourceforge.net
Re: [PyMOL] coordination bonds
I would like to show coordination bonds between Mg ion and its ligands (oxygens) as broken lines, the distance of Mg-O bond is between 1.9-2.2 A. Maia -- ___ PyMOL-users mailing list (PyMOL-users@lists.sourceforge.net) Info Page: https://lists.sourceforge.net/lists/listinfo/pymol-users Archives: http://www.mail-archive.com/pymol-users@lists.sourceforge.net
[ccp4bb] [Fwd: Is it possible for the Tris buffer to strip the Zn ions from the Zinc Finger motif of a protein?]
Original Message Subject: Is it possible for the Tris buffer to strip the Zn ions from the Zinc Finger motif of a protein? Date: Sun, 23 May 2010 08:45:55 -0600 From: Maia Cherney ch...@ualberta.ca To: ruheng rh_ibp2...@hotmail.com The complex can dissociate without loosing Zn. For example, if you dilute it too much. There is an equilibrium between the concentrations of complex and the individual components. Maia Date: Sat, 22 May 2010 11:17:41 +0800 From: rh_ibp2...@hotmail.com Subject: [ccp4bb] To: CCP4BB@JISCMAIL.AC.UK Dear all, Recently, I am working on a complex which includes two protein subunits. The interaction was based on the Zinc Finger motif of one protein. I co-purified the complex by nickel affinity column with one protein bearing a C terminal His tag and the other without any affinity tags. However, the complex was disassociated when applied to size exclusion chromatography. The buffer I use for SEC is 20mM Tris-HCl, 150mM NaCl, 1mM DTT, 5% Glycerol, pH 7.5, whearas the buffer I use for nickel affinity column is 50mM Na2HPO4, 10mM KH2PO4, 137mM NaCl, 2.7mM KCl, 10% Glycerol, pH7.4. So I am wondering is it possible for the Tris buffer to strip the Zn ions from the Zinc Finger motif of one protein that leads to the destruction of the complex? I will be very appreciated if anyone has some experience in such case and would like to share with me! Sincerely, Heng Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China 搜索本应是彩色的,快来体验新一代搜索引擎-必应,精美图片每天换哦! 立即试 用！ http://cn.bing.com/?form=CRMADS 聊天+搜索+邮箱 想要轻松出游,手机MSN帮你搞定! 立刻下载！ http://3g.msn.cn/
Re: [ccp4bb] Finding best model for molecular replacement
Try balbes. It needs only your sequence and your mtz file. http://www.ysbl.york.ac.uk/~fei/balbes/ Maia Paul Lindblom wrote: Hi everybody, I just crystallized a new project protein. How can I find a possible model for using molecular replacement? I have the sequence of my protein. Is it enough to make a sequence search in the pdb? Or is there another approach I can use? Thanks a lot, Paul
[PyMOL] [Fwd: Re: Symmetry Mates Problem]
Docking is very non-reliable. E. Krissinel (2009). /Crystal contacts as nature's docking solutions/. J. Comp. Chem., in press; published on-line 6 May 2009; DOI 10.1002/jcc.21303 Maia humayun scherrif wrote: Hello, Thank you for detailed explanation, surely it is helping me to sort out the possibilities. As per your query a) There are many references that the protein is a Hexamer, but I am considering, because the domain which I have got structure, interacts with other proteins to make a biological complex, their interaction could be important for biological hexamerization of the whole complex ( those interacting proteins also exist as hexamer in complex with my protein ) b) I coudnt find any hexameric homologue (although there are some good homologue structures but they mostly exist as dimer or monomer) c) the structure is not yet been solved and not reported as yet. So according your reply, does that mean the only possibility left is docking ? because others are not working for me at all. Thank you again for suggestions. On Wed, May 19, 2010 at 6:31 PM, Tsjerk Wassenaar tsje...@gmail.com mailto:tsje...@gmail.com wrote: Hi Humayun, Crystallograpic symmetries are often not of much help to construct biologically relevant complexes. Do you have (a) a reference of the hexameric structure, or (b) of a hexameric homologue, or (c) is it only known to form hexamers and is the structure still unsolved? In case of (a), the structure is likely to have a recipe to build the biological unit (possibly as REMARK 350 in the PDB file). In case of (b), you can try to fit copies of the structure onto each chain of the homologue, being aware that that will give you a crude approximation as starting point for further work. And in case of (c), you might want to consider doing some docking. Hope it helps, Tsjerk On Wed, May 19, 2010 at 10:26 AM, humayun scherrif hum@gmail.com mailto:hum@gmail.com wrote: Thank you all for the replies. The protein itself makes hexamer which is well documented and proved structural evidence from other cytoplasmic domains ( my structure is also a domain). I have run PISA, but the online PISA server didnt give me output like standalone PISA in CCP4 (result is mentioned below). Online PISA results show that there are not significant dimer interfaces and thus the trimer structure is because of only crystal packing result For homology modeling I didnt get any proper homologs which have hexameric assembly (I@ Bryn: I cant send you PDB id since its not submitted yet) Analysis of protein interfaces suggests that the following quaternary structures are stable in solution (I wonder the DGdiss is positive value, is it significant to make Hexamer assembly because I couldnt find any help to find out about the allowed values) .-.---.--- Set | No | Size Id ASA BSADGdiss | Formula +-+---+--- 1 | 1 | 60 19917.75536.3 3.8 | A(2)B(2)C(2) +-+---+--- 2 | 2 | 31 10722.92004.1 6.2 | ABC +-+---+--- 3 | 3 | 42 14004.23014.9 0.5 | A(2)B(2) | 4 | 134217.5 0.0 -0.0 | A +-+---+--- 4 | 5 | 247506.21003.3 7.0 |AB | 6 | 134217.5 0.0-0.0 |A +-+---+--- 5 | 7 | 257443.81000.8 6.8 | AB | 8 | 164282.4 0.0 -0.0 | A +-+---+--- 6 | 9 | 277556.51008.3 2.0 | A(2) | 10 | 184227.1 0.0 -0.0 |A | 11 | 134217.5 0.0 -0.0 |A '-'---'--- Waiting for your reply Thanks H On Wed, May 19, 2010 at 4:41 PM, Robert Brynmor Fenwick robert.fenw...@irbbarcelona.org mailto:robert.fenw...@irbbarcelona.org wrote: Also, if you would like to try homology modelling then that could work. However you would need a couple of hexamer strucutres to start with. It would probably take some tinkering with current tools. I would probably use an MD approach
Re: [ccp4bb] Is it possible to mutate a reversible epimerase into an inreversible one?
