Re: [ccp4bb] Aggregated protein for crystallization
Not sure if it will be helpful... but my protein is not the most stable protein, in fact, it does aggregate over time (most likely due to its 'sticky' nature). However, I still get crystals. The problem is the crystals are among the gunks and precipitates. Your case might be different since my protein does not elute out at void volume. Perhaps, try work faster? Sometimes protein aggregates over time rather than immediately. Allan Quoting Raji Edayathumangalam : Hi Folks, As crazy as it sounds, if you have crystallized and managed to solve the structure of a protein from aggregated protein, please could you share your experience. After many constructs, many many expression schemes and after the usual rigmarole of optimization that is also often discussed on ccp4bb (buffers, glycerol, salt concentrations, pH, detergent, additives etc.), I now have a decently expressing truncated construct for my protein (80 kDa) that is pure but aggregated (elutes in the void volume from a Superdex200 column). I am tempted to make a boatload of aggregated protein and set up some crystal trays (after perhaps testing by CD). So I'd like to hear from folks who have been successful in solving structures from aggregates when many many known and tested optimization methods still leave one with aggregated protein. Thanks. Raji -- Raji Edayathumangalam Instructor in Neurology, Harvard Medical School Research Associate, Brigham and Women's Hospital Visiting Research Scholar, Brandeis University -- Allan Pang PhD Student G35 Joseph Priestley Building Queen Mary University of London London E1 4NS Phone number: 02078828480 Twitter: @xerophytes
Re: [ccp4bb] Aggregated protein for crystallization
Dear Raji Running a blue-native gel with lanes in the presence and absence of a reducing agent could prove quite informative. DLS could also return a quick result on the particle distribution in your sample. In that case I would measure samples as fractionated from the superdex200 and compare the measurements after centrifuging the same samples at 100k x g for one hour. Best regards Savvas On 22 Feb 2012, at 00:21, Raji Edayathumangalam wrote: > Hi Folks, > > As crazy as it sounds, if you have crystallized and managed to solve the > structure of a protein from aggregated protein, please could you share your > experience. > > After many constructs, many many expression schemes and after the usual > rigmarole of optimization that is also often discussed on ccp4bb (buffers, > glycerol, salt concentrations, pH, detergent, additives etc.), I now have a > decently expressing truncated construct for my protein (80 kDa) that is pure > but aggregated (elutes in the void volume from a Superdex200 column). I am > tempted to make a boatload of aggregated protein and set up some crystal > trays (after perhaps testing by CD). So I'd like to hear from folks who have > been successful in solving structures from aggregates when many many known > and tested optimization methods still leave one with aggregated protein. > > Thanks. > Raji > > -- > Raji Edayathumangalam > Instructor in Neurology, Harvard Medical School > Research Associate, Brigham and Women's Hospital > Visiting Research Scholar, Brandeis University > >
Re: [ccp4bb] Putting Text into Movies
-BEGIN PGP SIGNED MESSAGE- Hash: SHA1 Hello Jacob, you may also like one of the many google hits with "mencoder add subtitles". Tim On 02/21/2012 04:59 PM, Jacob Keller wrote: > Dear Crystallographers, > > is there a good way to put text labels into movies from pymol or > otherwise? It would be helpful for conveying some ideas... > > Jacob > - -- - -- Dr Tim Gruene Institut fuer anorganische Chemie Tammannstr. 4 D-37077 Goettingen GPG Key ID = A46BEE1A -BEGIN PGP SIGNATURE- Version: GnuPG v1.4.10 (GNU/Linux) Comment: Using GnuPG with Mozilla - http://enigmail.mozdev.org/ iD8DBQFPRLWNUxlJ7aRr7hoRAnclAJ9Zxrm6+YFPE1G4mNVGzQZZoXCRyQCg52XQ ut6dp7nifTGJtflLte1f6Ks= =f1cY -END PGP SIGNATURE-
Re: [ccp4bb] Server or software for B factor analysis
Hi Dialing,
if you know some python you can use PyMOL.
