Re: [ccp4bb] Off topic: Crystal degredation with age
Hi Ed, A bit late on the subject I have collected atomic resolution data (around 0.97A) on both bovine and porcine phospholipase A2 crystals which at the time of data collection were between 10-16 years old. Crystallization setup was liquid-liquid diffusion in glass capillaries. Differently from the other stories, here there's no degradation involved...just rock-solid Xtals. In the paper we stated: 'Crystals are stable in the crystallization solution...' which I guess well reflects their behavior. Best wishes, Roberto On 5 Feb 2009, at 19:11, Edward Snell wrote: /lurk_mode_off /dumb_question_on Dear All, I was recently trying to find references on how age may degrade a crystal, i.e. grow them and use them or preserve them as fresh as possible. I seem to remember seeing a couple of papers on this but my memory is fading and I have been unable to locate them. Can anyone jog my memory or tell me if I'm imagining things? I've found plenty on the protein prep etc. but nothing on the crystal. Thanks, Eddie. Edward Snell Ph.D. Assistant Prof. Department of Structural Biology, SUNY Buffalo, Hauptman-Woodward Medical Research Institute 700 Ellicott Street, Buffalo, NY 14203-1102 Phone: (716) 898 8631 Fax: (716) 898 8660 Email: esn...@hwi.buffalo.edu Telepathy: 42.2 GHz Heisenberg was probably here!Crystallization, how quaint! /dumb_question_off /lurk_mode_on --- Roberto Steiner Randall Division of Cell and Molecular Biophysics New Hunt's House King's College London Guy's Campus London, SE1 1UL Phone +44 (0)20-7848-8216 Fax +44 (0)20-7848-6435 e-mail roberto.stei...@kcl.ac.uk
Re: [ccp4bb] Off topic: Crystal degredation with age
As written in Jovine, L., Djordjevic, S. Rhodes, D. (2000). “The crystal structure of yeast phenylalanine tRNA at 2.0 A resolution: cleavage by Mg(2+) in 15-year old crystals” J Mol Biol 301, 401-414. Furthermore, the possibility of a Mg2+-catalysed cleavage of the phosphodiester bond between H2U16 and H2U17 was also inferred from the electron density map of the orthorhombic form of the tRNAPhe crystals [Sussman et al 1978]. n.b. this chain break was seen 31 years ago, also, in fresh crystals. Sussman, J.L., Holbrook, S.R., Warrant, R.W., Church, G.M. Kim, S.- H. (1978). Crystal structure of yeast phenylalanine transfer RNA. I. crystallographic refinement J Mol Biol 123, 607-630. see Fig 9c: Residues 16 and 17. Note the discontinuity between phosphate 17 and ribose 17. This region is one of the weakest electron density regions. - Prof. Joel L. Sussmanjoel.suss...@weizmann.ac.il Pickman Prof. of Structural Biology +972 (8) 934 4531 - tel Department of Structural Biology +972 (8) 934 4159 - fax Weizmann Institute of Sciencewww.weizmann.ac.il/~joel Rehovot 76100 ISRAEL www.weizmann.ac.il/ISPC Proteopedia, www.proteopedia.org (because life has more than 2D) - On 5 Feb 2009, at 21:48, William G. Scott wrote: Some things improve with age. Here is one of my favorite stories: http://tinyurl.com/oldtrna The crystal structure of yeast phenylalanine tRNA at 2.0 Å resolution: cleavage by Mg2+ in 15-year old crystals Luca Jovine, Snezana Djordjevica and Daniela Rhodes We have re-determined the crystal structure of yeast tRNAPhe to 2.0 Å resolution using 15 year old crystals. The accuracy of the new structure, due both to higher resolution data and formerly unavailable refinement methods, consolidates the previous structural information, but also reveals novel details. In particular, the water structure around the tightly bound Mg2+ is now clearly resolved, and hence provides more accurate information on the geometry of the magnesium-binding sites and the role of water molecules in coordinating the metal ions to the tRNA. We have assigned a total of ten magnesium ions and identified a partly conserved geometry for high-affinity Mg2+ binding. In the electron density map there is also clear density for a spermine molecule binding in the major groove of the TΨC arm and also contacting a symmetry-related tRNA molecule. Interestingly, we have also found that two specific regions of the tRNA in the crystals are partially cleaved. The sites of hydrolysis are within the D and anticodon loops in the vicinity of Mg2+. On Feb 5, 2009, at 11:11 AM, Edward Snell wrote: /lurk_mode_off /dumb_question_on Dear All, I was recently trying to find references on how age may degrade a crystal, i.e. grow them and use them or preserve them as fresh as possible. I seem to remember seeing a couple of papers on this but my memory is fading and I have been unable to locate them. Can anyone jog my memory or tell me if I'm imagining things? I've found plenty on the protein prep etc. but nothing on the crystal. Thanks, Eddie. Edward Snell Ph.D. Assistant Prof. Department of Structural Biology, SUNY Buffalo, Hauptman-Woodward Medical Research Institute 700 Ellicott Street, Buffalo, NY 14203-1102 Phone: (716) 898 8631 Fax: (716) 898 8660 Email: esn...@hwi.buffalo.edu Telepathy: 42.2 GHz Heisenberg was probably here!Crystallization, how quaint! /dumb_question_off /lurk_mode_on
