[ccp4bb] Differentiating bound Mn Ca.
Dear all. I have recently solved a structure in-house, 2.8A, CuKa. I have a metal ion bound very obvious hepta-valent co-ordination, which would suggest either Ca or Mn. Neither was present in the crystallisation setup, but there was some Mg around, which has contaminants of both Ca Mn. At 2.8A, I don't really think I can reliably discriminate between 2.15A 2.36A distances to coordinating atoms ( http://tanna.bch.ed.ac.uk/newtargs_06.html). The B factors for refined Ca are 18, and Mn 30. The B-factors of coordinating atoms vary from... 18 30 - so no help there. I have a nice clear 6sigma anomalous difference peak, but then, according to http://skuld.bmsc.washington.edu/scatter/ both Ca (f ~1.3) and Mn (f ~2.8) scatter anomalously at that wavelength. The obvious solution is go to a synchrotron and scan around the Mn edge and see what happens, however, whilst waiting for beam time, is there any way I could... oh I don't know, use the peak in my anomalous difference Fourier to figure out what anomalous signal would be required to generate a peak of that size - a sort of back-transform??? Is this do-able, and if so, how would one go about it? Cheers, Dave -- --- David Briggs, PhD. Father Crystallographer www.dbriggs.talktalk.net iChat AIM ID: DBassophile --- Anyone who is capable of getting themselves made President should on no account be allowed to do the job. - Douglas Adams
Re: [ccp4bb] Differentiating bound Mn Ca.
Hey David, You can do Mn2+ identification by its anomalous diffraction using the Cu Kalpha radiation. Mn is an anomalous scatterer at Cu Kalpha (1.5418 A), despite being distant from its absorption edge (somewhere around 1.96 A if I remember well). I did this for a double-manganese bound ConA, have a look into THE JOURNAL OF BIOLOGICAL CHEMISTRY Vol. 275, No. 26, Issue of June 30, pp. 19778–19787, 2000 (you may need to use different programs for making your anomalous difference Patterson and Fourier maps these days). Julie. Kay Diederichs wrote: David Briggs wrote: Dear all. I have recently solved a structure in-house, 2.8A, CuKa. I have a metal ion bound very obvious hepta-valent co-ordination, which would suggest either Ca or Mn. Neither was present in the crystallisation setup, but there was some Mg around, which has contaminants of both Ca Mn. At 2.8A, I don't really think I can reliably discriminate between 2.15A 2.36A distances to coordinating atoms (http://tanna.bch.ed.ac.uk/newtargs_06.html http://tanna.bch.ed.ac.uk/newtargs_06.html). The B factors for refined Ca are 18, and Mn 30. The B-factors of coordinating atoms vary from... 18 30 - so no help there. I have a nice clear 6sigma anomalous difference peak, but then, according to http://skuld.bmsc.washington.edu/scatter/ both Ca (f ~1.3) and Mn (f ~2.8) scatter anomalously at that wavelength. The obvious solution is go to a synchrotron and scan around the Mn edge and see what happens, however, whilst waiting for beam time, is there any way I could... oh I don't know, use the peak in my anomalous difference Fourier to figure out what anomalous signal would be required to generate a peak of that size - a sort of back-transform??? Is this do-able, and if so, how would one go about it? Cheers, Dave Dave, f = 1.3 versus 2.8 sounds like quite a difference ... what is the anom peak height of the sulfurs in your structure? The f of sulfur at Cu Ka wavelength is 0.55 . So I'd expect the ratio of peakheights of your unknown metal divided by the average peakheight of sulfurs (of roughly 18-30 A**2 B-factor) to give you an idea of what you have. Of course this is no proof ... Are there any other anom scatterers in your structure? best, Kay -- Julie Bouckaert, PhD[EMAIL PROTECTED] VIB Project leader VIB Department of Molecular and Cellular Interactions, Vrije Universiteit Brussel Tel. 32-2-629-1988 Fax 32-2-6291963 ULTR, Building E 4.18 Vrije Universiteit Brussel Pleinlaan 2 1050 Brussel Belgium
[ccp4bb] imosflm bug for Mar 225 (3x3) detectors
There is a bug in the latest imosflm (v0.5.2) which prevents the
correct direct beam coordinates being read from the image header for
Mar 3x3 tiled CCDs (225mm square, as installed on ID23-1 and BM14 at
ESRF for example). This is the ONLY detector affected by this bug.
When the program cannot read the direct beam position, it sets it to
the physical centre of the detector. This generates a warning message
(an exclamation mark will appear in the bottom right corner of the
GUI, click on the exclamation mark to list the warning). The direct
beam coordinates will be set to 112.43, 112.43 (or values very close
to this).