# get C-alpha b-factors as list
from pymol import cmd, stored
stored.bfactors = []
cmd.iterate('name CA', 'stored.bfactors.append((b,resv))')
# min/max b-factors with residue number
print min(stored.bfactors)
print max(stored.bfactors)
# data for plotting
x = [resv for (b,resv) in stored.bfactors]
y = [bfor (b,resv) in stored.bfactors]
# plot to a pdf file with matplotlib
from matplotlib.pyplot import figure
from matplotlib.backends.backend_pdf import PdfPages
fig = figure()
sub = fig.add_subplot(111)
sub.plot(x, y)
pp = PdfPages('bfactors.pdf')
fig.savefig(pp, format='pdf')
pp.close()
Hope that helps.
Cheers,
Thomas
On 02/22/2012 05:04 AM, Dialing Pretty wrote:
Dear All,
Will you please tell me a server of software which can draw a curve for
the B factor of the atoms in a protein PDB file from the first residue
to the residue?Or a server or software by which we can easily order the
B factors of the atoms in the PDB file according to the B factor in
decrease or in increase? Or to get the residues with the highest B
factor and the lowest B factor?
Cheers,
Dialing
--
Thomas Holder
MPI for Developmental Biology
Spemannstr. 35
D-72076 Tübingen
[ccp4bb] scala and postref
Hi all, Does anyone have version(s) of scala and postref that "work together in tandem" and willing to share. Even older versions of these programs are fine with me. I can not seem to make the recent versions of the programs work together... even though there is a POSTREF option available in scala. Thanks for your consideration. Regards, Vijay Vijay S. Reddy, Ph.D. Associate Professor Department of Molecular Biology, TPC6 The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, CA 92037 E-mail: red...@scripps.edu WWW:http://www.scripps.edu/~reddyv VIPERdb:http://viperdb.scripps.edu Phone:(858) 784-8191 FAX:(858) 784-8896
Re: [ccp4bb] scala and postref
Hi Vijay, The obvious person to answer this is Phil Evans, but he is in New Zealand at the moment and may well not be reading Emails, so you might need to wait until he is back in the UK (1-2 weeks). I know that he felt the POSTREF option was not an important one to maintain, because post-refinement is carried out in MOSFLM, so I am not very surprised that it does not work. Cheers, Andrew On 22 Feb 2012, at 13:51, Vijay Reddy wrote: Hi all, Does anyone have version(s) of scala and postref that "work together in tandem" and willing to share. Even older versions of these programs are fine with me. I can not seem to make the recent versions of the programs work together... even though there is a POSTREF option available in scala. Thanks for your consideration. Regards, Vijay Vijay S. Reddy, Ph.D. Associate Professor Department of Molecular Biology, TPC6 The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, CA 92037 E-mail: red...@scripps.edu WWW:http://www.scripps.edu/~reddyv VIPERdb:http://viperdb.scripps.edu Phone:(858) 784-8191 FAX:(858) 784-8896
Re: [ccp4bb] residuewise rmsd for multiple chain superposition
Dear Sreetama, ProFit (http://www.bioinf.org.uk/software/profit/) does the RMS-by-residue calculation for multiple chain superposition. Avinash Punekar
Re: [ccp4bb] HKL2000 indexing problem
If you have tried all of the other things suggested by others (especially beam-center and direction of rotation), you can try the keyword 'weak level' in indexing box (see page 29 on HKL manual http://hkl-xray.com/sites/default/files/manual_online.pdf or http://www.hkl-xray.com/denzo-keywords-alphabetical-order). This has solved that very problem for me more than once. You will need to play with the value for your data try weak level 2, 5, 20, etc... I assume you don't have the darkest spots in the world, right? Joseph M. Watts, Ph.D. Syngenta Biotechnology, Inc. 3054 E. Cornwallis Road Durham, North Carolina. 27709 USA
[ccp4bb] CCPBioSim workshop: How to set up a Protein Simulation
One of our sister CCPs, CCPBioSim, is running a one-day workshop on "How to set up a Protein Simulation" on April 20th in Cambridge. It is aimed at people new to biological molecular dynamics simulations, and may be suitable for any crystallographers who want to have a go. It will use the (free) MD package Gromacs. Further details and registration are at: https://eventbooking.stfc.ac.uk/news-events/how-to-set-up-a-protein-simulation Cheers Martyn -- *** * * * Dr. Martyn Winn * * * * STFC Daresbury Laboratory, Daresbury, Warrington, WA4 4AD, U.K. * * Tel: +44 1925 603455E-mail: martyn.w...@stfc.ac.uk* * Fax: +44 1925 603634Skype name: martyn.winn * * URL: http://www.ccp4.ac.uk/martyn/ * ***
Re: [ccp4bb] Aggregated protein for crystallization