You absolutely right, I thought about it. Maia Marius Schmidt wrote: Interestingly, Maxwell's demon pops up here, wh... , don't do it. If you change the reaction rate in one direction 1000 times slower than in the other direction, then the reaction becomes practically irreversible. And the system might not be at equilibrium. Maia R. M. Garavito wrote: Vinson, As Dale and Randy pointed out, you cannot change the #916G of a reaction by mutation: enzyme, which is a catalyst, affects only the activation barrier (#916E double-dagger). You can just make it a better (or worse) catalyst which would allow the reaction to flow faster (or slower) towards equilibrium. Nature solves this problem very elegantly by taking a readily reversible enzyme, like an epimerase or isomerase, and coupling it to a much less reversible reaction which removes product quickly. Hence, the mass action is only in one direction. An example of such an arrangement is the triose phosphate isomerase (TIM)-glyceraldehyde 3-phosphate dehydrogenase (GAPDH) reaction pair. TIM is readily reversible (DHA = G3P), but G3P is rapidly converted to 1,3-diphosphoglycerate by GAPDH. The oxidation and phosphorylation reactions of GAPDH now make TIM work in one direction. Since many epimerases are very optimized enzymes, why not consider making a fusion with a second enzyme (like a reductase) to make the system flow in one direction. Of course, this depends on what you want to do with the product. Cheers, Michael // /R. Michael Garavito, Ph.D./ /Professor of Biochemistry Molecular Biology/ /513 Biochemistry Bldg. / /Michigan State University / /East Lansing, MI 48824-1319/ /Office:// //(517) 355-9724 Lab: (517) 353-9125/ /FAX: (517) 353-9334Email: rmgarav...@gmail.com mailto:garav...@gmail.com/ // On May 18, 2010, at 11:54 AM, Dale Tronrud wrote: Hi, I'm more of a Fourier coefficient kind of guy, but I thought that a #916G of zero simply corresponded to an equilibrium constant of one. You can certainly have reversible reactions with other equilibrium constants. In fact I think irreversible reactions are simply ones where the equilibrium constant is so far to one side that, in practice, the reaction always goes all the way to product. As Randy pointed out the enzyme cannot change the #916G (or the equilibrium constant). You could drive a reaction out of equilibrium by coupling it to some other reaction which itself is way out of equilibrium (such as ATP hydrolysis in the cell) but I don't think that's a simple mutation of your enzyme. ;-) Dale Tronrud On 05/18/10 00:31, Vinson LIANG wrote: Dear all, Sorry for this silly biochemistory question. Thing is that I have a reversible epimerase and I want to mutate it into an inreversible one. However, I have been told that the #916G of a reversible reaction is zero. Which direction the reaction goes depends only on the concentration of the substrate. So the conclusion is, A: I can mutate the epimerase into an inreversible one. But it has no influence on the reaction direction, and hence it has little mean. B: There is no way to change a reversible epimerase into an inversible one. Could somebody please give me some comment on the two conclution? Thank you all for your time. Best, Vinson Dr.habil. Marius Schmidt Asst. Professor University of Wisconsin-Milwaukee Department of Physics Room 454 1900 E. Kenwood Blvd. Milwaukee, WI 53211 phone: +1-414-229-4338 email: m-schm...@uwm.edu http://users.physik.tu-muenchen.de/marius/
Re: [ccp4bb] Native Gel Theory and Practice
That's interesting. Thanks. Maia Nadir T. Mrabet wrote: Maia speaks about native PAGE for which protein mobility (migration) depends on 3 different parameters as she states: charge, mass and shape. Blue native PAGE, which might be the answer to Jacob's question, is a 2D gel: Native in the first direction, then SDS-PAGE in the second one. You actually need both data to infer stoechiometry and subunit composition. Nadir Pr. Nadir T. Mrabet Structural Molecular Biochemistry Nutrigenex - INSERM U-954 Nancy University, School of Medicine 9, Avenue de la Foret de Haye, BP 184 54505 Vandoeuvre-les-Nancy Cedex France Phone: +33 (0)3.83.68.32.73 Fax: +33 (0)3.83.68.32.79 E-mail: Nadir.Mrabetat medecine.uhp-nancy.fr On 19/05/2010 13:01, Jürgen Bosch wrote: Not quite correct, look into Blue Native PAGE. There you can seperate natively by mass. Jürgen .. Jürgen Bosch Johns Hopkins Bloomberg School of Public Health Department of Biochemistry Molecular Biology Johns Hopkins Malaria Research Institute 615 North Wolfe Street, W8708 Baltimore, MD 21205 Phone: +1-410-614-4742 Lab: +1-410-614-4894 Fax: +1-410-955-3655 http://web.mac.com/bosch_lab/ On May 19, 2010, at 1:31, Maia Cherney ch...@ualberta.ca wrote: Dear Jacob, I offer you my opinion. Are you talking about electrophoresis? As far as I know it does not work for the mass. The velocity of a protein depends on the charge at a particular pH, the mass and shape of molecules etc. It's very difficult to take all these things into consideration. Otherwise this would be a very convenient method, much easier than the analytical centrifugation or gel-filtration that are usually used. However, electrophoresis does not work for mass determination. Besides, complex formation hugely depends on the protein concentration. If you dilute your mixture, your complexes might dissociate. There is equilibrium constant between different types of complexes. Maia Jacob Keller wrote: Dear Crystallographers, I am trying to optimize a native gel experiment of a two-protein complex, running the smallest-detectable amount of protein component A with varying amounts of component B. MWCharge MW/Charge A 22 -5-4308 B 17-24 -702 This experiment is partly to determine stoichiometry, but also to determine roughly the strength of the interaction. B definitely runs much faster than A alone, as predicted, but I am wondering what to expect with various oligomers. Should ABB run faster or slower than AB? What about AABB? Theoretically, AA should certainly run slower than A, and BB slower than B, simply because the mass/charge ratio is the same, but the overall mass is greater. But what happens when you have AAB, for example? There must be an equation relating the mass/charge and mass (and perhaps gel percentage) to the speed traveled in the gel--but what is the equation? Thanks for your consideration, Jacob *** Jacob Pearson Keller Northwestern University Medical Scientist Training Program Dallos Laboratory F. Searle 1-240 2240 Campus Drive Evanston IL 60208 lab: 847.491.2438 cel: 773.608.9185 email: j-kell...@northwestern.edu ***
Re: [ccp4bb] Is it possible to mutate a reversible epimerase into an inreversible one?
I think that it's possible to do a mutation that affects only one way of the reaction. You can mutate a residue that makes contacts only with the product of the direct way or only of the reverse way. Maia Randy Read wrote: Dear Vinson, I would agree with you on choice B. There are probably many ways to look at it. Here are two that come to me at the moment. 1. If the reaction is reversible, then there's no opportunity to put energy into the system to reduce its overall entropy. So a reversible epimerase would be like a Maxwell's demon, violating the second law of thermodynamics. 2. Reversible reactions obey the principle of microscopic reversibility, i.e. the reaction mechanism and the transition states are the same in both directions. There's no way for an enzyme to selectively reduce the transition state barrier going in one direction but not the other. Regards, Randy Read On 18 May 2010, at 08:31, Vinson LIANG wrote: Dear all, Sorry for this silly biochemistory question. Thing is that I have a reversible epimerase and I want to mutate it into an inreversible one. However, I have been told that the ΔG of a reversible reaction is zero. Which direction the reaction goes depends only on the concentration of the substrate. So the conclusion is, A: I can mutate the epimerase into an inreversible one. But it has no influence on the reaction direction, and hence it has little mean. B: There is no way to change a reversible epimerase into an inversible one. Could somebody please give me some comment on the two conclution? Thank you all for your time. Best, Vinson -- 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 RoadE-mail: rj...@cam.ac.uk mailto:rj...@cam.ac.uk Cambridge CB2 0XY, U.K. www-structmed.cimr.cam.ac.uk
Re: [ccp4bb] Is it possible to mutate a reversible epimerase into an inreversible one?
If you change the reaction rate in one direction 1000 times slower than in the other direction, then the reaction becomes practically irreversible. And the system might not be at equilibrium. Maia R. M. Garavito wrote: Vinson, As Dale and Randy pointed out, you cannot change the ΔG of a reaction by mutation: enzyme, which is a catalyst, affects only the activation barrier (ΔE double-dagger). You can just make it a better (or worse) catalyst which would allow the reaction to flow faster (or slower) towards equilibrium. Nature solves this problem very elegantly by taking a readily reversible enzyme, like an epimerase or isomerase, and coupling it to a much less reversible reaction which removes product quickly. Hence, the mass action is only in one direction. An example of such an arrangement is the triose phosphate isomerase (TIM)-glyceraldehyde 3-phosphate dehydrogenase (GAPDH) reaction pair. TIM is readily reversible (DHA = G3P), but G3P is rapidly converted to 1,3-diphosphoglycerate by GAPDH. The oxidation and phosphorylation reactions of GAPDH now make TIM work in one direction. Since many epimerases are very optimized enzymes, why not consider making a fusion with a second enzyme (like a reductase) to make the system flow in one direction. Of course, this depends on what you want to do with the product. Cheers, Michael // /R. Michael Garavito, Ph.D./ /Professor of Biochemistry Molecular Biology/ /513 Biochemistry Bldg. / /Michigan State University / /East Lansing, MI 48824-1319/ /Office:// //(517) 355-9724 Lab: (517) 353-9125/ /FAX: (517) 353-9334Email: rmgarav...@gmail.com mailto:garav...@gmail.com/ // On May 18, 2010, at 11:54 AM, Dale Tronrud wrote: Hi, I'm more of a Fourier coefficient kind of guy, but I thought that a ΔG of zero simply corresponded to an equilibrium constant of one. You can certainly have reversible reactions with other equilibrium constants. In fact I think irreversible reactions are simply ones where the equilibrium constant is so far to one side that, in practice, the reaction always goes all the way to product. As Randy pointed out the enzyme cannot change the ΔG (or the equilibrium constant). You could drive a reaction out of equilibrium by coupling it to some other reaction which itself is way out of equilibrium (such as ATP hydrolysis in the cell) but I don't think that's a simple mutation of your enzyme. ;-) Dale Tronrud On 05/18/10 00:31, Vinson LIANG wrote: Dear all, Sorry for this silly biochemistory question. Thing is that I have a reversible epimerase and I want to mutate it into an inreversible one. However, I have been told that the ΔG of a reversible reaction is zero. Which direction the reaction goes depends only on the concentration of the substrate. So the conclusion is, A: I can mutate the epimerase into an inreversible one. But it has no influence on the reaction direction, and hence it has little mean. B: There is no way to change a reversible epimerase into an inversible one. Could somebody please give me some comment on the two conclution? Thank you all for your time. Best, Vinson
Re: [ccp4bb] Is it possible to mutate a reversible epimerase into an inreversible one?