Re: [ccp4bb] Off topic: Crystal degredation with age
FWIW, many of the crystals used in classical neutron diffraction experiments were pretty elderly samples by the time data collection was initiated, partly to allow large crystals to grow ever larger, partly because of the mandatory deuterium exchange process and partly because the experiments lasted weeks to months. To be specific, I believe the crystal that led to the first endothiapepsin neutron structure was 11 years old before it began D2O-soaking prior to the neutron experiment. It diffracted pretty well, 2.1A I think... Cheers- Brad
Re: [ccp4bb] Off topic: Crystal degredation with age
For a counterexample, from Iwata's group, Horsefield et al. Succinate: quinone oxidoreductase Acta. Cryst.(2003). D59, 600-602: It proved critical to freeze the crystals within 72 h of crystallization set-up. Crystals that were frozen after this time limit showed no diffraction. This alteration in properties was apparent by the change in colour from deep orange to pale yellow that was observed in crystals more than four weeks old (Fig. 1c). The deep orange colour of the crystals is attributed to the presence of haem b within the protein. The loss or breakdown of haem, demonstrated by the change of colour in the crystals, could lead to structural instability and consequently loss of diffraction. I've heard the Fe-S clusters of E. coli SDH are oxygen-sensitive in the isolated enzyme. The mitochondrial counterpart is more stable, we've collected data from crystals 1-2 years old, and seen conversion (new crystals grow while old crystals dissolve) to a new space group with better diffraction (that was a within few months after setup). Edward Snell wrote: /lurk_mode_off /dumb_question_on Dear All, I was recently trying to find references on how age may degrade a crystal, i.e. grow them and use them or preserve them as fresh as possible. I seem to remember seeing a couple of papers on this but my memory is fading and I have been unable to locate them. Can anyone jog my memory or tell me if I'm imagining things? I've found plenty on the protein prep etc. but nothing on the crystal. Thanks, Eddie. Edward Snell Ph.D. Assistant Prof. Department of Structural Biology, SUNY Buffalo, Hauptman-Woodward Medical Research Institute 700 Ellicott Street, Buffalo, NY 14203-1102 Phone: (716) 898 8631 Fax: (716) 898 8660 Email: esn...@hwi.buffalo.edu Telepathy: 42.2 GHz Heisenberg was probably here!Crystallization, how quaint! /dumb_question_off /lurk_mode_on
Re: [ccp4bb] Off topic: Crystal degredation with age
I have to second that. I recently had crystals that will only grow after seeding and will live for exactly five days. On the sixth day, the same drop will have tracks of dissolved crystals left in every drop: they almost look like tire tracks. The crystals frozen on the fifth day diffract to 2.2 A, and I phased with these crystals using Se-Met, so they were definitely good on the fifth day. In this case, the structure did show why the crystals were so unstable, a whole domain (40% of the protein) was swerving several degrees without much crystal contacts holding it down. Actually, there was almost no crystal contacts in one dimension: these crystals should not have formed! So aging may be good until a certain day, and then it can turn really badly after that. Engin Edward A. Berry wrote: For a counterexample, from Iwata's group, Horsefield et al. Succinate: quinone oxidoreductase Acta. Cryst.