Re: [ccp4bb] Confusion about space group nomenclature
On Tue, 2014-04-29 at 16:12 +0200, Bernhard Rupp wrote: Response to off-board mail: How about [calling them] non-centro-symmetric space groups, as I often tell my students? Almost, but not exact enough. The 65 are only a subset of non-centrosymmetric space groups: Not all enantiogenic (not elements of the 65-set) space groups are centrosymmetric. Simplest example Pm. According to above definition Pm (and many more lacking a center of inversion) would be a ok space group for chiral motifs. (when a space group has the 'center at ' annotation in the Tables, it has a coi and is a centrosymmetric space group). This implies that there are actually three types of crystal structures (cf. Flack): (a) chiral (non-centrosymmetric) crystal structures (b) centrosymmetric crystal structures (c) achiral non-centrosymmetric crystal structures And just as a reminder, the substructure inversion for 3 members of the 65 is not about the origin (0,0,0): I41, I4122, F4132 are their own enantiomorph, so for them there is no enantiomorphic pair (eg. I41 and I43), in fact no separate space group I43 is even necessary - look at the SG diagram #80 - both, 41 and 43 axes appear in the same SG. (2005 Erice paper of George explains more) Enough yet? Cheers, BR Not quite, here's a table giving the complete list of the 3 types: http://pd.chem.ucl.ac.uk/pdnn/symm3/allsgp.htm The table heading states: Space groups possessing a point of inversion are termed *centrosymmetric*; these are shown in the table in red. Some space groups have no symmetry element that can change the handedness of an object; these are termed *enantiomorphic* space groups and are shown in magenta. i.e. your (a) set are the magenta ones, your (b) set are the red ones and your (c) set are the remaining black ones. -- Ian
Re: [ccp4bb] shelxCDE in ccp4i
-BEGIN PGP SIGNED MESSAGE- Hash: SHA1 Dear Mirek, can you check shelx c/d/e are actually in your path when you start ccp4i? You can open a terminal and type 'shelxc' to check and then start ccp4i from the very same terminal - are shelx c/d/e still greyed out? Best, Tim On 04/30/2014 01:31 AM, Cygler, Miroslaw wrote: Hi, I have shelx programs installed on my computer but only shelxs shows as active on the ccp4i interface. How to activate shelxCDE? Thanks, Mirek - -- - -- Dr Tim Gruene Institut fuer anorganische Chemie Tammannstr. 4 D-37077 Goettingen GPG Key ID = A46BEE1A -BEGIN PGP SIGNATURE- Version: GnuPG v1.4.12 (GNU/Linux) Comment: Using GnuPG with Icedove - http://www.enigmail.net/ iD8DBQFTYMdXUxlJ7aRr7hoRAo0iAKC8v+mHDJStiOpzsTeiWsyRjQYmAQCeL1sR DRx+JRvCVkV9xnzkyr8EQlY= =EwXv -END PGP SIGNATURE-
[ccp4bb] activation of ShelxC/D/E
Dear All I am facing problem in installation of ShelxCDE on CCP4 6.4.0. I have installed CCP4 and all the programs are running successfully. I copied and pasted ShelxCDE executables into CCP4 bin. Earlier also I adopted the same procedure to activate ShelxCDE on CCp4 interface and it worked. But this time its not working. Please suggest me how to solve this issue. Thanking you in advance. Best regards Shanti Pal
Re: [ccp4bb] activation of ShelxC/D/E
-BEGIN PGP SIGNED MESSAGE- Hash: SHA1 Dear Shanti Pal, when you download the shelx binaries the executable flag is not necessarily set. Can you check with 'ls -l $CBIN/shelxc' if this is the case? You can set it with 'chmod +x $CBIN/shelxc'. Best, Tim On 04/30/2014 11:51 AM, Shanti Pal Gangwar wrote: Dear All I am facing problem in installation of ShelxCDE on CCP4 6.4.0. I have installed CCP4 and all the programs are running successfully. I copied and pasted ShelxCDE executables into CCP4 bin. Earlier also I adopted the same procedure to activate ShelxCDE on CCp4 interface and it worked. But this time its not working. Please suggest me how to solve this issue. Thanking you in advance. Best regards Shanti Pal - -- - -- Dr Tim Gruene Institut fuer anorganische Chemie Tammannstr. 4 D-37077 Goettingen GPG Key ID = A46BEE1A -BEGIN PGP SIGNATURE- Version: GnuPG v1.4.12 (GNU/Linux) Comment: Using GnuPG with Icedove - http://www.enigmail.net/ iD8DBQFTYMfwUxlJ7aRr7hoRAsHlAKCwS96CX+qtOhXv6+ReIRWMAAJmHwCgkr+6 ceM6G31HQibsAMgZ9nNKyTI= =6scZ -END PGP SIGNATURE-
[ccp4bb] metals disapear
Hi all, Has anyone experienced catalytic metal ions disappearing during data collection ? If so, is there a way of preventing it? D. Dean Derbyshire Senior Research Scientist [cid:image001.jpg@01CF6470.5FA976D0] Box 1086 SE-141 22 Huddinge SWEDEN Visit: Lunastigen 7 Direct: +46 8 54683219 www.medivir.comhttp://www.medivir.com -- This transmission is intended for the person to whom or the entity to which it is addressed and may contain information that is privileged, confidential and exempt from disclosure under applicable law. If you are not the intended recipient, please be notified that any dissemination, distribution or copying is strictly prohibited. If you have received this transmission in error, please notify us immediately. Thank you for your cooperation.