some more thoughts, Do a cryo-EM imaging, it will be ideal than DLS. if the particle sizes are uniform i would think your protein in that state might be useful. cheers Padayatti On Tue, Feb 21, 2012 at 6:21 PM, Raji Edayathumangalam wrote: > Hi Folks, > > As crazy as it sounds, if you have crystallized and managed to solve the > structure of a protein from aggregated protein, please could you share your > experience. > > After many constructs, many many expression schemes and after the usual > rigmarole of optimization that is also often discussed on ccp4bb (buffers, > glycerol, salt concentrations, pH, detergent, additives etc.), I now have a > decently expressing truncated construct for my protein (80 kDa) that is pure > but aggregated (elutes in the void volume from a Superdex200 column). I am > tempted to make a boatload of aggregated protein and set up some crystal > trays (after perhaps testing by CD). So I'd like to hear from folks who have > been successful in solving structures from aggregates when many many known > and tested optimization methods still leave one with aggregated protein. > > Thanks. > Raji > > -- > Raji Edayathumangalam > Instructor in Neurology, Harvard Medical School > Research Associate, Brigham and Women's Hospital > Visiting Research Scholar, Brandeis University > > -- Pius S Padayatti,PhD, Phone: 216-658-4528
[ccp4bb] OPENING: Genentech, Research Associate/Senior Research Associate in Purification and Crystallization
Dear All, We have an open position at Genentech in the Structural Biology group for an Associate/Senior Associate in our protein purification and crystallization laboratory (Job #: 380684). Prior experience in purification and/or crystallization is required. Preference will be given to candidates with a track record of success in challenging systems such as membrane proteins, multi-domain proteins, or protein:protein (or protein:nucleic acid) complexes. Please encourage qualified candidates to apply to the Genentech website (details below). All the best, Jim * James Kiefer, Ph.D. * Structural Biology Genentech, Inc. 1 DNA Way, Mailstop 27 South San Francisco, CA 94080-4990 -- Here is the official posting: Job requisition number: 380684 RA/SRA – Crystallography, Macromolecular Who we are: At the Roche Group, about 80,000 people across 150 countries are pushing back the frontiers of healthcare. Working together, we've become one of the world's leading research-focused healthcare groups. A member of the Roche Group, Genentech has been at the forefront of the biotechnology industry for more than 30 years, using human genetic information to develop novel medicines for serious and life-threatening diseases. The headquarters for Roche pharmaceutical operations in the United States, Genentech has multiple therapies on the market for cancer and other serious illnesses. Please take this opportunity to learn about Genentech, where we believe that our employees are our most important asset and are dedicated to remaining a great place to work. The Position: E2/E3Position available for a Research Associate to join the Structural Biology Department involved in exploring structural and functional properties of proteins of therapeutic interest. The successful candidate will be responsible for the purification and characterization of proteins for structural analysis by X-ray crystallography. Day-to-day activities include designing and optimizing protein expression and purification strategies for challenging structural biology targets. Additional experience in the crystallization, cloning, expression and/or use of automation methods for protein purification and characterization would be considered strong assets. Who you are: A B.S. or M.S. in biochemistry, molecular biology or related discipline and 5 (five) or more years) of experience in areas relating to protein biochemistry and crystallization are required. Experience with crystallization automation and HPLC or FPLC purification systems is preferred. The successful candidate must be motivated, capable of working independently, and enjoy working in a collaborative setting. Strong analytical, communication and organizational skills are highly desirable. To apply visit www.gene.com /careers. Genentech is an Equal Opportunity Employer with a commitment to diversity. All individuals are encouraged to apply.
Re: [ccp4bb] effects of salt on twinned crystals
Forgot to mention, that this 2.5-3A diffracting crystal was the same one that I have been unable to index and suspect are twinned due to the presence of these other fused crystals in the same drop. Thanks for any input.