Sounds like a good explanation. Thank you. Maia Dale Tronrud wrote: If you change the reaction rate in one direction 1000 times slower then the reaction rate in the other direction will also be 1000 times slower and the equilibrium will be in exactly the same place. You can't make the transition state less stable when approached from the left without making it less stable when approached from the right. Dale Tronrud On 05/18/10 12:34, Maia Cherney wrote: If you change the reaction rate in one direction 1000 times slower than in the other direction, then the reaction becomes practically irreversible. And the system might not be at equilibrium. Maia R. M. Garavito wrote: Vinson, As Dale and Randy pointed out, you cannot change the ΔG of a reaction by mutation: enzyme, which is a catalyst, affects only the activation barrier (ΔE double-dagger). You can just make it a better (or worse) catalyst which would allow the reaction to flow faster (or slower) towards equilibrium. Nature solves this problem very elegantly by taking a readily reversible enzyme, like an epimerase or isomerase, and coupling it to a much less reversible reaction which removes product quickly. Hence, the mass action is only in one direction. An example of such an arrangement is the triose phosphate isomerase (TIM)-glyceraldehyde 3-phosphate dehydrogenase (GAPDH) reaction pair. TIM is readily reversible (DHA = G3P), but G3P is rapidly converted to 1,3-diphosphoglycerate by GAPDH. The oxidation and phosphorylation reactions of GAPDH now make TIM work in one direction. Since many epimerases are very optimized enzymes, why not consider making a fusion with a second enzyme (like a reductase) to make the system flow in one direction. Of course, this depends on what you want to do with the product. Cheers, Michael // /R. Michael Garavito, Ph.D./ /Professor of Biochemistry Molecular Biology/ /513 Biochemistry Bldg. / /Michigan State University / /East Lansing, MI 48824-1319/ /Office:// //(517) 355-9724 Lab: (517) 353-9125/ /FAX: (517) 353-9334Email: rmgarav...@gmail.com mailto:garav...@gmail.com/ // On May 18, 2010, at 11:54 AM, Dale Tronrud wrote: Hi, I'm more of a Fourier coefficient kind of guy, but I thought that a ΔG of zero simply corresponded to an equilibrium constant of one. You can certainly have reversible reactions with other equilibrium constants. In fact I think irreversible reactions are simply ones where the equilibrium constant is so far to one side that, in practice, the reaction always goes all the way to product. As Randy pointed out the enzyme cannot change the ΔG (or the equilibrium constant). You could drive a reaction out of equilibrium by coupling it to some other reaction which itself is way out of equilibrium (such as ATP hydrolysis in the cell) but I don't think that's a simple mutation of your enzyme. ;-) Dale Tronrud On 05/18/10 00:31, Vinson LIANG wrote: Dear all, Sorry for this silly biochemistory question. Thing is that I have a reversible epimerase and I want to mutate it into an inreversible one. However, I have been told that the ΔG of a reversible reaction is zero. Which direction the reaction goes depends only on the concentration of the substrate. So the conclusion is, A: I can mutate the epimerase into an inreversible one. But it has no influence on the reaction direction, and hence it has little mean. B: There is no way to change a reversible epimerase into an inversible one. Could somebody please give me some comment on the two conclution? Thank you all for your time. Best, Vinson
Re: [ccp4bb] Native Gel Theory and Practice
Dear Jacob, I offer you my opinion. Are you talking about electrophoresis? As far as I know it does not work for the mass. The velocity of a protein depends on the charge at a particular pH, the mass and shape of molecules etc. It's very difficult to take all these things into consideration. Otherwise this would be a very convenient method, much easier than the analytical centrifugation or gel-filtration that are usually used. However, electrophoresis does not work for mass determination. Besides, complex formation hugely depends on the protein concentration. If you dilute your mixture, your complexes might dissociate. There is equilibrium constant between different types of complexes. Maia Jacob Keller wrote: Dear Crystallographers, I am trying to optimize a native gel experiment of a two-protein complex, running the smallest-detectable amount of protein component A with varying amounts of component B. MWCharge MW/Charge A 22 -5-4308 B 17-24 -702 This experiment is partly to determine stoichiometry, but also to determine roughly the strength of the interaction. B definitely runs much faster than A alone, as predicted, but I am wondering what to expect with various oligomers. Should ABB run faster or slower than AB? What about AABB? Theoretically, AA should certainly run slower than A, and BB slower than B, simply because the mass/charge ratio is the same, but the overall mass is greater. But what happens when you have AAB, for example? There must be an equation relating the mass/charge and mass (and perhaps gel percentage) to the speed traveled in the gel--but what is the equation? Thanks for your consideration, Jacob *** Jacob Pearson Keller Northwestern University Medical Scientist Training Program Dallos Laboratory F. Searle 1-240 2240 Campus Drive Evanston IL 60208 lab: 847.491.2438 cel: 773.608.9185 email: j-kell...@northwestern.edu ***
Re: [ccp4bb] Unexplained density
I think cacodilate is less likely because it's negatively charged as the carboxy groups that surround the density. I think it's Zn or it may be another (endogenous) metal ion like Ca. You need to look at the coordination. Maia David Schuller wrote: The figures would be more helpful if you told us what each map contour represented, and at what contour level. I assume blue is 2fo-fc and green+red is fo-fc, positive and negative. It would be even more helpful to have the maps to turn over in 3D, but I understand you would probably be reluctant share that. The density looks much too strong to be mere water. Acidic surface residues may bind zinc. The metals might appear at multiple alternate locations, if the occupancies are less than 1.0. Synchrotron anomalous maps would be helpful in seeing Zinc, whose K edge is at 1.28 A. Multiple alternate locations are likely to show up at special symmetry positions. I have the impression this may be a special position in your map, but I can't be certain (2-fold axis?). Zinc and cacodylate may form cage complexes which can be bound between multiple surface acid resides (personal observation, not published). The complexes I have seen have one zinc at each pole with three cacodylates around the equator. Your density does not look especially similar to the complexes I have seen. Cacodylate also has a pretty good peak on anomalous maps, and since it's peak is at 1.04 A, anomalous maps at multiple wavelengths could distinguish between arsenic and zinc. On 04/29/10 17:38, Daniel Bonsor wrote: Hello again I currently have some unexplained density in my structure. As you can hopefully see from the images (see file), the density is dumbbell shaped. Whatever it is, it is coordinated by Asp and Glu residues. To me it looks like each lobe is a ring structure. The crystallization condition was: 6.5% PEG 8K, 10mM ZnSO4, 100mM sodium cacodylate pH 6.5, 100mM Am2SO4, 1% glycerol, with 20% glycerol as cryo. Protein was originally in 50mM Tris, 50mM NaCl pH 7.5. I originally placed a single Zn at the center of each lobe. Though after refmac, the Zn was displaced to one side. Two zincs in each dumbbell may have worked, but I am dubious about two zinc atoms being 4A apart and there is still some unexplained density. Are there any possible cyclization reactions of Tris, cacodylate or glycerol may have undergone to explain the density? Or is it simply a highly ordered water network? Or is there some other explanation? Thanks in advance Dan
Re: [ccp4bb] program align does not see libcxa.so.5
Hi bb,
when I try to run al3 (align) I get the error message
error while loading shared libraries: libcxa.so.5: cannot open shared
object file: No such file or directory
In fact, this file exists. How can I tell al3 where to look for this file?
Maia
Ed Pozharski wrote:
On Fri, 2010-04-30 at 13:35 +0100, Nicholas Keep wrote:
If anyone has a piece of software that would do this it would be
great.
How about this (this is a single line)
---
grep 'ATOM\|HETATM' file1.pdb file2.pdb |grep -v REMARK | cut -d: -f 2 |
cut -c 13-54 | sort | awk 'BEGIN {FIELDWIDTHS = 14 28; pt=} {if(pt==
$1) print pr,$2; pt=$1; pr=$0;}' | awk 'BEGIN {FIELDWIDTHS = 14 4 8 8 8
5 8 8 8} {printf %s %8.4f\n,$1,sqrt(($3-$7)^2+($4-$8)^2+($5-$9)^2);}'
| awk 'BEGIN {FIELDWIDTHS = 4 1 3 1 1 5 9} {printf %s %s %s %s %s\n,
$3,$5,$6,$1,$7;}'
---
The output is not sorted (sort isn't friendly to the idea of sorting
alphabetically and numerically at the same time). And awk means
gawk - mawk (Ubuntu default) doesn't allow fixed field selection
which is key to dealing with files that have alternate conformers).
HTH,
Ed.
Re: [ccp4bb] program align does not see libcxa.so.5
Dear James, Tim.
the command below (setenv LD_LIBRARY_PATH
${LD_LIBRARY_PATH}:/path/of/directory/which/contains/thelib)
worked for me.
