[ccp4bb] Diffraction image viewer with display of resolution circles
Dear CCP4 users, I'm looking for the diffraction image viewer, which will be able to display of resolution circles and export it to new image. I tried use idiffdisp, but after choose of the Show/clear resolution circles, there is no action. Images were collected using Rayonics MX-225 detector - maybe detector format is a problem? Best regards, Rafal |--| |Rafal Dolot, Ph.D.| | | |Polish Academy of Sciences| |Centre of Molecular and Macromolecular Studies| |Department of Bioorganic Chemistry| |Sienkiewicza 112 | |90-363 Lodz, Poland | |Phone: +48(42)6803215 | |Cell: +48 502897781 | |--|
Re: [ccp4bb] Diffraction image viewer with display of resolution circles
Hi Rafal, have you try ADXV ? I never try to use idiffdisplay, but i know that ADXV can display resolution circles. If your software is able to open the image, but can't show the circles. I think it's not really a problem with the image format, but with the information in the Header of your image. Normally you can find information about the resolution, and detector position etc... If you detector control software didn't writte a complet Header, or if your image viewer is not able to read the Header it could be the source of your problem. Hope to help you. Nicolas Le 23/05/13 10:16, Rafal Dolot a écrit : Dear CCP4 users, I'm looking for the diffraction image viewer, which will be able to display of resolution circles and export it to new image. I tried use idiffdisp, but after choose of the Show/clear resolution circles, there is no action. Images were collected using Rayonics MX-225 detector - maybe detector format is a problem? Best regards, Rafal |--| |Rafal Dolot, Ph.D.| | | |Polish Academy of Sciences| |Centre of Molecular and Macromolecular Studies| |Department of Bioorganic Chemistry| |Sienkiewicza 112 | |90-363 Lodz, Poland | |Phone: +48(42)6803215 | |Cell: +48 502897781 | |--| -- This message has been scanned for viruses and dangerous content by MailScanner, and is believed to be clean.
Re: [ccp4bb] Diffraction image viewer with display of resolution circles
iMosflm can certainly do this for you. You might have to screen grab to capture the resolution rings though. Tony. On 23 May 2013, at 09:27, Rafal Dolot rdo...@cbmm.lodz.pl wrote: Dear CCP4 users, I'm looking for the diffraction image viewer, which will be able to display of resolution circles and export it to new image. I tried use idiffdisp, but after choose of the Show/clear resolution circles, there is no action. Images were collected using Rayonics MX-225 detector - maybe detector format is a problem? Best regards, Rafal |--| |Rafal Dolot, Ph.D.| | | |Polish Academy of Sciences| |Centre of Molecular and Macromolecular Studies| |Department of Bioorganic Chemistry| |Sienkiewicza 112 | |90-363 Lodz, Poland | |Phone: +48(42)6803215 | |Cell: +48 502897781 | |--|
Re: [ccp4bb] Diffraction image viewer with display of resolution circles
Photoshop? A. Sent from my iPad On 23 May 2013, at 10:16, Rafal Dolot rdo...@cbmm.lodz.pl wrote: Dear CCP4 users, I'm looking for the diffraction image viewer, which will be able to display of resolution circles and export it to new image. I tried use idiffdisp, but after choose of the Show/clear resolution circles, there is no action. Images were collected using Rayonics MX-225 detector - maybe detector format is a problem? Best regards, Rafal |--| |Rafal Dolot, Ph.D.| | | |Polish Academy of Sciences| |Centre of Molecular and Macromolecular Studies| |Department of Bioorganic Chemistry| |Sienkiewicza 112 | |90-363 Lodz, Poland | |Phone: +48(42)6803215 | |Cell: +48 502897781 | |--|
Re: [ccp4bb] Diffraction image viewer with display of resolution circles
Phenix has a diffraction viewer, too. You may want to try that. Ciao, Sebastiano -- Sebastiano Pasqualato Crystallography Unit Department of Experimental Oncology European Institute of Oncology via Adamello, 16 Milan, Italy On 23/mag/2013, at 10:16, Rafal Dolot rdo...@cbmm.lodz.pl wrote: Dear CCP4 users, I'm looking for the diffraction image viewer, which will be able to display of resolution circles and export it to new image. I tried use idiffdisp, but after choose of the Show/clear resolution circles, there is no action. Images were collected using Rayonics MX-225 detector - maybe detector format is a problem? Best regards, Rafal |--| |Rafal Dolot, Ph.D.| | | |Polish Academy of Sciences| |Centre of Molecular and Macromolecular Studies| |Department of Bioorganic Chemistry| |Sienkiewicza 112 | |90-363 Lodz, Poland | |Phone: +48(42)6803215 | |Cell: +48 502897781 | |--|
