Re: [ccp4bb] frm2frm
You need to register at their site first, I suppose. Have you tried to ask them for the software ? At 06:06 26-10-2011, khuchtumur bumerdene wrote: Hello, Does anyone know where I could download frm2frm utility from Bruker? Is it even possible to do so? Industry and Medicine Applied Crystallography Macromolecular Crystallography Unit ___ Phones : (351-21) 446-9100 Ext. 1669 (351-21) 446-9669 (direct) Fax : (351-21) 441-1277 or 443-3644 email : mat...@itqb.unl.pt http://www.itqb.unl.pt/research/biological-chemistry/industry-and-medicine-applied-crystallography http://www.itqb.unl.pt/labs/macromolecular-crystallography-unit Mailing address : Instituto de Tecnologia Quimica e Biologica Apartado 127 2781-901 OEIRAS Portugal
Re: [ccp4bb] IUCr committees, depositing images
Since when has the cost of any project been limited by the cost of hardware? Someone has to /implement /this -- and make a career out of it; thunderingly absent from this thread has been the chorus of volunteers who will write the grant. phx On 25/10/2011 21:10, Herbert J. Bernstein wrote: To be fair to those concerned about cost, a more conservative estimate from the NSF RDLM workshop last summer in Princeton is $1,000 to $3,000 per terabyte per year for long term storage allowing for overhead in moderate-sized institutions such as the PDB. Larger entities, such as Google are able to do it for much lower annual costs in the range of $100 to $300 per terabyte per year. Indeed, if this becomes a serious effort, one might wish to consider involving the large storage farm businesses such as Google and Amazon. They might be willing to help support science partially in exchange for eyeballs going to their sites. Regards, H. J. Bernstein At 1:56 PM -0600 10/25/11, James Stroud wrote: On Oct 24, 2011, at 3:56 PM, James Holton wrote: The PDB only gets about 8000 depositions per year Just to put this into dollars. If each dataset is about 17 GB in size, then that's about 14 TB of storage that needs to come online every year to store the raw data for every structure. A two second search reveals that Newegg has a 3GB hitachi for $200. So that's about $1000 / year of storage for the raw data behind PDB deposits. James
Re: [ccp4bb] COOT not connected to PHENIX
-BEGIN PGP SIGNED MESSAGE- Hash: SHA1 Grand - with every python script one has to distribute a specific python version. More food for my prejudice against python ;-) Tim On 10/26/2011 12:15 AM, Paul Emsley wrote: FYI, from version 0.7, I will not distribute binaries without python. Paul. On 25/10/11 17:02, Yuri wrote: I installed the version with python embedded in coot And it works!! Thanks a lot!! On Tue, 25 Oct 2011 11:41:38 -0700, Nathaniel Echols wrote: On Tue, Oct 25, 2011 at 11:40 AM, Yuriyuri.pom...@ufl.edu wrote: Now here comes the stupid question... How do I fix it? Install a different coot version or is it something in my architecture? Install a different Coot. If you're downloading from Paul Emsley's page, you need a package with python in the file name. I have no idea whether the Linux binaries distributed by CCP4 have Python or not (the Mac version definitely does). -Nat - -- - -- Dr Tim Gruene Institut fuer anorganische Chemie Tammannstr. 4 D-37077 Goettingen GPG Key ID = A46BEE1A -BEGIN PGP SIGNATURE- Version: GnuPG v1.4.10 (GNU/Linux) Comment: Using GnuPG with Mozilla - http://enigmail.mozdev.org/ iD8DBQFOp8XQUxlJ7aRr7hoRAm9tAKCFFKmPIS+W7puqehbnJYuFhhQR5wCfUo+G ZAeWO1qzOhhW2xXTqLgPMpI= =53xa -END PGP SIGNATURE-
Re: [ccp4bb] frm2frm
-BEGIN PGP SIGNED MESSAGE- Hash: SHA1 Good luck - I registered with Bruker one or two years ago and have not heard a reply since ... Tim On 10/26/2011 09:32 AM, Pedro M. Matias wrote: You need to register at their site first, I suppose. Have you tried to ask them for the software ? At 06:06 26-10-2011, khuchtumur bumerdene wrote: Hello, Does anyone know where I could download frm2frm utility from Bruker? Is it even possible to do so? Industry and Medicine Applied Crystallography Macromolecular Crystallography Unit ___ Phones : (351-21) 446-9100 Ext. 1669 (351-21) 446-9669 (direct) Fax : (351-21) 441-1277 or 443-3644 email : mat...@itqb.unl.pt http://www.itqb.unl.pt/research/biological-chemistry/industry-and-medicine-applied-crystallography http://www.itqb.unl.pt/labs/macromolecular-crystallography-unit Mailing address : Instituto de Tecnologia Quimica e Biologica Apartado 127 2781-901 OEIRAS Portugal - -- - -- Dr Tim Gruene Institut fuer anorganische Chemie Tammannstr. 4 D-37077 Goettingen GPG Key ID = A46BEE1A -BEGIN PGP SIGNATURE- Version: GnuPG v1.4.10 (GNU/Linux) Comment: Using GnuPG with Mozilla - http://enigmail.mozdev.org/ iD8DBQFOp8YTUxlJ7aRr7hoRAofzAJ4oXdDvXvSue+aSHZbeA8M0X1rsOgCgzSdf BMOH2FVyTRX0uy/+i3+tyFI= =e2Vg -END PGP SIGNATURE-
Re: [ccp4bb] IUCr committees, depositing images
Hi James, Just to pick up on your point about the Pilatus detectors. Yesterday in 2 hours of giving a beamline a workout (admittedly with Thaumatin) we acquired 400 + GB of data*. Now I appreciate that this is not really routine operation, but it does raise an interesting point - if you have loaded a sample and centred it, collected test shots and decided it's not that great, why not collect anyway as it may later prove to be useful? Bzzt. 2 minutes or less later you have a full data set, and barely even time to go get a cup of tea. This does to some extent move the goalposts, as you can acquire far more data than you need. You never know, you may learn something interesting from it - perhaps it has different symmetry or packing? What it does mean is if we can have a method of tagging this data there may be massively more opportunity to get also-ran data sets for methods development types. What it also means however is that the cost of curating this data is then an order of magnitude higher. Also moving it around is also rather more painful. Anyhow, I would try to avoid dismissing the effect that new continuous readout detectors will have on data rates, from experience it is pretty substantial. Cheerio, Graeme *by data here what I mean is images, rather than information which is rather more time consuming to acquire. I would argue you get that from processing / analysing the data... On 24 October 2011 22:56, James Holton jmhol...@lbl.gov wrote: The Pilatus is fast, but or decades now we have had detectors that can read out in ~1s. This means that you can collect a typical ~100 image dataset in a few minutes (if flux is not limiting). Since there are ~150 beamlines currently operating around the world and they are open about 200 days/year, we should be collecting ~20,000,000 datasets each year. We're not. The PDB only gets about 8000 depositions per year, which means either we throw away 99.96% of our images, or we don't actually collect images anywhere near the ultimate capacity of the equipment we have. In my estimation, both of these play about equal roles, with ~50-fold attrition between ultimate data collection capacity and actual collected data, and another ~50 fold attrition between collected data sets and published structures. Personally, I think this means that the time it takes to collect the final dataset is not rate-limiting in a typical structural biology project/paper. This does not mean that the dataset is of little value. Quite the opposite! About 3000x more time and energy is expended preparing for the final dataset than is spent collecting it, and these efforts require experimental feedback. The trick is figuring out how best to compress the data used to solve a structure for archival storage. Do the previous data sets count? Or should the compression be lossy about such historical details? Does the stuff between the spots matter? After all, h,k,l,F,sigF is really just a form of data compression. In fact, there is no such thing as raw data. Even raw diffraction images are a simplification of the signals that came out of the detector electronics. But we round-off and average over a lot of things to remove noise. Largely because noise is difficult to compress. The question of how much compression is too much compression depends on which information (aka noise) you think could be important in the future. When it comes to fine-sliced data, such as that from Pilatus, the main reason why it doesn't compress very well is not because of the spots, but the background. It occupies thousands of times more pixels than the spots. Yes, there is diffuse scattering information in the background pixels, but this kind of data is MUCH smoother than the spot data (by definition), and therefore is optimally stored in larger pixels. Last year, I messed around a bit with applying different compression protocols to the spots and the background, and found that ~30 fold compression can be easily achieved if you apply h264 to the background and store the spots with lossless png compression: http://bl831.als.lbl.gov/~jamesh/lossy_compression/ I think these results speak to the relative information content of the spots and the pixels between them. Perhaps at least the online version of archived images could be in some sort of lossy-background format? With the real images in some sort of slower storage (like a room full of tapes that are available upon request)? Would 30-fold compression make the storage of image data tractable enough for some entity like the PDB to be able to afford it? I go to a lot of methods meetings, and it pains me to see the most brilliant minds in the field starved for interesting data sets. The problem is that it is very easy to get people to send you data that is so bad that it can't be solved by any software imaginable (I've got piles of that!). As a developer, what you really need is a right answer
Re: [ccp4bb] IUCr committees, depositing images
The archiving of all raw data and subsequently making it public is something that the large facilities are currently debating whether to do. Here at the ESRF we store user data for only 6 months (and I believe that it is available longer on tape) and we already have trouble with capacity. My personal view is that facilities should take the lead on this - for MX we already have a very good archiving system - ISPyB - also running at Diamond. ISPyB stores lots of meta data and jpgs of the raw images but not the images themselves but a link to the location of the data with an option to download if still available. My preferred option would be to store all academically funded data and then make it publicly available after say 2-5 years (this will no doubt spark another debate on time limits, special dispensation etc). What needs to be thought about is how to order the data and how to make sure that the correct meta data are stored with each data set - this will rely heavily on user input at the time of the experiment rather than gathering together data sets for depositions much later. As already mentioned, this type of resource could be extremely useful for developers and also as a general scientific resource. Smells like an EU grant to me. Cheers, Matt. On 26/10/2011 10:21, Frank von Delft wrote: Since when has the cost of any project been limited by the cost of hardware? Someone has to /implement /this -- and make a career out of it; thunderingly absent from this thread has been the chorus of volunteers who will write the grant. phx -- Matthew Bowler Structural Biology Group European Synchrotron Radiation Facility B.P. 220, 6 rue Jules Horowitz F-38043 GRENOBLE CEDEX FRANCE === Tel: +33 (0) 4.76.88.29.28 Fax: +33 (0) 4.76.88.29.04 http://go.esrf.eu/MX http://go.esrf.eu/Bowler ===
Re: [ccp4bb] frm2frm
-BEGIN PGP SIGNED MESSAGE- Hash: SHA1 Dear list, I just re-registered at http://www.brukersupport.com/Login.aspx?ReturnUrl=%2fdefault.aspx, and supported by a notifying email to Martin Adam (martin.a...@bruker-axs.de) the registration succeeded within a couple of minutes. Tim On 10/26/2011 10:34 AM, Tim Gruene wrote: Good luck - I registered with Bruker one or two years ago and have not heard a reply since ... Tim On 10/26/2011 09:32 AM, Pedro M. Matias wrote: You need to register at their site first, I suppose. Have you tried to ask them for the software ? At 06:06 26-10-2011, khuchtumur bumerdene wrote: Hello, Does anyone know where I could download frm2frm utility from Bruker? Is it even possible to do so? Industry and Medicine Applied Crystallography Macromolecular Crystallography Unit ___ Phones : (351-21) 446-9100 Ext. 1669 (351-21) 446-9669 (direct) Fax : (351-21) 441-1277 or 443-3644 email : mat...@itqb.unl.pt http://www.itqb.unl.pt/research/biological-chemistry/industry-and-medicine-applied-crystallography http://www.itqb.unl.pt/labs/macromolecular-crystallography-unit Mailing address : Instituto de Tecnologia Quimica e Biologica Apartado 127 2781-901 OEIRAS Portugal - -- - -- Dr Tim Gruene Institut fuer anorganische Chemie Tammannstr. 4 D-37077 Goettingen GPG Key ID = A46BEE1A -BEGIN PGP SIGNATURE- Version: GnuPG v1.4.10 (GNU/Linux) Comment: Using GnuPG with Mozilla - http://enigmail.mozdev.org/ iD8DBQFOp88PUxlJ7aRr7hoRAnlhAKD+6+9qCP/WClVAQlyeoiQ/x1OgbwCeOUYQ 2YabkSUBjUaUR0xtzbqrbkM= =jasv -END PGP SIGNATURE-
Re: [ccp4bb] IUCr committees, depositing images