Thanks a lot.
Maia
James Holton wrote:
for tcsh, the command is:
setenv LD_LIBRARY_PATH
${LD_LIBRARY_PATH}:/path/of/directory/which/contains/thelib
-James Holton
TCSH Scientist
On 4/30/2010 12:13 PM, Tim Gruene wrote:
one-time solution:
In the terminal from which you want to start the program, type
export
LD_LIBRARY_PATH=$LD_LIBRARY_PATH:/path/of/directory/which/contains/thelib
then start the program.
This is valid for a POSIX-compliant shell (bash, ksh, zsh, sh,...).
If you use (t)csh, the syntax is different, probably something like
set LD_LIBRARY_PATH
$LD_LIBRARY_PATH:/path/of/directory/which/contains/thelib
but [flame] you shouldn't use tcsh, it's outdated - the welcome page
of tcsh.org
was last edited nearly six years ago[/flame] (don't take this comment
seriously
unless you like nerdish flame wars - I hope someone on the list does
;-) )
long-term solution:
a) put the above command into your shell's start-up script
b) ask the author of the program or whoever compiled it to use the
switch
'-static-intel'. This doesn't alter the functionality of the program
and makes
it independent of the intel libraries which most people (notably
non-developers)
probably don't have on their system.
Cheers, Tim
On Fri, Apr 30, 2010 at 12:53:49PM -0600, Maia Cherney wrote:
Hi bb,
when I try to run al3 (align) I get the error message
error while loading shared libraries: libcxa.so.5: cannot open shared
object file: No such file or directory
In fact, this file exists. How can I tell al3 where to look for this
file?
Maia
Ed Pozharski wrote:
On Fri, 2010-04-30 at 13:35 +0100, Nicholas Keep wrote:
If anyone has a piece of software that would do this it would be
great.
How about this (this is a single line)
---
grep 'ATOM\|HETATM' file1.pdb file2.pdb |grep -v REMARK | cut -d:
-f 2 |
cut -c 13-54 | sort | awk 'BEGIN {FIELDWIDTHS = 14 28; pt=}
{if(pt==
$1) print pr,$2; pt=$1; pr=$0;}' | awk 'BEGIN {FIELDWIDTHS = 14 4
8 8 8
5 8 8 8} {printf %s
%8.4f\n,$1,sqrt(($3-$7)^2+($4-$8)^2+($5-$9)^2);}'
| awk 'BEGIN {FIELDWIDTHS = 4 1 3 1 1 5 9} {printf %s %s %s %s
%s\n,
$3,$5,$6,$1,$7;}'
---
The output is not sorted (sort isn't friendly to the idea of sorting
alphabetically and numerically at the same time). And awk means
gawk - mawk (Ubuntu default) doesn't allow fixed field selection
which is key to dealing with files that have alternate conformers).
HTH,
Ed.
Re: [ccp4bb] where to find the shell's startup script
Tim, I have a difficulty to find my shell's startup script. After I
upgraded my ubuntu to 9.04 version,
my cshrc and .cshrc files don't work, and a s a result, a have to give
the full path to make some programs work.
How do I find the startup script/
Maia
Tim Gruene wrote:
one-time solution:
In the terminal from which you want to start the program, type
export
LD_LIBRARY_PATH=$LD_LIBRARY_PATH:/path/of/directory/which/contains/thelib
then start the program.
This is valid for a POSIX-compliant shell (bash, ksh, zsh, sh,...).
If you use (t)csh, the syntax is different, probably something like
set LD_LIBRARY_PATH $LD_LIBRARY_PATH:/path/of/directory/which/contains/thelib
but [flame] you shouldn't use tcsh, it's outdated - the welcome page of tcsh.org
was last edited nearly six years ago[/flame] (don't take this comment seriously
unless you like nerdish flame wars - I hope someone on the list does ;-) )
long-term solution:
a) put the above command into your shell's start-up script
b) ask the author of the program or whoever compiled it to use the switch
'-static-intel'. This doesn't alter the functionality of the program and makes
it independent of the intel libraries which most people (notably non-developers)
probably don't have on their system.
Cheers, Tim
On Fri, Apr 30, 2010 at 12:53:49PM -0600, Maia Cherney wrote:
Hi bb,
when I try to run al3 (align) I get the error message
error while loading shared libraries: libcxa.so.5: cannot open shared
object file: No such file or directory
In fact, this file exists. How can I tell al3 where to look for this file?
Maia
Ed Pozharski wrote:
On Fri, 2010-04-30 at 13:35 +0100, Nicholas Keep wrote:
If anyone has a piece of software that would do this it would be
great.
How about this (this is a single line)
---
grep 'ATOM\|HETATM' file1.pdb file2.pdb |grep -v REMARK | cut -d: -f 2 |
cut -c 13-54 | sort | awk 'BEGIN {FIELDWIDTHS = 14 28; pt=} {if(pt==
$1) print pr,$2; pt=$1; pr=$0;}' | awk 'BEGIN {FIELDWIDTHS = 14 4 8 8 8
5 8 8 8} {printf %s %8.4f\n,$1,sqrt(($3-$7)^2+($4-$8)^2+($5-$9)^2);}'
| awk 'BEGIN {FIELDWIDTHS = 4 1 3 1 1 5 9} {printf %s %s %s %s %s\n,
$3,$5,$6,$1,$7;}'
---
The output is not sorted (sort isn't friendly to the idea of sorting
alphabetically and numerically at the same time). And awk means
gawk - mawk (Ubuntu default) doesn't allow fixed field selection
which is key to dealing with files that have alternate conformers).
HTH,
Ed.
Re: [ccp4bb] Unexplained density
It's hard to see clearly the density, but judging from the abundance of carboxy groups, it may be a metal. Maia Daniel Bonsor wrote: Hello again I currently have some unexplained density in my structure. As you can hopefully see from the images (see file), the density is dumbbell shaped. Whatever it is, it is coordinated by Asp and Glu residues. To me it looks like each lobe is a ring structure. The crystallization condition was: 6.5% PEG 8K, 10mM ZnSO4, 100mM sodium cacodylate pH 6.5, 100mM Am2SO4, 1% glycerol, with 20% glycerol as cryo. Protein was originally in 50mM Tris, 50mM NaCl pH 7.5. I originally placed a single Zn at the center of each lobe. Though after refmac, the Zn was displaced to one side. Two zincs in each dumbbell may have worked, but I am dubious about two zinc atoms being 4A apart and there is still some unexplained density. Are there any possible cyclization reactions of Tris, cacodylate or glycerol may have undergone to explain the density? Or is it simply a highly ordered water network? Or is there some other explanation? Thanks in advance Dan
Re: [PyMOL] PyMOL Plugin Architecture
Thank, Robert, Jason. Finally I figured this out with your help. Maia Robert Campbell wrote: Hi Maia, Just to add to what Jason said: On Thu, 22 Apr 2010 17:16:41 -0400, Jason Vertrees jason.vertr...@schrodinger.com wrote: 2. Which plugin allows me to move one structure relative to another. (I want to do manual docking of one structure into another). You don't need a plugin to move objects. Just use SHIFT-middle-mouse button drag, while in editing mode. If you're starting from one object (eg. protein and ligand come from one file) then split them up: In case it isn't obvious to you, you can switch between viewing mode and editing mode simply by left clicking in the bottom-right region of the main PyMOL window where the mouse button functions are described. Cheers, Rob -- ___ PyMOL-users mailing list (PyMOL-users@lists.sourceforge.net) Info Page: https://lists.sourceforge.net/lists/listinfo/pymol-users Archives: http://www.mail-archive.com/pymol-users@lists.sourceforge.net
Re: [PyMOL] pymol and freemol
Hi David, I downloaded apbs and pdb2pqr, but I don't know how to install them on my ubuntu 9.04. Is it possible to make an easy to install release? For now, could you please send me the instruction. Maia David Hall wrote: In my incentive build, freemol is present with the following readme: This is a binary distribution of the FreeMOL Open-Source Compilation. The FreeMOL project home page can be found at: http://bioinformatics.org/freemol Complete source code for building FreeMOL and its various components can be browsed via: http://www.bioinformatics.org/websvn/listing.php?repname=freemol and fetched via: svn co svn://bioinformatics.org/svnroot/freemol/trunk Licensing information can be found in LICENSE.txt If you have any questions about FreeMOL, please contact: Warren L. DeLano, Ph.D. DeLano Scientific LLC war...@delsci.com Thank You! I don't have any instructions how to incorporate this into building from pymol trunk necessarily though. In the incentive builds, the freemol folder is not inside ext or modules or anything, but instead is a top level directory like those. You will have to build the sources of the things that come with it (apbs, mengine, mpeg_encode, pdb2pqr). I think pymol autodetects the presence of freemol based on this grep output on the pymol source. modules/pymol/__init__.py:# auto-detect bundled FREEMOL (if present) modules/pymol/__init__.py:if not os.environ.has_key(FREEMOL): modules/pymol/__init__.py:test_path = os.path.join(os.environ['PYMOL_PATH'],freemol) modules/pymol/__init__.py:os.environ['FREEMOL'] = test_path modules/pymol/__init__.py:# include FREEMOL's libpy in sys.path (if present) modules/pymol/__init__.py:if os.environ.has_key(FREEMOL): modules/pymol/__init__.py:freemol_libpy = os.path.join(os.environ['FREEMOL'],libpy) modules/pymol/__init__.py:if os.path.isdir(freemol_libpy): modules/pymol/__init__.py:if freemol_libpy not in sys.path: modules/pymol/__init__.py:sys.path.append(freemol_libpy) Hopefully this all helps. -David - Original Message From: Maia Cherney ch...