The physical centre of the image can be several mm from the true
direct beam position and this will almost certainly cause the indexing
to fail !
The fix is very simple, and requires a change to the file session.tcl.
The original code reads:
# Only set beam position if image is from reliable detector
set header_beam_x [$a_dom selectNodes normalize-space(//beam_x)]
set header_beam_y [$a_dom selectNodes normalize-space(//beam_y)]
if {[lsearch [list QUAD Q210 Q315 Q420 SAPPHIRE JUPITER SATURN
MERCURY MARCCD MARMOSAIC MOSAIC4] $detector_model] -1 ||
$detector_manufacturer == CBF } {
updateSetting beam_x $header_beam_x 1 1 Images
updateSetting beam_y $header_beam_y 1 1 Images
updateSetting backstop_x $header_beam_x 1 1 Images
updateSetting backstop_y $header_beam_y 1 1 Images
} else {
The required change is to replace MARMOSAIC with MOSAIC3.
This is the ONLY change required.
As this is part of the Tcl for the GUI, NO recompilation is required.
If you do not have access to the source code, a (messy) workaround is
to read an image into ipmosflm (which will read the direct beam
coordinates correctly), make a note of the values, then enter these
values into imosflm (at the head of the Images page).
Andrew Leslie and Harry Powell
Re: [ccp4bb] Differentiating bound Mn Ca.
Hi David, You can use Sheldrick's Calcium Bond Valence Sum to descriminate between metals (see Muller, P., Kopke, S., and Sheldrick, G. M. (2003) Acta Crystallogr., Sect. D: Biol. Crystallogr. 59, 32-37) even at low resolution. I have had good success with this method combined with estimation of anomalous contribution scaled against the S atoms as suggested by Kay. Have a look at gratuitous self plug Graham, S. C., Bond, C. S., Freeman, H. C., and Guss, J. M. (2005) Biochemistry 44, 13820-13836. /gratuitous self plug for plenty of examples. To do the anomalous difference calculations I just integrated a 2x2x2 A box around the metal atom and around the S atoms (in MAPMAN) then calculated the ratio of anomalous difference... Cheers, Stephen On 4/16/07, David Briggs [EMAIL PROTECTED] wrote: Dear all. I have recently solved a structure in-house, 2.8A, CuKa. I have a metal ion bound very obvious hepta-valent co-ordination, which would suggest either Ca or Mn. Neither was present in the crystallisation setup, but there was some Mg around, which has contaminants of both Ca Mn. At 2.8A, I don't really think I can reliably discriminate between 2.15A 2.36A distances to coordinating atoms (http://tanna.bch.ed.ac.uk/newtargs_06.html ). The B factors for refined Ca are 18, and Mn 30. The B-factors of coordinating atoms vary from... 18 30 - so no help there. I have a nice clear 6sigma anomalous difference peak, but then, according to http://skuld.bmsc.washington.edu/scatter/ both Ca (f ~1.3) and Mn (f ~2.8) scatter anomalously at that wavelength. The obvious solution is go to a synchrotron and scan around the Mn edge and see what happens, however, whilst waiting for beam time, is there any way I could... oh I don't know, use the peak in my anomalous difference Fourier to figure out what anomalous signal would be required to generate a peak of that size - a sort of back-transform??? Is this do-able, and if so, how would one go about it? Cheers, Dave -- --- David Briggs, PhD. Father Crystallographer www.dbriggs.talktalk.net iChat AIM ID: DBassophile --- Anyone who is capable of getting themselves made President should on no account be allowed to do the job. - Douglas Adams -- Dr Stephen Graham Nuffield Medical Fellow Division of Structural Biology Wellcome Trust Centre for Human Genetics Roosevelt Drive Oxford OX3 7BN United Kingdom Phone: +44 1865 287 549
[ccp4bb] Team Leader in Structure-Based Drug Design
Team Leader in Structure-Based Drug Design Institute of Cancer Research, London The Institute of Cancer Research is seeking to appoint a Team Leader, who will spearhead the establishment and development of high-throughput approaches to structure-based drug design within The Institute of Cancer Research, and build a thriving research team developing and applying structure-based drug design techniques to therapeutic targets of relevance to the treatment of cancer. This new post will be a joint appointment between the Section of Structural Biology and the Cancer Research UK Centre for Cancer Therapeutics to establish in-house expertise in structure-based design. The successful applicant will have practical expertise in X-ray crystallography with a proven track record in structure determination and analysis of protein-ligand complexes, and a strong research interest in developing and applying high-throughput experimental and virtual ligand screening techniques. The Team Leader will be expected to interact closely with biologists and medicinal chemists throughout The Institute, and contribute to the development and management of emerging drug programmes. Further information is available on http://www.icr.ac.uk/jobs/(ref HAD135). For informal enquiries please contact either David Barford ([EMAIL PROTECTED]) or Laurence Pearl ([EMAIL PROTECTED]). ___ David Barford Institute of Cancer Research 237 Fulham Road London, SW3 6JB United Kingdom Tel. +44 (0)20 7153 5420 FAX +44 (0)20 7153 5457 E. mail: [EMAIL PROTECTED] http://www.icr.ac.uk/research/research_sections/structural_biology/teams/barford_team/index.shtml The Institute of Cancer Research: Royal Cancer Hospital, a charitable Company Limited by Guarantee, Registered in England under Company No. 534147 with its Registered Office at 123 Old Brompton Road, London SW7 3RP. This e-mail message is confidential and for use by the addressee only. If the message is received by anyone other than the addressee, please return the message to the sender by replying to it and then delete the message from your computer and network.