Re: [ccp4bb] effects of salt on twinned crystals
I read your description of the crystals and Figure 4 of the following paper came to mind. Post-crystallization treatment in lower PEG eventually allowed them to tease the bundles apart. 1. MacRae, I. J. & Doudna, J. A. An unusual case of pseudo-merohedral twinning in orthorhombic crystals of Dicer. urn:issn:0907-4449 63, 993–999 (2007). F On Feb 22, 2012, at 10:59 AM, Peter Hsu wrote: > Forgot to mention, that this 2.5-3A diffracting crystal was the same one that > I have been unable to index and suspect are twinned due to the presence of > these other fused crystals in the same drop. > > Thanks for any input. - Francis E. Reyes M.Sc. 215 UCB University of Colorado at Boulder
[ccp4bb] Postdoc/Software Developer - Harvard Medical School
*Scientific Web Service Developer Position* (Harvard Medical School, Boston MA) Description: The SBGrid Consortium (www.sbgrid.org) has an immediate opening for an enthusiastic scientific web service developer. The role involves developing, updating and maintaining high-throughput structural biology work-flows in an academic environment. Existing, published SBGrid web applications have processed hundreds of structural biology problems utilising hundreds of thousands of CPU hours through the Open Science Grid. The position includes developing new work-flows and applications in collaboration with academic researchers, as well as ensuring that the existing applications continue to serve SBGrid members. The role involves working in a team based environment. Skills: - Fluency in python - Familiarity with django, MySQL, HTML, CSS and JavaScript - Understanding of massively parallel computational grids - Experience with administrating web servers and related services - An interest in Bioinformatics - PhD in Computational, Structural Biology or Bioinformatics strongly preferred. To apply please e-mail your CV, a short cover letter, and three references to sli...@hkl.hms.harvard.edu. -- Piotrek Sliz Assistant Professor in Pediatrics Lecturer on BCMP Harvard Medical School Center for Molecular and Cellular Dynamics http://hkl.hms.harvard.edu Voice: (617) 299-1455
[ccp4bb] Crystal twinning
Hi all. I have a 'learning' question based on recent thread where crystal twinning is mentioned. With the current computational methods, what types of twinnig can and cannot be solved with computers? Thank you. Theresa
[ccp4bb] Lysozyme fusion protein-bacteria
Dear All, We are working with proteins with a lot of surface hydrophobicity and hence solubility is a big issue. We so far have tried expressing them as fusion proteins.The strategy has yielded soluble protein but most of the protein elutes in the void volume on a gel-filtration colum. This brings me to my question, "Are there vectors which have an inactive lysozyme fusion tag, which could be used for expression in E.coli?". Many thanks to all in anticipation, Vasan
Re: [ccp4bb] Crystal twinning
Dear Theresa, all types of twinning - merohedral, pseudo-merohedral and non-merohedral ones - can be solved given somewhat favorable conditions. In small molecule crystallography, it's quite common, especially for (pseudo) merohedral twins, it becomes increasingly popular for macromolecules as well. However, if your crystal is a split one with many domains, you should possibly look for a better crystal. You need to treat your data correctly: - In cases of (pseudo) merohedral twinning: Integrate in the lower - correct - space group. Not all methods of structure solution work well with twins, especially S-SAD - which is very sensitive towards data errors - and those methods that require several crystals. For refinement, the free R set has to contain also reflections related by twinning, and the random R value will go down, so don't be fooled. - In case of a non-merohedral twin, particular attention should be paid to overlapping reflections, they should be left out at first for indexing and treated special in integration. Scaling is also more difficult. While non-merohedral twins can be dealt with, it is not (yet?) common practice for macromolecules. Best wishes Andrea. Am 22.02.2012 19:56, schrieb Theresa H. Hsu: Hi all. I have a 'learning' question based on recent thread where crystal twinning is mentioned. With the current computational methods, what types of twinnig can and cannot be solved with computers? Thank you. Theresa ath...@shelx.uni-ac.gwdg.de http://shelx.uni-ac.gwdg.de/~athorn
[ccp4bb] Fwd: [ccp4bb] Aggregated protein for crystallization