@ualberta.ca To: Jason Vertrees jason.vertr...@schrodinger.com Cc: pymol-users@lists.sourceforge.net Sent: Wed, January 20, 2010 9:37:46 PM Subject: Re: [PyMOL] new ideas Thanks, But I could not find the download site for FreeMol. Could somebody send me a link to FreeMol? Maia Jason Vertrees wrote: Maia, You can already move structures independently. If you install PyMOL+FreeMOL you can have access to the MMFF-enabled PyMOL that will allow you to do small molecule cleanup and editing. Try putting your mouse into Editing Mode and moving atoms around. It's not hard. Check this out: http://pymolwiki.org/index.php/Molecular_Sculpting for some help. Regards, -- Jason On Wed, Jan 20, 2010 at 11:52 AM, Maia Cherney wrote: Hi Jason I need such features. I open two different pdbs and I want to manually move one structure relative to another. I want to dock one structure into another or superpose 2 structures manually. Is it possible? Or I want to move a fragment of the molecule relative to the rest of the molecule. Is it possible? Maia -- Throughout its 18-year history, RSA Conference consistently attracts the world's best and brightest in the field, creating opportunities for Conference attendees to learn about information security's most important issues through interactions with peers, luminaries and emerging and established companies. http://p.sf.net/sfu/rsaconf-dev2dev ___ PyMOL-users mailing list (PyMOL-users@lists.sourceforge.net) Info Page: https://lists.sourceforge.net/lists/listinfo/pymol-users Archives: http://www.mail-archive.com/pymol-users@lists.sourceforge.net -- Download Intel#174; Parallel Studio Eval Try the new software tools for yourself. Speed compiling, find bugs proactively, and fine-tune applications for parallel performance. See why Intel Parallel Studio got high marks during beta. http://p.sf.net/sfu/intel-sw-dev ___ PyMOL-users mailing list (PyMOL-users@lists.sourceforge.net) Info Page: https://lists.sourceforge.net/lists/listinfo/pymol-users Archives: http://www.mail-archive.com/pymol-users@lists.sourceforge.net
Re: [ccp4bb] programs to check the structure of DNA
I know two programs; 3DNA by Lu and curves by Lavery and Sklenar. 3DNA is easier to use, it can also make input for the Curves. Maia Alessandra Pesce wrote: Dear All, I am looking for available programs and/or websites able to check the structure of DNA in DNA-protein complexes. I need to evaluate the DNA bending, its backbone distortion, the alteration of the base pairing, and its interaction with the protein. The aim is to compare in easy and quick way different conformation of DNA in different DNA-protein complexes. Can anybody help me? Thanks in advance. Cheers Alessandra * Alessandra Pesce, PhD Department of Physics University of Genova Via Dodecaneso 33 16146 Genova, Italy Tel.(DIFI) ++39 010 353 6243 | 6309 e-mail pe...@ge.infm.it mailto:pe...@ge.infm.it *
Re: [ccp4bb] protein degradation during concentration for crystallization trials
Hi, MBP tag: 1.there might be a TEV cleavage site in your MBP variant. 2. your protein needs salt to stay in solution 3. your protein forms aggregates with MBP GST tag: you probably concentrate a protease together with your protein. You need a protease inhibitor kit to take care of different types of proteases. It looks like a His tag would be a better option for you. Maia vikrant saa wrote: Dear all I am working on purification of 14 kd protein(pI 8.3, basic protein) that has MBP(maltose binding protein, 45 kd,) tag, and same protein in other vector(pGEX-KT) that has GST tag. During affinity purification in both cases I used 300mM nacl and 50 mM tris, pH 7.5 buffer throughout the purification.I do on column cleavage with TEV to remove the tag (for crystallization purpose). Purification with MBP tag: After cleavage MBP also appear in elution fraction along with cleaved protein of my interest. To remove MBP(pI 5.0) from protein of my interest I have to do anion exchange chr. with DEAE resin(weak anion exchanger) with buffer of pH7.3. Hence I do dialysis against the buffer 20mM nacl and 20 mM tris, ph 7.3. in cold room. I have few problems: 1) why the MBP elute with protein of my interest ( I tried at low salt concentration also but still elute). 2) During dialysis my protein of interest precipitate. 3) If I tried FPLC, protein of interest and MBP elute in same fraction with superdex 200 column. Problem with GST tag purification: it give me impure protein even after enough washes with high salt concentration buffer (upto 2 molar). When i concentrate protein in CENTRICON (Millipore, centrifugation 4000rpm at 4 degree, 300mM NaCl and 50 mm Tris buffer with 5mM BME) it degrade very fast and probably aggregate as smear obtain on SDS page below the size of protein after concentration (even protease inhibitor not very much helpful) and band intensity of protein of interest almost remain same before and after concentration step. Please send me your valuable suggestion to overcome to these difficulty. I have also tried with some additives such as sucrose, glycerol, PBS buffer. With regards -- $$ VIKRANT Junior Research fellow Cancer Research Institute Advanced Centre for Treatment, Research and Education in Cancer (ACTREC), Kharghar, Navi Mumbai, India $$$ ** ** Send free SMS to your Friends on Mobile from your Yahoo! Messenger. Download Now! http://messenger.yahoo.com/download.php
Re: [COOT] pointer distances, recording water
Hi Paul, the format of the file got screwed. I am resending the file in the attachment. I would like to repeat the description of the problem. There is a difference in recording of water molecules in coot and in phenix. The example is given in the attached pdb file. I apologize for the wrong subject (pointer distances), as this was my previous request to make pointer distances available to atoms of symmetry molecules. Maia Maia Cherney wrote: Thank you, Paul. There is also difference in recording water. Should it be ATOM or HETATM? Phenix makes it HETATM, but coot makes it ATOM. Here is an example that was saved after adding one water molecule in coot (after refinement in phenix). HETATM10472 O HOH S1252 75.727 6.379 70.023 1.00 31.17 O HETATM10473 O HOH S1253 40.692 -0.062 54.840 1.00 37.64 O TER10474 HOH S1253 ATOM 10475 O HOH S1254 75.842 2.043 56.024 1.00 35.64 O TER10476 HOH S1254 END Maia Paul Emsley wrote: Maia Cherney wrote: Hi Paul, Coot saves pdb files with TER card. The problem is the TER card takes the next atom number. ATOM 2303 C4 ADP A 288 33.070 4.240 14.161 1.00 11.08 C TER 2304 ADP A 288 ATOM 2305 O SER B -1 The same model after phenix gives no number to the TER card. ATOM 2303 C4 ADP A 288 33.070 4.240 14.161 1.00 11.08 C TER ATOM 2304 O SER B -1 As a result, the number of atoms in the pdb files is different. That is confusing. Could you please fix this problem. Maia Hi Maia, It seems to us that this is an issue with Phenix, not Coot or mmdb. Have a look here: http://www.wwpdb.org/documentation/format32/sect9.html#TER The serial number of the TER record is one number greater than the serial number of the ATOM/HETATM preceding the TER Regards, Paul. water.pdb Description: Protein Databank data
Re: [ccp4bb] ccp4 vs. phenix special position atoms
Hi Pavel, you should add to the explanation what /==1 and !=1 are, as the majority of people don't know. / == : equal != : not equal Maia / / Pavel Afonine wrote: Hi Regina, this subject was discussed on PHENIX bulletin board some time ago: http://www.phenix-online.org/pipermail/phenixbb/2009-December/003074.html I think this answers your question; please let me know otherwise. Also, I'm not sure what you mean by (except that refinement will fail if the special position atoms are not a separate group in refinement). If something fails in phenix.refine then I would to know about it so I (or someone else) can fix it asap. We can discuss it off-list. Pavel. On 3/23/10 1:37 PM, Regina Kettering wrote: Hello all. I have a refinement that includes atoms at special positions, am unsure how to delineate them in CCP4 vs. PHENIX programs. According to the information, for CCP4 you can reduce the occupancy by the appropriate amount, depending on the axis (2-fold to 0.5, 3-fold to 0.33, etc). However, I have not been able to determine whether PHENIX uses this same convention (except that refinement will fail if the special position atoms are not a separate group in refinement). My refinement uses PHENIX TLS, so I would rather continue using PHENIX. Thanks in advance. Regina Kettering
Re: [ccp4bb] Soaking at pH 4.0 ?