Re: [ccp4bb] metals disapear
-BEGIN PGP SIGNED MESSAGE- Hash: SHA1 Dear Dean, this is probably a very common observation: X-rays produce reducing electrons and as you reduce a metal I imagine it does not like its chemical environment as much as it did highly charged. Everything you can do to avoid radiation damage should help you prevent the ion to disappear: - - optimise your strategy to collect a minimal amount of data - - add vitamin C - - cool below 100K - - collect at short wavelength When your ion is intended to be used for phasing there are of course restraints limiting the choice. Regards, Tim On 04/30/2014 12:33 PM, Dean Derbyshire wrote: Hi all, Has anyone experienced catalytic metal ions disappearing during data collection ? If so, is there a way of preventing it? D. Dean Derbyshire Senior Research Scientist [cid:image001.jpg@01CF6470.5FA976D0] Box 1086 SE-141 22 Huddinge SWEDEN Visit: Lunastigen 7 Direct: +46 8 54683219 www.medivir.comhttp://www.medivir.com -- This transmission is intended for the person to whom or the entity to which it is addressed and may contain information that is privileged, confidential and exempt from disclosure under applicable law. If you are not the intended recipient, please be notified that any dissemination, distribution or copying is strictly prohibited. If you have received this transmission in error, please notify us immediately. Thank you for your cooperation. - -- - -- Dr Tim Gruene Institut fuer anorganische Chemie Tammannstr. 4 D-37077 Goettingen GPG Key ID = A46BEE1A -BEGIN PGP SIGNATURE- Version: GnuPG v1.4.12 (GNU/Linux) Comment: Using GnuPG with Icedove - http://www.enigmail.net/ iD8DBQFTYNSPUxlJ7aRr7hoRAr7WAKCzC7FzqTkcVLILovmIL74OUQlsWQCgg2Yr xZgDCvIlf5YEWHLTDLiKcRc= =tp4F -END PGP SIGNATURE-
[ccp4bb] anomalous signal
Dear all I am working on a metalloprotein which probably contains Ca at its active site..The sulfur containing amino acid constitutes almost 5.4% of the total amino acid residues of this protein..I have collected the data at home source (CuKalpha=1.54A)..Since f'' of Sulfur is 0.56 and that of Ca is 1.28 we can always expect some anomalous signal out of the data..My question is ..how we will know if the anomalous signal is coming out of Sulfur or from Calcium ?? is there any method through which we can get to know the identity of the scattering molecule through the data..Can FFT anomalous map from CCP4 is of any help in this direction, if yes then please tell me how to interpret the output from this.. -- Regards Faisal School of Life Sciences JNU
Re: [ccp4bb] metals disapear
Can the radiation damage gurus comment on this ? I know there is a problem with radiation damage changing the valence state of metals, but I don't remember hearing about the metal actually being lost due to radiation damage. Is this really common ? Thanks, Andrew On 30 Apr 2014, at 11:46, Tim Gruene t...@shelx.uni-ac.gwdg.de wrote: -BEGIN PGP SIGNED MESSAGE- Hash: SHA1 Dear Dean, this is probably a very common observation: X-rays produce reducing electrons and as you reduce a metal I imagine it does not like its chemical environment as much as it did highly charged. Everything you can do to avoid radiation damage should help you prevent the ion to disappear: - - optimise your strategy to collect a minimal amount of data - - add vitamin C - - cool below 100K - - collect at short wavelength When your ion is intended to be used for phasing