[ccp4bb] cryo condition
Dear all Can anybody tell me the appropriate cryo condition for the crystals obtained in 2M Ammonium sulphate and tris pH8.5..I tried to add 10% glycerol to it but still the ice ring is forming.. Thanx in advance -- Regards Faisal School of Life Sciences JNU
Re: [ccp4bb] cryo condition
80% saturated Li2SO4 On Thu, 23 May 2013 11:42:09 +0200, Faisal Tarique faisaltari...@gmail.com wrote: Dear all Can anybody tell me the appropriate cryo condition for the crystals obtained in 2M Ammonium sulphate and tris pH8.5..I tried to add 10% glycerol to it but still the ice ring is forming.. Thanx in advance -- Enrico A. Stura D.Phil. (Oxon) ,Tel: 33 (0)1 69 08 4302 Office Room 19, Bat.152, Tel: 33 (0)1 69 08 9449Lab http://www-dsv.cea.fr/ibitecs/simopro/ltmb/cristallogenese LTMB, SIMOPRO, IBiTec-S, CE Saclay, 91191 Gif-sur-Yvette, FRANCE http://scholar.google.com/citations?hl=enuser=Kvm06WIoPAsCpagesize=100sortby=pubdate http://www.chem.gla.ac.uk/protein/mirror/stura/index2.html e-mail: est...@cea.fr Fax: 33 (0)1 69 08 90 71
Re: [ccp4bb] cryo condition
Hi, Paraton oil worked nicely for me for these conditions. Boaz Boaz Shaanan, Ph.D. Dept. of Life Sciences Ben-Gurion University of the Negev Beer-Sheva 84105 Israel E-mail: bshaa...@bgu.ac.il Phone: 972-8-647-2220Skype: boaz.shaanan Fax: 972-8-647-2992 or 972-8-646-1710 From: CCP4 bulletin board [CCP4BB@JISCMAIL.AC.UK] on behalf of Faisal Tarique [faisaltari...@gmail.com] Sent: Thursday, May 23, 2013 12:42 PM To: CCP4BB@JISCMAIL.AC.UK Subject: [ccp4bb] cryo condition Dear all Can anybody tell me the appropriate cryo condition for the crystals obtained in 2M Ammonium sulphate and tris pH8.5..I tried to add 10% glycerol to it but still the ice ring is forming.. Thanx in advance -- Regards Faisal School of Life Sciences JNU
[ccp4bb] Post Doctoral Positions in membrane protein structure and function at the Karolinska Institutet, Sweden
Dear bulletin board, A two-year post-doctoral position is available from September 2013 at the Department of Medical Biochemistry and Biophysics (MBB), Karolinska Institutet, Stockholm. The project deals primarily with over-expression, purification, crystallization and structure determination of human membrane proteins involved in the generation of lipid mediators of inflammation, belonging to the MAPEG (membrane associated proteins in eicosanoid and glutathione metabolism) superfamily. The successful candidate will join a multi-disciplinary research team and the position represents an opportunity for a postdoctoral scientist to develop his/her expertise as well as broaden his/her experience. The broad interests of the lab are in membrane protein structure and function. We will combine structural techniques such as macromolecular crystallography with biophysical and biochemical methods such as enzyme kinetics to investigate the structural bases of the catalytic mechanisms. The lab has all necessary equipment and access to facilities needed for the project. We have periodic access to beamlines at ESRF (France), Diamond (UK) and Bessy (Germany). Qualifications: The candidates should hold a Ph. D. degree, must be motivated and a have strong background in biochemistry or structural biology. Experience with protein expression, purification and biochemical characterization is required. Prior experience in membrane protein biochemistry will be added advantages. The application should contain a single PDF document including: Letter of intention stating the scientific interests, technical skills, and previous experience of the candidate. C.V. including date and field of (expected) graduation and list of publications. The contact information of at least two references. Please send your electronic application to: agnes.rinaldo-matt...@ki.se before 1st of Aug 2013.