This raises an important point. The new continuous readout detectors such as the Pilatus for beamlines or the Bruker Photon for in-house use enable the crystal to be rotated at constant velocity, eliminating the mechanical errors associated with 'stop and go' data collection. Storing their data in 'frames' is an artifical construction that is currently required for the established data integration programs but is in fact throwing away information. Maybe in 10 years time 'frames' will be as obsolete as punched cards! George On Wed, Oct 26, 2011 at 09:39:40AM +0100, Graeme Winter wrote: Hi James, Just to pick up on your point about the Pilatus detectors. Yesterday in 2 hours of giving a beamline a workout (admittedly with Thaumatin) we acquired 400 + GB of data*. Now I appreciate that this is not really routine operation, but it does raise an interesting point - if you have loaded a sample and centred it, collected test shots and decided it's not that great, why not collect anyway as it may later prove to be useful? Bzzt. 2 minutes or less later you have a full data set, and barely even time to go get a cup of tea. This does to some extent move the goalposts, as you can acquire far more data than you need. You never know, you may learn something interesting from it - perhaps it has different symmetry or packing? What it does mean is if we can have a method of tagging this data there may be massively more opportunity to get also-ran data sets for methods development types. What it also means however is that the cost of curating this data is then an order of magnitude higher. Also moving it around is also rather more painful. Anyhow, I would try to avoid dismissing the effect that new continuous readout detectors will have on data rates, from experience it is pretty substantial. Cheerio, Graeme *by data here what I mean is images, rather than information which is rather more time consuming to acquire. I would argue you get that from processing / analysing the data... On 24 October 2011 22:56, James Holton jmhol...@lbl.gov wrote: The Pilatus is fast, but or decades now we have had detectors that can read out in ~1s. This means that you can collect a typical ~100 image dataset in a few minutes (if flux is not limiting). Since there are ~150 beamlines currently operating around the world and they are open about 200 days/year, we should be collecting ~20,000,000 datasets each year. We're not. The PDB only gets about 8000 depositions per year, which means either we throw away 99.96% of our images, or we don't actually collect images anywhere near the ultimate capacity of the equipment we have. In my estimation, both of these play about equal roles, with ~50-fold attrition between ultimate data collection capacity and actual collected data, and another ~50 fold attrition between collected data sets and published structures. Personally, I think this means that the time it takes to collect the final dataset is not rate-limiting in a typical structural biology project/paper. This does not mean that the dataset is of little value. Quite the opposite! About 3000x more time and energy is expended preparing for the final dataset than is spent collecting it, and these efforts require experimental feedback. The trick is figuring out how best to compress the data used to solve a structure for archival storage. Do the previous data sets count? Or should the compression be lossy about such historical details? Does the stuff between the spots matter? After all, h,k,l,F,sigF is really just a form of data compression. In fact, there is no such thing as raw data. Even raw diffraction images are a simplification of the signals that came out of the detector electronics. But we round-off and average over a lot of things to remove noise. Largely because noise is difficult to compress. The question of how much compression is too much compression depends on which information (aka noise) you think could be important in the future. When it comes to fine-sliced data, such as that from Pilatus, the main reason why it doesn't compress very well is not because of the spots, but the background. It occupies thousands of times more pixels than the spots. Yes, there is diffuse scattering information in the background pixels, but this kind of data is MUCH smoother than the spot data (by definition), and therefore is optimally stored in larger pixels. Last year, I messed around a bit with applying different compression protocols to the spots and the background, and found that ~30 fold compression can be easily achieved if you apply h264 to the background and store the spots with lossless png compression: http://bl831.als.lbl.gov/~jamesh/lossy_compression/ I think these results speak to the relative information content of the spots and the pixels between them. Perhaps at least the
Re: [ccp4bb] IUCr committees, depositing images
Dear James, Good analysis! You bring up important points. On 10/24/11 23:56, James Holton wrote: The Pilatus is fast, but or decades now we have had detectors that can read out in ~1s. This means that you can collect a typical ~100 image dataset in a few minutes (if flux is not limiting). Since there are ~150 beamlines currently operating around the world and they are open about 200 days/year, we should be collecting ~20,000,000 datasets each year. We're not. The PDB only gets about 8000 depositions per year, which means either we throw away 99.96% of our images, or we don't actually collect images anywhere near the ultimate capacity of the equipment we have. In my estimation, both of these play about equal roles, with ~50-fold attrition between ultimate data collection capacity and actual collected data, and another ~50 fold attrition between collected data sets and published structures. Your estimation says: we collect 1/50 * 20,000,000 = 400,000 data sets of which only 8,000 get deposited. An average Pilatus data set (0.1 degree scan) takes about 4 Gb (compressed, without loosing information. EVAL can read those!). Storing the 8,000 data sets, as James Stroud mentions, can not be the problem. It is the 392,000 other data sets that we have to find a home for. That would be 1568 Tb and would cost 49,000 $/year. This may be a slight overestimation, but it shows us the problems we face if we want to store ALL raw data. Even if we would find a way to store all these data, how would we set up a useful data base? If we store all data by name, date and beamline, we will in the end inevitable be drowning is a sea of information. It is very unlikely that the very interesting data sets will ever be found and used. It would be much more useful if every data sets would be annotated by the user or beam line scientist. Like: impossible to index, bad data from integration step, overlap, diffuse streaks etc. Such information could be part of the meta data. This however, takes time and may not fit the eagerness to get results from one of the other data sets recorded at the same synchrotron trip. I am afraid that just throwing data sets in a big pool, will not be very useful. Loes. -- __ Dr. Loes Kroon-Batenburg Dept. of Crystal and Structural Chemistry Bijvoet Center for Biomolecular Research Utrecht University Padualaan 8, 3584 CH Utrecht The Netherlands E-mail : l.m.j.kroon-batenb...@uu.nl phone : +31-30-2532865 fax: +31-30-2533940 __
[ccp4bb] Program announcement: Nautilus v0.2
New version of Nautilus (my nucleic acid building program). Main changes: - Now available for both OSX(x86) and Linux. - It no longer corrupts the residue names of any existing non-nucleic acid model you feed in. - Most cases where the output model clashes with itself have been fixed. - I've done some (fairly conservative) optimisations. It's now very fast. Very, very fast. Faster than something quite fast moving not entirely slowly. It is available from here: http://www.ysbl.york.ac.uk/~cowtan/nautilus/nautilus.html .. 'Nautilus' is a program for automatic model building of nucleotide structures in electron density maps. It will trace a map with no model, extend an existing model, or add nucleotide chains to an existing non-nucleotide model. 'nautilus' does not currently perform refinement - you will need to refine and recycle for further model building yourself. Neither does it assign sequence - the model is built as ploy-U. This is an alpha release. It may not work at all. It has only been tested on synthetic data with simulated errors. The API will change significantly in subsequent releases.
Re: [ccp4bb] IUCr committees, depositing images
Dear Frank, re 'who will write the grant?'. This is not as easy as it sounds, would that it were! There are two possible business plans:- Option 1. Specifically for MX is the PDB as the first and foremost candidate to seek such additional funds for full diffraction data deposition for each future PDB deposiition entry. This business plan possibility is best answered by PDB/EBI (eg Gerard Kleywegt has answered this in the negative thus far at the CCP4 January 2010). Option 2 The Journals that host the publications could add the cost to the subscriber and/or the author according to their funding model. As an example and as a start a draft business plan has been written by one of us [JRH] for IUCr Acta Cryst E; this seemed attractive because of its simpler 'author pays' financing. This proposed business plan is now with IUCr Journals to digest and hopefully refine. Initial indications are that Acta Cryst C would be perceived by IUCr Journals as a better place to start considering this in detail, as it involves fewer crystal structures than Acta E and would thus be more manageable. The overall advantage of the responsibility being with Journals as we see it is that it encourages such 'archiving of data with literature' across all crystallography related techniques (single crystal, SAXS, SANS, Electron crystallography etc) and fields (Biology, Chemistry, Materials, Condensed Matter Physics etc) ie not just one technique and field, although obviously biology is dear to our hearts here in the CCP4bb. Yours sincerely, John and Tom John Helliwell and Tom Terwilliger On Wed, Oct 26, 2011 at 9:21 AM, Frank von Delft frank.vonde...@sgc.ox.ac.uk wrote: Since when has the cost of any project been limited by the cost of hardware? Someone has to implement this -- and make a career out of it; thunderingly absent from this thread has been the chorus of volunteers who will write the grant. phx On 25/10/2011 21:10, Herbert J. Bernstein wrote: To be fair to those concerned about cost, a more conservative estimate from the NSF RDLM workshop last summer in Princeton is $1,000 to $3,000 per terabyte per year for long term storage allowing for overhead in moderate-sized institutions such as the PDB. Larger entities, such as Google are able to do it for much lower annual costs in the range of $100 to $300 per terabyte per year. Indeed, if this becomes a serious effort, one might wish to consider involving the large storage farm businesses such as Google and Amazon. They might be willing to help support science partially in exchange for eyeballs going to their sites. Regards, H. J. Bernstein At 1:56 PM -0600 10/25/11, James Stroud wrote: On Oct 24, 2011, at 3:56 PM, James Holton wrote: The PDB only gets about 8000 depositions per year Just to put this into dollars. If each dataset is about 17 GB in size, then that's about 14 TB of storage that needs to come online every year to store the raw data for every structure. A two second search reveals that Newegg has a 3GB hitachi for $200. So that's about $1000 / year of storage for the raw data behind PDB deposits. James -- Professor John R Helliwell DSc
Re: [ccp4bb] COOT not connected to PHENIX
On Wed, 2011-10-26 at 10:33 +0200, Tim Gruene wrote: with every python script one has to distribute a specific python version ... and with every program one has to distribute binaries for every platform... more food for my prejudice against software ;-) This really is not about python, it's about distributing with or without dependencies. And you are absolutely right about that: for example, ccp4-6.2.0 comes with python2.6.7 embedded, and, if one goes with defaults and downloads coot with it, python2.6 in coot's lib folder. Same with phenix - you get python2.7 with it and python2.4 with pymol0.99 that comes with it. By the way, I already have another pymol that I compiled myself (1.4) and the one from ubuntu repositories (1.2). Except for the latter, each carries its own copy of whichever python it needs. Every single python avatar takes 50-100Mb of space, which is fortunately not in short supply. This is why the right way to distribute *nix software is to distribute software itself and ask the end-user to get all the dependencies (not that hard these days). It is fully understood, of course, that people that do this for living find it more troublesome to deal with me whining about how their software is screwing up my matplotlib than to just give me another python copy. What's an extra 50Mb between friends ;-) Cheers, Ed. -- I'd jump in myself, if I weren't so good at whistling. Julian, King of Lemurs
[ccp4bb] Question about drawing disulfide bonds in Pymol
Dear All, I have a question regarding making disulfide bonds in Pymol. The overall structure is in cartoon representation. When I display the disulfide bond residues as stick models, the cysteine residues sitting on the beta strand have a gap from the actual beta-strand. If I check off the flat sheet option, all the beta-strands look very wavy, but the cysteines are actually on the beta strands. I want to just change the two beta-strands with cysteine residues on it as wavy strands and keep other beta-strands flat looking. Does anyone know a way to do this or has better ways to draw individual residues on beta-strands. Thanks! jiyuan Jiyuan Ke, Ph.D. Research Scientist Van Andel Research Institute 333 Bostwick Ave NE Grand Rapids, MI 49503 The information transmitted is intended only for the person or entity to which it is addressed and may contain confidential and/or privileged material. Any review, retransmission, dissemination or other use of, or taking of any action in reliance upon, this information by persons or entities other than the intended recipient is prohibited. If you received this in error, please contact the sender and delete the material from any computer.