there are of course restraints limiting the choice. Regards, Tim On 04/30/2014 12:33 PM, Dean Derbyshire wrote: Hi all, Has anyone experienced catalytic metal ions disappearing during data collection ? If so, is there a way of preventing it? D. Dean Derbyshire Senior Research Scientist [cid:image001.jpg@01CF6470.5FA976D0] Box 1086 SE-141 22 Huddinge SWEDEN Visit: Lunastigen 7 Direct: +46 8 54683219 www.medivir.comhttp://www.medivir.com -- This transmission is intended for the person to whom or the entity to which it is addressed and may contain information that is privileged, confidential and exempt from disclosure under applicable law. If you are not the intended recipient, please be notified that any dissemination, distribution or copying is strictly prohibited. If you have received this transmission in error, please notify us immediately. Thank you for your cooperation. - -- - -- Dr Tim Gruene Institut fuer anorganische Chemie Tammannstr. 4 D-37077 Goettingen GPG Key ID = A46BEE1A -BEGIN PGP SIGNATURE- Version: GnuPG v1.4.12 (GNU/Linux) Comment: Using GnuPG with Icedove - http://www.enigmail.net/ iD8DBQFTYNSPUxlJ7aRr7hoRAr7WAKCzC7FzqTkcVLILovmIL74OUQlsWQCgg2Yr xZgDCvIlf5YEWHLTDLiKcRc= =tp4F -END PGP SIGNATURE-
Re: [ccp4bb] anomalous signal
Dear Faisal, you could use Phenix to do an f’’ refinement against the data like described here: Liu, Q., Liu, Q. Hendrickson, W. A. (2013). Acta Cryst. D69, 1314-1332. best regards, Tobias On 30 Apr 2014, at 14:01, Faisal Tarique faisaltari...@gmail.com wrote: Dear all I am working on a metalloprotein which probably contains Ca at its active site..The sulfur containing amino acid constitutes almost 5.4% of the total amino acid residues of this protein..I have collected the data at home source (CuKalpha=1.54A)..Since f'' of Sulfur is 0.56 and that of Ca is 1.28 we can always expect some anomalous signal out of the data..My question is ..how we will know if the anomalous signal is coming out of Sulfur or from Calcium ?? is there any method through which we can get to know the identity of the scattering molecule through the data..Can FFT anomalous map from CCP4 is of any help in this direction, if yes then please tell me how to interpret the output from this.. -- Regards Faisal School of Life Sciences JNU
Re: [ccp4bb] metals disapear
Attenuate the beam and take many more frames such that you get a full dataset, albeit at slightly lower resolution, before the metals go away. In other words, reduce dose/degPhi. JPK From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of Dean Derbyshire Sent: Wednesday, April 30, 2014 6:33 AM To: CCP4BB@JISCMAIL.AC.UK Subject: [ccp4bb] metals disapear Hi all, Has anyone experienced catalytic metal ions disappearing during data collection ? If so, is there a way of preventing it? D. Dean Derbyshire Senior Research Scientist [cid:image001.jpg@01CF6450.2D6A0D80] Box 1086 SE-141 22 Huddinge SWEDEN Visit: Lunastigen 7 Direct: +46 8 54683219 www.medivir.comhttp://www.medivir.com -- This transmission is intended for the person to whom or the entity to which it is addressed and may contain information that is privileged, confidential and exempt from disclosure under applicable law. If you are not the intended recipient, please be notified that any dissemination, distribution or copying is strictly prohibited. If you have received this transmission in error, please notify us immediately. Thank you for your cooperation.