Re: [ccp4bb] cryo condition
30% glycerol or 25% glucose should be sufficient for 1-2 M ammonium sulfate. Roger Rowlett On May 23, 2013 5:42 AM, Faisal Tarique faisaltari...@gmail.com wrote: Dear all Can anybody tell me the appropriate cryo condition for the crystals obtained in 2M Ammonium sulphate and tris pH8.5..I tried to add 10% glycerol to it but still the ice ring is forming.. Thanx in advance -- Regards Faisal School of Life Sciences JNU
Re: [ccp4bb] cryo condition
I would try 2.5M or 3M sodium malonate pH 7 Janet Newman Principal Scientist / Director, Collaborative Crystallisation Centre CSIRO Material Science and Engineering 343 Royal Parade Parkville. VIC. 3052 Australia Tel +613 9662 7326 Email janet.new...@csiro.au From: CCP4 bulletin board [CCP4BB@JISCMAIL.AC.UK] on behalf of Roger Rowlett [rrowl...@colgate.edu] Sent: 23 May 2013 21:21 To: CCP4BB@JISCMAIL.AC.UK Subject: Re: [ccp4bb] cryo condition 30% glycerol or 25% glucose should be sufficient for 1-2 M ammonium sulfate. Roger Rowlett On May 23, 2013 5:42 AM, Faisal Tarique faisaltari...@gmail.commailto:faisaltari...@gmail.com wrote: Dear all Can anybody tell me the appropriate cryo condition for the crystals obtained in 2M Ammonium sulphate and tris pH8.5..I tried to add 10% glycerol to it but still the ice ring is forming.. Thanx in advance -- Regards Faisal School of Life Sciences JNU
Re: [ccp4bb] cryo condition
Hi Faisal, 3 - 3.5M Ammonium sulphate with your buffer should work too. Take a small loop. Jan -- Jan Abendroth Emerald BioStructures Seattle / Bainbridge Island WA, USA home: Jan.Abendroth_at_gmail.com work: JAbendroth_at_embios.com http://www.emeraldbiostructures.com On May 23, 2013, at 2:42 AM, Faisal Tarique faisaltari...@gmail.com wrote: Dear all Can anybody tell me the appropriate cryo condition for the crystals obtained in 2M Ammonium sulphate and tris pH8.5..I tried to add 10% glycerol to it but still the ice ring is forming.. Thanx in advance -- Regards Faisal School of Life Sciences JNU
Re: [ccp4bb] cryo condition
Hi Faisal, On 2013-05-23 11:42, Faisal Tarique wrote: Dear all Can anybody tell me the appropriate cryo condition for the crystals obtained in 2M Ammonium sulphate and tris pH8.5..I tried to add 10% glycerol to it but still the ice ring is forming.. Thanx in advance-- Regards Faisal School of Life Sciences JNU -- Matthew Bowler Synchrotron Science Group European Molecular Biology Laboratory BP 181, 6 rue Jules Horowitz 38042 Grenoble Cedex 9 France === Tel: +33 (0) 4.76.20.76.37 Fax: +33 (0) 4.76.88.29.04 http://www.embl.fr/ ===
Re: [ccp4bb] cryo condition
Hi Faisal, if your solvent channels are smaller than 40A in the largest dimension (most are) you can use a mesh loop to pick up the crystal and then wick away all of the mother liquor. You can then flash cool your crystal without having to transfer the crystal to another solution. Good luck, Matt On 2013-05-23 11:42, Faisal Tarique wrote: Dear all Can anybody tell me the appropriate cryo condition for the crystals obtained in 2M Ammonium sulphate and tris pH8.5..I tried to add 10% glycerol to it but still the ice ring is forming.. Thanx in advance-- Regards Faisal School of Life Sciences JNU -- Matthew Bowler Synchrotron Science Group European Molecular Biology Laboratory BP 181, 6 rue Jules Horowitz 38042 Grenoble Cedex 9 France === Tel: +33 (0) 4.76.20.76.37 Fax: +33 (0) 4.76.88.29.04 http://www.embl.fr/ ===
Re: [ccp4bb] Diffraction image viewer with display of resolution circles
The version of ADXV we have has some nice resolution ring features. There is a mode with the typical 5 rings of evenly spaced resolution. There is Anchor1 which draws a circle about the beam centre and through a selected point. There is Pick3 which constructs a circle from 3 selected points. Anchor1 is good for tweaking the beam centre from ice rings. -- === All Things Serve the Beam === David J. Schuller modern man in a post-modern world MacCHESS, Cornell University schul...@cornell.edu
Re: [ccp4bb] cryo condition
Just to follow up, the paper that was attached is based on the far more original work by Garman and Mitchell Elspeth Garman and Edward Mitchell. (1996) Glycerol concentrations required for cryoprotection of 50 typical protein crystallisation solutions. J.Appl. Cryst. 29, 584-587. And there is also a further paper Glycerol concentrations required for the successful vitrification of cocktail conditions in a high-throughput crystallization screen.Kempkes R, Stofko E, Lam K, Snell EH.(2003) Acta Cryst D 64, 287-301. There are a few other papers on high salt concentrations as a cryoprotectant including: Todd Holyoak, Timothy D. Fenn, Mark A. Wilson, Aaron G. Moulin, Dagmar Ringe and Gregory A. Petsko. (2003) Malonate: a versatile cryoprotectant and stabilizing solution for salt-grown macromolecular crystals. Acta D. 59, 2356-2358. Cheers, Eddie. Edward Snell Ph.D. Assistant Prof. Department of Structural Biology, SUNY Buffalo, Senior Scientist, Hauptman-Woodward Medical Research Institute 700 Ellicott Street, Buffalo, NY 14203-1102 Phone: (716) 898 8631 Fax: (716) 898 8660 Skype: eddie.snell Email: esn...