Re: [ccp4bb] Crystals Disappearing Overnight
Thank you all for getting back! I will set up the drops using microbatch method. Regarding the temp, I set them up in lab and put them in incubator. The lab temp may be slightly higher. So are they not stable at lower temp? Or its the shock? So how can I take care of the temp issue? Thanks again! Maria On Saturday, May 3, 2014, Bob Cudney b...@hrmail.com wrote: Hello Maria, Check to see if there might have been a temperature change between the time the crystals were present and when the crystals disappeared. If your sample has temperature dependent solubility, in this relatively low ionic strength condition, a temperature change could mean the difference between the presence and absence of crystals. That being said, if the experiment is returned to the temperature that produced the crystals, the crystals should/might reappear. If your drop is made by mixing 1 part of protein with 1 part of reagent the initial drop concentration would be 10 mM Tris, 150 mM NaCl, 2.5% w/v PEG 8,000, 50 mM Sodium cacodylate. Is this a vapor diffusion experiment? If yes, then the reservoir would be 5% w/v PEG 8,000, 100 mM Sodium cacodylate. The ionic strength of your drop would initially be higher than the ionic strength in your reservoir. This means water vapor leaves the reservoir and vapor diffuses into the drop, lowering the protein and reagent concentration in your drop. This decrease in relative supersaturation could dissolve a crystal. Your set up would be a reserve vapor diffusion. You say the crystals appeared right after setting the experiment so your crystallization is essentially a batch experiment. Therefore you might want to change your set up from a vapor diffusion to a microbatch experiment under oil. If you need more information about how to perform a microbatch experiment, let me know and I’ll explain. Hope this helps. Kind Regards, Bob Cudney Hampton Research 34 Journey Aliso Viejo, CA 92656-3317 USA Telephone 1 949 425 1321 Extension 200 Fax 1 949 425 1611 E-mail b...@hrmail.com Web www.hamptonresearch.com From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of dusky dew Sent: Friday, May 02, 2014 4:39 AM To: CCP4BB@JISCMAIL.AC.UK Subject: [ccp4bb] Crystals Disappearing Overnight Dear All, I am trying to crystallize a protein with Adenosine. My protein is in 20 mM Tris, 300 mM NaCl and the crystals appear in a condition with 5 percent PEG8K, 0.1 M Sodium Cacodylate. The protein is incubated with adenosine for 1/2 hr before setting the drop. The crystals appear right after the drop is set but unfortunately they dissolve overnight. The plate is kept at 16 degree. Could anyone elaborate on this. Is it possibly occurring because Adenosine has stability issues. Thanks for your suggestions. ~ Maria
[ccp4bb] Post Doctoral Positions in Membrane Protein Structure and Function at the National Institutes of Health (NIH)
Dear all, Please circulate to those who may be interested - The research group of Dr. Anirban Banerjee at the National Institutes of Health (NIH) is seeking candidates for postdoctoral fellows . 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 reconstitution based biochemistry, electrophysiology together with synthetic chemistry to investigate the structural bases of the mechanisms of a number of membrane proteins. The lab has state-of-the-art equipment and facilities. We have dedicated periodic access to the 22-ID beamline at the APS, for crystallographers at the NIH. The DC Metro area is a wonderful place to live, the weather is pleasant and is a vibrant international community. In terms of scientific opportunities, the NIH is absolutely unique in bringing scientists from a virtually all corner of biomedical sciences in one campus. The Cell Biology and Metabolism program is part of the National Institute of Child Health and Human Development (NICHD). Our lab also has an affiliation to the National Institute of Neurological Disorders and Stroke (NINDS). 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 or insect/mammalian expression systems will be added advantages. Interested candidates can send an e-mail to anirban.baner...@nih.gov with a CV and a summary of previous research experience and future interests. Disclaimers: The NIH is dedicated to building a diverse community in its training and employment programs and this position is subject to a background investigation. Thanks very much. Best, Anirban
[ccp4bb] INSTRUCT Training Course announcement: ISBio2014 in Caparica, Portugal
2nd Call for applications to the INSTRUCT Training Course Integrative Structural Biology tools for the study of protein-ligand interactions The course will take place at FCT-Universidade NOVA de Lisboa, in the campus of Caparica, Portugal, from 6-12 October 2014 COURSE DETAILS REGISTRATION: http://eventos.fct.unl.pt/isbio2014 http://www.structuralbiology.eu/support/whats-on/calendar-events/integrative-structural-biology-tools-study-protein-ligand APPLICATION DEADLINE: June 1, 2014 (attendance is limited to 16 participants; PhD students and young Post-docs will be given priority) OBJECTIVE OF THE COURSE: to illustrate the added value of a structural biology approach to the study of protein-ligand interactions by combining four core methodologies, X-ray Crystallography, NMR, Carbohydrate Microarrays and ITC from a hands-on perspective, giving emphasis to their limitations and complementarities. To achieve this, and to better illustrate the integrative context, the students will receive training by working in a study-case, a protein that recognizes and binds to carbohydrates, which will be the focus of the several complementary methodologies. Besides the hands-on training there will be theory lectures where the main theoretical aspects of each technique will be explained always with an emphasis on the complementarity with the other techniques. APPLICATIONS AND COURSE FEE: The detailed registration info and the online application form are available at http://eventos.fct.unl.pt/isbio2014/pages/registration-info Fee is €350 and includes attendance, course documentation, coffee breaks (all course days), lunches (all course days), course dinner (Oct 7) and accommodation (single room inc. breakfast in Lisboa Almada Hotel; check-in: Oct 5; check-out: Oct 12). SUPPORTED BY: INSTRUCT, REQUIMTE-FCT-UNL, CIISA-FMV-UL Please, forward this information to anyone who might be interested. We are looking forward to welcome you in Caparica (and the beaches nearby...) Best regards, The Organizers Ana Luisa Carvalho Eurico Cabrita Assistant Researcher of Associate Lab REQUIMTE/CQFB@FCT-UNL *** Biologia Estrutural - Cristalografia de Raios-X (Gab 6.34) Dep. Quimica, FCT-UNL 2829-516 Caparica Portugal Phone: 00351212948300 (ext. 10940) Fax: 00351212948550 http://docentes.fct.unl.pt/almc/biocv http://xtal.dq.fct.unl.pt/ *** Post-Doctoral opportunity: http://xtal.dq.fct.unl.pt/PCISBIO/Job_announcements.html INSTRUCT Training Course in Caparica, from 6 to 12 October: ISBio2014 (registration open until June, 1st) Visit the Portuguese Centre for Integrated Structural Biology: http://xtal.dq.fct.unl.pt/PCISBIO and learn how you can benefit from all the infrastructures and skills! And join us in the IYCr2014 celebrations in Caparica!