Could you transfer your crystals in a higher pH buffer? Maia Paul Lindblom wrote: Hi, I am trying to soak some sugars in my crystals, but the cystallization condition has a pH of 4.0. Does anybody has any experience with acidic oxidation in such a case. I think I can´t avoid oxidation at this pH? Any suggestions, or do I have to screen for other conditions? Thanks, P.
Re: [ccp4bb] units of f0, f', f''
Thanks, I was actually joking because I was a little annoyed about the discussion, but then I realized that this discussion is great, (now I will not forget the units of electron density) and it's still not resolved. You said the charge/mass ratio and the density of that, but other people said the electron density is number of electrons (not charge) divided by volume (1/A^3). Maia H. Raaijmakers wrote: Maia, Usually we live in a macroscopic world and usually gravity is the most important force. In x-ray diffraction the charge/mass ratio is the most important paramater (and the density of that). Hans Maia Cherney schreef: Hi all, Usually density means mass divided by volume. The mass of an electron is known. Then it will be no arguments. Maia
Re: [ccp4bb] units of f0, f', f''
Hi all, Usually density means mass divided by volume. The mass of an electron is known. Then it will be no arguments. Maia Ian Tickle wrote: I'm not aware that anyone has suggested the notation rho e/Å^3. I think you misunderstood my point, I certainly didn't mean to imply that anyone had suggested or used that notation, quite the opposite in fact. My point was that you said that you use the term 'electron density' to define two different things either at the same time or on different occasions, but that to resolve the ambiguity you use labels such as 'e/Å^3' or 'sigma/Å^3' attached to the values. My point was that if I needed to use these quantities in equations then the rules of algebra require that distinguishable symbols (e.g. rho and rho') be assigned, otherwise I would be forced into the highly undesirable situation of labelling the symbols with their units in the equations in the way you describe in order to distinguish them. Then in my 'Notation' section my definitions of rho rho' would need to be different in some way, again in order to distinguish them: I could not simply call both of them 'electron density' as you appear to be doing. The question of whether your units of electron density are '1/Å^3' or 'e/Å^3' clearly comes down to definition, nothing more. If we can't agree on the definition then we are surely not going to agree on the units! Actually we don't need to agree on the definition: as long as I know what precisely your set of definitions is, I can make the appropriate adjustments to my units you can do the same if you know my definitions; it just makes life so much easier if we can agree to use the same definitions! Again it comes down to the importance of having a 'Notation' section so everyone knows exactly what the definitions in use are. My definition of electron density is number of electrons per unit volume which I happen to find convenient and for which the appropriate units are '1/Å^3'. In order for your choice of units 'e/Å^3' to be appropriate then your definition would have to be electric charge per unit volume, then you need to include the conversion factor 'e' (charge ! on the electron) in order to convert from my number of electrons to your electric charge, otherwise your values will all be very small (around 10^-19 in SI units). I would prefer to call this quantity electric charge density since electron density to me implies density of electrons not density of charge. I just happen to think that it's easier to avoid conversion factors unless they're essential. Exactly the same thing of course happens with the scattering factor: I'm using what I believe is the standard definition (i.e. the one given in International Tables), namely the ratio of scattered amplitude to that for a free electron which clearly must be unitless. So I would say 'f = 10' or whatever. I take it that you would say 'f = 10e'. Assuming that to be the case, then it means you must be using a different definition consistent with the inclusion of the conversion factor 'e', namely that the scattering factor is the equivalent point electric charge, i.e. the point charge that would scatter the same X-ray amplitude as the atom. I've not seen the scattering factor defined in that way before: it's somewhat more convoluted than the standard definition but still usable. The question remains of course - why would you not want to stick to the standard definitions? BTW I assume your 'sigma/Å^3' was a slip and you intended to write just 'sigma' since sigma(rho) must have the same units as rho (being its RMS value), i.e. 1/Å^3, so in your second kind of e.d. map rho/sigma(rho) is dimensionless (and therefore unitless). However since rho and sigma(rho) have identical units I don't see how their ratio rho/sigma(rho) can have units of 'sigma', as you seem to imply if I've understood correctly? What I'm more concerned about is when you assign a numerical value to a quantity. Take the equation E=MC^2. The equation is true regardless of how you measure your energy, mass, and speed. It is when you say that M = 42 that it becomes important to unambiguously label 42 with its units. It is when you are given a mass equal to 42 newtons, the speed of light in furlongs/fortnight, and asked to calculate the energy in calories that you have to track your units carefully and perform all the proper conversions to calculate the number of calories. I can only agree with you there, but I never suggested or implied that a mass value (or speed or energy) should be given without the appropriate units specification, or that one should not take great care to track the units conversions. Actually many equations in crystallography are not as friendly as this one since they have conversion factors built into their standard formulations. With the conversion factor built in you are then restricted to use the units that were assumed.
Re: [COOT] moving atoms
Hi Paul, I can not move an atom or a group of atoms in the ligand (rotate/translate zone). The ctrl-click does not work (never works for me on several computers including Mac, PC etc). To move an atom or several specific atoms should not be so difficult. It looks like coot choses how many atoms to move in the chain. For a ligand, it moves the whole thing, but not its part. General torsion sometimes is ok, but with some ligands (or some bonds?) the whole ligand rotates around one bond (I want only part of the ligand to rotate around this bond). What is the problem with this? I like the way torsion works in xfit. Just click on *_2 _*atoms of a bond and the whole part of the molecule (next to the second click) rotates around this bond. I use coot 0.6, revision 2540 with ubuntu. Maia
Re: [PyMOL] new ideas
Hi Jason, Just to remind you. You promised to put back FreeMol and let us know about it. I am waiting. I need to move structures independently. Maia Jason Vertrees wrote: Maia, You can already move structures independently. If you install PyMOL+FreeMOL you can have access to the MMFF-enabled PyMOL that will allow you to do small molecule cleanup and editing. Try putting your mouse into Editing Mode and moving atoms around. It's not hard. Check this out: http://pymolwiki.org/index.php/Molecular_Sculpting for some help. Regards, -- Jason On Wed, Jan 20, 2010 at 11:52 AM, Maia Cherney ch...@ualberta.ca wrote: Hi Jason I need such features. I open two different pdbs and I want to manually move one structure relative to another. I want to dock one structure into another or superpose 2 structures manually. Is it possible? Or I want to move a fragment of the molecule relative to the rest of the molecule. Is it possible? Maia -- The Planet: dedicated and managed hosting, cloud storage, colocation Stay online with enterprise data centers and the best network in the business Choose flexible plans and management services without long-term contracts Personal 24x7 support from experience hosting pros just a phone call away. http://p.sf.net/sfu/theplanet-com ___ PyMOL-users mailing list (PyMOL-users@lists.sourceforge.net) Info Page: https://lists.sourceforge.net/lists/listinfo/pymol-users Archives: http://www.mail-archive.com/pymol-users@lists.sourceforge.net
Re: [COOT] Coot, delete a chain
