[ccp4bb] Post-Doctoral Position Available
POST-DOCTORAL POSITION DATE OF AVAILABILITY: starting as soon as possible, for 2 years. TITLE: structural biology of complexes involved in tumor reversion. POSITION: the position is opened in the department of Integrated Structural biology, IGBMC, located in Strasbourg-Illkirch (France) in the team of Jean Cavarelli (Structural biology of epigenetic targets). JOB DESCRIPTION: Our main scientific goal is to identify, characterize and analyze at the structural level, functional complexes involving selected protein targets to study the molecular switches and signals that induce biological activity. The aim of the proposed research project is to better understand the biology of a multi-functional protein which is strongly involved in the process of tumor reversion. The purpose of the structural study is to decipher at the molecular level the interactions between this protein and different biological partners. The structural work is part of a collaborative project involving three groups. JOB REQUIREMENTS: The applicant should hold a PhD in structural biology and must have a strong background in protein biochemistry (cloning, purification and biochemical/biophysical characterization). Experience in X-ray crystallography would be an advantage. SCIENTIFIC ENVIRONMENT: IGBMC is one of the leading European centers of biomedical research, devoted to the study of higher eukaryotic genomes and to the control of genetic expression as well as the analysis of the function of genes and proteins. The department of Integrated Structural Biology aims at understanding the mechanisms of cellular processes by correlating the structural and the functional properties of their macromolecular components. The post-doc will benefit from a technical environment including, the Structural Biology and Genomics Technology (SBGT) platform. The SBGT platform includes offers access to state of the art equipments for robotized cloning and expression tests, large scale prokaryotic and eukaryotic sample production and purification facilities, biophysical characterization, high-throughput crystallization screening, electron microscopy, X-ray and NMR facilities. TO APPLY: please send a CV and the names of two referees to Pr. Jean Cavarelli: c...@igbmc.fr
Re: [ccp4bb] Anyway to change the toolbar position in coot?
That's great. The 3D screen we got is only 23 inch. A full screen GUI like in the game will certainly be helpful. Nian Huang, Ph.D. Dept of Biochemistry UT Southwestern Medical Center Dallas, TX 75390 On Fri, Dec 31, 2010 at 5:00 PM, Nian Huang wrote: > Dear all, > The python version coot has two nicely made toolbars. I can change the > position or get rid of one of them by going to the preferences. In the > current wide screen dominant world, the other toolbar takes too much > precious horizontal space. Is there a way that I can make it vertical? > Thanks. > > Nian Huang, Ph.D. > Dept of Biochemistry > UT Southwestern Medical Center > Dallas, TX 75390 >
Re: [ccp4bb] SDS and IMAC
Especially if you're dealing with lysate, I suspect the best way to do it is with magnetic Ni beads that you lift up and out of the gunk, to help avoid false positives from aggregating stuff that SDS/urea/guan would all "elute". But why do you want X to remain on the column/beads? Removing Y but not X probably requires knowing more about the nature of their interaction than you do at this point? (e.g. will it take high salt, pH change, low (1M) urea, mild detergents etc) = Phoebe A. Rice Dept. of Biochemistry & Molecular Biology The University of Chicago phone 773 834 1723 http://bmb.bsd.uchicago.edu/Faculty_and_Research/01_Faculty/01_Faculty_Alphabetically.php?faculty_id=123 http://www.rsc.org/shop/books/2008/9780854042722.asp Original message >Date: Thu, 23 Dec 2010 09:11:31 -0600 >From: CCP4 bulletin board (on behalf of Jacob Keller >) >Subject: [ccp4bb] SDS and IMAC >To: CCP4BB@JISCMAIL.AC.UK > >Dear Crystallographers, > >I am interested in doing a type of pull-down experiment by >immobilizing protein X on IMAC resin, flowing a large volume of dilute >lysate containing protein Y over it, then adding some concentrated >agent (solid SDS perhaps) to some more of the same lysate, and running >that over the column to elute protein Y off protein X, without eluting >X off the column. I am afraid from past experience that SDS might >knock X off the column, presumably depending on the concentration. I >do not care about the folding state of Y--I will just be running a >PAGE gel anyway. Does anyone know either what is the minimal >concentration of SDS for robustly unfolding proteins/breaking up >interactions (and whether that concentration is safe for IMAC), or >what would be a good alternative agent to do the same? > >Thanks in advance for your help, > >Jacob Keller > >*** >Jacob Pearson Keller >Northwestern University >Medical Scientist Training Program >cel: 773.608.9185 >email: j-kell...@northwestern.edu >***
[ccp4bb] Only four weeks left to apply for RapiData 2011, our course on Data Collection and Structure Solving at the NSLS.