Re: [ccp4bb] frm2frm
This thread may be relevant http://www.mail-archive.com/ccp4bb@jiscmail.ac.uk/msg18422.html On Wed, 2011-10-26 at 15:06 +1000, khuchtumur bumerdene wrote: Hello, Does anyone know where I could download frm2frm utility from Bruker? Is it even possible to do so? -- Hurry up before we all come back to our senses! Julian, King of Lemurs
Re: [ccp4bb] IUCr committees, depositing images
Dear John and colleagues, There seem to be a set a centrifugal forces at play within this thread that are distracting us from a sensible path of concrete action by throwing decoys in every conceivable direction, e.g. * Pilatus detectors spew out such a volume of data that we can't possibly archive it all - does that mean that because the 5th generation of Dectris detectors will be able to write one billion images a second and catch every scattered photon individually, we should not try and archive more information than is given by the current merged structure factor data? That seems a complete failure of reasoning to me: there must be a sensible form of raw data archiving that would stand between those two extremes and would retain much more information that the current merged data but would step back from the enormous degree of oversampling of the raw diffraction pattern that the Pilatus and its successors are capable of. * It is all going to cost an awful lot of money, therefore we need a team of grant writers to raise its hand and volunteer to apply for resources from one or more funding agencies - there again there is an avoidance of the feasible by invocation of the impossible. The IUCr Forum already has an outline of a feasibility study that would cost only a small amount of joined-up thinking and book-keeping around already stored information, so let us not use the inaccessibility of federal or EC funding as a scarecrow to justify not even trying what is proposed there. And the idea that someone needs to decide to stake his/her career on this undertaking seems totally overblown. Several people have already pointed out that the sets of images that would need to be archived would be a very small subset of the bulk of datasets that are being held on the storage systems of synchrotron sources. What needs to be done, as already described, is to be able to refer to those few datasets that gave rise to the integrated data against which deposited structures were refined (or, in some cases, solved by experimental phasing), to give them special status in terms of making them visible and accessible on-line at the same time as the pdb entry itself (rather than after the statutory 2-5 years that would apply to all the rest, probably in a more off-line form), and to maintain that accessibility for ever, with a link from the pdb entry and perhaps from the associated publication. It seems unlikely that this would involve the mobilisation of such large resources as to require either a human sacrifice (of the poor person whose life would be staked on this gamble) or writing a grant application, with the indefinite postponement of action and the loss of motivation this would imply. Coming back to the more technical issue of bloated datasets, it is a scientific problem that must be amenable to rational analysis to decide on a sensible form of compression of overly-verbose sets of thin-sliced, perhaps low-exposure images that would already retain a large fraction, if not all, of the extra information on which we would wish future improved versions of processing programs to cut their teeth, for a long time to come. This approach would seem preferable to stoking up irrational fears of not being able to cope with the most exaggerated predictions of the volumes of data to archive, and thus doing nothing at all. I very much hope that the can do spirit that marked the final discussions of the DDDWG (Diffraction Data Deposition Working Group) in Madrid will emerge on top of all the counter-arguments that consist in moving the goal posts to prove that the initial goal is unreachable. With best wishes, Gerard. -- On Wed, Oct 26, 2011 at 02:18:25PM +0100, John R Helliwell wrote: Dear Frank, re 'who will write the grant?'. This is not as easy as it sounds, would that it were! There are two possible business plans:- Option 1. Specifically for MX is the PDB as the first and foremost candidate to seek such additional funds for full diffraction data deposition for each future PDB deposiition entry. This business plan possibility is best answered by PDB/EBI (eg Gerard Kleywegt has answered this in the negative thus far at the CCP4 January 2010). Option 2 The Journals that host the publications could add the cost to the subscriber and/or the author according to their funding model. As an example and as a start a draft business plan has been written by one of us [JRH] for IUCr Acta Cryst E; this seemed attractive because of its simpler 'author pays' financing. This proposed business plan is now with IUCr Journals to digest and hopefully refine. Initial indications are that Acta Cryst C would be perceived by IUCr Journals as a better place to start considering this in detail, as it involves fewer crystal structures than Acta E and would thus be more manageable. The overall advantage of the responsibility being with Journals as we see it is that it
[ccp4bb] Structural biologist position at Constellation Pharmaceuticals
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Re: [ccp4bb] Question about drawing disulfide bonds in Pymol
On Wed, 2011-10-26 at 10:15 -0400, Ke, Jiyuan wrote: flat sheet option IIUC, the set command in pymol allows per-selection application, i.e. if you try this in the command line instead of checking the option in the menu set cartoon_flat_sheets, 0, blah where blah is your selection. -- Oh, suddenly throwing a giraffe into a volcano to make water is crazy? Julian, King of Lemurs
Re: [ccp4bb] IUCr committees, depositing images
Could you perhaps use the principle of capture storage that is used by wild-life photographers with high-speed cameras? The principle is that the movie is written to the same area of memory, jumping back to the beginning when it is full (this part is not essential, but it makes the principle clear). Then, when the photographer takes his finger off the trigger, the last x seconds is permanently stored. So you keep your wits about you, and press the metaphorical store button just *after *you have got the movie in the can so to speak Just a thought Patrick On Wed, Oct 26, 2011 at 2:18 PM, John R Helliwell jrhelliw...@gmail.comwrote: Dear Frank, re 'who will write the grant?'. This is not as easy as it sounds, would that it were! There are two possible business plans:- Option 1. Specifically for MX is the PDB as the first and foremost candidate to seek such additional funds for full diffraction data deposition for each future PDB deposiition entry. This business plan possibility is best answered by PDB/EBI (eg Gerard Kleywegt has answered this in the negative thus far at the CCP4 January 2010). Option 2 The Journals that host the publications could add the cost to the subscriber and/or the author according to their funding model. As an example and as a start a draft business plan has been written by one of us [JRH] for IUCr Acta Cryst E; this seemed attractive because of its simpler 'author pays' financing. This proposed business plan is now with IUCr Journals to digest and hopefully refine. Initial indications are that Acta Cryst C would be perceived by IUCr Journals as a better place to start considering this in detail, as it involves fewer crystal structures than Acta E and would thus be more manageable. The overall advantage of the responsibility being with Journals as we see it is that it encourages such 'archiving of data with literature' across all crystallography related techniques (single crystal, SAXS, SANS, Electron crystallography etc) and fields (Biology, Chemistry, Materials, Condensed Matter Physics etc) ie not just one technique and field, although obviously biology is dear to our hearts here in the CCP4bb. Yours sincerely, John and Tom John Helliwell and Tom Terwilliger On Wed, Oct 26, 2011 at 9:21 AM, Frank von Delft frank.vonde...@sgc.ox.ac.uk wrote: Since when has the cost of any project been limited by the cost of hardware? Someone has to implement this -- and make a career out of it; thunderingly absent from this thread has been the chorus of volunteers who will write the grant. phx On 25/10/2011 21:10, Herbert J. Bernstein wrote: To be fair to those concerned about cost, a more conservative estimate from the NSF RDLM workshop last summer in Princeton is $1,000 to $3,000 per terabyte per year for long term storage allowing for overhead in moderate-sized institutions such as the PDB. Larger entities, such as Google are able to do it for much lower annual costs in the range of $100 to $300 per terabyte per year. Indeed, if this becomes a serious effort, one might wish to consider involving the large storage farm businesses such as Google and Amazon. They might be willing to help support science partially in exchange for eyeballs going to their sites. Regards, H. J. Bernstein At 1:56 PM -0600 10/25/11, James Stroud wrote: On Oct 24, 2011, at 3:56 PM, James Holton wrote: The PDB only gets about 8000 depositions per year Just to put this into dollars. If each dataset is about 17 GB in size, then that's about 14 TB of storage that needs to come online every year to store the raw data for every structure. A two second search reveals that Newegg has a 3GB hitachi for $200. So that's about $1000 / year of storage for the raw data behind PDB deposits. James -- Professor John R Helliwell DSc -- patr...@douglas.co.ukDouglas Instruments Ltd. Douglas House, East Garston, Hungerford, Berkshire, RG17 7HD, UK Directors: Peter Baldock, Patrick Shaw Stewart http://www.douglas.co.uk Tel: 44 (0) 148-864-9090US toll-free 1-877-225-2034 Regd. England 2177994, VAT Reg. GB 480 7371 36
Re: [ccp4bb] IUCr committees, depositing images
I just want to jump in to state that I am ALL FOR the notion of depositing the images that go with the structure factors and the refined structure. Through the years, I have been interviewing folks about the strange satellite diffraction they saw, but ignored, used the mains that they could integrate and deposited that structure, does not help me to justify the existance of modulated protein crystals to reviewers. But if I could go and retrieve those images, and reanalyze with new methods. Dream come true. Reviewers convinced. On Wed, Oct 26, 2011 at 10:59 AM, Patrick Shaw Stewart patr...@douglas.co.uk wrote: Could you perhaps use the principle of capture storage that is used by wild-life photographers with high-speed cameras? The principle is that the movie is written to the same area of memory, jumping back to the beginning when it is full (this part is not essential, but it makes the principle clear). Then, when the photographer takes his finger off the trigger, the last x seconds is permanently stored. So you keep your wits about you, and press the metaphorical store button just after you have got the movie in the can so to speak Just a thought Patrick On Wed, Oct 26, 2011 at 2:18 PM, John R Helliwell jrhelliw...@gmail.com wrote: Dear Frank, re 'who will write the grant?'. This is not as easy as it sounds, would that it were! There are two possible business plans:- Option 1. Specifically for MX is the PDB as the first and foremost candidate to seek such additional funds for full diffraction data deposition for each future PDB deposiition entry. This business plan possibility is best answered by PDB/EBI (eg Gerard Kleywegt has answered this in the negative thus far at the CCP4 January 2010). Option 2 The Journals that host the publications could add the cost to the subscriber and/or the author according to their funding model. As an example and as a start a draft business plan has been written by one of us [JRH] for IUCr Acta Cryst E; this seemed attractive because of its simpler 'author pays' financing. This proposed business plan is now with IUCr Journals to digest and hopefully refine. Initial indications are that Acta Cryst C would be perceived by IUCr Journals as a better place to start considering this in detail, as it involves fewer crystal structures than Acta E and would thus be more manageable. The overall advantage of the responsibility being with Journals as we see it is that it encourages such 'archiving of data with literature' across all crystallography related techniques (single crystal, SAXS, SANS, Electron crystallography etc) and fields (Biology, Chemistry, Materials, Condensed Matter Physics etc) ie not just one technique and field, although obviously biology is dear to our hearts here in the CCP4bb. Yours sincerely, John and Tom John Helliwell and Tom Terwilliger On Wed, Oct 26, 2011 at 9:21 AM, Frank von Delft frank.vonde...@sgc.ox.ac.uk wrote: Since when has the cost of any project been limited by the cost of hardware? Someone has to implement this -- and make a career out of it; thunderingly absent from this thread has been the chorus of volunteers who will write the grant. phx On 25/10/2011 21:10, Herbert J. Bernstein wrote: To be fair to those concerned about cost, a more conservative estimate from the NSF RDLM workshop last summer in Princeton is $1,000 to $3,000 per terabyte per year for long term storage allowing for overhead in moderate-sized institutions such as the PDB. Larger entities, such as Google are able to do it for much lower annual costs in the range of $100 to $300 per terabyte per year. Indeed, if this becomes a serious effort, one might wish to consider involving the large storage farm businesses such as Google and Amazon. They might be willing to help support science partially in exchange for eyeballs going to their sites. Regards, H. J. Bernstein At 1:56 PM -0600 10/25/11, James Stroud wrote: On Oct 24, 2011, at 3:56 PM, James Holton wrote: The PDB only gets about 8000 depositions per year Just to put this into dollars. If each dataset is about 17 GB in size, then that's about 14 TB of storage that needs to come online every year to store the raw data for every structure. A two second search reveals that Newegg has a 3GB hitachi for $200. So that's about $1000 / year of storage for the raw data behind PDB deposits. James -- Professor John R Helliwell DSc -- patr...@douglas.co.uk Douglas Instruments Ltd. Douglas House, East Garston, Hungerford, Berkshire, RG17 7HD, UK Directors: Peter Baldock, Patrick Shaw Stewart http://www.douglas.co.uk Tel: 44 (0) 148-864-9090 US toll-free 1-877-225-2034 Regd. England 2177994, VAT Reg. GB 480 7371 36
Re: [ccp4bb] IUCr committees, depositing images