@hwi.buffalo.edu Telepathy: 42.2 GHz Heisenberg was probably here! -Original Message- From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of Thibaut Crepin Sent: Thursday, May 23, 2013 6:24 AM To: CCP4BB@JISCMAIL.AC.UK Subject: Re: [ccp4bb] cryo condition Dear Faisal, this paper can be really useful. Regards Thibaut On 23/05/2013 11:42, Faisal Tarique wrote: Dear all Can anybody tell me the appropriate cryo condition for the crystals obtained in 2M Ammonium sulphate and tris pH8.5..I tried to add 10% glycerol to it but still the ice ring is forming.. Thanx in advance -- Regards Faisal School of Life Sciences JNU
Re: [ccp4bb] cryo condition
L-proline works well with ammonium sulfate: http://www.ncbi.nlm.nih.gov/pubmed/22868767 Sent from Jack's iPad On May 23, 2013, at 4:42 AM, Faisal Tarique faisaltari...@gmail.com wrote: Dear all Can anybody tell me the appropriate cryo condition for the crystals obtained in 2M Ammonium sulphate and tris pH8.5..I tried to add 10% glycerol to it but still the ice ring is forming.. Thanx in advance -- Regards Faisal School of Life Sciences JNU
Re: [ccp4bb] cryo condition
Matt, with this technique, how do you prevent crystal from drying up (other than doing it fast)? I know Thorne's group does this trick under oil. If you take no extra precautions, do you have an estimate of how often diffraction is destroyed by this? On the other hand, it's quite possible that what destroys resolution when crystals dry up is increase in concentration of non-volatile mother liquor components, which shouldn't be happening here to the same degree. Cheers, Ed. On Thu, 2013-05-23 at 14:38 +0200, Matthew BOWLER wrote: Hi Faisal, if your solvent channels are smaller than 40A in the largest dimension (most are) you can use a mesh loop to pick up the crystal and then wick away all of the mother liquor. You can then flash cool your crystal without having to transfer the crystal to another solution. Good luck, Matt -- After much deep and profound brain things inside my head, I have decided to thank you for bringing peace to our home. Julian, King of Lemurs
[ccp4bb] Organic solvents for ligand solubilisation
Dear CCP4BB followers, We are currently trying to obtain ligand-bound complexes for one of our proteins by soaking and/or co-crystallisation. We have had prior success for this protein, but using a different class of ligands. The new ligand (in DMSO) remains in solution (more or less) when mixed with the reservoir/cryoprotectant, and the diffraction pattern survives the soaking nicely. Annoyingly though, all we see are density peaks that match the size of DMSO and become more pronounced when increasing [DMSO] (to help solubilisation of the ligand). Soaking times varied between minutes to 16 hours. Kd was measured ( ~ 10 uM) in the solution state. We have tried pyridine (which keeps the ligand in solution, but kills diffraction in an instant), and DMF (which doesn't keep the ligand in solution when mixing with cryoprotectant). I am wondering whether the community has suggestions for alternative organic solvents that have been used to solubilise hydrophobic ligands, and are reasonably gentle to the protein crystal. Thank you. Klaus === Klaus Fütterer, Ph.D. Reader in Structural Biology Undergraduate Admissions School of Biosciences P: +44-(0)-121-414 5895 University of Birmingham F: +44-(0)-121-414 5925 Edgbaston E: k.futte...@bham.ac.uk Birmingham, B15 2TT, UK W: http://tinyurl.com/futterer-lab ===
Re: [ccp4bb] Organic solvents for ligand solubilisation
Hi Klaus, - small molecular weight PEG's e.g. 200 instead of DMSO, has the advantage of also helping to cryo protect - Methanol (only for dispensing the compound into wells) then allow to evaporate and simply add your cryo-protected crystals, the hope is that sufficient of your ligand goes into solution - different method but also efficient, use your protein as solubilizer . Figure out the lowest concentration of DMSO under which your ligand is soluble (maybe 0.5%) in solution at say 50 µM then take your diluted protein and mix it with the ligand, then concentrate to the required concentration needed for crystallization Jürgen On May 23, 2013, at 9:36 AM, Klaus Fütterer wrote: Dear CCP4BB followers, We are currently trying to obtain ligand-bound complexes for one of our proteins by soaking and/or co-crystallisation. We have had prior success for this protein, but using a different class of ligands. The new ligand (in DMSO) remains in solution (more or less) when mixed with the reservoir/cryoprotectant, and the diffraction pattern survives the soaking nicely. Annoyingly though, all we see are density