We are offering RapiData 2011, the thirteenth offering of our popular course: Rapid Data Collection and Structure Solving at the NSLS: A Practical Course in Macromolecular X-Ray Diffraction Measurement The course will be held 3-8 April 2011. Students could be at any level from advanced undergraduate to full professor. The course should accommodate 48 students total. All students are encouraged to bring their own specimens for data collection, and to bring old data for the data-reduction and structure-solving tutorials. Please read the Course Announcement at http://www.bnl.gov/RapiData/. You'll see that many experts in the field will be available for lectures and tutorials. You'll find the application materials on the Course Application tab at this site. For the ninth time we will hold a short lecture course on the fundamentals of crystallography for roughly five hours on Sunday 3 April. The body of the RapiData course really requires that students have a healthy knowledge of crystallography. For potential students who have some experience but are shaky about fundamentals, this course will help. There will be a small additional fee for the fundamentals course, to pay for Saturday night accomodations and food on Sunday morning and noon. Latin American Scientists: Several scholarships are available, from the International Union of Crystallography, to pay partial travel and subsistence costs for Latin-American students and junior faculty (under 40 yrs). Please apply for the course, and then contact R. Sweet (sw...@bnl.gov) if you are interested in applying for a scholarship. In accordance with the standards of the International Union of Crystallography, we observe the basic policy of non-discrimination, affirming the right and freedom of scientists to associate in international scientific activity without regard to such factors as citizenship, religion, creed, political stance, ethnic origin, race, colour, language, age, or gender, in accordance with the Statutes of the International Council for Science. At this course no barriers will exist beyond the application procedure that would prevent the participation of bona fide scientists. Please apply or send your students to our course, Bob Sweet, Sal Sclafani, and Alex Soares Course Announcement at http://www.bnl.gov/RapiData/ = Robert M. Sweet E-Dress: sw...@bnl.gov Group Leader, PXRR: Macromolecular ^ (that's L Crystallography Research Resource at NSLSnot 1) http://px.nsls.bnl.gov/ Biology Dept Brookhaven Nat'l Lab. Phones: Upton, NY 11973631 344 3401 (Office) U.S.A. 631 344 2741 (Facsimile) =
[ccp4bb] Postdoctoral position in Brisbane Australia
Sorry for posting this again, but the web address of the full ad (see below) has changed. The deadline is 10 January. Happy New Year! POSTDOCTORAL POSITION IN MACROMOLECULAR CRYSTALLOGRAPHY, UNIVERSITY OF QUEENSLAND, BRISBANE, AUSTRALIA Applications are invited for a post-doctoral position in macromolecular crystallography in the laboratory of Prof Bostjan Kobe at the School of Chemistry and Molecular Biosciences (SCMB) and the Institute for Molecular Bioscience (IMB), University of Queensland, Brisbane, Australia. The main area of study will involve characterization of three-dimensional structures of proteins and protein complexes involved in innate immunity pathways. Experience in molecular biology, protein purification, macromolecular crystallography and protein/protein interaction analysis is desirable. The salary will be according to qualifications and experience, starting at AUD$67,958. The position is available from January 2011. University of Queensland is one of Australia's top universities. SCMB combines the disciplines of chemistry, biochemistry & molecular biology, microbiology and parasitology into a single academic unit. The laboratory is located in a recently refurbished area of SCMB. SCMB and IMB are situated in the beautiful St. Lucia campus on the Brisbane River. State-of-the art equipment is available for all aspects of the work, including FR-E X-ray generator /R-axis IV++ and Saturn 944 CCD X-ray detectors, Mosquito crystallization robot and the Rock Imager imaging system. There is regular access to the Australian Synchrotron. Brisbane has been voted Australia's most liveable city and has a fantastic subtropical climate. It is only a short drive from Australia's best beaches and provides opportunities for all sorts of outdoors and cultural activities. For further information please see http://www.seek.com.au/Job/postdoctoral-research-fellow/in/brisbane/18804467 or contact Bostjan Kobe (telephone: +617-3365-2132, email b.k...@uq.edu.au). Please forward applications, including a cover letter, curriculum vitae and contact details for three referees, by by 10 January 2011, to Bostjan Kobe by email (b.k...@uq.edu.au) or post (Bostjan Kobe, SCMB, University of Queensland, St. Lucia, Queensland 4072, Australia). --- Bostjan Kobe ARC Federation Fellow Professor of Structural Biology School of Chemistry and Molecular Biosciences > and Institute for Molecular Bioscience (Division of Chemistry and Structural > Biology) and Centre for Infectious Disease Research Cooper Road University of Queensland Brisbane, Queensland 4072 Australia Phone: +61 7 3365 2132 Fax: +61 7 3365 4699 E-mail: b.k...@uq.edu.au URL: http://www.scmb.uq.edu.au/staff/bostjan-kobe Office: Building 76 Room 329 Notice: If you receive this e-mail by mistake, please notify me, and do not make any use of its contents. I do not waive any privilege, confidentiality or copyright associated with it. Unless stated otherwise, this e-mail represents only the views of the Sender and not the views of The University of Queensland.