Is anyone seriously questioning whether we should archive the images used for published structures? That amount of space is trivial, could be implemented just as another link in the PDB website, and would be really helpful in some cases. One person could set it up in a day! You could just make it a policy: no images, no PDB submission, no publishing! Jacob On Wed, Oct 26, 2011 at 11:15 AM, Gloria Borgstahl gborgst...@gmail.com wrote: I just want to jump in to state that I am ALL FOR the notion of depositing the images that go with the structure factors and the refined structure. Through the years, I have been interviewing folks about the strange satellite diffraction they saw, but ignored, used the mains that they could integrate and deposited that structure, does not help me to justify the existance of modulated protein crystals to reviewers. But if I could go and retrieve those images, and reanalyze with new methods. Dream come true. Reviewers convinced. On Wed, Oct 26, 2011 at 10:59 AM, Patrick Shaw Stewart patr...@douglas.co.uk wrote: Could you perhaps use the principle of capture storage that is used by wild-life photographers with high-speed cameras? The principle is that the movie is written to the same area of memory, jumping back to the beginning when it is full (this part is not essential, but it makes the principle clear). Then, when the photographer takes his finger off the trigger, the last x seconds is permanently stored. So you keep your wits about you, and press the metaphorical store button just after you have got the movie in the can so to speak Just a thought Patrick On Wed, Oct 26, 2011 at 2:18 PM, John R Helliwell jrhelliw...@gmail.com wrote: Dear Frank, re 'who will write the grant?'. This is not as easy as it sounds, would that it were! There are two possible business plans:- Option 1. Specifically for MX is the PDB as the first and foremost candidate to seek such additional funds for full diffraction data deposition for each future PDB deposiition entry. This business plan possibility is best answered by PDB/EBI (eg Gerard Kleywegt has answered this in the negative thus far at the CCP4 January 2010). Option 2 The Journals that host the publications could add the cost to the subscriber and/or the author according to their funding model. As an example and as a start a draft business plan has been written by one of us [JRH] for IUCr Acta Cryst E; this seemed attractive because of its simpler 'author pays' financing. This proposed business plan is now with IUCr Journals to digest and hopefully refine. Initial indications are that Acta Cryst C would be perceived by IUCr Journals as a better place to start considering this in detail, as it involves fewer crystal structures than Acta E and would thus be more manageable. The overall advantage of the responsibility being with Journals as we see it is that it encourages such 'archiving of data with literature' across all crystallography related techniques (single crystal, SAXS, SANS, Electron crystallography etc) and fields (Biology, Chemistry, Materials, Condensed Matter Physics etc) ie not just one technique and field, although obviously biology is dear to our hearts here in the CCP4bb. Yours sincerely, John and Tom John Helliwell and Tom Terwilliger On Wed, Oct 26, 2011 at 9:21 AM, Frank von Delft frank.vonde...@sgc.ox.ac.uk wrote: Since when has the cost of any project been limited by the cost of hardware? Someone has to implement this -- and make a career out of it; thunderingly absent from this thread has been the chorus of volunteers who will write the grant. phx On 25/10/2011 21:10, Herbert J. Bernstein wrote: To be fair to those concerned about cost, a more conservative estimate from the NSF RDLM workshop last summer in Princeton is $1,000 to $3,000 per terabyte per year for long term storage allowing for overhead in moderate-sized institutions such as the PDB. Larger entities, such as Google are able to do it for much lower annual costs in the range of $100 to $300 per terabyte per year. Indeed, if this becomes a serious effort, one might wish to consider involving the large storage farm businesses such as Google and Amazon. They might be willing to help support science partially in exchange for eyeballs going to their sites. Regards, H. J. Bernstein At 1:56 PM -0600 10/25/11, James Stroud wrote: On Oct 24, 2011, at 3:56 PM, James Holton wrote: The PDB only gets about 8000 depositions per year Just to put this into dollars. If each dataset is about 17 GB in size, then that's about 14 TB of storage that needs to come online every year to store the raw data for every structure. A two second search reveals that Newegg has a 3GB hitachi for $200. So that's about $1000 / year of storage for the raw data behind PDB deposits. James -- Professor John R Helliwell DSc --
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[ccp4bb] cryo protection
Hi All, I have run into a very sensitive crystals system when it comes to cryo protecting them. I have run through the usual suspects and trays are going to be setup with a cryo protectant as part of crystallization cocktail. The one problem that seems to be occurring is that the crystals crack as soon as they are transfered out of the original drop. I am running out of ideas and really would love some new ones. Thanks in advance. Len Leonard Thomas Ph.D. Macromolecular Crystallography Laboratory Manager University of Oklahoma Department of Chemistry and Biochemistry Stephenson Life Sciences Research Center 101 Stephenson Parkway Norman, OK 73019-5251 lmtho...@ou.edu http://barlywine.chem.ou.edu Office: (405)325-1126 Lab: (405)325-7571
Re: [ccp4bb] IUCr committees, depositing images
Cool - we've found our volunteer!! On 26/10/2011 17:28, Jacob Keller wrote: Is anyone seriously questioning whether we should archive the images used for published structures? That amount of space is trivial, could be implemented just as another link in the PDB website, and would be really helpful in some cases. One person could set it up in a day! You could just make it a policy: no images, no PDB submission, no publishing! Jacob On Wed, Oct 26, 2011 at 11:15 AM, Gloria Borgstahlgborgst...@gmail.com wrote: I just want to jump in to state that I am ALL FOR the notion of depositing the images that go with the structure factors and the refined structure. Through the years, I have been interviewing folks about the strange satellite diffraction they saw, but ignored, used the mains that they could integrate and deposited that structure, does not help me to justify the existance of modulated protein crystals to reviewers. But if I could go and retrieve those images, and reanalyze with new methods. Dream come true. Reviewers convinced. On Wed, Oct 26, 2011 at 10:59 AM, Patrick Shaw Stewart patr...@douglas.co.uk wrote: Could you perhaps use the principle of capture storage that is used by wild-life photographers with high-speed cameras? The principle is that the movie is written to the same area of memory, jumping back to the beginning when it is full (this part is not essential, but it makes the principle clear). Then, when the photographer takes his finger off the trigger, the last x seconds is permanently stored. So you keep your wits about you, and press the metaphorical store button just after you have got the movie in the can so to speak Just a thought Patrick On Wed, Oct 26, 2011 at 2:18 PM, John R Helliwelljrhelliw...@gmail.com wrote: Dear Frank, re 'who will write the grant?'. This is not as easy as it sounds, would that it were! There are two possible business plans:- Option 1. Specifically for MX is the PDB as the first and foremost candidate to seek such additional funds for full diffraction data deposition for each future PDB deposiition entry. This business plan possibility is best answered by PDB/EBI (eg Gerard Kleywegt has answered this in the negative thus far at the CCP4 January 2010). Option 2 The Journals that host the publications could add the cost to the subscriber and/or the author according to their funding model. As an example and as a start a draft business plan has been written by one of us [JRH] for IUCr Acta Cryst E; this seemed attractive because of its simpler 'author pays' financing. This proposed business plan is now with IUCr Journals to digest and hopefully refine. Initial indications are that Acta Cryst C would be perceived by IUCr Journals as a better place to start considering this in detail, as it involves fewer crystal structures than Acta E and would thus be more manageable. The overall advantage of the responsibility being with Journals as we see it is that it encourages such 'archiving of data with literature' across all crystallography related techniques (single crystal, SAXS, SANS, Electron crystallography etc) and fields (Biology, Chemistry, Materials, Condensed Matter Physics etc) ie not just one technique and field, although obviously biology is dear to our hearts here in the CCP4bb. Yours sincerely, John and Tom John Helliwell and Tom Terwilliger On Wed, Oct 26, 2011 at 9:21 AM, Frank von Delft frank.vonde...@sgc.ox.ac.uk wrote: Since when has the cost of any project been limited by the cost of hardware? Someone has to implement this -- and make a career out of it; thunderingly absent from this thread has been the chorus of volunteers who will write the grant. phx On 25/10/2011 21:10, Herbert J. Bernstein wrote: To be fair to those concerned about cost, a more conservative estimate from the NSF RDLM workshop last summer in Princeton is $1,000 to $3,000 per terabyte per year for long term storage allowing for overhead in moderate-sized institutions such as the PDB. Larger entities, such as Google are able to do it for much lower annual costs in the range of $100 to $300 per terabyte per year. Indeed, if this becomes a serious effort, one might wish to consider involving the large storage farm businesses such as Google and Amazon. They might be willing to help support science partially in exchange for eyeballs going to their sites. Regards, H. J. Bernstein At 1:56 PM -0600 10/25/11, James Stroud wrote: On Oct 24, 2011, at 3:56 PM, James Holton wrote: The PDB only gets about 8000 depositions per year Just to put this into dollars. If each dataset is about 17 GB in size, then that's about 14 TB of storage that needs to come online every year to store the raw data for every structure. A two second search reveals that Newegg has a 3GB hitachi for $200. So that's about $1000 / year of storage for the raw data behind PDB deposits. James -- Professor John R Helliwell DSc -- patr...@douglas.co.ukDouglas Instruments
Re: [ccp4bb] IUCr committees, depositing images
Touche! But alas, I have no access to the PDB's server, so... JPK On Wed, Oct 26, 2011 at 11:54 AM, Frank von Delft frank.vonde...@sgc.ox.ac.uk wrote: Cool - we've found our volunteer!! On 26/10/2011 17:28, Jacob Keller wrote: Is anyone seriously questioning whether we should archive the images used for published structures? That amount of space is trivial, could be implemented just as another link in the PDB website, and would be really helpful in some cases. One person could set it up in a day! You could just make it a policy: no images, no PDB submission, no publishing! Jacob On Wed, Oct 26, 2011 at 11:15 AM, Gloria Borgstahlgborgst...@gmail.com wrote: I just want to jump in to state that I am ALL FOR the notion of depositing the images that go with the structure factors and the refined structure. Through the years, I have been interviewing folks about the strange satellite diffraction they saw, but ignored, used the mains that they could integrate and deposited that structure, does not help me to justify the existance of modulated protein crystals to reviewers. But if I could go and retrieve those images, and reanalyze with new methods. Dream come true. Reviewers convinced. On Wed, Oct 26, 2011 at 10:59 AM, Patrick Shaw Stewart patr...@douglas.co.uk wrote: Could you perhaps use the principle of capture storage that is used by wild-life photographers with high-speed cameras? The principle is that the movie is written to the same area of memory, jumping back to the beginning when it is full (this part is not essential, but it makes the principle clear). Then, when the photographer takes his finger off the trigger, the last x seconds is permanently stored. So you keep your wits about you, and press the metaphorical store button just after you have got the movie in the can so to speak Just a thought Patrick On Wed, Oct 26, 2011 at 2:18 PM, John R Helliwelljrhelliw...@gmail.com wrote: Dear Frank, re 'who will write the grant?'. This is not as easy as it sounds, would that it were! There are two possible business plans:- Option 1. Specifically for MX is the PDB as the first and foremost candidate to seek such additional funds for full diffraction data deposition for each future PDB deposiition entry. This business plan possibility is best answered by PDB/EBI (eg Gerard Kleywegt has answered this in the negative thus far at the CCP4 January 2010). Option 2 The Journals that host the publications could add the cost to the subscriber and/or the author according to their funding model. As an example and as a start a draft business plan has been written by one of us [JRH] for IUCr Acta Cryst E; this seemed attractive because of its simpler 'author pays' financing. This proposed business plan is now with IUCr Journals to digest and hopefully refine. Initial indications are that Acta Cryst C would be perceived by IUCr Journals as a better place to start considering this in detail, as it involves fewer crystal structures than Acta E and would thus be more manageable. The overall advantage of the responsibility being with Journals as we see it is that it encourages such 'archiving of data with literature' across all crystallography related techniques (single crystal, SAXS, SANS, Electron crystallography etc) and fields (Biology, Chemistry, Materials, Condensed Matter Physics etc) ie not just one technique and field, although obviously biology is dear to our hearts here in the CCP4bb. Yours sincerely, John and Tom John Helliwell and Tom Terwilliger On Wed, Oct 26, 2011 at 9:21 AM, Frank von Delft frank.vonde...@sgc.ox.ac.uk wrote: Since when has the cost of any project been limited by the cost of hardware? Someone has to implement this -- and make a career out of it; thunderingly absent from this thread has been the chorus of volunteers who will write the grant. phx On 25/10/2011 21:10, Herbert J. Bernstein wrote: To be fair to those concerned about cost, a more conservative estimate from the NSF RDLM workshop last summer in Princeton is $1,000 to $3,000 per terabyte per year for long term storage allowing for overhead in moderate-sized institutions such as the PDB. Larger entities, such as Google are able to do it for much lower annual costs in the range of $100 to $300 per terabyte per year. Indeed, if this becomes a serious effort, one might wish to consider involving the large storage farm businesses such as Google and Amazon. They might be willing to help support science partially in exchange for eyeballs going to their sites. Regards, H. J. Bernstein At 1:56 PM -0600 10/25/11, James Stroud wrote: On Oct 24, 2011, at 3:56 PM, James Holton wrote: The PDB only gets about 8000 depositions per year Just to put this into dollars. If each dataset is about 17 GB in size, then that's about 14 TB of storage that needs to come online every year to store the raw data for every
Re: [ccp4bb] cryo protection
you may have thought of this already, but you could try cryoprotection in the drop itself. i.e. slowly adding cryoprotectant to the reservoir, or replacing the reservoir bit by bit with solution containing cryoprotectant, and then adding small volumes to the side of the drop - for example, exchanging 20% volume cryo-solution with the reservoir, letting equilibrate with the unchanged drop for a few hours, then add 20% volume to the drop from the reservoir (i.e. 0.2 ul if the drop is 1 ul), then add another 20% of cryo to the reservoir, equilibrate a few hrs, etc. - the idea being to very slowly change the drop conditions and minimise risk of cracking. Of course, you may need patience and many drops of crystals, not just many crystals in a few drops, until you find the cryoprotectant where the crystals do not crack and still diffract, if you are successful at all... but if it works, you can just harvest from the equilibrated drop and directly flash-cool Mark J van Raaij Laboratorio M-4 Dpto de Estructura de Macromoleculas Centro Nacional de Biotecnologia - CSIC c/Darwin 3 E-28049 Madrid, Spain tel. (+34) 91 585 4616 http://www.cnb.csic.es/content/research/macromolecular/mvraaij On 26 Oct 2011, at 18:46, Leonard Thomas wrote: Hi All, I have run into a very sensitive crystals system when it comes to cryo protecting them. I have run through the usual suspects and trays are going to be setup with a cryo protectant as part of crystallization cocktail. The one problem that seems to be occurring is that the crystals crack as soon as they are transfered out of the original drop. I am running out of ideas and really would love some new ones. Thanks in advance. Len Leonard Thomas Ph.D. Macromolecular Crystallography Laboratory Manager University of Oklahoma Department of Chemistry and Biochemistry Stephenson Life Sciences Research Center 101 Stephenson Parkway Norman, OK 73019-5251 lmtho...@ou.edu http://barlywine.chem.ou.edu Office: (405)325-1126 Lab: (405)325-7571
Re: [ccp4bb] cryo protection