[ccp4bb] Pilatus and Strategy wrt Radiation Damage
Dear Pilatus/Radiation Damage Cognoscenti, I read a few years ago, before the advent of Pilatus detectors, that the best strategy was a sort of compromise between number of images and detector readout noise overhead. I have heard that Pilatus detectors, however, have essentially no readout noise, so I am wondering whether strategies have changed in light of this, i.e., is the best practice now to collect as many images as possible at lowest exposure possible? JPK *** Jacob Pearson Keller, PhD Looger Lab/HHMI Janelia Farms Research Campus 19700 Helix Dr, Ashburn, VA 20147 email: kell...@janelia.hhmi.org ***
Re: [ccp4bb] metals disapear
Dear Dean (and Andrew), I am certainly no radiation damage guru, but at the RD8 Workshop in Hamburg 3 weeks ago (http://www.rd-eight.org/RD8-01/) I heard a talk by Pernille Harris about the photoreduction by X-rays of Cu(II) to Cu(I) in a Cu-insulin crystal. Tim's description couldn't be more appropriate: Cu(II) likes e.g. square-planar or square-bipyramidal coordination, while Cu(I) likes tetrahedral coordination; so a Cu(II) atom photoreduced to a Cu(I) might find itself acutely uncomfortable in its previous surroundings and want to rearrange its ligands. If its presence in the protein is the outcome of an evolutionary choice to enable it to carry out e.g. some electron transport function, a rearrangement upon a change of oxidation state can be expected to be be well choreographed, and the ligand rearrangement quite orderly. However if the catalytic role of that metal is predicated on a fixed oxidation state, the environment of the photoreduced metal atom could become highly disordered: that atom and the surrounding ones would then end up with very high B-factors and would look as if they had disappeared. Do get in touch with the real gurus, though, as listed on the RD8 page indicated above. With best wishes, Gerard. -- On Wed, Apr 30, 2014 at 01:08:59PM +0100, Andrew Leslie wrote: Can the radiation damage gurus comment on this ? I know there is a problem with radiation damage changing the valence state of metals, but I don't remember hearing about the metal actually being lost due to radiation damage. Is this really common ? Thanks, Andrew On 30 Apr 2014, at 11:46, Tim Gruene t...@shelx.uni-ac.gwdg.de wrote: -BEGIN PGP SIGNED MESSAGE- Hash: SHA1 Dear Dean, this is probably a very common observation: X-rays produce reducing electrons and as you reduce a metal I imagine it does not like its chemical environment as much as it did highly charged. Everything you can do to avoid radiation damage should help you prevent the ion to disappear: - - optimise your strategy to collect a minimal amount of data - - add vitamin C - - cool below 100K - - collect at short wavelength When your ion is intended to be used for phasing there are of course restraints limiting the choice. Regards, Tim On 04/30/2014 12:33 PM, Dean Derbyshire wrote: Hi all, Has anyone experienced catalytic metal ions disappearing during data collection ? If so, is there a way of preventing it? D. Dean Derbyshire Senior Research Scientist [cid:image001.jpg@01CF6470.5FA976D0] Box 1086 SE-141 22 Huddinge SWEDEN Visit: Lunastigen 7 Direct: +46 8 54683219 www.medivir.comhttp://www.medivir.com -- This transmission is intended for the person to whom or the entity to which it is addressed and may contain information that is privileged, confidential and exempt from disclosure under applicable law. If you are not the intended recipient, please be notified that any dissemination, distribution or copying is strictly prohibited. If you have received this transmission in error, please notify us immediately. Thank you for your cooperation. - -- - -- Dr Tim Gruene Institut fuer anorganische Chemie Tammannstr. 4 D-37077 Goettingen GPG Key ID = A46BEE1A -BEGIN PGP SIGNATURE- Version: GnuPG v1.4.12 (GNU/Linux) Comment: Using GnuPG with Icedove - http://www.enigmail.net/ iD8DBQFTYNSPUxlJ7aRr7hoRAr7WAKCzC7FzqTkcVLILovmIL74OUQlsWQCgg2Yr xZgDCvIlf5YEWHLTDLiKcRc= =tp4F -END PGP SIGNATURE- -- === * * * Gerard Bricogne g...@globalphasing.com * * * * Global Phasing Ltd. * * Sheraton House, Castle Park Tel: +44-(0)1223-353033 * * Cambridge CB3 0AX, UK Fax: +44-(0)1223-366889 * * * ===
Re: [ccp4bb] Pilatus and Strategy wrt Radiation Damage