peaks that match the size of DMSO and become more pronounced when increasing [DMSO] (to help solubilisation of the ligand). Soaking times varied between minutes to 16 hours. Kd was measured ( ~ 10 uM) in the solution state. We have tried pyridine (which keeps the ligand in solution, but kills diffraction in an instant), and DMF (which doesn't keep the ligand in solution when mixing with cryoprotectant). I am wondering whether the community has suggestions for alternative organic solvents that have been used to solubilise hydrophobic ligands, and are reasonably gentle to the protein crystal. Thank you. Klaus === Klaus Fütterer, Ph.D. Reader in Structural Biology Undergraduate Admissions School of Biosciences P: +44-(0)-121-414 5895 University of Birmingham F: +44-(0)-121-414 5925 Edgbaston E: k.futte...@bham.ac.ukmailto:k.futte...@bham.ac.uk Birmingham, B15 2TT, UK W: http://tinyurl.com/futterer-lab === .. Jürgen Bosch Johns Hopkins University Bloomberg School of Public Health Department of Biochemistry Molecular Biology Johns Hopkins Malaria Research Institute 615 North Wolfe Street, W8708 Baltimore, MD 21205 Office: +1-410-614-4742 Lab: +1-410-614-4894 Fax: +1-410-955-2926 http://lupo.jhsph.edu
Re: [ccp4bb] Organic solvents for ligand solubilisation
Dear Klaus, We've had a lot of luck with ethylene glycol and isopropanol, using them successfully in cases where DMSO affects diffraction. Best wishes Richard = Dr Richard Bayliss, Reader in Structural Biology Department of Biochemistry Henry Wellcome Building University of Leicester Lancaster Road, Leicester LE1 9HN Tel: 0116 2297100 Web: http://www2.le.ac.uk/departments/biochemistry/staff/richard-bayliss/research Elite Without Being Elitist Times Higher Awards Winner 2007, 2008, 2009, 2010, 2011 Follow us on Twitter http://twitter.com/uniofleicester On 23 May 2013, at 14:36, Klaus Fütterer wrote: Dear CCP4BB followers, We are currently trying to obtain ligand-bound complexes for one of our proteins by soaking and/or co-crystallisation. We have had prior success for this protein, but using a different class of ligands. The new ligand (in DMSO) remains in solution (more or less) when mixed with the reservoir/cryoprotectant, and the diffraction pattern survives the soaking nicely. Annoyingly though, all we see are density peaks that match the size of DMSO and become more pronounced when increasing [DMSO] (to help solubilisation of the ligand). Soaking times varied between minutes to 16 hours. Kd was measured ( ~ 10 uM) in the solution state. We have tried pyridine (which keeps the ligand in solution, but kills diffraction in an instant), and DMF (which doesn't keep the ligand in solution when mixing with cryoprotectant). I am wondering whether the community has suggestions for alternative organic solvents that have been used to solubilise hydrophobic ligands, and are reasonably gentle to the protein crystal. Thank you. Klaus === Klaus Fütterer, Ph.D. Reader in Structural Biology Undergraduate Admissions School of Biosciences P: +44-(0)-121-414 5895 University of Birmingham F: +44-(0)-121-414 5925 Edgbaston E: k.futte...@bham.ac.ukmailto:k.futte...@bham.ac.uk Birmingham, B15 2TT, UK W: http://tinyurl.com/futterer-lab ===
Re: [ccp4bb] cryo condition
Hi Ed, good question. I have found that you have a good 30 seconds to remove the surrounding liquid - so while you have to do it fast you have enough time that it doesn't need a robot and even a malcoordinate such as myself can do it. I'm afraid that I have no estimate for how often diffraction is lost this way more info here http://scripts.iucr.org/cgi-bin/paper?S0907444911031210 I started looking at this when doing dehydration experiments where it has long been observed that you can directly cool the crystals after dehydration - I have a feeling that 'over drying' a crystal leads to a complete loss of lattice order - I have no evidence for this but all systems that we have looked at that benefit from dehydration, or not, come to quite a sharp cutoff where suddenly there is no more diffraction, one can often recover diffraction by rehydration. This could of course be due to too high a concentration of mother liquor but quite often it occurs at relative humidity values Cheers, Matt. On 2013-05-23 15:32, Ed Pozharski wrote: Matt, with this technique, how do you prevent crystal from drying up (other than doing it fast)? I know Thorne's group does this trick under oil. If you take no extra precautions, do you have an estimate of how often diffraction is destroyed by this? On the other hand, it's quite possible that what destroys resolution when crystals dry up is increase in concentration of non-volatile mother liquor components, which shouldn't be happening here to the same degree. Cheers, Ed. On Thu, 2013-05-23 at 14:38 +0200, Matthew BOWLER wrote: Hi Faisal, if your solvent channels are smaller than 40A in the largest dimension (most are) you can use a mesh loop to pick up the crystal and then wick away all of the mother liquor. You can then flash cool your crystal without having to transfer the crystal to another solution. Good luck, Matt -- Matthew Bowler Synchrotron Science Group European Molecular Biology Laboratory BP 181, 6 rue Jules Horowitz 38042 Grenoble Cedex 9 France === Tel: +33 (0) 4.76.20.76.37 Fax: +33 (0) 4.76.88.29.04 http://www.embl.fr/ ===