Re: [ccp4bb] Differentiating bound Mn Ca.
In cases like this I use the S atoms to calibrate the peak height. Of course it isnt definitive a) it is near the noise level, and b) peak height is very dependent on B factor.. But the ratio might distinguish between an atom with an f of 1.3 or f=2.8 Eleanor David Briggs wrote: Dear all. I have recently solved a structure in-house, 2.8A, CuKa. I have a metal ion bound very obvious hepta-valent co-ordination, which would suggest either Ca or Mn. Neither was present in the crystallisation setup, but there was some Mg around, which has contaminants of both Ca Mn. At 2.8A, I don't really think I can reliably discriminate between 2.15A 2.36A distances to coordinating atoms (http://tanna.bch.ed.ac.uk/newtargs_06.html http://tanna.bch.ed.ac.uk/newtargs_06.html). The B factors for refined Ca are 18, and Mn 30. The B-factors of coordinating atoms vary from... 18 30 - so no help there. I have a nice clear 6sigma anomalous difference peak, but then, according to http://skuld.bmsc.washington.edu/scatter/ both Ca (f ~1.3) and Mn (f ~2.8) scatter anomalously at that wavelength. The obvious solution is go to a synchrotron and scan around the Mn edge and see what happens, however, whilst waiting for beam time, is there any way I could... oh I don't know, use the peak in my anomalous difference Fourier to figure out what anomalous signal would be required to generate a peak of that size - a sort of back-transform??? Is this do-able, and if so, how would one go about it? Cheers, Dave -- --- David Briggs, PhD. Father Crystallographer www.dbriggs.talktalk.net http://www.dbriggs.talktalk.net iChat AIM ID: DBassophile --- Anyone who is capable of getting themselves made President should on no account be allowed to do the job. - Douglas Adams
Re: [ccp4bb] Differentiating bound Mn Ca.
Although the peak height of S atoms can be used as an internal yardstick, one has to worry about differences in occupancy and possibly hetergeneous sites (i.e. Ca, Mn and Mg) which can confuse the interpretation of the results. On Mon, 16 Apr 2007, Eleanor Dodson wrote: In cases like this I use the S atoms to calibrate the peak height. Of course it isnt definitive a) it is near the noise level, and b) peak height is very dependent on B factor.. But the ratio might distinguish between an atom with an f of 1.3 or f=2.8 Eleanor David Briggs wrote: Dear all. I have recently solved a structure in-house, 2.8A, CuKa. I have a metal ion bound very obvious hepta-valent co-ordination, which would suggest either Ca or Mn. Neither was present in the crystallisation setup, but there was some Mg around, which has contaminants of both Ca Mn. At 2.8A, I don't really think I can reliably discriminate between 2.15A 2.36A distances to coordinating atoms (http://tanna.bch.ed.ac.uk/newtargs_06.html http://tanna.bch.ed.ac.uk/newtargs_06.html). The B factors for refined Ca are 18, and Mn 30. The B-factors of coordinating atoms vary from... 18 30 - so no help there. I have a nice clear 6sigma anomalous difference peak, but then, according to http://skuld.bmsc.washington.edu/scatter/ both Ca (f ~1.3) and Mn (f ~2.8) scatter anomalously at that wavelength. The obvious solution is go to a synchrotron and scan around the Mn edge and see what happens, however, whilst waiting for beam time, is there any way I could... oh I don't know, use the peak in my anomalous difference Fourier to figure out what anomalous signal would be required to generate a peak of that size - a sort of back-transform??? Is this do-able, and if so, how would one go about it? Cheers, Dave -- --- David Briggs, PhD. Father Crystallographer www.dbriggs.talktalk.net http://www.dbriggs.talktalk.net iChat AIM ID: DBassophile --- Anyone who is capable of getting themselves made President should on no account be allowed to do the job. - Douglas Adams
Re: [ccp4bb] Selenomethionine reduction v.s. disulfide bond
Hello Tiancen, The pundits often suggest keeping the selenium reduced (and indeed, it's not a bad idea) however if you're worried about disulphides - I would say that they take precedence over the selenium. If you work reasonably fast you should be able to have the best of both worlds - have the selenium safe and the disulphides oxidized. Not to mention that there always are the good old heavy atom derivatives to fall back on if the selenium data does not pan out :) Artem
Re: [ccp4bb] salt or protein?