Hi Jacob, I'll take a first crack as I am sure many will follow. It is true that with CCD detectors one has to be careful how small an oscillation range to use for a frame before read noise starts to eat into the data quality. Pilatus offers two major new features - is fast and is photon counting as opposed to integrating detector. The speed allows to collect data without a shutter and it is very important as it can dramatically improve data quality. Now there are fast CCD detectors as well on the market. Being a photon counter, Pilatus has no read noise which, as you have pointed out, allows you to collect as thin a frame as you want. However, it is if you consider the detector only. In reality, if you go very thin and very fast, you may not have enough flux to record the data. Also, even once we get rid of the shutter, there are still other sources of instabilities and they do affect the fast data collection adversely. One could try going (very) thin sliced and somewhat slow but there is another gotcha there. Most rotation stages used for rotating the sample crystal, do not like going extremely slow which would be the case with thin frames and long exposure times. In this case the speed may not remain as constant as we would like during data collection. I think there was a publication from Diamond Synchrotron discussing strategies of data collection with Pilatus. We've done a little bit of systematic studies as well and while things may well be sample- and facility-dependent, ~0.2 degree frames with ~0.2 sec exposure time seemed to make good compromise between above-mentioned issues. Here I would like to emphasize again - there certainly will be samples which will benefit from somewhat different parameters. Hope it helps, Cheers, N. Ruslan Sanishvili (Nukri) Macromolecular Crystallographer GM/CA@APS X-ray Science Division, ANL 9700 S. Cass Ave. Lemont, IL 60439 Tel: (630)252-0665 Fax: (630)252-0667 rsanishv...@anl.gov From: CCP4 bulletin board [CCP4BB@jiscmail.ac.uk] on behalf of Keller, Jacob [kell...@janelia.hhmi.org] Sent: Wednesday, April 30, 2014 7:49 AM To: CCP4BB@jiscmail.ac.uk Subject: [ccp4bb] Pilatus and Strategy wrt Radiation Damage Dear Pilatus/Radiation Damage Cognoscenti, I read a few years ago, before the advent of Pilatus detectors, that the best strategy was a sort of compromise between number of images and detector readout noise overhead. I have heard that Pilatus detectors, however, have essentially no readout noise, so I am wondering whether strategies have changed in light of this, i.e., is the best practice now to collect as many images as possible at lowest exposure possible? JPK *** Jacob Pearson Keller, PhD Looger Lab/HHMI Janelia Farms Research Campus 19700 Helix Dr, Ashburn, VA 20147 email: kell...@janelia.hhmi.org ***
Re: [ccp4bb] metals disapear
My comments: Such observation is very uncommon for metals involved in catalysis by proteins. I have seen quite a few such structures involving Mg, Ca, Fe, Mn, Zn and most of the radiation damage was not at the catalytic metal. In case of Fe once I noticed slight shift in the position of the Fe ion upon exposure. The only metal ion that was significantly affected is Hg, and this was observed in multiple cases. We published results of radiation damage as a function of temperature going down up to 15K. There was some overall reduction of radiation damage, by about factor of 1.7; however, the most of the impact was away from the catalytic site. As a result, relative radiation damage was MORE concentrated at the catalytic site. Metals (Ca and Mn) were not particularly affected at any temperature. We observed that nitrate and iodine scavenge radicals help reduce specific radiation damage. However, increased X-ray absorption by iodine does makes overall situation worse and impact of nitrate was observed only at relatively low doses (up to 2 MGy). Kmetko at all. (2011) were negative about potential of using scavengers in general, including ascorbic acid. Data collection wavelength does not matter! It is an urban legend that shorter wavelength will help. I remember it being debunked two decades ago, and somehow it is still alive. Zbyszek Otwinowski -BEGIN PGP SIGNED MESSAGE- Hash: SHA1 Dear Dean, this is probably a very common observation: X-rays produce reducing electrons and as you reduce a metal I imagine it does not like its chemical environment as much as it did highly charged. Everything you can do to avoid radiation damage should help you prevent the ion to disappear: - - optimise your strategy to collect a minimal amount of data - - add vitamin C - - cool below 100K - - collect at short wavelength When your ion is intended to be used for phasing there are of course restraints limiting the choice. Regards, Tim On 04/30/2014 12:33 PM, Dean Derbyshire wrote: Hi all, Has anyone experienced catalytic metal ions disappearing during data collection ? If so, is there a way of preventing it? D. Dean Derbyshire Senior Research Scientist [cid:image001.jpg@01CF6470.5FA976D0] Box 1086 SE-141 22 Huddinge SWEDEN Visit: Lunastigen 7 Direct: +46 8 54683219 www.medivir.comhttp://www.medivir.com -- This transmission is intended for the person to whom or the entity to which it is addressed and may contain information that is privileged, confidential and exempt from disclosure under applicable law. If you are not the intended recipient, please be notified that any dissemination, distribution or copying is strictly prohibited. If you have received this transmission in error, please notify us immediately. Thank you for your cooperation. - -- - -- Dr Tim Gruene Institut fuer anorganische Chemie Tammannstr. 