(2003). D59, 600-602: It proved critical to freeze the crystals within 72 h of crystallization set-up. Crystals that were frozen after this time limit showed no diffraction. This alteration in properties was apparent by the change in colour from deep orange to pale yellow that was observed in crystals more than four weeks old (Fig. 1c). The deep orange colour of the crystals is attributed to the presence of haem b within the protein. The loss or breakdown of haem, demonstrated by the change of colour in the crystals, could lead to structural instability and consequently loss of diffraction. I've heard the Fe-S clusters of E. coli SDH are oxygen-sensitive in the isolated enzyme. The mitochondrial counterpart is more stable, we've collected data from crystals 1-2 years old, and seen conversion (new crystals grow while old crystals dissolve) to a new space group with better diffraction (that was a within few months after setup). Edward Snell wrote: /lurk_mode_off /dumb_question_on Dear All, I was recently trying to find references on how age may degrade a crystal, i.e. grow them and use them or preserve them as fresh as possible. I seem to remember seeing a couple of papers on this but my memory is fading and I have been unable to locate them. Can anyone jog my memory or tell me if I'm imagining things? I've found plenty on the protein prep etc. but nothing on the crystal. Thanks, Eddie. Edward Snell Ph.D. Assistant Prof. Department of Structural Biology, SUNY Buffalo, Hauptman-Woodward Medical Research Institute 700 Ellicott Street, Buffalo, NY 14203-1102 Phone: (716) 898 8631 Fax: (716) 898 8660 Email: esn...@hwi.buffalo.edu Telepathy: 42.2 GHz Heisenberg was probably here!Crystallization, how quaint! /dumb_question_off /lurk_mode_on
Re: [ccp4bb] Off topic: Crystal degredation with age
Some things improve with age. Here is one of my favorite stories: http://tinyurl.com/oldtrna The crystal structure of yeast phenylalanine tRNA at 2.0 Å resolution: cleavage by Mg2+ in 15-year old crystals Luca Jovine, Snezana Djordjevica and Daniela Rhodes We have re-determined the crystal structure of yeast tRNAPhe to 2.0 Å resolution using 15 year old crystals. The accuracy of the new structure, due both to higher resolution data and formerly unavailable refinement methods, consolidates the previous structural information, but also reveals novel details. In particular, the water structure around the tightly bound Mg2+ is now clearly resolved, and hence provides more accurate information on the geometry of the magnesium- binding sites and the role of water molecules in coordinating the metal ions to the tRNA. We have assigned a total of ten magnesium ions and identified a partly conserved geometry for high-affinity Mg2+ binding. In the electron density map there is also clear density for a spermine molecule binding in the major groove of the TΨC arm and also contacting a symmetry-related tRNA molecule. Interestingly, we have also found that two specific regions of the tRNA in the crystals are partially cleaved. The sites of hydrolysis are within the D and anticodon loops in the vicinity of Mg2+. On Feb 5, 2009, at 11:11 AM, Edward Snell wrote: /lurk_mode_off /dumb_question_on Dear All, I was recently trying to find references on how age may degrade a crystal, i.e. grow them and use them or preserve them as fresh as possible. I seem to remember seeing a couple of papers on this but my memory is fading and I have been unable to locate them. Can anyone jog my memory or tell me if I'm imagining things? I've found plenty on the protein prep etc. but nothing on the crystal. Thanks, Eddie. Edward Snell Ph.D. Assistant Prof. Department of Structural Biology, SUNY Buffalo, Hauptman-Woodward Medical Research Institute 700 Ellicott Street, Buffalo, NY 14203-1102 Phone: (716) 898 8631 Fax: (716) 898 8660 Email: esn...@hwi.buffalo.edu Telepathy: 42.2 GHz Heisenberg was probably here!Crystallization, how quaint! /dumb_question_off /lurk_mode_on
Re: [ccp4bb] Off topic: Crystal degredation with age
Here's another very similar case: http://www.ncbi.nlm.nih.gov/pubmed/12270703 On Thu, Feb 5, 2009 at 11:48 AM, William G. Scott wgsc...@chemistry.ucsc.edu wrote: Some things improve with age. Here is one of my favorite stories: http://tinyurl.com/oldtrna The crystal structure of yeast phenylalanine tRNA at 2.0 Å resolution: cleavage by Mg2+ in 15-year old crystals Luca Jovine, Snezana Djordjevica and Daniela Rhodes We have re-determined the crystal structure of yeast tRNAPhe to 2.0 Å resolution using 15 year old crystals. The accuracy of the new structure, due both to higher resolution data and formerly unavailable refinement methods, consolidates the previous structural information, but also reveals novel details. In particular, the water structure around the tightly bound Mg2+ is now clearly resolved, and hence provides more accurate information on