Re: [ccp4bb] cryo condition
I keep sending mails by accident today - apologies for the spam. The last sentence of my should read: This could of course be due to too high a concentration of mother liquor but quite often it occurs at relative humidity values where the concentration of the mother liquor components will not have increased by very much. Cheers, Matt. On 2013-05-23 15:32, Ed Pozharski wrote: Matt, with this technique, how do you prevent crystal from drying up (other than doing it fast)? I know Thorne's group does this trick under oil. If you take no extra precautions, do you have an estimate of how often diffraction is destroyed by this? On the other hand, it's quite possible that what destroys resolution when crystals dry up is increase in concentration of non-volatile mother liquor components, which shouldn't be happening here to the same degree. Cheers, Ed. On Thu, 2013-05-23 at 14:38 +0200, Matthew BOWLER wrote: Hi Faisal, if your solvent channels are smaller than 40A in the largest dimension (most are) you can use a mesh loop to pick up the crystal and then wick away all of the mother liquor. You can then flash cool your crystal without having to transfer the crystal to another solution. Good luck, Matt -- Matthew Bowler Synchrotron Science Group European Molecular Biology Laboratory BP 181, 6 rue Jules Horowitz 38042 Grenoble Cedex 9 France === Tel: +33 (0) 4.76.20.76.37 Fax: +33 (0) 4.76.88.29.04 http://www.embl.fr/ ===
Re: [ccp4bb] Organic solvents for ligand solubilisation
Dear Klaus If your compound can be dried, you can add it to the drop as a powder from the tip of a acupuncture needle. At least this works for many of the insoluble heavy metal derivatives. We have also successfully resuspended the powder in the optimal buffer of choice and added it as insoluble power in solution. cheers Preben On 5/23/13 3:36 PM, Klaus Fütterer wrote: Dear CCP4BB followers, We are currently trying to obtain ligand-bound complexes for one of our proteins by soaking and/or co-crystallisation. We have had prior success for this protein, but using a different class of ligands. The new ligand (in DMSO) remains in solution (more or less) when mixed with the reservoir/cryoprotectant, and the diffraction pattern survives the soaking nicely. Annoyingly though, all we see are density peaks that match the size of DMSO and become more pronounced when increasing [DMSO] (to help solubilisation of the ligand). Soaking times varied between minutes to 16 hours. Kd was measured ( ~ 10 uM) in the solution state. We have tried pyridine (which keeps the ligand in solution, but kills diffraction in an instant), and DMF (which doesn't keep the ligand in solution when mixing with cryoprotectant). I am wondering whether the community has suggestions for alternative organic solvents that have been used to solubilise hydrophobic ligands, and are reasonably gentle to the protein crystal. Thank you. Klaus === Klaus Fütterer, Ph.D. Reader in Structural Biology Undergraduate Admissions School of Biosciences P: +44-(0)-121-414 5895 University of Birmingham F: +44-(0)-121-414 5925 Edgbaston E: k.futte...@bham.ac.uk mailto:k.futte...@bham.ac.uk Birmingham, B15 2TT, UK W: http://tinyurl.com/futterer-lab === -- J. Preben Morth, Ph.D Group Leader Membrane Transport Group Nordic EMBL Partnership Centre for Molecular Medicine Norway (NCMM) University of Oslo P.O.Box 1137 Blindern 0318 Oslo, Norway Email: j.p.mo...@ncmm.uio.no Tel: +47 2284 0794 http://www.jpmorth.dk
[ccp4bb] Postdoctoral position available at University of Wisconsin at Madison
You are invited to apply for a postdoctoral position in the structural biology lab of McArdle Cancer Institute of University of Wisconsin at Madison. We are interested in understanding the structural basis of protein phosphatase 2A (PP2A) regulation and its cross-talk to cellular and viral kinases in controlling signaling proteins involved in cell cycle, DNA damage response, and cell metabolism. One important aspect of the study is to elucidate mechanisms of protein chaperones in modulating the function of diverse PP2A complexes, kinases and nuclear receptors in response to cellular and environmental signals using structural biology techniques in combination of biochemical and biophysical methods and cell biology approaches. Initial results of this specific aspect of study can be found in our recent publications (Nat. Commun., 4:1699, 