This was meant to Raji, So here it goes to all. -- Forwarded message -- From: Zhang, Zhen Date: Wed, Feb 22, 2012 at 12:15 PM Subject: RE: [ccp4bb] Aggregated protein for crystallization To: Pius Padayatti Hi Pius, I have done exactly that. I have one protein eluted at void volume of S200 column. The MALS experiment estimates more than 100 copies of monomer in the aggregate. Against my belief, the protein crystallized and diffracted to 2.3A and the resolution was improved to 1.6A later. It turns out that the crystallization buffer breaks the aggregate to dimer and crystallized it from there. I used the lower concentration of the crystallization buffer to run the sizing column and the protein was eluted at reasonable elution time for dimer even though the profile looks ugly and the purified protein is not crystallizable any more. I guess the aggregate somehow protects the folding of the protein and releases the protein slowly to the protein crystal in the right buffer condition. So I think you should setup crystallization trials with your aggregate protein. We cannot search hundreds of conditions for running sizing column. So why not let crystallization trials find that for you? Good luck. Zhen Marasco Laboratory Cancer Immunology and AIDS Dana Farber Cancer institute http://www.marascolab.org/ -Original Message- From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of Pius Padayatti Sent: Wednesday, February 22, 2012 11:55 AM To: CCP4BB@JISCMAIL.AC.UK Subject: Re: [ccp4bb] Aggregated protein for crystallization some more thoughts, Do a cryo-EM imaging, it will be ideal than DLS. if the particle sizes are uniform i would think your protein in that state might be useful. cheers Padayatti On Tue, Feb 21, 2012 at 6:21 PM, Raji Edayathumangalam wrote: > Hi Folks, > > As crazy as it sounds, if you have crystallized and managed to solve the > structure of a protein from aggregated protein, please could you share your > experience. > > After many constructs, many many expression schemes and after the usual > rigmarole of optimization that is also often discussed on ccp4bb (buffers, > glycerol, salt concentrations, pH, detergent, additives etc.), I now have a > decently expressing truncated construct for my protein (80 kDa) that is pure > but aggregated (elutes in the void volume from a Superdex200 column). I am > tempted to make a boatload of aggregated protein and set up some crystal > trays (after perhaps testing by CD). So I'd like to hear from folks who have > been successful in solving structures from aggregates when many many known > and tested optimization methods still leave one with aggregated protein. > > Thanks. > Raji > > -- > Raji Edayathumangalam > Instructor in Neurology, Harvard Medical School > Research Associate, Brigham and Women's Hospital > Visiting Research Scholar, Brandeis University > > -- Pius S Padayatti,PhD, Phone: 216-658-4528 The information in this e-mail is intended only for the person to whom it is addressed. If you believe this e-mail was sent to you in error and the e-mail contains patient information, please contact the Partners Compliance HelpLine at http://www.partners.org/complianceline . If the e-mail was sent to you in error but does not contain patient information, please contact the sender and properly dispose of the e-mail. -- Pius S Padayatti,PhD, Phone: 216-658-4528
Re: [ccp4bb] Lysozyme fusion protein-bacteria
Although this is not the answer to your question, Bryan Matthews papers show In T4 lysozyme A mutation at A73 to AAA is shown to make extensive core-repacking in T4lysozyme that makes the enzyme inactive. Is the issue here is to use lysozyme for bacterial cell breakage while your fusion protein remain inactive? Bacterial cells can be otherwise disrupted easily So like i mentioned Poteete and co workers (Rennell eta al., 1991, systematic mutation of bacteriophage t4 lysozyme JMB 222:67-87) mutated 164 sites with 13 different amino acids 50 % of the sites all 13 susb. left the protein like wild type. So the A73 mutation mentioned above can be incorporated into your constructs? padayatti On Wed, Feb 22, 2012 at 2:25 PM, R.Srinivasan wrote: > Dear All, > > We are working with proteins with a lot of surface hydrophobicity and > hence solubility is a big issue. We so far have tried expressing them as > fusion proteins.The strategy has yielded soluble protein but most of the > protein elutes in the void volume on a gel-filtration colum. > > This brings me to my question, "Are there vectors which have an > inactive lysozyme fusion tag, which could be used for expression in > E.coli?". > > > Many thanks to all in anticipation, > Vasan > > -- Pius S Padayatti,PhD, Phone: 216-658-4528