Multiple overlapping salt lattices can sometimes look like protein diffraction, as long as you're looking in only two dimensions. However, if you can find the dominant rings, you should be able to discriminate since the c-spacing of salt would nearly always be pretty small. Consider powder patterns, and you should see what I mean. Generally speaking, if your crystals visually appear to be single, and give huge salt peaks - then they're probably salt. Exceptions - large, non-diffracting protein crystals that have small salt crystals stuck to them. Ultimately, you can use the 'stick a fork in it' method: stick a needle in your rod-like crystals and push. If you hear a crack, and see sharp clean edges on the break - it's salt. If you feel the crystal 'give' and see bending or pitting - it's probably protein. Good luck! Artem Hi All! I have been trying to screen for my protein crystals, from the crystals grown in 0.5 M Ammonium Sulphate, 1.0M Lithium Sulphate Monohydrate in 0.1 M TriSodium Citrate Buffer dihydrate Buffer at pH 5.6. Two different kinds of crystals observed: rod shaped and thin platy ones. Whenever I am trying to collect data from the rod shaped ones I am getting dominantly salt patterns but also spots at 15 A resolution bin repeatedly, where some of the spots very much look like from protein! what can be the reason for this? if anyone has come across similar situation please help. with regards, Sreeram Mahesh Research Student Prof S Ramakumar's lab PHYSICS department IISc Bangalore-560 012. ph:080-2293 2718. mobile: 9241145183. -- This message has been scanned for viruses and dangerous content by MailScanner, and is believed to be clean.
[ccp4bb] imgCIF workshop (new series) at BSR 2007
Third imgCIF workshop (new series) at BSR 2007 in Manchester and at Diamond: The Management of Synchrotron Image Data: Changes to the imgCIF dictionary and software, interaction with NeXus Sponsored by DOE under grant ER64212-1027708-0011962, NSF under grant DBI-0610407. You are cordially invited to a CBF/imgCIF workshop in two lunch sessions at BSR 2007 in Manchester and at Diamond. There will be a working lunch on 17 August as a breakout session to the BSR2007 meeting during a visit to Diamond Light Source. The meeting will be held at 12:30 in room 1.17 of Diamond House. The lunch is open to those not attending the main BSR2007 meeting, though places are limited. The major topics discussed at the Diamond session will be recent changes in the imgCIF dictionary and software and the interaction with NeXus. For those who cannot make it to the Diamond session or who want to get an introduction to the subject, there will also be a working lunch during the BSR meeting in Manchester. The time and room will be announced on the MEDSBIO.org web site and at the meeting. For further information and to register please contact Herbert J. Bernstein and Alun Ashton at [EMAIL PROTECTED] -- = Herbert J. Bernstein, Professor of Computer Science Dowling College, Kramer Science Center, KSC 121 Idle Hour Blvd, Oakdale, NY, 11769 +1-631-244-3035 [EMAIL PROTECTED] =
[ccp4bb] Openings for Phaser team
There are two positions available for people to join the team developing Phaser, funded by the NIH grant that supports the development of the Phenix package. (Of course, the same version of Phaser goes into CCP4 as well!) If you're interested, please look at http://www.jobs.ac.uk/jobfiles/ZE333.html. -- Randy J. Read Department of Haematology, University of Cambridge Cambridge Institute for Medical Research Tel: + 44 1223 336500 Wellcome Trust/MRC Building Fax: + 44 1223 336827 Hills RoadE-mail: [EMAIL PROTECTED] Cambridge CB2 0XY, U.K. www-structmed.cimr.cam.ac.uk
Re: [ccp4bb] salt or protein?