[ccp4bb] Postdoctoral position in GPCR structural biology
An NIH R01-funded postdoctoral position in G protein-coupled receptor structural biology is available in the Pioszak laboratory in the Department of Biochemistry and Molecular Biology at the University of Oklahoma Health Sciences Center. The position will involve biochemical, pharmacological, and X-ray crystallographic studies of peptide-activated class B GPCRs. The Pioszak lab is housed in the modern S.L. Young Biomedical Research Center on the expanding medical school campus adjacent to the downtown Oklahoma City area. We have state-of-the-art equipment for protein expression and purification, biophysical characterization of proteins (ITC, DSC, CD), robotic crystallization (ARI Gryphon), and in-house X-ray data collection (Rigaku system), as well as access to synchrotron macromolecular crystallography beam lines. Appplicants should hold a recent Ph.D. in Biochemistry, Pharmacology, or related area. Experience with molecular biology techniques, protein expression and purification, protein-ligand interaction methods, cell culture, and X-ray crystallography would be an advantage, but experience in all of these areas is not required. Salary to be commensurate with qualifications and experience. To apply, please send a cover letter, CV, and the names and contact information for 3 references to augen-pios...@ouhsc.edumailto:augen-pios...@ouhsc.edu. The University of Oklahoma is an Affirmative Action/Equal Opportunity Employer. - Augen A. Pioszak, Ph.D. Assistant Professor Department of Biochemistry and Molecular Biology University of Oklahoma Health Sciences Center Letter address (office): 975 N.E. 10th Street, BRC 462B P.O. Box 26901 Oklahoma City, OK 73126-0901 Shipping address (lab): 975 N.E. 10th Street, BRC 464 Oklahoma City, OK 73104 Phone: 405-271-2401 Fax: 405-271-3910 Email: augen-pios...@ouhsc.edumailto:augen-pios...@ouhsc.edu http://www.oumedicine.com/department-of-biochemistry-and-molecular-biology/faculty/augen-pioszak-ph-d- -
[ccp4bb] ADVX in remote acces is a nightmare
Hi all, I'm currently trying to collect data remotely (me sitting at home, crystals at Diamond Light Source). This involves accessing the beamline computer system via a NX machine system, thus opening a virtual desktop of the remote machine on my local computer (PC running W7) Now, I'm trying to look at the images via ADXV, which works fine on site, but close to impossible here. When I load the file in ADXV the main window is white, no images (all the menus are alive, it's reacting to the position of the pointer, giving resolution etc...). Sometimes (but not always!), I can see the image once I've dragged another window onto the ADVX window, this seems to refresh it and then I can work with it fine. But as of now, that happens very only rarely... thus I cannot see my images.. Is there a trick/workaround ? Thanks... (beamtime is running by..) Cedric
Re: [ccp4bb] ADVX in remote acces is a nightmare
Once upon a time, I got this from Andy Arvai: What if the Image Window doesn't refresh correctly under NX? This is a bug in NX and should be fixed in newer versions of the NX server. Depending on the version of NX you have try adding one of the following options: /usr/NX/etc/node.cfg AgentExtraOptions=-defer 0 /etc/nxserver/node.conf AGENT_EXTRA_OPTIONS_X=-defer 0 If this doesn't work try setting the Link Type to LAN. This may be slower, but may also work. Finally, try using adxv -nopixmap HTH! -James Holton MAD Scientist On 5/3/2014 2:34 PM, Cedric Govaerts wrote: Hi all, I'm currently trying to collect data remotely (me sitting at home, crystals at Diamond Light Source). This involves accessing the beamline computer system via a NX machine system, thus opening a virtual desktop of the remote machine on my local computer (PC running W7) Now, I'm trying to look at the images via ADXV, which works fine on site, but close to impossible here. When I load the file in ADXV the main window is white, no images (all the menus are alive, it's reacting to the position of the pointer, giving resolution etc...). Sometimes (but not always!), I can see the image once I've dragged another window onto the ADVX window, this seems to refresh it and then I can work with it fine. But as of now, that happens very only rarely... thus I cannot see my images.. Is there a trick/workaround ? Thanks... (beamtime is running by..) Cedric