-BEGIN PGP SIGNED MESSAGE- Hash: SHA1 Dear Len, just to be on the safe side, my list of 'usual suspects' includes - - glycerol/PEG400 - - LiCl et al at high concentration - - Butanediol - - sugars (glucose/ fructose) - - oil - - NaMalonate - - MPD ... you mention cracking upon transferring the crystal. - - do you use a pipet for transfer? - - addition of cryo TO the drop? - - did you try slow (several minutes - 1hr) / quick addition of cryo protectant - - seeding into slightly different conditions/additive screens - - seeding into cryo conditions ... How about collecting data at room temperature? Hope this list contains some new ideas. Best wishes, Tim On 10/26/2011 06:46 PM, Leonard Thomas wrote: Hi All, I have run into a very sensitive crystals system when it comes to cryo protecting them. I have run through the usual suspects and trays are going to be setup with a cryo protectant as part of crystallization cocktail. The one problem that seems to be occurring is that the crystals crack as soon as they are transfered out of the original drop. I am running out of ideas and really would love some new ones. Thanks in advance. Len Leonard Thomas Ph.D. Macromolecular Crystallography Laboratory Manager University of Oklahoma Department of Chemistry and Biochemistry Stephenson Life Sciences Research Center 101 Stephenson Parkway Norman, OK 73019-5251 lmtho...@ou.edu http://barlywine.chem.ou.edu Office: (405)325-1126 Lab: (405)325-7571 - -- - -- Dr Tim Gruene Institut fuer anorganische Chemie Tammannstr. 4 D-37077 Goettingen GPG Key ID = A46BEE1A -BEGIN PGP SIGNATURE- Version: GnuPG v1.4.10 (GNU/Linux) Comment: Using GnuPG with Mozilla - http://enigmail.mozdev.org/ iD8DBQFOqD5+UxlJ7aRr7hoRAlFlAJ9b4ieJzoX5J6RRce85Si05d/pkFACdEQGR GAh002nC1bod6VCBolOv3pQ= =IO/R -END PGP SIGNATURE-
Re: [ccp4bb] cryo protection
Len, We have run into this problem from time to time, and it is very frustrating. Here are some things to try, some of which you may have done already: Grow crystals in a small percentage of the cryoprotectant (e.g., 5-10% glycerol). This often allows crystal transfer into a cryo drop without cracking. (Almost never works for us, though.) Do your crystal transfers in the cold room. This slows evaporation markedly, and may prevent crystal cracking. (This works for us some of the time.) Transfer your crystals to gradually higher cryoprotectant concentrations (e.g., to 15% glycerol, then 30% glycerol). (Fiddly, and the crystals get handled a lot, but often works.) Use different cryoprotectants. We almost always have fewer cracking issues with glucose than glycerol, but YMMV. Avoid transferring the crystal from the drop at all. Just add cryoprotectant to the drop. Even better, add cryoprotectant to the drop gradually, while keeping the drop humidified over well solution. This is our "No-fail" method (this is usually, but not always successful): http://capsicum.colgate.edu/chwiki/tiki-index.php?page=Mounting+Protein+Crystals#No_fail_cryoprotection We typically use glucose in this method, but in principle you could try glycerol, MPD, PEG-400, or sodium formate, etc. Otherwise, you can try to grow out of a cryo condition that doesn't need extra cryoprotectant (been there done that) or give up and shoot at room temp in-house. Cheers, ___ Roger S. Rowlett Gordon Dorothy Kline Professor Department of Chemistry Colgate University 13 Oak Drive Hamilton, NY 13346 tel: (315)-228-7245 ofc: (315)-228-7395 fax: (315)-228-7935 email: rrowl...@colgate.edu On 10/26/2011 12:46 PM, Leonard Thomas wrote: Hi All, I have run into a very sensitive crystals system when it comes to cryo protecting them. I have run through the usual suspects and trays are going to be setup with a cryo protectant as part of crystallization cocktail. The one problem that seems to be occurring is that the crystals crack as soon as they are transfered out of the original drop. I am running out of ideas and really would love some new ones. Thanks in advance. Len Leonard Thomas Ph.D. Macromolecular Crystallography Laboratory Manager University of Oklahoma Department of Chemistry and Biochemistry Stephenson Life Sciences Research Center 101 Stephenson Parkway Norman, OK 73019-5251 lmtho...@ou.edu http://barlywine.chem.ou.edu Office: (405)325-1126 Lab: (405)325-7571
Re: [ccp4bb] Question about drawing disulfide bonds in Pymol
Hi, I think it should also work with the cartoon side chain helper option which adjust the cartoon slightly to prevent such situations. Christian Am Mittwoch 26 Oktober 2011 16:15:55 schrieb Ke, Jiyuan: Dear All, I have a question regarding making disulfide bonds in Pymol. The overall structure is in cartoon representation. When I display the disulfide bond residues as stick models, the cysteine residues sitting on the beta strand have a gap from the actual beta-strand. If I check off the flat sheet option, all the beta-strands look very wavy, but the cysteines are actually on the beta strands. I want to just change the two beta-strands with cysteine residues on it as wavy strands and keep other beta-strands flat looking. Does anyone know a way to do this or has better ways to draw individual residues on beta-strands. Thanks! jiyuan Jiyuan Ke, Ph.D. Research Scientist Van Andel Research Institute 333 Bostwick Ave NE Grand Rapids, MI 49503 The information transmitted is intended only for the person or entity to which it is addressed and may contain confidential and/or privileged material. Any review, retransmission, dissemination or other use of, or taking of any action in reliance upon, this information by persons or entities other than the intended recipient is prohibited. If you received this in error, please contact the sender and delete the material from any computer.
[ccp4bb] cryo protection
Dear Len This is a classic sign of osmotic shock. You can try matching the osmotic pressure of the mother liquor and the cryoprotectant buffer. For a protocol see Acta Cryst D (1999) 55, 1649 section 6.4 Good luck Best wishes Elspeth -Original Message- From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of Leonard Thomas Sent: 26 October 2011 17:46 To: CCP4BB@JISCMAIL.AC.UK Subject: [ccp4bb] cryo protection Hi All, I have run into a very sensitive crystals system when it comes to cryo protecting them. I have run through the usual suspects and trays are going to be setup with a cryo protectant as part of crystallization cocktail. The one problem that seems to be occurring is that the crystals crack as soon as they are transfered out of the original drop. I am running out of ideas and really would love some new ones. Thanks in advance. Len Leonard Thomas Ph.D. Macromolecular Crystallography Laboratory Manager University of Oklahoma Department of Chemistry and Biochemistry Stephenson Life Sciences Research Center 101 Stephenson Parkway Norman, OK 73019-5251 lmtho...@ou.edu http://barlywine.chem.ou.edu Office: (405)325-1126 Lab: (405)325-7571
Re: [ccp4bb] cryo protection
Hello Leonard, one thing to test is whether transferring your crystals to a drop containing simply well solution also causes cracking. If yes, then the possibility exists that the absence of protein in solution is causing the trouble. In that case, you can transfer the crystals to oil: you'll be transferring the solution (with protein) in which the crystal grew as well, and slowly remove it without adding anything 'different'. However, if your crystals crack simply because they are mechanically fragile, then the oil may actually be worse. Filip On Wed, Oct 26, 2011 at 9:46 AM, Leonard Thomas lmtho...@ou.edu wrote: Hi All, I have run into a very sensitive crystals system when it comes to cryo protecting them. I have run through the usual suspects and trays are going to be setup with a cryo protectant as part of crystallization cocktail. The one problem that seems to be occurring is that the crystals crack as soon as they are transfered out of the original drop. I am running out of ideas and really would love some new ones. Thanks in advance. Len Leonard Thomas Ph.D. Macromolecular Crystallography Laboratory Manager University of Oklahoma Department of Chemistry and Biochemistry Stephenson Life Sciences Research Center 101 Stephenson Parkway Norman, OK 73019-5251 lmtho...@ou.edu http://barlywine.chem.ou.edu Office: (405)325-1126 Lab: (405)325-7571 -- Filip Van Petegem, PhD Assistant Professor The University of British Columbia Dept. of Biochemistry and Molecular Biology 2350 Health Sciences Mall - Rm 2.356 Vancouver, V6T 1Z3 phone: +1 604 827 4267 email: filip.vanpete...@gmail.com http://crg.ubc.ca/VanPetegem/
Re: [ccp4bb] cryo protection
You can also try to crosslink before transferring to cryo. From: Filip Van Petegem filip.vanpete...@gmail.com To: CCP4BB@JISCMAIL.AC.UK CCP4BB@JISCMAIL.AC.UK Sent: Wed Oct 26 13:19:16 2011 Subject: Re: [ccp4bb] cryo protection Hello Leonard, one thing to test is whether transferring your crystals to a drop containing simply well solution also causes cracking. If yes, then the possibility exists that the absence of protein in solution is causing the trouble. In that case, you can transfer the crystals to oil: you'll be transferring the solution (with protein) in which the crystal grew as well, and slowly remove it without adding anything 'different'. However, if your crystals crack simply because they are mechanically fragile, then the oil may actually be worse. Filip On Wed, Oct 26, 2011 at 9:46 AM, Leonard Thomas lmtho...@ou.edumailto:lmtho...@ou.edu wrote: Hi All, I have run into a very sensitive crystals system when it comes to cryo protecting them. I have run through the usual suspects and trays are going to be setup with a cryo protectant as part of crystallization cocktail. The one problem that seems to be occurring is that the crystals crack as soon as they are transfered out of the original drop. I am running out of ideas and really would love some new ones. Thanks in advance. Len Leonard Thomas Ph.D. Macromolecular Crystallography Laboratory Manager University of Oklahoma Department of Chemistry and Biochemistry Stephenson Life Sciences Research Center 101 Stephenson Parkway Norman, OK 73019-5251 lmtho...@ou.edumailto:lmtho...@ou.edu http://barlywine.chem.ou.edu Office: (405)325-1126tel:%28405%29325-1126 Lab: (405)325-7571tel:%28405%29325-7571 -- Filip Van Petegem, PhD Assistant Professor The University of British Columbia Dept. of Biochemistry and Molecular Biology 2350 Health Sciences Mall - Rm 2.356 Vancouver, V6T 1Z3 phone: +1 604 827 4267 email: filip.vanpete...@gmail.commailto:filip.vanpete...@gmail.com http://crg.ubc.ca/VanPetegem/
Re: [ccp4bb] cryo protection
Hi Len; I was having exactly the same problem with my crystals, but when we grow the crystals in presence of increasing concentration of Glycerol and MPD starting from 0.5 to 10%. The crystal doesn't appear after 3% of Glycerol or MPD but the one which appear in 2.5 to 3 % were much resistant to cracking than the original crystals. Good luck Bashir On Wed, October 26, 2011 18:46, Leonard Thomas wrote: Hi All, I have run into a very sensitive crystals system when it comes to cryo protecting them. I have run through the usual suspects and trays are going to be setup with a cryo protectant as part of crystallization cocktail. The one problem that seems to be occurring is that the crystals crack as soon as they are transfered out of the original drop. I am running out of ideas and really would love some new ones. Thanks in advance. Len Leonard Thomas Ph.D. Macromolecular Crystallography Laboratory Manager University of Oklahoma Department of Chemistry and Biochemistry Stephenson Life Sciences Research Center 101 Stephenson Parkway Norman, OK 73019-5251 lmtho...@ou.edu http://barlywine.chem.ou.edu Office: (405)325-1126 Lab: (405)325-7571 -- Muhammad Bashir Khan ** Department for Structural and Computational Biology Max F. Perutz Laboratories University of Vienna Campus Vienna Biocenter 5 A-1030 Vienna Austria Austria Phone: +43(1)427752224 Fax: +43(1)42779522
Re: [ccp4bb] IUCr committees, depositing images
Dear Colleagues, Gerard strikes a very useful note in pleading for a can-do approach. Part of going from can-do to actually-done is to make realistic estimates of the costs of doing and then to adjust plans appropriately to do what can be afforded now and to work towards doing as much of what remains undone as has sufficient benefit to justify the costs. We appear to be in a fortunate situation in which some portion of the raw data behind a signficant portion of the studies released in the PDB could probably be retained for some significant period of time and be made available for further analysis. It would seem wise to explore these possibilities and try to optimize the approaches used -- e.g. to consider moves towards well documented formats, and retention of critical metadata with such data to help in future analysis. Please do not let the perfect be the enemy of the good. Regards, Herbert = Herbert J. Bernstein, Professor of Computer Science Dowling College, Kramer Science Center, KSC 121 Idle Hour Blvd, Oakdale, NY, 11769 +1-631-244-3035 y...@dowling.edu = On Wed, 26 Oct 2011, Gerard Bricogne wrote: Dear John and colleagues, There seem to be a set a centrifugal forces at play within this thread that are distracting us from a sensible path of concrete action by throwing decoys in every conceivable direction, e.g. * Pilatus detectors spew out such a volume of data that we can't possibly archive it all - does that mean that because the 5th generation of Dectris detectors will be able to write one billion images a second and catch every scattered photon individually, we should not try and archive more information than is given by the current merged structure factor data? That seems a complete failure of reasoning to me: there must be a sensible form of raw data archiving that would stand between those two extremes and would retain much more information that the current merged data but would step back from the enormous degree of oversampling of the raw diffraction pattern that the Pilatus and its successors are capable of. * It is all going to cost an awful lot of money, therefore we need a team of grant writers to raise its hand and volunteer to apply for resources from one or more funding agencies - there again there is an avoidance of the feasible by invocation of the impossible. The IUCr Forum already has an outline of a feasibility study that would cost only a small amount of joined-up thinking and book-keeping around already stored information, so let us not use the inaccessibility of federal or EC funding as a scarecrow to justify not even trying what is proposed there. And the idea that someone needs to decide to stake his/her career on this undertaking seems totally overblown. Several people have already pointed out that the sets of images that would need to be archived would be a very small subset of the bulk of datasets that are being held on the storage systems of synchrotron sources. What needs to be done, as already described, is to be able to refer to those few datasets that gave rise to the integrated data against which deposited structures were refined (or, in some cases, solved by experimental phasing), to give them special status in terms of making them visible and accessible on-line at the same time as the pdb entry itself (rather than after the statutory 2-5 years that would apply to all the rest, probably in a more off-line form), and to maintain that accessibility for ever, with a link from the pdb entry and perhaps from the associated publication. It seems unlikely that this would involve the mobilisation of such large resources as to require either a human sacrifice (of the poor person whose life would be staked on this gamble) or writing a grant application, with the indefinite postponement of action and the loss of motivation this would imply. Coming back to the more technical issue of bloated datasets, it is a scientific problem that must be amenable to rational analysis to decide on a sensible form of compression of overly-verbose sets of thin-sliced, perhaps low-exposure images that would already retain a large fraction, if not all, of the extra information on which we would wish future improved versions of processing programs to cut their teeth, for a long time to come. This approach would seem preferable to stoking up irrational fears of not being able to cope with the most exaggerated predictions of the volumes of data to archive, and thus doing nothing at all. I very much hope that the can do spirit that marked the final discussions of the DDDWG (Diffraction Data Deposition Working Group) in Madrid will emerge on top of all the counter-arguments that consist in moving the goal posts to prove that the initial goal is unreachable. With best wishes, Gerard.