Re: [ccp4bb] help with kinase purification
Hi Neeraj, A few points to add to Artem's excellent advice: If you would like to purify an untagged construct of your kinase, you could try an ATP affinity column. The caveats to this are 1.) the ATP resins can be expensive, and therefore it may be impractical to make a column with a high binding capacity, and 2.) Lots of proteins bind ATP, so your fractions might not be so clean (preceding your ATP column with an AS precipitation could help). Artem mentioned that often eukaryotic kinases are expressed in insect cells, and also touched on the fact that they can be phosphorylated by other kinases within your expression system. It may be worth a try to see if you can get your protein to express in a bacterial system, because most bacteria (notably laboratory strains of E.coli) lack S/T/Y kinases. Autophosphorylation, however, cannot really be avoided. Certain competent E.coli are pretty good for expressing eukaryotic proteins. You might try the BL21 CodonPlus Rosetta strain (available commercially - novagen I think, or maybe Stratagene) because it has a plasmid that expresses a number of tRNAs that correspond to RILP codons that are common in eukaryotes, but rare in E.coli. Good luck, Mike Thompson - Original Message - From: "Artem Evdokimov" To: CCP4BB@JISCMAIL.AC.UK Sent: Saturday, January 1, 2011 6:56:03 AM GMT -08:00 US/Canada Pacific Subject: Re: [ccp4bb] help with kinase purification Each protein is different, but there are indeed similarities within classes. It can be dangerous/unproductive to generalize -- you will find out experimentally whether you can assume things or not. Depending on the expression method and the design of your construct you may or may not have a tag. Historically, a lot of eukaryotic protein kinases tend to be expressed in insect cells with a GST or His-GST tag. Protein kinases tend to favorably express in insect cells but the GST thing is in my experience more of a historical trend than a real requirement - I've expressed quite a few kinases with a simple His-tag. So, we can perhaps assume your first purification is a tag of some sort, and that you can cleave the GST away (His-tag typically does not have to be cleaved for crystallization, though it can certainly help). Depending on the purity and homogeneity of the enzyme you may want to try an orthogonal second step - a dye affinity resin or an ion exchanger, followed by size exclusion if necessary. If your kinase is autophosphorylated or if it's phosphorylated by another kinase you may have a mixed population of phosphates which can often be resolved by careful ion exchange chromatography (monoQ typically) either alone or in combination with phosphatase treatment. Note that phosphatases may not remove all phosphates from a mature folded protein - if you have phsophorylation and you *must* have a completely non-phosphorylated protein you may have to try co-expression with a suitable phosphatase. If your first purification step is 'generic' (like an AS cut or a crude ion exchange) nothing much changes for the following steps except you will have less pure starting material and therefore may have to do more total steps (and perhaps add another technique into the mix like HIC etc.). Good luck, Artem On Sat, Jan 1, 2011 at 1:03 AM, Neeraj Kapoor < nkap...@mail.rockefeller.edu > wrote: Hi All, A very happy new year to everyone. I am beginning to purify a novel kinase and was wondering if there's a standard protocol / method that can be utilized to begin purification of a kinase. I am new to the field of kinases and would appreciate any help with respect to the precautions / pitfalls while dealing with kinase purification. Thanks and have a great year ahead! Neeraj -- Michael C. Thompson Graduate Student Biochemistry & Molecular Biology Division Department of Chemistry & Biochemistry University of California, Los Angeles mi...@chem.ucla.edu
[ccp4bb] PDB deposition
Dear all, Last year, when I deposited PDBs using ADIT, immediately I got an automatic confirmation email. And after 1 day, I got the responses from the PDBj Annotator. And within 3 days, all the edition and validation of my structures were finished with the help of member from the wwPDB. I greatly appreciate the enthusiasm of PDB Data Annotation Staff. However, one weeks ago, I also deposited several PDBs using ADIT, I only got the automatic confirmation email but I have not received any responses from the PDBj Annotator yet. I am just afraid something wrong with my deposition? Could anybody let me know if there were any changes in PDB deposition regulation? How long will be the processing time? Thank you a lot for your advices I look forward to hearing from you
Re: [ccp4bb] PDB deposition
Dear all, Thank to your replies, I am so released now! Thank you a lot for all responses. Best regards.
Re: [ccp4bb] PDB deposition
Dear all, I have just received response from the PDBj Annotator through CCP4BB My PDBs will be processed within two weeks As some of you mentioned, the reasons of delaying processing time is due to holiday season and the extremely large amount of depositions recently Again, thank you all of you for responses Especially, thank you two members of the PDBj Annotator who nicely responded me. Best regards