4 D-37077 Goettingen GPG Key ID = A46BEE1A -BEGIN PGP SIGNATURE- Version: GnuPG v1.4.12 (GNU/Linux) Comment: Using GnuPG with Icedove - http://www.enigmail.net/ iD8DBQFTYNSPUxlJ7aRr7hoRAr7WAKCzC7FzqTkcVLILovmIL74OUQlsWQCgg2Yr xZgDCvIlf5YEWHLTDLiKcRc= =tp4F -END PGP SIGNATURE- Zbyszek Otwinowski UT Southwestern Medical Center at Dallas 5323 Harry Hines Blvd. Dallas, TX 75390-8816 Tel. 214-645-6385 Fax. 214-645-6353
Re: [ccp4bb] anomalous signal
Well - this is pretty common, and really doesnt matter. I just let SHELXC/D give an occupancy to its anom scatterer solutions and assume that the stronger one is Ca. But in fact it wont matter at all for the phasing, and IF the experiment works it is easy to sort out S from Ca in the final map. On 30 April 2014 13:21, Tobias Weinert tobias.wein...@psi.ch wrote: Dear Faisal, you could use Phenix to do an f’’ refinement against the data like described here: Liu, Q., Liu, Q. Hendrickson, W. A. (2013). Acta Cryst. D69, 1314-1332. best regards, Tobias On 30 Apr 2014, at 14:01, Faisal Tarique faisaltari...@gmail.com wrote: Dear all I am working on a metalloprotein which probably contains Ca at its active site..The sulfur containing amino acid constitutes almost 5.4% of the total amino acid residues of this protein..I have collected the data at home source (CuKalpha=1.54A)..Since f'' of Sulfur is 0.56 and that of Ca is 1.28 we can always expect some anomalous signal out of the data..My question is ..how we will know if the anomalous signal is coming out of Sulfur or from Calcium ?? is there any method through which we can get to know the identity of the scattering molecule through the data..Can FFT anomalous map from CCP4 is of any help in this direction, if yes then please tell me how to interpret the output from this.. -- Regards Faisal School of Life Sciences JNU
Re: [ccp4bb] Pilatus and Strategy wrt Radiation Damage
Hi Marcus Mueller (from Dectris, who develop and manufacture the Pilatus) did some work on this a couple of years ago and determined that an oscillation angle ~ 0.5x the mosaicity of the crystal (using the XDS value of mosaicity, which is not the same as Mosflm's); the abstract says - The results show that fine ’-slicing can substantially improve scaling statistics and anomalous signal provided that the rotation angle is comparable to half the crystal mosaicity. Acta Cryst. (2012). D68, 42-56[ doi:10.1107/S0907444911049833 ] Optimal fine -slicing for single-photon-counting pixel detectors M. Mueller, M. Wang and C. Schulze-Briese My reading of this is that there is still a place for strategy calculations. On 30 Apr 2014, at Wed30 Apr 15:06, Sanishvili, Ruslan wrote: Hi Jacob, I'll take a first crack as I am sure many will follow. It is true that with CCD detectors one has to be careful how small an oscillation range to use for a frame before read noise starts to eat into the data quality. Pilatus offers two major new features - is fast and is photon counting as opposed to integrating detector. The speed allows to collect data without a shutter and it is very important as it can dramatically improve data quality. Now there are fast CCD detectors as well on the market. Being a photon counter, Pilatus has no read noise which, as you have pointed out, allows you to collect as thin a frame as you want. However, it is if you consider the detector only. In reality, if you go very thin and very fast, you may not have enough flux to record the data. Also, even once we get rid of the shutter, there are still other sources of instabilities and they do affect the fast data collection adversely. One could try going (very) thin sliced and somewhat slow but there is another gotcha there. Most rotation stages used for rotating the sample crystal, do not like going extremely slow which would be the case with thin frames and long exposure times. In this case the speed may not remain as constant as we would like during data collection. I think there was a publication from Diamond Synchrotron discussing strategies of data collection with Pilatus. We've done a little bit of systematic studies as well and while things may well be sample- and facility-dependent, ~0.2 degree frames with ~0.2 sec exposure time seemed to make good compromise between above-mentioned issues. Here I would like to emphasize again - there certainly will be samples which will benefit from somewhat different parameters. Hope it helps, Cheers, N. Ruslan Sanishvili (Nukri) Macromolecular Crystallographer GM/CA@APS X-ray Science Division, ANL 9700 S. Cass Ave. Lemont, IL 60439 Tel: (630)252-0665 Fax: (630)252-0667 rsanishv...