Re: [ccp4bb] IUCr committees, depositing images
On Oct 26, 2011, at 9:59 AM, Patrick Shaw Stewart wrote: The principle is that the movie is written to the same area of memory, jumping back to the beginning when it is full (this part is not essential, but it makes the principle clear). Then, when the photographer takes his finger off the trigger, the last x seconds is permanently stored. So you keep your wits about you, and press the metaphorical store button just after you have got the movie in the can so to speak This idea seems equivalent to only storing permanently those datasets that actually yield structures worthy of deposition. James
Re: [ccp4bb] cryo protection
Another possibility (other than those already mentioned) is to try freezing without a cryoprotectant, by fishing the crystals out onto a mesh and removing all the mother liquor. The following paper has some details: Direct cryocooling of naked crystals: are cryoprotection agents always necessary? Erika Pellegrini, Dario Pianoa, and Matthew W. Bowlera Acta Cryst. (2011). D67, 902906 -- Andrew Purkiss X-ray Laboratory Manager Cancer Research UK London Research Institute. Quoting Leonard Thomas lmtho...@ou.edu: Hi All, I have run into a very sensitive crystals system when it comes to cryo protecting them. I have run through the usual suspects and trays are going to be setup with a cryo protectant as part of crystallization cocktail. The one problem that seems to be occurring is that the crystals crack as soon as they are transfered out of the original drop. I am running out of ideas and really would love some new ones. Thanks in advance. Len Leonard Thomas Ph.D. Macromolecular Crystallography Laboratory Manager University of Oklahoma Department of Chemistry and Biochemistry Stephenson Life Sciences Research Center 101 Stephenson Parkway Norman, OK 73019-5251 lmtho...@ou.edu http://barlywine.chem.ou.edu Office: (405)325-1126 Lab: (405)325-7571 This message was sent using IMP, the Internet Messaging Program.
Re: [ccp4bb] cryo protection
A good number of things to try. Just a little more info that was asked for. The crystals are grown in Peg 3350 over a range of pH values using Bis-Tris Propane. The are coming out of 2 different salt conditions. My feeling is it is an osmolality problem though I also observed cracking when going into a separately made well solution. I will look it trying a number of suggestions given. Cheers, Len Leonard Thomas Ph.D. Macromolecular Crystallography Laboratory Manager University of Oklahoma Department of Chemistry and Biochemistry Stephenson Life Sciences Research Center 101 Stephenson Parkway Norman, OK 73019-5251 lmtho...@ou.edu http://barlywine.chem.ou.edu Office: (405)325-1126 Lab: (405)325-7571 On Oct 26, 2011, at 12:54 PM, Muhammed bashir Khan wrote: Hi Len; I was having exactly the same problem with my crystals, but when we grow the crystals in presence of increasing concentration of Glycerol and MPD starting from 0.5 to 10%. The crystal doesn't appear after 3% of Glycerol or MPD but the one which appear in 2.5 to 3 % were much resistant to cracking than the original crystals. Good luck Bashir On Wed, October 26, 2011 18:46, Leonard Thomas wrote: Hi All, I have run into a very sensitive crystals system when it comes to cryo protecting them. I have run through the usual suspects and trays are going to be setup with a cryo protectant as part of crystallization cocktail. The one problem that seems to be occurring is that the crystals crack as soon as they are transfered out of the original drop. I am running out of ideas and really would love some new ones. Thanks in advance. Len Leonard Thomas Ph.D. Macromolecular Crystallography Laboratory Manager University of Oklahoma Department of Chemistry and Biochemistry Stephenson Life Sciences Research Center 101 Stephenson Parkway Norman, OK 73019-5251 lmtho...@ou.edu http://barlywine.chem.ou.edu Office: (405)325-1126 Lab: (405)325-7571 -- Muhammad Bashir Khan ** Department for Structural and Computational Biology Max F. Perutz Laboratories University of Vienna Campus Vienna Biocenter 5 A-1030 Vienna Austria Austria Phone: +43(1)427752224 Fax: +43(1)42779522
Re: [ccp4bb] cryo protection
Len, May be you have already done this. I would closely check my crystallization conditions and also check the pH of the cryo. In some cases, during cryoprotection the pH of the original drop may drastically different than the cryo solution. Also, sometime back, we were exploring different cryoprotectant conditions for sensitive crystal and came across this - http://www.xtals.org/crystal_cryo.pdf by Artem. HTH Harkewal On Wed, 26 Oct 2011 11:46:08 -0500, Leonard Thomas lmtho...@ou.edu wrote: Hi All, I have run into a very sensitive crystals system when it comes to cryo protecting them. I have run through the usual suspects and trays are going to be setup with a cryo protectant as part of crystallization cocktail. The one problem that seems to be occurring is that the crystals crack as soon as they are transfered out of the original drop. I am running out of ideas and really would love some new ones. Thanks in advance. Len Leonard Thomas Ph.D. Macromolecular Crystallography Laboratory Manager University of Oklahoma Department of Chemistry and Biochemistry Stephenson Life Sciences Research Center 101 Stephenson Parkway Norman, OK 73019-5251 lmtho...@ou.edu http://barlywine.chem.ou.edu Office: (405)325-1126 Lab: (405)325-7571
Re: [ccp4bb] cryo protection
One small point: Just make sure that you are not too off from the contents of the protein solution. Sometimes protein solution may have a high amount of salt or things like that and we forget to include atleast half of this concentration into the cryo solution. This could easily crack the crystals depending on the concentration and the type of compounds in it. Regards, Mathews -Original Message- From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of Leonard Thomas Sent: Wednesday, October 26, 2011 11:57 AM To: CCP4BB@JISCMAIL.AC.UK Subject: Re: [ccp4bb] cryo protection A good number of things to try. Just a little more info that was asked for. The crystals are grown in Peg 3350 over a range of pH values using Bis-Tris Propane. The are coming out of 2 different salt conditions. My feeling is it is an osmolality problem though I also observed cracking when going into a separately made well solution. I will look it trying a number of suggestions given. Cheers, Len Leonard Thomas Ph.D. Macromolecular Crystallography Laboratory Manager University of Oklahoma Department of Chemistry and Biochemistry Stephenson Life Sciences Research Center 101 Stephenson Parkway Norman, OK 73019-5251 lmtho...@ou.edu http://barlywine.chem.ou.edu Office: (405)325-1126 Lab: (405)325-7571 On Oct 26, 2011, at 12:54 PM, Muhammed bashir Khan wrote: Hi Len; I was having exactly the same problem with my crystals, but when we grow the crystals in presence of increasing concentration of Glycerol and MPD starting from 0.5 to 10%. The crystal doesn't appear after 3% of Glycerol or MPD but the one which appear in 2.5 to 3 % were much resistant to cracking than the original crystals. Good luck Bashir On Wed, October 26, 2011 18:46, Leonard Thomas wrote: Hi All, I have run into a very sensitive crystals system when it comes to cryo protecting them. I have run through the usual suspects and trays are going to be setup with a cryo protectant as part of crystallization cocktail. The one problem that seems to be occurring is that the crystals crack as soon as they are transfered out of the original drop. I am running out of ideas and really would love some new ones. Thanks in advance. Len Leonard Thomas Ph.D. Macromolecular Crystallography Laboratory Manager University of Oklahoma Department of Chemistry and Biochemistry Stephenson Life Sciences Research Center 101 Stephenson Parkway Norman, OK 73019-5251 lmtho...@ou.edu http://barlywine.chem.ou.edu Office: (405)325-1126 Lab: (405)325-7571 -- Muhammad Bashir Khan ** Department for Structural and Computational Biology Max F. Perutz Laboratories University of Vienna Campus Vienna Biocenter 5 A-1030 Vienna Austria Austria Phone: +43(1)427752224 Fax: +43(1)42779522
Re: [ccp4bb] cryo protection
One more thing you could try: high pressure cryo-cooling. Se any of a number of paperas by Chae Un Kim; e.g. http://www.ncbi.nlm.nih.gov/pubmed/17452791 Acta Crystallogr D Biol Crystallogr. http://www.ncbi.nlm.nih.gov/pubmed/17452791# 2007 May;63(Pt 5):653-9. Epub 2007 Apr 21. On 10/26/11 12:46, Leonard Thomas wrote: Hi All, I have run into a very sensitive crystals system when it comes to cryo protecting them. I have run through the usual suspects and trays are going to be setup with a cryo protectant as part of crystallization cocktail. The one problem that seems to be occurring is that the crystals crack as soon as they are transfered out of the original drop. I am running out of ideas and really would love some new ones. Thanks in advance. Len Leonard Thomas Ph.D. Macromolecular Crystallography Laboratory Manager University of Oklahoma Department of Chemistry and Biochemistry Stephenson Life Sciences Research Center 101 Stephenson Parkway Norman, OK 73019-5251 lmtho...@ou.edu http://barlywine.chem.ou.edu Office: (405)325-1126 Lab: (405)325-7571 -- === All Things Serve the Beam === David J. Schuller modern man in a post-modern world MacCHESS, Cornell University schul...@cornell.edu
Re: [ccp4bb] IUCr committees, depositing images
I have been nominated by the IUCr synchrotron commission (thanks colleagues!) to represent them for this issue. However, at the moment, this is a personal view. 1. For archiving raw diffraction image data for structures in the PDB, it should be the responsibility of the worldwide PDB. They are by far the best place to do it and as Jacob says the space requirements are trivial. Gerard K's negative statement at CCP4-2010 sounds rather ex cathedra (in increasing order of influence/power do we have the Pope, US president, the Bond Market and finally Gerard K?). Did he make the statement in a formal presentation or in the bar? More seriously, I am sure he had good reasons (e.g. PDB priorities) if he did make this statement. It would be nice if Gerard could provide some explanation. 2. I agree with the can do attitude at Madrid as supported by Gerard B. Setting up something as best one can with existing enthusiasts will get the ball rolling, provide some immediate benefit and allow subsequent improvements. 3. Ideally the data to be deposited should include all stages e.g. raw images, corrected images, MIR/SAD/MAD images, unmerged integrated intensities, scaled, merged etc. Plus the metadata, software versions used for the various stages. Worrying too much about all of this should not of course prevent a start being made. (An aside. I put the corrected in quotes because the raw images have fewer errors. The subsequent processing for detector distortions etc. depend on an imperfect model for the detector. I don't like the phrase data correction). 4. Doing this for PDB depositions would then provide a basis for other data which did not result in PDB depositions. There seems to be a view that the archiving of this should be the responsibility of the synchrotrons which generated the data. This should be possible for some synchrotrons (e.g. Diamond) where there is pressure in any case from their funders to archive all data generated at the facility. However not all synchrotrons will be able to do this. There is also the issue of data collected at home sources. Presumably it will require a few willing synchrotrons to pioneer this in a coordinated way. Hopefully others will then follow. I don't think we can expect the PDB to archive the 99.96% of the data which did not result in structures. 5. My view is that for data in the PDB the same release rules should apply for the images as for the other data. For other data, the funders of the research might want to define release rules. However, we can make suggestions! 6. Looking to the future, there is FEL data coming along, both single molecule and nano-crystals (assuming the FEL delivers for these areas). 7. I agree with Gerard B - as far as I see it, the highest future benefit of having archived raw images will result from being able to reprocess datasets from samples containing multiple lattices My view is that all crystals are, to a greater or lesser extent, subject to this. We just might not see it easily as the detector resolution or beam divergence is inadequate. Just think we could have several structures (one from each lattice) each with less disorder rather than just one average structure. Not sure whether Gloria's modulated structures would be as ubiquitous but her argument is along the same lines. Regards Colin -Original Message- From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of Herbert J. Bernstein Sent: 26 October 2011 18:55 To: ccp4bb Subject: Re: [ccp4bb] IUCr committees, depositing images Dear Colleagues, Gerard strikes a very useful note in pleading for a can-do approach. Part of going from can-do to actually-done is to make realistic estimates of the costs of doing and then to adjust plans appropriately to do what can be afforded now and to work towards doing as much of what remains undone as has sufficient benefit to justify the costs. We appear to be in a fortunate situation in which some portion of the raw data behind a signficant portion of the studies released in the PDB could probably be retained for some significant period of time and be made available for further analysis. It would seem wise to explore these possibilities and try to optimize the approaches used -- e.g. to consider moves towards well documented formats, and retention of critical metadata with such data to help in future analysis. Please do not let the perfect be the enemy of the good. Regards, Herbert = Herbert J. Bernstein, Professor of Computer Science Dowling College, Kramer Science Center, KSC 121 Idle Hour Blvd, Oakdale, NY, 11769 +1-631-244-3035 y...@dowling.edu = On Wed, 26 Oct 2011, Gerard Bricogne wrote: Dear John and colleagues, There seem to be a set a centrifugal forces at play within this thread
Re: [ccp4bb] cryo protection
For some ideas on cryocrystallography, one can watch an online webinar on the subject: http://www.rigaku.com/protein/webinar-001.html Maybe some unbiased folks can comment? ;) Jim From: CCP4 bulletin board [CCP4BB@JISCMAIL.AC.UK] on behalf of Leonard Thomas [lmtho...@ou.edu] Sent: Wednesday, October 26, 2011 11:46 AM To: CCP4BB@JISCMAIL.AC.UK Subject: [ccp4bb] cryo protection Hi All, I have run into a very sensitive crystals system when it comes to cryo protecting them. I have run through the usual suspects and trays are ...