2013; Molecular Cell 41, 1–12, 2011). The lab is equipped with a state-of-the-art setup for x-ray crystallographic studies, including crystallography robot, and have convenient monthly access to powerful x-ray beamlines at APS. The researcher will work in a multidisciplinary setup, including cell biology, biophysics, proteomics, and protein biochemistry, in addition to protein crystallography. Candidates should hold a PhD in protein biochemistry and crystallography and have a good working knowledge of molecular biology techniques. The candidates should be highly motivated, goal-driven, and dedicated individuals who can think critically and work both independently and collaboratively with good verbal and written communication skills. It is also important that the candidates are creative in problem solving and have a strong desire to seek answers to questions related to the molecular basis of cellular processes. Experience in cell culture techniques and writing scientific research papers would be advantageous. Eligibility to training grant is appealing, but not essential. This is a full-time position with a starting salary of $39,264 – 44,340 per year depending on skills and experience. Review of applications will begin immediately and continue until the positions are filled. To apply: Please send a cover letter, CV and the names and contact information for three references to Dr. Yongna Xing, 1400 University Avenue, McArdle Laboratory for Cancer Research, University of Wisconsin-Madison-Madison, WI 53706. Email: yx...@oncology.wisc.edu.
[ccp4bb] Final Announcement: BioSAS Training Workshop, ACA 2013
ACA BioSAS Training Workshop WK.01 Biological Small Angle Solution Scattering Theory and Practice 2013 ACA meeting in Hawaii There is still Room. Reserve your space now! The early registration discount ends May 31*. Small angle solution scattering (SAS) has recently experienced a dramatic increase in popularity within the structural biology community. As molecular biological systems of interest become ever increasingly complex, SAS is providing valuable insight into structure and conformational behavior in solution. The availability of synchrotron radiation and neutron sources, commercial lab-source SAXS instrumentation, low-noise detectors, powerful computing hardware, and better algorithms has made the technique accessible to a much larger audience than ever before. This dual-track workshop brings together leading beamline scientists as well as experts in laboratory-based BioSAXS sources to provide a unique, practical HOW TO course in SAS data collection, processing, and interpretation. Track A: Getting Started in Biological Small Angle Solution Scattering: your practical HOW TO guide. Topics: Basics of scattering, Guinier analysis, monodispersity aggregation, Kratky plots, folding flexibility, molecular mass determination, P(r) function, sample preparation and characterization tips, shape reconstruction, using lab sources, neutron scattering basics, publishing your first data: what you should know. Speakers: Angela Chriswell (Rigaku), Javier Perez (Soleil), Srinivas Chakravarthy (APS), Jill Trewhella (U. Sydney), Richard Gillilan (MacCHESS, Cornell U.), Andreas Keilbach (Anton Paar), Juergen Graf (Incoatec), (Bruker speaker, to be confirmed), and others to be confirmed. Track B: Using Advanced Methods in Biological Small Angle Solution Scattering Topics: evaluating SAXS data, molecular mass, flexibility, modeling software: FOXS, MES, FoxDOCK, AllosMod-FOXS, strategy, inference-free data, similarity maps, contrast matched heavy atom labeling, ensemble analysis, NMR+SAXS, ab initio modeling at the residue level, building a lab source for BioSAXS. Speakers: Robert Rambo, Michal Hammel, Greg Hura (ALS), Kushol Gupta (Perelman School of Medicine, U. Penn) Andreas Keilbach (Anton Paar), Juergen Graf (Incoatec), (Bruker speaker, to be confirmed), Angela Chriswell (Rigaku), and others to be confirmed. Workshop format: serious attention will be given to hands-on practical exercises. After the Workshop: Several highly relevant BioSAXS-related sessions are scheduled for the beginning of the regular ACA meeting to immediately follow this workshop: TR.01 (Probing Biological Structure With HT SAS), 09.01 Dynamic Flexible Structures in Biomolecules, 09.03 Membrane Protein Scattering. Students of this workshop will be in an excellent position to take full advantage these cutting-edge presentations and are thus encouraged to attend and exchange ideas with the many SAS experts in attendance. For more information: www.amercrystalassn.org/wk.01 *To Register: www.amercrystalassn.org/2013-registration Note: students only attending the training Workshop do not need to pay the full ACA registration fee. To register for just the workshop, fill out and return the ACA registration form (PDF file), paying the workshop fee only.