Hi Sreeram, assuming you have plenty of those crystals, why don't you loop a few pass them through a drop of your reservoir for washing and load them on a SDS gel ? Juergen Sreeram Mahesh wrote: Hi All! I have been trying to screen for my protein crystals, from the crystals grown in 0.5 M Ammonium Sulphate, 1.0M Lithium Sulphate Monohydrate in 0.1 M TriSodium Citrate Buffer dihydrate Buffer at pH 5.6. Two different kinds of crystals observed: rod shaped and thin platy ones. Whenever I am trying to collect data from the rod shaped ones I am getting dominantly salt patterns but also spots at 15 A resolution bin repeatedly, where some of the spots very much look like from protein! what can be the reason for this? if anyone has come across similar situation please help. with regards, Sreeram Mahesh Research Student Prof S Ramakumar's lab PHYSICS department IISc Bangalore-560 012. ph:080-2293 2718. mobile: 9241145183. -- Jürgen Bosch University of Washington Dept. of Biochemistry, K-426 1705 NE Pacific Street Seattle, WA 98195 Box 357742 Phone: +1-206-616-4510 FAX: +1-206-685-7002
[ccp4bb] FOM label in SigmaA
We need to run phase combination twice with SigmaA via CCP4i and encountered some confusion in FOM column label assignment. First, two sets of MAD phases are combined, we have output figure of merit, WCMB, from the input FOM_mad1 and FOM_mad2. Second time, we wanted to combine the Fc and PHIC obtained from a partial helical model with the combined MAD phases. This time the GUI would not let us choose WCMB as the input FOM label, rather only FOM_mad1 and FOM_mad2 were allowed. Is this a feature in SigmaA trying to avoid potential confusion between the same FOM label for input and output, or a bug in the GUI? Thanks for any help. Huiying - Huiying Li, Ph. D Department of Molecular Biology and Biochemistry Natural Sciences I, Rm 2443 University of California at Irvine Irvine, CA 92697, USA Tel: 949-824-4322(or -1953); Fax: 949-824-3280 email: [EMAIL PROTECTED] --
[ccp4bb] Crystallization Course: focus on membrane proteins 2007
Crystallization: focus on membrane proteins. http://www.nsls.bnl.gov/newsroom/events/workshops/2007/crys/ The Crystallization Course at Brookhaven National Laboratory this year will focus on membrane proteins. Sponsored by the IUCr the purpose of the course is to provide participants with hands-on experience of a variety of crystal growth methods. The course will address both conventional and non-conventional methods in membrane proteins. Optimization methods will include application of oils, novel nucleating agents, detergents, crystallization in lipid cubic phase, crystallization with gels , microfluidics, and more... Introductory lectures will precede the practical sessions. Time for discussions and informal meeting speakers and tutors is scheduled. Speakers and tutors: Marcia Amstrong (Qiagen) Neer Asherie (Yeshiva University) Bert van den Berg (Massachussets University) Troy Burke (GE Healthcare)Mark Elliot (Emerald BioSystems), TBC Ingo Grotjohann (Arizona State University), TBC Trevor Harvard (Precision Detectors) Rustem Ismagilov (Chicago University), TBC Liang Li ( Chicago University), TBC Patrick J. Loll (Drexel University) Marie-Claude Marchand (Qiagen), TBC Abel Moreno (Universidad Nacional Autonoma de Mexico) Gweneth Nneji (Birbeck University of London) Peter Nollert (Emerald BioSystems) Michael C. Wiener (University of Virginia), TBC Limited Travel assitance is provided by the International Union of Crystallography. Please check the Course site for further details or contact: Vivian Stojanoff Brookhaven National Laboratory National Synchrotron Light Source Bldg725D Upton NY 11973 USA Phone: +1 631 344 8375 (office) +1 631 344 5836 (X6A beam line) Fax:+1 631 344 3238 email: [EMAIL PROTECTED], http://protein.nsls.bnl.gov
[ccp4bb] phosphorylation of protein is troublesome or not for crystallization
Dear all, Below is a question my friend asked me, but I have never worked on phosphorylated proteins. Has anyone worked on crystallizing phosphorylated proteins and can you comment on it? Thanks Rongjin Guan -- I would like to ask how feasible it is to crystalize protein containing post-translational modification, such as phosphorylation. To my limited knowledge, I think the heterogenicity of modified and unmodified proteins causes the major obstacle in the crystal structure. Is there any specific method to crystalized modified proteins (like phosphorylated proteins ) these days?