Re: [ccp4bb] surface residue mutation
Dear all, Thanks for your response to my quarry about surface residue mutation. I tried the SERp Server, but did not get satisfactory clue. This software classified our protein as difficult to crystallize, however we have crystallized and solved the structure also. But our crystal form is not good for soaking experiments. Wondering is there any other software. Thanks Prem On Thu, Feb 16, 2012 at 3:56 PM, Joel Tyndall wrote: > Steve Kent has published a few more (at least 1 other) since HIV... > > 3ODV http://www.rcsb.org/pdb/explore/explore.do?structureId=3ODV > > Total chemical synthesis and X-ray structure of kaliotoxin by racemic > protein crystallography. > > Pentelute, B.L., Mandal, K., Gates, Z.P., Sawaya, M.R., Yeates, T.O., > Kent, S.B., > > Journal: (2010) Chem.Commun.(Camb.) 46: 8174-8176 > > _ > Joel Tyndall, PhD > > Senior Lecturer in Medicinal Chemistry > National School of Pharmacy > University of Otago > PO Box 56 Dunedin 9054 > New Zealand > Skype: jtyndall > http://www.researcherid.com/rid/C-2803-2008 > Pukeka Matua > Te Kura Taiwhanga Putaiao > Te Whare Wananga o Otago > Pouaka Poutapeta 56 Otepoti 9054 > Aotearoa > > Ph / Waea +64 3 4797293 > Fax / Waeawhakaahua +64 3 4797034 > > > -Original Message- > From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of > Jacob Keller > Sent: Thursday, 16 February 2012 7:36 a.m. > To: CCP4BB@JISCMAIL.AC.UK > Subject: Re: [ccp4bb] surface residue mutation > > Right on the money! > > JPK > > On Wed, Feb 15, 2012 at 12:28 PM, David Schuller > wrote: > > On 02/15/12 12:41, Jacob Keller wrote: > >>> > >>> Are there any all-D proteins out there, of known structure or > >>> otherwise? If so, do enantiomer-specific catalyses become inverted? > >>> > >>> JPK > >>> > > I looked a little harder, and at least one D-enantiomeric protein was > > an > > enzyme: > > > > Total chemical synthesis of a D-enzyme: the enatiomers of HIV-1 > > protease show demonstration of reciprocal chiral substrate specificty > > R.C. deL. Milton, S.C.F. Milton, S.B.H. Kent (1992) Science 256(5062) > > 1445-1448. > > > > I guess that answers your question. > > > > > > -- > > == > > = > > All Things Serve the Beam > > == > > = > > David J. Schuller > > modern man in a post-modern world > > MacCHESS, Cornell University > > schul...@cornell.edu > > > > -- > *** > Jacob Pearson Keller > Northwestern University > Medical Scientist Training Program > email: j-kell...@northwestern.edu > *** > -- Prem S. Kaushal Post Doctoral Fellow School of Medicine Department of Pharmacology Case Western Reserve University Cleveland OHIO 44106 2164328447
Re: [ccp4bb] effects of salt on twinned crystals
hi did u have other crystals grew in different conditions in your additive screen ? in my case, i did the additve screen for the condition i got twinned crystals i got crystals grew in few conditions one of them is salt but it still gave me twinned crystals then i tried NDSB-221 and the crystals turned out non-twin hope it helps by the way, even though its twinned crystal should be ok to solve the structure Mary From: CCP4 bulletin board [CCP4BB@JISCMAIL.AC.UK] on behalf of Francis E Reyes [francis.re...@colorado.edu] Sent: Thursday, February 23, 2012 4:06 AM To: CCP4BB@JISCMAIL.AC.UK Subject: Re: [ccp4bb] effects of salt on twinned crystals I read your description of the crystals and Figure 4 of the following paper came to mind. Post-crystallization treatment in lower PEG eventually allowed them to tease the bundles apart. 1. MacRae, I. J. & Doudna, J. A. An unusual case of pseudo-merohedral twinning in orthorhombic crystals of Dicer. urn:issn:0907-4449 63, 993–999 (2007). F On Feb 22, 2012, at 10:59 AM, Peter Hsu wrote: > Forgot to mention, that this 2.5-3A diffracting crystal was the same one that > I have been unable to index and suspect are twinned due to the presence of > these other fused crystals in the same drop. > > Thanks for any input. - Francis E. Reyes M.Sc. 215 UCB University of Colorado at Boulder
Re: [ccp4bb] Aggregated protein for crystallization