[ccp4bb] Postdoctoral Position at King's College London
A three-year postdoctoral position in the field of mechanistic enzymology is immediately available in the Steiner Laboratory of King’s College London. The salary is £33,193 per annum inclusive of London allowance. The project aims at studying the intriguing biological process of cofactor-independent oxygenation catalysis (1). To understand how dioxygen chemistry takes place in a cofactor-less manner we will use enzymes for which we have recently obtained structural information in various catalytically relevant states (2). The ideal candidate has a strong biochemistry/chemistry background, extensive experience in structural biology focused on enzyme mechanisms and an interest in molecular dynamics (MD). The MD work will be carried out in collaboration with Prof. Ceccarelli of the University of Cagliari, Italy, where the postdoc will be able to spend some time to improve his/her skills in MD techniques. The postdoc will also be in close contact with the Manchester group of Prof. Scrutton where complementary spectroscopic and kinetic studies will be carried out. This project will equip the post-holder with a rare and sought-after skill-set as well as a comprehensive overview of an inter-disciplinary study. To apply for this position go to http://www.kcl.ac.uk/depsta/pertra/vacancy/external/pers_detail.php?jobindex=10871. Please make sure you quote the reference number G6/JKA/772/11-JT. Informal enquiries are welcome at roberto.stei...@kcl.ac.ukmailto:roberto.stei...@kcl.ac.uk. The closing date for this application is 24 Nov. 2011. (1) Fetzner S. and Steiner RA (2010). Cofactor-independent oxidases and oxygenases. Appl. Microbiol. Biotechnol. 86, 791-804. (2) Steiner RA, Janssen HJ, Roversi P, Oakley OJ, Fetzner S (2010). Structural basis for cofactor independent dioxygenation of N-heteroaromatic compounds at the •/•-hydrolase fold. Proc. Natl. Acad. Sci. USA, 107, 657-662. Roberto Steiner, PhD Group Leader Randall Division of Cell and Molecular Biophysics King's College London Room 3.10A New Hunt's House Guy's Campus SE1 1UL, London, UK Tel 0044-20-78488216 Fax 0044-20-78486435 roberto.stei...@kcl.ac.ukmailto:roberto.stei...@kcl.ac.uk
Re: [ccp4bb] IUCr committees, depositing images
Dear George, dear all, I was just trying to summarize my point of view regarding this important issue when I got your e-mail, that reflects exactly my own opinion! Martin Dr. Martin Martinez-Ripoll Research Professor xmar...@iqfr.csic.es Department of Crystallography Structural Biology www.xtal.iqfr.csic.es Telf.: +34 917459550 Consejo Superior de Investigaciones Científicas Spanish National Research Council www.csic.es -Mensaje original- De: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] En nombre de George M. Sheldrick Enviado el: miércoles, 26 de octubre de 2011 11:52 Para: CCP4BB@JISCMAIL.AC.UK Asunto: Re: [ccp4bb] IUCr committees, depositing images This raises an important point. The new continuous readout detectors such as the Pilatus for beamlines or the Bruker Photon for in-house use enable the crystal to be rotated at constant velocity, eliminating the mechanical errors associated with 'stop and go' data collection. Storing their data in 'frames' is an artifical construction that is currently required for the established data integration programs but is in fact throwing away information. Maybe in 10 years time 'frames' will be as obsolete as punched cards! George On Wed, Oct 26, 2011 at 09:39:40AM +0100, Graeme Winter wrote: Hi James, Just to pick up on your point about the Pilatus detectors. Yesterday in 2 hours of giving a beamline a workout (admittedly with Thaumatin) we acquired 400 + GB of data*. Now I appreciate that this is not really routine operation, but it does raise an interesting point - if you have loaded a sample and centred it, collected test shots and decided it's not that great, why not collect anyway as it may later prove to be useful? Bzzt. 2 minutes or less later you have a full data set, and barely even time to go get a cup of tea. This does to some extent move the goalposts, as you can acquire far more data than you need. You never know, you may learn something interesting from it - perhaps it has different symmetry or packing? What it does mean is if we can have a method of tagging this data there may be massively more opportunity to get also-ran data sets for methods development types. What it also means however is that the cost of curating this data is then an order of magnitude higher. Also moving it around is also rather more painful. Anyhow, I would try to avoid dismissing the effect that new continuous readout detectors will have on data rates, from experience it is pretty substantial. Cheerio, Graeme *by data here what I mean is images, rather than information which is rather more time consuming to acquire. I would argue you get that from processing / analysing the data... On 24 October 2011 22:56, James Holton jmhol...@lbl.gov wrote: The Pilatus is fast, but or decades now we have had detectors that can read out in ~1s. This means that you can collect a typical ~100 image dataset in a few minutes (if flux is not limiting). Since there are ~150 beamlines currently operating around the world and they are open about 200 days/year, we should be collecting ~20,000,000 datasets each year. We're not. The PDB only gets about 8000 depositions per year, which means either we throw away 99.96% of our images, or we don't actually collect images anywhere near the ultimate capacity of the equipment we have. In my estimation, both of these play about equal roles, with ~50-fold attrition between ultimate data collection capacity and actual collected data, and another ~50 fold attrition between collected data sets and published structures. Personally, I think this means that the time it takes to collect the final dataset is not rate-limiting in a typical structural biology project/paper. This does not mean that the dataset is of little value. Quite the opposite! About 3000x more time and energy is expended preparing for the final dataset than is spent collecting it, and these efforts require experimental feedback. The trick is figuring out how best to compress the data used to solve a structure for archival storage. Do the previous data sets count? Or should the compression be lossy about such historical details? Does the stuff between the spots matter? After all, h,k,l,F,sigF is really just a form of data compression. In fact, there is no such thing as raw data. Even raw diffraction images are a simplification of the signals that came out of the detector electronics. But we round-off and average over a lot of things to remove noise. Largely because noise is difficult to compress. The question of how much compression is too much compression depends on which information (aka noise) you think could be important in the future. When it comes to fine-sliced data, such as that from Pilatus, the main reason why it doesn't compress very well is not because of the spots, but the background. It
Re: [ccp4bb] IUCr committees, depositing images
Dear Colin, Thank you for accepting the heavy burden of responsibility your colleagues have thrown onto your shoulders ;-) . It is great that you are entering this discussion, and I am grateful for the support you are bringing to the notion of starting something at ground level and learning from it, rather that staying in the realm of conjecture and axiomatics, or entering the virility contest as to whose beamline will make raw data archiving most impossible. One small point, however, about your statement regarding multiple lattices, that ... all crystals are, to a greater or lesser extent, subject to this. We just might not see it easily as the detector resolution or beam divergence is inadequate. Just think we could have several structures (one from each lattice) each with less disorder rather than just one average structure. I am not sure that what you describe in your last sentence is a realistic prospect, nor that it would in any case constitute the main advantage of better dealing with multiple lattices. The most important consequence of their multiplicity is that their spots overlap and corrupt each other's intensities, so that the main benefit of improved processing would be to mitigate that mutual corruption, first by correctly flagging overlaps, then by partially trying to resolve those overlaps internally as much as scaling procedures will allow (one could call that non-merohedral detwinning - it is done e.g. by small-molecule softeware), and finally by adapting refinement protocols to recognise that they may have to refine against measurements that are a mixture of several intensities, to a degree and according to a pattern that varies from one observation to another (unlike regular twinning). Currently, if a main lattice can be identified and indexed, one tends to integrate the spots it successfully indexes, and to abstain from worrying about the accidental corruption of the resulting intensities by accidental overlaps with spots of the other lattices (whose existence is promptly forgotten). It is the undoing of that corruption that would bring the main benefit, not the fact that one could see several variants of the structure by fitting the data attached to the various lattices: that would be possible only if overlaps were negligible. The prospects for improving electron density maps by reprocessing raw images in the future are therefore considerable for mainstream structures, not just as a way of perhaps teasing interestingly different structures from each lattice in infrequent cases. I apologise if I have laboured this point, but I am concerned that every slight slip of the pen that makes the benefits of future reprocessing look as if they will just contribute to splitting hairs does a disservice to this crucial discussion (and hence, potentially, to the community) by belittling the importance and urgency of the task. With best wishes, Gerard (B.) -- On Wed, Oct 26, 2011 at 07:58:51PM +, Colin Nave wrote: I have been nominated by the IUCr synchrotron commission (thanks colleagues!) to represent them for this issue. However, at the moment, this is a personal view. 1. For archiving raw diffraction image data for structures in the PDB, it should be the responsibility of the worldwide PDB. They are by far the best place to do it and as Jacob says the space requirements are trivial. Gerard K's negative statement at CCP4-2010 sounds rather ex cathedra (in increasing order of influence/power do we have the Pope, US president, the Bond Market and finally Gerard K?). Did he make the statement in a formal presentation or in the bar? More seriously, I am sure he had good reasons (e.g. PDB priorities) if he did make this statement. It would be nice if Gerard could provide some explanation. 2. I agree with the can do attitude at Madrid as supported by Gerard B. Setting up something as best one can with existing enthusiasts will get the ball rolling, provide some immediate benefit and allow subsequent improvements. 3. Ideally the data to be deposited should include all stages e.g. raw images, corrected images, MIR/SAD/MAD images, unmerged integrated intensities, scaled, merged etc. Plus the metadata, software versions used for the various stages. Worrying too much about all of this should not of course prevent a start being made. (An aside. I put the corrected in quotes because the raw images have fewer errors. The subsequent processing for detector distortions etc. depend on an imperfect model for the detector. I don't like the phrase data correction). 4. Doing this for PDB depositions would then provide a basis for other data which did not result in PDB depositions. There seems to be a view that the archiving of this should be the responsibility of the synchrotrons which generated the data. This should be possible for some synchrotrons (e.g. Diamond) where there
Re: [ccp4bb] COOT not connected to PHENIX
I experienced this same issue a while ago. When I attempt to reinstall coot using Fink (instructions on Bill Scott's coot page), I receive this error in Terminal: WARNING: While resolving dependency nose-py27 for package numpy-py27-1.5.1-1, package nose-py27 was not found. Reading build dependency for numpy-py27-1.5.1-1... WARNING: While resolving dependency nose-py27 for package numpy-py27-1.5.1-1, package nose-py27 was not found. Can't resolve dependency nose-py27 for package numpy-py27-1.5.1-1 (no matching packages/versions found) Exiting with failure. I had originally downloaded ccp4-6.2.0 (Mac version), and experienced no problems until I tried to open coot from PHENIX. Any suggestions on how to resolve this issue? Jaime From: CCP4 bulletin board [CCP4BB@JISCMAIL.AC.UK] on behalf of Ed Pozharski [epozh...@umaryland.edu] Sent: Wednesday, October 26, 2011 9:24 AM To: CCP4BB@JISCMAIL.AC.UK Subject: Re: [ccp4bb] COOT not connected to PHENIX On Wed, 2011-10-26 at 10:33 +0200, Tim Gruene wrote: with every python script one has to distribute a specific python version ... and with every program one has to distribute binaries for every platform... more food for my prejudice against software ;-) This really is not about python, it's about distributing with or without dependencies. And you are absolutely right about that: for example, ccp4-6.2.0 comes with python2.6.7 embedded, and, if one goes with defaults and downloads coot with it, python2.6 in coot's lib folder. Same with phenix - you get python2.7 with it and python2.4 with pymol0.99 that comes with it. By the way, I already have another pymol that I compiled myself (1.4) and the one from ubuntu repositories (1.2). Except for the latter, each carries its own copy of whichever python it needs. Every single python avatar takes 50-100Mb of space, which is fortunately not in short supply. This is why the right way to distribute *nix software is to distribute software itself and ask the end-user to get all the dependencies (not that hard these days). It is fully understood, of course, that people that do this for living find it more troublesome to deal with me whining about how their software is screwing up my matplotlib than to just give me another python copy. What's an extra 50Mb between friends ;-) Cheers, Ed. -- I'd jump in myself, if I weren't so good at whistling. Julian, King of Lemurs
Re: [ccp4bb] IUCr committees, depositing images