the geometry of the magnesium-binding sites and the role of water molecules in coordinating the metal ions to the tRNA. We have assigned a total of ten magnesium ions and identified a partly conserved geometry for high-affinity Mg2+ binding. In the electron density map there is also clear density for a spermine molecule binding in the major groove of the TΨC arm and also contacting a symmetry-related tRNA molecule. Interestingly, we have also found that two specific regions of the tRNA in the crystals are partially cleaved. The sites of hydrolysis are within the D and anticodon loops in the vicinity of Mg2+. On Feb 5, 2009, at 11:11 AM, Edward Snell wrote: /lurk_mode_off /dumb_question_on Dear All, I was recently trying to find references on how age may degrade a crystal, i.e. grow them and use them or preserve them as fresh as possible. I seem to remember seeing a couple of papers on this but my memory is fading and I have been unable to locate them. Can anyone jog my memory or tell me if I'm imagining things? I've found plenty on the protein prep etc. but nothing on the crystal. Thanks, Eddie. Edward Snell Ph.D. Assistant Prof. Department of Structural Biology, SUNY Buffalo, Hauptman-Woodward Medical Research Institute 700 Ellicott Street, Buffalo, NY 14203-1102 Phone: (716) 898 8631 Fax: (716) 898 8660 Email: esn...@hwi.buffalo.edu Telepathy: 42.2 GHz Heisenberg was probably here!Crystallization, how quaint! /dumb_question_off /lurk_mode_on
Re: [ccp4bb] Off topic: Crystal degredation with age
I had a structure that was done with crystals that were about a year old. Initial crystals appeared in a less then a day and diffracted very poorly. In trying to make room for more trays I reexamined the old trays before throwing them out and low and behold nice well diffracting crystals. Of course these are probably more the exception then the rule. Len Leonard Thomas Ph. D. Macromolecular Crystallography Laboratory Manager University of Oklahoma Department of Chemistry and Biochemistry 620 Parrington Oval Norman, OK 73032 lmtho...@ou.edu Office: 405-325-1126 Lab: 405-325-7571 On Feb 5, 2009, at 2:05 PM, Nathaniel Echols wrote: Here's another very similar case: http://www.ncbi.nlm.nih.gov/pubmed/12270703 On Thu, Feb 5, 2009 at 11:48 AM, William G. Scott wgsc...@chemistry.ucsc.edu wrote: Some things improve with age. Here is one of my favorite stories: http://tinyurl.com/oldtrna The crystal structure of yeast phenylalanine tRNA at 2.0 Å resolution: cleavage by Mg2+ in 15-year old crystals Luca Jovine, Snezana Djordjevica and Daniela Rhodes We have re-determined the crystal structure of yeast tRNAPhe to 2.0 Å resolution using 15 year old crystals. The accuracy of the new structure, due both to higher resolution data and formerly unavailable refinement methods, consolidates the previous structural information, but also reveals novel details. In particular, the water structure around the tightly bound Mg2+ is now clearly resolved, and hence provides more accurate information on the geometry of the magnesium-binding sites and the role of water molecules in coordinating the metal ions to the tRNA. We have assigned a total of ten magnesium ions and identified a partly conserved geometry for high-affinity Mg2+ binding. In the electron density map there is also clear density for a spermine molecule binding in the major groove of the TΨC arm and also contacting a symmetry-related tRNA molecule. Interestingly, we have also found that two specific regions of the tRNA in the crystals are partially cleaved. The sites of hydrolysis are within the D and anticodon loops in the vicinity of Mg2+. On Feb 5, 2009, at 11:11 AM, Edward Snell wrote: /lurk_mode_off /dumb_question_on Dear All, I was recently trying to find references on how age may degrade a crystal, i.e. grow them and use them or preserve them as fresh as possible. I seem to remember seeing a couple of papers on this but my memory is fading and I have been unable to locate them. Can anyone jog my memory or tell me if I'm imagining things? I've found plenty on the protein prep etc. but nothing on the crystal. Thanks, Eddie. Edward Snell Ph.D. Assistant Prof. Department of Structural Biology, SUNY Buffalo, Hauptman-Woodward Medical Research Institute 700 Ellicott Street, Buffalo, NY 14203-1102 Phone: (716) 898 8631 Fax: (716) 898 8660 Email: esn...@hwi.buffalo.edu Telepathy: 42.2 GHz Heisenberg was probably here!Crystallization, how quaint! /dumb_question_off /lurk_mode_on