If you haven't done so already, I would screen buffer conditions (pH, salt concentration, glycerol, strongly reducing conditions, ligands, detergents) by DLS to see if you can reduce aggregation. You might get lucky by setting up crystallization plates, but chances are you won't get very useful information from them, especially if your aggregated protein is soluble. There are fluorescent dyes sensitive to aggregation state such as ANS (anilinonaphthalene-8-sulfonate) or Nile Red. I have not used them myself and would like to hear if others have found them useful for screening buffer conditions. Ho Ho Leung Ng University of Hawaii at Manoa Assistant Professor, Department of Chemistry h...@hawaii.edu
[ccp4bb] Position Available in Australia
Postdoctoral Research Fellow Department of Biochemistry and Molecular Biology, Faculty of Medicine, Nursing and Health Sciences http://research.med.monash.edu.au/rossjohn/ Prof. Jamie Rossjohn, NHMRC Australia Fellow, is seeking highly motivated and talented post-doctoral scientists to join his research group. The selected candidate will work in the area of Infection and Immunity. The applicants should hold a PhD in Biochemistry or Molecular Biology with an interest in Protein Chemistry. Candidates with a promising track record in the relevant areas and a proven publication record in international journals are encouraged to apply. Appointment will be made at a level appropriate to the successful applicant's qualifications, experience and in accordance with classification standards for each level. Salary range: Level A $55,332 - $75,095 Duration: The position is for 1 year Location:Clayton Campus Contact: Margaret Billsmargaret.bi...@monash.edu Jennifer Huynh jennifer.hu...@monash.edu Closing Date:30th March 2012 -- Margaret Bills Laboratory Manager Protein Crystallography Unit Dept. of Biochemistry, SOBS B76 G14 Monash University Wellington Rd Clayton Victoria Australia. 03 99029238 margaret.bi...@monash.edu
Re: [ccp4bb] Aggregated protein for crystallization
> You might get lucky by setting up crystallization plates, but chances are you won't get very useful information from them, especially if your aggregated protein is soluble. I seem to fail to understand how crystallization plates would give information in the not-special case of protein aggregates NOT being soluble? BR Ho Ho Leung Ng University of Hawaii at Manoa Assistant Professor, Department of Chemistry h...@hawaii.edu
Re: [ccp4bb] Server or software for B factor analysis
Why not just use PROCHECK program?
在 2012年2月22日 下午6:24，Thomas Holder 写道：
> Hi Dialing,
>
> if you know some python you can use PyMOL.
>
> # get C-alpha b-factors as list
> from pymol import cmd, stored
> stored.bfactors = []
> cmd.iterate('name CA', 'stored.bfactors.append((b,resv))')
>
> # min/max b-factors with residue number
> print min(stored.bfactors)
> print max(stored.bfactors)
>
> # data for plotting
> x = [resv for (b,resv) in stored.bfactors]
> y = [bfor (b,resv) in stored.bfactors]
>
> # plot to a pdf file with matplotlib
> from matplotlib.pyplot import figure
> from matplotlib.backends.backend_pdf import PdfPages
> fig = figure()
> sub = fig.add_subplot(111)
> sub.plot(x, y)
> pp = PdfPages('bfactors.pdf')
> fig.savefig(pp, format='pdf')
> pp.close()
>
> Hope that helps.
>
> Cheers,
> Thomas
>
>
> On 02/22/2012 05:04 AM, Dialing Pretty wrote:
>>
>> Dear All,
>>
>> Will you please tell me a server of software which can draw a curve for
>> the B factor of the atoms in a protein PDB file from the first residue
>> to the residue?Or a server or software by which we can easily order the
>> B factors of the atoms in the PDB file according to the B factor in
>> decrease or in increase? Or to get the residues with the highest B
>> factor and the lowest B factor?
>>
>> Cheers,
>>
>> Dialing
>
>
> --
> Thomas Holder
> MPI for Developmental Biology
> Spemannstr. 35
> D-72076 Tübingen
--
Cheng Chen, Ph.D. Candidate
Laboratory of Structural Biology
Life Science Building,Tsinghua University
Beijing 100084
China
Tel:+86-10-62772291
Fax:+86-10-62773145
E-mail:che...@xtal.tsinghua.edu.cn
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邮编：100084
Re: [ccp4bb] R-mergd-F
Hi all, Thanks for providing the solution. Regards, ARKO On Wed, Feb 22, 2012 at 10:14 AM, arka chakraborty < arko.chakrabort...@gmail.com> wrote: > Hi all , > > I will like to know which program we can use to calculate R-mergd-F( not > Rmerge) between two data sets, or more generally R factor between two data > sets. > > Thanks in advance, > > Regards, > > ARKO > > -- > > *ARKA CHAKRABORTY* > *CAS in Crystallography and Biophysics* > *University of Madras* > *Chennai,India* > > -- *ARKA CHAKRABORTY* *CAS in Crystallography and Biophysics* *University of Madras* *Chennai,India*