Dear George, Martin I don't understand the point that one is throwing away information by storing in frames. If the frames have sufficiently fine intervals (given by some sampling theorem consideration) I can't see how one loses information. Can one of you explain? Thanks Colin -Original Message- From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of Martin M. Ripoll Sent: 26 October 2011 22:50 To: ccp4bb Subject: Re: [ccp4bb] IUCr committees, depositing images Dear George, dear all, I was just trying to summarize my point of view regarding this important issue when I got your e-mail, that reflects exactly my own opinion! Martin Dr. Martin Martinez-Ripoll Research Professor xmar...@iqfr.csic.es Department of Crystallography Structural Biology www.xtal.iqfr.csic.es Telf.: +34 917459550 Consejo Superior de Investigaciones Científicas Spanish National Research Council www.csic.es -Mensaje original- De: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] En nombre de George M. Sheldrick Enviado el: miércoles, 26 de octubre de 2011 11:52 Para: CCP4BB@JISCMAIL.AC.UK Asunto: Re: [ccp4bb] IUCr committees, depositing images This raises an important point. The new continuous readout detectors such as the Pilatus for beamlines or the Bruker Photon for in-house use enable the crystal to be rotated at constant velocity, eliminating the mechanical errors associated with 'stop and go' data collection. Storing their data in 'frames' is an artifical construction that is currently required for the established data integration programs but is in fact throwing away information. Maybe in 10 years time 'frames' will be as obsolete as punched cards! George On Wed, Oct 26, 2011 at 09:39:40AM +0100, Graeme Winter wrote: Hi James, Just to pick up on your point about the Pilatus detectors. Yesterday in 2 hours of giving a beamline a workout (admittedly with Thaumatin) we acquired 400 + GB of data*. Now I appreciate that this is not really routine operation, but it does raise an interesting point - if you have loaded a sample and centred it, collected test shots and decided it's not that great, why not collect anyway as it may later prove to be useful? Bzzt. 2 minutes or less later you have a full data set, and barely even time to go get a cup of tea. This does to some extent move the goalposts, as you can acquire far more data than you need. You never know, you may learn something interesting from it - perhaps it has different symmetry or packing? What it does mean is if we can have a method of tagging this data there may be massively more opportunity to get also-ran data sets for methods development types. What it also means however is that the cost of curating this data is then an order of magnitude higher. Also moving it around is also rather more painful. Anyhow, I would try to avoid dismissing the effect that new continuous readout detectors will have on data rates, from experience it is pretty substantial. Cheerio, Graeme *by data here what I mean is images, rather than information which is rather more time consuming to acquire. I would argue you get that from processing / analysing the data... On 24 October 2011 22:56, James Holton jmhol...@lbl.gov wrote: The Pilatus is fast, but or decades now we have had detectors that can read out in ~1s. This means that you can collect a typical ~100 image dataset in a few minutes (if flux is not limiting). Since there are ~150 beamlines currently operating around the world and they are open about 200 days/year, we should be collecting ~20,000,000 datasets each year. We're not. The PDB only gets about 8000 depositions per year, which means either we throw away 99.96% of our images, or we don't actually collect images anywhere near the ultimate capacity of the equipment we have. In my estimation, both of these play about equal roles, with ~50-fold attrition between ultimate data collection capacity and actual collected data, and another ~50 fold attrition between collected data sets and published structures. Personally, I think this means that the time it takes to collect the final dataset is not rate-limiting in a typical structural biology project/paper. This does not mean that the dataset is of little value. Quite the opposite! About 3000x more time and energy is expended preparing for the final dataset than is spent collecting it, and these efforts require experimental feedback. The trick is figuring out how best to compress the data used to solve a structure for archival storage. Do the previous data sets count? Or should the compression be lossy about such historical details? Does the stuff between the spots matter? After all, h,k,l,F,sigF is really just a form of data compression. In fact, there is no such thing as raw data. Even raw diffraction images are a simplification of the
[ccp4bb] Off-topic: DSF thermo cycler low temp limit
We are trying to determine Tm value of rather unstable proteins with a Tm in the mid 20 C range using DSF/thermofluor. Our Stratagene thermocycler has a low temp limit of 25 C (Peltier). I called the company and they said it's a 'hardware limit' that cannot be changed. 1) Has anyone been able to 'hotwire' the MX3000/3005 to go below this limit? 2) Are there other thermofluor machines that allow lower starting temperatures (say 4, 10 or 15 C)? Best regards, -Reggie
Re: [ccp4bb] IUCr committees, depositing images
Dear Gerard Yes, perhaps I was getting a bit carried away with the possibilities. Although I believe that, with high resolution detectors and low divergence beams, one should be able to separate out the various lattices it is not really relevant to the main issue - getting the best from existing data. The point I made about correcting data probably comes in a similar category - taking the opportunity to air a favourite subject. Regards Colin PS. While here though I realise one of my points was a bit unclear. Point 5 should be 5. My view is that for data in the PDB the same release rules should apply for the images as for the other data. For data not (yet) in the PDB, the funders of the research might want to define release rules. However, we can make suggestions! The original had For other data rather than For data not (yet) in the PDB -Original Message- From: Gerard Bricogne [mailto:g...@globalphasing.com] Sent: 26 October 2011 23:23 To: Nave, Colin (DLSLtd,RAL,DIA) Cc: ccp4bb Subject: Re: [ccp4bb] IUCr committees, depositing images Dear Colin, Thank you for accepting the heavy burden of responsibility your colleagues have thrown onto your shoulders ;-) . It is great that you are entering this discussion, and I am grateful for the support you are bringing to the notion of starting something at ground level and learning from it, rather that staying in the realm of conjecture and axiomatics, or entering the virility contest as to whose beamline will make raw data archiving most impossible. One small point, however, about your statement regarding multiple lattices, that ... all crystals are, to a greater or lesser extent, subject to this. We just might not see it easily as the detector resolution or beam divergence is inadequate. Just think we could have several structures (one from each lattice) each with less disorder rather than just one average structure. I am not sure that what you describe in your last sentence is a realistic prospect, nor that it would in any case constitute the main advantage of better dealing with multiple lattices. The most important consequence of their multiplicity is that their spots overlap and corrupt each other's intensities, so that the main benefit of improved processing would be to mitigate that mutual corruption, first by correctly flagging overlaps, then by partially trying to resolve those overlaps internally as much as scaling procedures will allow (one could call that non-merohedral detwinning - it is done e.g. by small-molecule softeware), and finally by adapting refinement protocols to recognise that they may have to refine against measurements that are a mixture of several intensities, to a degree and according to a pattern that varies from one observation to another (unlike regular twinning). Currently, if a main lattice can be identified and indexed, one tends to integrate the spots it successfully indexes, and to abstain from worrying about the accidental corruption of the resulting intensities by accidental overlaps with spots of the other lattices (whose existence is promptly forgotten). It is the undoing of that corruption that would bring the main benefit, not the fact that one could see several variants of the structure by fitting the data attached to the various lattices: that would be possible only if overlaps were negligible. The prospects for improving electron density maps by reprocessing raw images in the future are therefore considerable for mainstream structures, not just as a way of perhaps teasing interestingly different structures from each lattice in infrequent cases. I apologise if I have laboured this point, but I am concerned that every slight slip of the pen that makes the benefits of future reprocessing look as if they will just contribute to splitting hairs does a disservice to this crucial discussion (and hence, potentially, to the community) by belittling the importance and urgency of the task. With best wishes, Gerard (B.) -- On Wed, Oct 26, 2011 at 07:58:51PM +, Colin Nave wrote: I have been nominated by the IUCr synchrotron commission (thanks colleagues!) to represent them for this issue. However, at the moment, this is a personal view. 1. For archiving raw diffraction image data for structures in the PDB, it should be the responsibility of the worldwide PDB. They are by far the best place to do it and as Jacob says the space requirements are trivial. Gerard K's negative statement at CCP4-2010 sounds rather ex cathedra (in increasing order of influence/power do we have the Pope, US president, the Bond Market and finally Gerard K?). Did he make the statement in a formal presentation or in the bar? More seriously, I am sure he had good reasons (e.g. PDB priorities) if he did make this statement. It would be nice if Gerard could provide some explanation. 2. I agree with the can do attitude at Madrid as
[ccp4bb] unsubscribe ccp4bb
Re: [ccp4bb] data processing problem with ice rings
Hi there, Thank you for all your suggestions and generous help, I tried some methods you guys mentioned and learned something new . I really appreciate it. With Kay's help,(after exclusion of the ice rings he found that the data are P1, not P2(1). ) I got my structure solved, there are four copies in the AU, I cut the high resolution to 2.4, and now the R/Rfree is 0.2254/0.2648, this is not the final result. I'm still working on it. Thank you so much. Best Regards, Tiantian
Re: [ccp4bb] cryo protection
Hey Len, I had this problem, too. As you know, my favorite first try is always fomblin (no need to mix anything). I had quite a bit success in stubborn cases to inject about 4uL fomblin through the tape on top of the drop and then looping crystals through the oil layer. You can wick the mother liquor off and try them right away or continue manipulation under oil Cheers, Jens On Wed, 2011-10-26 at 11:46 -0500, Leonard Thomas wrote: Hi All, I have run into a very sensitive crystals system when it comes to cryo protecting them. I have run through the usual suspects and trays are going to be setup with a cryo protectant as part of crystallization cocktail. The one problem that seems to be occurring is that the crystals crack as soon as they are transfered out of the original drop. I am running out of ideas and really would love some new ones. Thanks in advance. Len Leonard Thomas Ph.D. Macromolecular Crystallography Laboratory Manager University of Oklahoma Department of Chemistry and Biochemistry Stephenson Life Sciences Research Center 101 Stephenson Parkway Norman, OK 73019-5251 lmtho...@ou.edu http://barlywine.chem.ou.edu Office: (405)325-1126 Lab: (405)325-7571
Re: [ccp4bb] Off-topic: DSF thermo cycler low temp limit
CD spec with Pelletier is an option too 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 Oct 26, 2011, at 19:08, Reginald McNulty rmcnu...@uci.edu wrote: We are trying to determine Tm value of rather unstable proteins with a Tm in the mid 20 C range using DSF/thermofluor. Our Stratagene thermocycler has a low temp limit of 25 C (Peltier). I called the company and they said it's a 'hardware limit' that cannot be changed. 1) Has anyone been able to 'hotwire' the MX3000/3005 to go below this limit? 2) Are there other thermofluor machines that allow lower starting temperatures (say 4, 10 or 15 C)? Best regards, -Reggie
[ccp4bb] twist angle bwtween monomers
Hi all, Does anybody know of any software to calculate the twist angle between two monomers in a dimeric assembly. Or calculate manually. Thank you in advance. Sincerely Debajyoti
Re: [ccp4bb] twist angle bwtween monomers
Assuming you are dealing with a pure twist, isn't the polar rotation angle reported by lsqkab or superpose what you are looking for? On Thu, 2011-10-27 at 04:16 +, Debajyoti Dutta wrote: Hi all, Does anybody know of any software to calculate the twist angle between two monomers in a dimeric assembly. Or calculate manually. Thank you in advance. Sincerely Debajyoti Treat yourself at a restaurant, spa, resort and much more with Rediff Deal ho jaye!
Re: [ccp4bb] cryo protection
I have always been a fan of oil, which has already been suggested. Have you tried that? Cross-linking has already been suggested, and these are some good protocols: Lusty (1999) J. Appl. Crystallogr. 32, 106-112. McWhirter, et al. (1999) PNAS USA 96, 8408-8413. In the latter paper the crystals cracked immediately upon breaking the seal on the cover slip (limbo trays). The cross linker was introduced with a Hamilton syringe via a pre-cut and grease-filled hole in the crystallization chamber. That way there were no mechanical vibrations at all. You do NOT need to add it directly to the drop. Gluteraldehyde has sufficient vapor pressure to permeate slowly into it. After a few days, the crystals were incredibly robust, and gave the best diffraction. The only problem with gluteraldehyde is if you have primary amines in your buffer, such as tris. If that is the case, you can usually substitute bis-tris, or just use a different kind of crosslinker. Another trick I like if you have a cryo component in the mother liquor (protein counts) is to just let the drop dry up slowly. You can keep sampling it with a small loop (removes ~1 nl) until you see it flash-cool clear. Then, if the crystals survived, you can flash-cool them in the dried-down mother liquor. This worked for me once with crystals that just didn't want to transfer into anything. -James Holton MAD Scientist On 10/26/2011 9:46 AM, Leonard Thomas wrote: Hi All, I have run into a very sensitive crystals system when it comes to cryo protecting them. I have run through the usual suspects and trays are going to be setup with a cryo protectant as part of crystallization cocktail. The one problem that seems to be occurring is that the crystals crack as soon as they are transfered out of the original drop. I am running out of ideas and really would love some new ones. Thanks in advance. Len Leonard Thomas Ph.D. Macromolecular Crystallography Laboratory Manager University of Oklahoma Department of Chemistry and Biochemistry Stephenson Life Sciences Research Center 101 Stephenson Parkway Norman, OK 73019-5251 lmtho...@ou.edu http://barlywine.chem.ou.edu Office: (405)325-1126 Lab: (405)325-7571