@anl.gov From: CCP4 bulletin board [CCP4BB@jiscmail.ac.uk] on behalf of Keller, Jacob [kell...@janelia.hhmi.org] Sent: Wednesday, April 30, 2014 7:49 AM To: CCP4BB@jiscmail.ac.uk Subject: [ccp4bb] Pilatus and Strategy wrt Radiation Damage Dear Pilatus/Radiation Damage Cognoscenti, I read a few years ago, before the advent of Pilatus detectors, that the best strategy was a sort of compromise between number of images and detector readout noise overhead. I have heard that Pilatus detectors, however, have essentially no readout noise, so I am wondering whether strategies have changed in light of this, i.e., is the best practice now to collect as many images as possible at lowest exposure possible? JPK *** Jacob Pearson Keller, PhD Looger Lab/HHMI Janelia Farms Research Campus 19700 Helix Dr, Ashburn, VA 20147 email: kell...@janelia.hhmi.org *** Harry -- ** note change of address ** Dr Harry Powell, MRC Laboratory of Molecular Biology, Francis Crick Avenue, Cambridge Biomedical Campus, Cambridge, CB2 0QH Chairman of European Crystallographic Association SIG9 (Crystallographic Computing)
[ccp4bb] NWCW 2014: Last day to register at early registration rates
-BEGIN PGP SIGNED MESSAGE- Hash: SHA1 Northwest Crystallography Workshop http://oregonstate.edu/conferences/event/nwcw2014/ June 20-22, 2014 The (Pacific) Northwest Crystallography Workshop is a regional gathering of people who are interested in macromolecular structure determination but folk from anywhere are welcome. This year it will be held at Oregon State University in Corvallis Oregon. Today is the last day to register at the early registration rates of $75/$100 for students/others. Tomorrow the prices will rise by $25. We encourage you to sign up today! We will continue to accept abstracts until May 16th, but please try to get them in ASAP. Registration is not linked to abstract submission so you can register today and submit an abstract later. Next week, however, we will begin to define the speaking schedule. Dale E. Tronrud and P. Andrew Karplus Department of Biochemistry and Biophysics 2011 ALS Bldg Oregon State University Corvallis, OR 97331 -BEGIN PGP SIGNATURE- Version: GnuPG v2.0.22 (MingW32) Comment: Using GnuPG with Thunderbird - http://www.enigmail.net/ iEYEARECAAYFAlNhKJgACgkQU5C0gGfAG13b5wCfdlIVFbwJlZX2MPzXLdSODCpm nxwAoL19Va3igMKmXQhtkBmBf+7rkPsk =U+SM -END PGP SIGNATURE-
Re: [ccp4bb] anomalous signal
I myself have never seen a separate peak that was a single sulfur atom. I've seen, in a moderately-radiation-damaged dataset, little shards of anomalous scattering density in Phaser-generated LLG maps. They were in the interior of the protein, and the closest possible atoms were sulfurs. They were not in positions suggestive of Fourier truncation ripples, so I hypothesized that these sulfurs were ionized and landed at the nearest convenient location, and left it at that. JPK
Re: [ccp4bb] metals disapear
Dear Dean An example, albeit not a metal, can be found here:- http://journals.iucr.org/s/issues/2007/01/00/xh5011/xh5011.pdf Such specific damage has a long history:- http://www.sciencedirect.com/science/article/pii/0022024888903223 An X-ray sensitive metals centre is the Mn5Ca OEC of PS II and details can found here :- http://m.pnas.org/content/102/34/12047.long But metals in proteins can also be robust to X-rays and eg verified by parallel non damaging neutron crystallographic studies such as on MnCa concanabalin A . Best wishes John Prof John R Helliwell DSc On 30 Apr 2014, at 11:33, Dean Derbyshire dean.derbysh...@medivir.com wrote: Hi all, Has anyone experienced catalytic metal ions disappearing during data collection ? If so, is there a way of preventing it? D. Dean Derbyshire Senior Research Scientist image001.jpg Box 1086 SE-141 22 Huddinge SWEDEN Visit: Lunastigen 7 Direct: +46 8 54683219 www.medivir.com -- This transmission is intended for the person to whom or the entity to which it is addressed and may contain information that is privileged, confidential and exempt from disclosure under applicable law. If you are not the intended recipient, please be notified that any dissemination, distribution or copying is strictly prohibited. If you have received this transmission in error, please notify us immediately. Thank you for your cooperation.