Thanks, Judit. That is exactly what I wanted. But it has not worked for me yet. To delete a chain in coot (also it would be nice to have a similar function in Pymol, too). The graphics programs make easily new chains, but it's difficult to delete them without going to a text editor. Also, again, I tried to delete all waters at once in coot, it's difficult. You need map etc. Maia Debreczeni, Judit wrote: How about this: 1. click on the bin in the toolbar and select Delete zone 2. go to the first residue in the chain (in the Go to atom window or with ^g), click on the residue 3. similarly, go to the last one and click that - Coot should delete the chain (it will take a while...) It that seems a little fiddly, try this: --- (define (delete-chain imol chain-id) (if (and (string? chain-id) and (number? imol)) (let* ((n-res (chain-n-residues chain-id imol)) (start (seqnum-from-serial-number imol chain-id 0)) (end (seqnum-from-serial-number imol chain-id (- n-res 1 (delete-residue-range imol chain-id start end --- where imol is the molecule number and chain-id is the chain you want to delete. using it like e.g.: (delete-chain 0 A) JED. How nice would it be to be able to delete chains/residues in the go to atom window... or renumber residues and change chain IDs and copy fragments by simple drag-and-drop... sigh. -- AstraZeneca UK Limited is a company incorporated in England and Wales with registered number: 03674842 and a registered office at 15 Stanhope Gate, London W1K 1LN. Confidentiality Notice: This message is private and may contain confidential, proprietary and legally privileged 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 unauthorised use or disclosure of the contents of this message is not permitted and may be unlawful. Disclaimer: Email messages may be subject to delays, interception, non-delivery and unauthorised alterations. Therefore, information expressed in this message is not given or endorsed by AstraZeneca UK Limited unless otherwise notified by an authorised representative independent of this message. No contractual relationship is created by this message by any person unless specifically indicated by agreement in writing other than email. Monitoring: AstraZeneca UK Limited may monitor email traffic data and content for the purposes of the prevention and detection of crime, ensuring the security of our computer systems and checking Compliance with our Code of Conduct and Policies. -Original Message- From: Mailing list for users of COOT Crystallographic Software [mailto:c...@jiscmail.ac.uk] On Behalf Of Maia Cherney Sent: 31 January 2010 20:07 To: COOT@JISCMAIL.AC.UK Subject: Re: [COOT] Coot, delete a chain Hi all, Is there a way to delete the whole chain of a complex? Or all solvent atoms at once? Maia
Re: [COOT] Coot, delete a chain
Thanks, Judit. It worked for everything, the whole chain, a part of a chain and for all waters. Debreczeni, Judit wrote: How about this: 1. click on the bin in the toolbar and select Delete zone 2. go to the first residue in the chain (in the Go to atom window or with ^g), click on the residue 3. similarly, go to the last one and click that - Coot should delete the chain (it will take a while...) It that seems a little fiddly, try this: --- (define (delete-chain imol chain-id) (if (and (string? chain-id) and (number? imol)) (let* ((n-res (chain-n-residues chain-id imol)) (start (seqnum-from-serial-number imol chain-id 0)) (end (seqnum-from-serial-number imol chain-id (- n-res 1 (delete-residue-range imol chain-id start end --- where imol is the molecule number and chain-id is the chain you want to delete. using it like e.g.: (delete-chain 0 A) JED. How nice would it be to be able to delete chains/residues in the go to atom window... or renumber residues and change chain IDs and copy fragments by simple drag-and-drop... sigh. -- AstraZeneca UK Limited is a company incorporated in England and Wales with registered number: 03674842 and a registered office at 15 Stanhope Gate, London W1K 1LN. Confidentiality Notice: This message is private and may contain confidential, proprietary and legally privileged 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 unauthorised use or disclosure of the contents of this message is not permitted and may be unlawful. Disclaimer: Email messages may be subject to delays, interception, non-delivery and unauthorised alterations. Therefore, information expressed in this message is not given or endorsed by AstraZeneca UK Limited unless otherwise notified by an authorised representative independent of this message. No contractual relationship is created by this message by any person unless specifically indicated by agreement in writing other than email. Monitoring: AstraZeneca UK Limited may monitor email traffic data and content for the purposes of the prevention and detection of crime, ensuring the security of our computer systems and checking Compliance with our Code of Conduct and Policies. -Original Message- From: Mailing list for users of COOT Crystallographic Software [mailto:c...@jiscmail.ac.uk] On Behalf Of Maia Cherney Sent: 31 January 2010 20:07 To: COOT@JISCMAIL.AC.UK Subject: Re: [COOT] Coot, delete a chain Hi all, Is there a way to delete the whole chain of a complex? Or all solvent atoms at once? Maia
Re: [COOT] Coot, delete a chain
Hi all, Is there a way to delete the whole chain of a complex? Or all solvent atoms at once? Maia
Re: [PyMOL] new ideas
Hi Jason I need such features. I open two different pdbs and I want to manually move one structure relative to another. I want to dock one structure into another or superpose 2 structures manually. Is it possible? Or I want to move a fragment of the molecule relative to the rest of the molecule. Is it possible? Maia -- Throughout its 18-year history, RSA Conference consistently attracts the world's best and brightest in the field, creating opportunities for Conference attendees to learn about information security's most important issues through interactions with peers, luminaries and emerging and established companies. http://p.sf.net/sfu/rsaconf-dev2dev ___ PyMOL-users mailing list (PyMOL-users@lists.sourceforge.net) Info Page: https://lists.sourceforge.net/lists/listinfo/pymol-users Archives: http://www.mail-archive.com/pymol-users@lists.sourceforge.net
Re: [PyMOL] new ideas
Thanks, But I could not find the download site for FreeMol. Could somebody send me a link to FreeMol? Maia Jason Vertrees wrote: Maia, You can already move structures independently. If you install PyMOL+FreeMOL you can have access to the MMFF-enabled PyMOL that will allow you to do small molecule cleanup and editing. Try putting your mouse into Editing Mode and moving atoms around. It's not hard. Check this out: http://pymolwiki.org/index.php/Molecular_Sculpting for some help. Regards, -- Jason On Wed, Jan 20, 2010 at 11:52 AM, Maia Cherney ch...@ualberta.ca wrote: Hi Jason I need such features. I open two different pdbs and I want to manually move one structure relative to another. I want to dock one structure into another or superpose 2 structures manually. Is it possible? Or I want to move a fragment of the molecule relative to the rest of the molecule. Is it possible? Maia -- Throughout its 18-year history, RSA Conference consistently attracts the world's best and brightest in the field, creating opportunities for Conference attendees to learn about information security's most important issues through interactions with peers, luminaries and emerging and established companies. http://p.sf.net/sfu/rsaconf-dev2dev ___ PyMOL-users mailing list (PyMOL-users@lists.sourceforge.net) Info Page: https://lists.sourceforge.net/lists/listinfo/pymol-users Archives: http://www.mail-archive.com/pymol-users@lists.sourceforge.net
Re: [COOT] Release 0.6
Thank you, guys, for the new release. I suggested some time ago to include Go to atom button in the bar beside Display manager, as this is one of the most used buttons. Paul liked the idea, but I don't see it in the new release. Maia Kevin Cowtan wrote: Looks like 2540: http://www.biop.ox.ac.uk/coot/devel/svn.log Tim Fenn wrote: On Fri, 4 Dec 2009 02:56:49 + Paul Emsley paul.ems...@bioch.ox.ac.uk wrote: We are please to announce the release of coot-0.6 Source here: http://www.biop.ox.ac.uk/coot/software/source/releases/coot-0.6.tar.gz Binaries from here: http://www.biop.ox.ac.uk/coot/software/binaries/releases/ Grats! Which SVN tag corresponds to the 0.6 release? -Tim -- - Tim Fenn f...@stanford.edu Stanford University, School of Medicine James H. Clark Center 318 Campus Drive, Room E300 Stanford, CA 94305-5432 Phone: (650) 736-1714 FAX: (650) 736-1961 -
Re: [COOT] Ramachandran plot
Hi Paul, I just figured out the movements in the plot areas. Please, disregard my question. Maia
Re: [PyMOL] [Solved] Upgraded computer, Pymol usable again. :^)
Hi John, I have a similar problem with my ubuntu 8.10. When I use a command window, I get the error message: Traceback (most recent call last): File /var/lib/python-support/python2.5/pymol//__init__.py, line 167, in module import pymol File /var/lib/python-support/python2.5/pymol/__init__.py, line 412, in module from pymol import _cmd ImportError: /var/lib/python-support/python2.5/pymol/_cmd.so: undefined symbol: PyUnicodeUCS4_SetDefaultEncoding I have installed another PyMol in a different place. When I give a full path to that PyMol, I get it open for a split second a dissapear with a message PyMOL(TM) Incentive Product - Copyright (C) 2006 DeLano Scientific LLC. A current PyMOL Maintenance and/or Support Subscription may be required for legal use of this Build beyond a finite honor-system evaluation period. Please visit http://www.pymol.org/funding.html for more information. This PyMOL Executable Build incorporates Open-Source PyMOL 0.99rc1. OpenGL graphics engine: GL_VENDOR: NVIDIA Corporation GL_RENDERER: GeForce 9800 GTX+/PCI/SSE2 GL_VERSION: 2.1.2 NVIDIA 177.82 Segmentation fault I can open PyMol only from the applications/education menu. I checked the Visual Effects, they are none. Maia John Ladasky wrote: Hello again, With some hints from Ubuntu Forums, I tracked down and corrected my problem. Hopefully this will be of help to someone else. The fancy new desktop rendering schemes which are part of Ubuntu v.8, but which were not part