Re: [ccp4bb] Call for Proposals: IMAGINE, a New Neutron Crystallography Diffractometer
This should be your first highlight to NSF - and he should see it today/tomorrow From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of Meilleur, Flora Sent: Thursday, May 23, 2013 3:54 PM To: CCP4BB@JISCMAIL.AC.UK Subject: [ccp4bb] Call for Proposals: IMAGINE, a New Neutron Crystallography Diffractometer IMAGINE, Neutron Crystallography Diffractometer High Flux Isotope Reactor (HFIR), Oak Ridge National Laboratory Call for proposals for experiments anticipated to run from August through December 2013 You are invited to apply for beam time on the neutron quasi-Laue diffractometer IMAGINE, at the High Flux Isotope Reactor. Proposal will be accepted via the web-based proposal system until NOON Wednesday, July 31, 2013. This call is for experiments anticipated to run from August through December 2013. Please see the attached flyer for additional information. For technical information about the capabilities of IMAGINE go to neutrons.ornl.gov/imagine/http://neutrons.ornl.gov/imagine/ or contact Flora Meilleur, meille...@ornl.gov, or Andrey Kovalevsky, kovalevsk...@ornl.gov. Flora Meilleur, Ph. D Assistant Professor, Molecular and Structural Biochemistry North Carolina State University IMAGINE lead scientist, Neutron Sciences Directorate Oak Ridge National Laboratory Phone: 865-242-5747
Re: [ccp4bb] Call for Proposals: IMAGINE, a New Neutron Crystallography Diffractometer
Is this Laue diffraction in the sense that the neutrons are a spectrum of energies, or does Laue mean something else here? Jacob On Thu, May 23, 2013 at 3:53 PM, Meilleur, Flora meille...@ornl.gov wrote: IMAGINE, Neutron Crystallography Diffractometer High Flux Isotope Reactor (HFIR), Oak Ridge National Laboratory *Call for proposals for experiments anticipated to run from August through December 2013* ** ** You are invited to apply for beam time on the neutron quasi-Laue diffractometer IMAGINE, at the High Flux Isotope Reactor. Proposal will be accepted via the web-based proposal system until NOON Wednesday, July 31, 2013. This call is for experiments anticipated to run from August through December 2013. ** ** Please see the attached flyer for additional information. For technical information about the capabilities of IMAGINE go to neutrons.ornl.gov/imagine/ or contact Flora Meilleur, meille...@ornl.gov, or Andrey Kovalevsky, kovalevsk...@ornl.gov. ** ** Flora Meilleur, Ph. D Assistant Professor, Molecular and Structural Biochemistry North Carolina State University IMAGINE lead scientist, Neutron Sciences Directorate Oak Ridge National Laboratory Phone: 865-242-5747 ** ** -- *** Jacob Pearson Keller, PhD Looger Lab/HHMI Janelia Farms Research Campus 19700 Helix Dr, Ashburn, VA 20147 email: kell...@janelia.hhmi.org ***
Re: [ccp4bb] Improve diffraction ...any ideas?
Dear Umri, I think the main problem is co-crystallization. What I would do is crystallize protein and antibody separately and then soak protein crystals into reservoir solution containing antibody or vice versa. And do try to get crystals from different conditions which may alter the space group and thereby improve diffraction quality, hopefully. All the best, Rajiv