Re: [ccp4bb] metals disapear
The question about metal sensitivity to radiation cannot be answered in general; it needs to be discussed in specific chemical coordination context. Agreed. The author of the original question should have provided more details about the metal in question, about the samples and the way experiment was carried out. The advice about using helical scan is horrible in this context. The diffraction collected by such method represents state of crystal exposed to constant high dose. If anything, the helical scan method is more suitable to study radiation damaged state of the crystal. I am afraid the advise was horribly misunderstood. A crystal during helical data collection doesn't have to be exposed to constant high dose. The exposure level is selected by the experimenter and is not intrinsic feature of helical data collection. The advise comprised a four-step program and it started by determining (in the first step) a lower dose that would still allow making valid enzymologic (or mechanistic) conclusions. Then further steps instructed how to lower this dose even more by using multiple crystals (if necessary due to poor crystals quality - 2nd step), or by spreading the same low dose over a larger volume using helical data collection - step 4. I suspect the misunderstanding is based on a misconception that if one is using helical data collection, one necessarily is using small beam and high intensity, but it is not so at all. The beam size to be used is dictated by the size of the well-diffracting volume (advised to determine in step #3). If one has large well-diffracting volume, large beam can be used for helical data collection as well (if the volume is larger than the beam size). Hope it clarifies things little better. Cheers, N. Ruslan Sanishvili (Nukri) Macromolecular Crystallographer GM/CA@APS X-ray Science Division, ANL 9700 S. Cass Ave. Lemont, IL 60439 Tel: (630)252-0665 Fax: (630)252-0667 rsanishv...@anl.gov From: zbys...@work.swmed.edu [zbys...@work.swmed.edu] Sent: Wednesday, April 30, 2014 10:33 AM To: Sanishvili, Ruslan Cc: ccp4bb@jiscmail.ac.uk Subject: Re: [ccp4bb] metals disapear If metal ion will be sensitive to radiation depends on its redox chemistry and not its X-ray properties. For a metal to be affected by radiation dose it needs to be reduced by free radicals. However, such metals are rarely (by gene counts or deposits in PDB) present in catalytic sites of enzymes. The most frequently occurring metal ions in catalytic sites e.g. Mg++, Ca++, Zn++, Fe++, Mn++ lack oxidation state to which they could be reduced by a single radical. For this reason these metals tend to be very stable upon radiation and they are last to go. Copper is the counterexample where radiation sensitivity is much more likely to be expected. Unfortunately, the original question and part of the discussion involved generic category of metals involved in catalysis, rather then specific one. Magnesium is by far the the most frequently encounter metal in catalytic sites. In redox reaction the most frequent cofactors are not metals, but NAD or FAD. Iron is most often present in Fe-S clusters rather than as standalone Fe++ ion. These clusters are structurally diverse and do not necessarily participate in catalysis. If they have similar or diverse sensitivity to radiation is not clear to me. The question about metal sensitivity to radiation cannot be answered in general; it needs to be discussed in specific chemical coordination context. Separate issue: The advice about using helical scan is horrible in this context. The diffraction collected by such method represents state of crystal exposed to constant high dose. If anything, the helical scan method is more suitable to study radiation damaged state of the crystal. Zbyszek Otwinowski Dear Dean, You have already received excellent insight into radiation effects on metals. From personal experience, it doesn't take long for the metal occupancy to go down to 80%. Of course it is not anywhere near disappearing but then again, we don't know the details of your data collection and of how disappeared are your disappeared metals. I will only add that you can use modern approaches to data collection to minimize the adverse effects of radiation. 1. Do not chase the highest resolution and try to get what is enough to make your enzymologic statements valid. I.e. use less dose. 2. If your crystals diffract poorly, consider using several crystals to merge few sets of underexposed data. 3. For each crystal, use some sort of grid scanning (we call it rastering) to estimate the crystal quality. It is implemented on many beamlines worldwide. I will insert a shameless plug here and others can follow the suite... See Hilgart et al., pages 717-722 here: http://journals.iucr.org/s/issues/2011/05/00/issconts.html 4. Select the better diffracting regions of the crystal and use vector or helical data collection