of Ubuntu v.6, use 3D graphics to grow, shrink, and move windows and icons. Apparently, this conflicts with the way that PyMol wants to talk to OpenGL. The solution is to tell the desktop to behave simply. From the Ubuntu main menu, select System Preferences Appearance Visual Effects. Change the setting from Normal to None. Then PyMol works as expected! -- Come build with us! The BlackBerry(R) Developer Conference in SF, CA is the only developer event you need to attend this year. Jumpstart your developing skills, take BlackBerry mobile applications to market and stay ahead of the curve. Join us from November 9 - 12, 2009. Register now! http://p.sf.net/sfu/devconference ___ PyMOL-users mailing list (PyMOL-users@lists.sourceforge.net) Info Page: https://lists.sourceforge.net/lists/listinfo/pymol-users Archives: http://www.mail-archive.com/pymol-users@lists.sourceforge.net -- Come build with us! The BlackBerry(R) Developer Conference in SF, CA is the only developer event you need to attend this year. Jumpstart your developing skills, take BlackBerry mobile applications to market and stay ahead of the curve. Join us from November 9 - 12, 2009. Register now! http://p.sf.net/sfu/devconference ___ PyMOL-users mailing list (PyMOL-users@lists.sourceforge.net) Info Page: https://lists.sourceforge.net/lists/listinfo/pymol-users Archives: http://www.mail-archive.com/pymol-users@lists.sourceforge.net
Re: [PyMOL] Alignment Objects
Hi, Is it posible in pymol to select all residues of a certain type automatically, like all arginines or all lysines + arginines in a chain? Maia -- Let Crystal Reports handle the reporting - Free Crystal Reports 2008 30-Day trial. Simplify your report design, integration and deployment - and focus on what you do best, core application coding. Discover what's new with Crystal Reports now. http://p.sf.net/sfu/bobj-july ___ PyMOL-users mailing list (PyMOL-users@lists.sourceforge.net) Info Page: https://lists.sourceforge.net/lists/listinfo/pymol-users Archives: http://www.mail-archive.com/pymol-users@lists.sourceforge.net
[PyMOL] pymol does not open from a terminal
Hi, Could you please help me with a pymol problem. I have ubuntu linux and I have pymol in the /usr/bin (comes with ubuntu installation) and also one in the /programs/linux directory. But I can't open pymol in a terminal (both installations), only from applications/education menu. I get this error from /usr/bin/pymol: Traceback (most recent call last): File /var/lib/python-support/python2.5/pymol//__init__.py, line 167, in module import pymol File /var/lib/python-support/python2.5/pymol/__init__.py, line 412, in module from pymol import _cmd ImportError: /var/lib/python-support/python2.5/pymol/_cmd.so: undefined symbol: PyUnicodeUCS4_SetDefaultEncoding or this error from the programs/linux/pymol Traceback (most recent call last): File string, line 1, in ? File /raid/programs/linux/pymol-0.99rc1/modules/pymol/__init__.py, line 353, in ? import _cmd ImportError: libpng.so.3: cannot open shared object file: No such file or directory Could anybody ,please, advise me how to fix it. -- Let Crystal Reports handle the reporting - Free Crystal Reports 2008 30-Day trial. Simplify your report design, integration and deployment - and focus on what you do best, core application coding. Discover what's new with Crystal Reports now. http://p.sf.net/sfu/bobj-july ___ PyMOL-users mailing list (PyMOL-users@lists.sourceforge.net) Info Page: https://lists.sourceforge.net/lists/listinfo/pymol-users Archives: http://www.mail-archive.com/pymol-users@lists.sourceforge.net
Re: [COOT] Novel maps...
Hi all, Is it possible to change residue or atom occupancies in coot? Maia
Re: [COOT] mouse buttons and arrow keys [was Re: problems of version 0.6]
Dear Paul, thank you for your reply. I thought for several days that you ignored my wishes, but you have not. Actually, a lot of people would be happy with the middle button translation. Your guess is right. I do prop up my head (as many others), or I hold a cup of coffee (as many others). So, how to enable the middle mouse drag for translate? (Chording on the mouse is fine with me, too). Thanks for the arrow keys for translate. Maia Paul Emsley wrote: Dear Maia, Maia Cherney wrote: I would like to use only one-button keys, so that I could use only one hand when fitting. Why!? Because the other one is needed to prop up your head, maybe? :-) http://linuxoutlaws.com/files/dan-jaunty-party-thumb.jpg I find using my left hand for keyboarding makes things way faster than clicking on icons with the right hand on the mouse. Well, now help me with key-bindings, please. I need one or two buttons on the mouse for map dragging. Pymol and x-fit use the middle button for translation. It confuses me all the time, when I work in coot after working in pymol. Do you have any unused two-button combinations on the mouse? If it's not possible to touch the mouse buttons in the key-bindings, please help me to program arrow key for translation. Could you give me a script for that, I can't write it myself. There are no two-button combinations (chording) on the mouse. I have avoided them so far - it seems an esoteric interface and there have been better ways of providing the necessary functionality. Middle mouse drag for translate could be enabled as an option, I suppose. As for the arrow keys for translate, add the following to your ~/.coot file. You might need to tweak zsc. Be slightly careful not to overly dwell with you finger on the button if you do not have a fast processor/graphics card and have a fast key repeat. Paul. (let* ((zsc 0.02) (screen-coords-nudge (lambda (tvm nudge ori) (map (lambda (e) (* nudge (apply + (map * e ori tvm))) (f (lambda (axes) (let ((nudge (* (zoom-factor) zsc)) (rc (rotation-centre)) (tvm (transpose-simple-matrix (view-matrix (apply set-rotation-centre (map + rc (screen-coords-nudge tvm nudge axes))) (add-key-binding Translate UpUp(lambda () (f '(0 1 0 (add-key-binding Translate Down Down (lambda () (f '(0 -1 0 (add-key-binding Translate Left Left (lambda () (f '(-1 0 0 (add-key-binding Translate Right Right (lambda () (f '( 1 0 0)
Re: [COOT] problems of version 0.6
Paul, Bernhard, Thank you for your reply. I am using version: 0.6-pre-1 (revision 1950) [with guile 1.6.8 embedded]. I will upgrade it with gtk2 in the name. Thank you for working on the torsion thing. Well, now help me with key-bindings, please. I need one or two buttons on the mouse for map dragging. Pymol and x-fit use the middle button for translation. It confuses me all the time, when I work in coot after working in pymol. Do you have any unused two-button combinations on the mouse? If it's not possible to touch the mouse buttons in the key-bindings, please help me to program arrow key for translation. Could you give me a script for that, I can't write it myself. Maia Paul Emsley wrote: Maia Cherney wrote: The general torsion function is not as good as the torsion in old X-fit. Are you or have you ever been a member of the von Delft Pressure Group? In x-fit you click on two atoms of a single bond and you can rotate the rest of the molecule around that bond. This bond can be in the middle of the main chain etc. It's a very useful function, if you need to rotate a polypeptide around a single bond. Are you planning to do a similar thing? Yes - we intend to have that before version 1.0. Paul.
Re: [COOT] problems of version 0.6
Thank you, Bernhard The buttons I like to change are internally assigned. For example, I would prefer to use key b to move backwards through residues of a chain. Right now you need two hands for that (shift-space). To drag the map, again, you need two hands (ctrl-left mouse). I would like to have some button assignments included in the preferences. There are so many keys on the key board, why do you need to combine several keys to do one function? Maia Bernhard Lohkamp wrote: I tried today the new version (-0.6) and see some problems. In general it helps if you can give revision numbers as things move quickly (especially in the pre-releases). 1. Simple mutate for nucleotides. The program makes Gr, Cr etc. in DNA (should be Gd, Cd etc). And it does not mutate to thymine at all, because there is no Tr. Where can I say that it's a DNA? Not entirely sure what happens there. Maybe Paul knows more. 2. Where is pukka puckers...?? They got (temporarily!?) removed (hidden) due to some problems. If you still keen in using them (no warranties) use the scripting function: Scheme: (pukka-puckers? imol) Python: pukka_puckers_qm(imol) My wish list: I am waiting for a version where users can reprogram keys. I would like to use only one-button keys, so that I could use only one hand (or better, one finger) when fitting. This is already in place (although you cannot change the internally assigned button). For some scheme examples: http://strucbio.biologie.uni-konstanz.de/ccp4wiki/index.php/COOT#Example_11:_Paul_Emsley.27s_Key_Bindings or in python: http://strucbio.biologie.uni-konstanz.de/ccp4wiki/index.php/COOT#Example_1:_Bernhard_Lohkamp.27s_Key_Bindings Hope this helps, B *** Dr. Bernhard Lohkamp Assistant Professor Div. Molecular Structural Biology Dept. of Medical Biochemistry and Biophysics (MBB) Karolinska Institutet S-17177 Stockholm Sweden phone: (+46) 08-52487673 fax: (+46) 08-327626 email: bernhard.lohk...@ki.se
