[ccp4bb] disordered helix
Dear all, I have solved the structure of my protein by molecular replacement at 2.9A, with Rfactor and Rfree 18 and 22 respectively. Overall everything seems fine, its a two domain protein NTD and CTD, the NTD have high average B factor compared to CTD. A helix of NTD seems to be disordered, I tried different geometric weights but the refined structure does not seem to follow proper geometry for this helix. The B-factor of this helix is very high compared to overall B factor for NTD and omit map shows only some partial density in this region( off course not conclusive). All the homologous structure have helix in this region although with high B-factor. Should I submit the current pdb or need more refinement? thanks and regards Atul Kumar
[ccp4bb] Post-doctoral position at the LMB, Cambridge, UK
MRC Laboratory of Molecular Biology, Cambridge, UK Post-doctoral Scientist Position We seek to appoint a postdoctoral researcher to the Division of Structural Studies to undertake protein crystallographic analysis of the anaphase promoting complex (APC/C) and its subunits. This is an Investigator Scientist position funded by Cancer Research UK for 3 years and is located at the MRC Laboratory of Molecular Biology in Cambridge, UK within the group of David Barford. The Investigator Scientist will join a team investigating the structure and mechanism of the anaphase promoting complex (APC/C), a large macro-molecular assembly that regulates progression through the cell cycle. The project will focus on crystal structure determination of the whole APC/C and APC/C subunits and sub-complexes. This project extends our existing 11 Å resolution structure determined using single particle electron microscopy and takes advantage of our recombinant over-expression system for reconstituting APC/C (da Fonseca et al., (2011) and Schreiber et al., (2011)). The project would suit a highly motivated and committed scientist interested in molecular mechanisms of the cell cycle, cancer biology and structural biology. The successful applicant will hold at least a PhD (or equivalent) in a biological/physical science. Experience with protein crystallography is essential and experience with protein crystallisation and purification would be an advantage. The post is available from 1st October 2013 (or earlier). The starting salary will be set within £26,282 to £31,686. However, the full salary range for a position of this type is from £26,282 - £37,040. Informal enquires may be made to David Barford (david.barf...@icr.ac.ukmailto:david.barf...@icr.ac.uk). 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] disordered helix
Hard to say without seeing the maps and experimenting. My first check would be to set the NTD occupancies to 0.0 - refine the CTD alone, then look at the maps in COOT. Or maybe let an automatic modelling building program such as Buccaneer try to rebuild the NTD section, with starting phases from the CTD. Eleanor On 13 May 2013 09:04, atul kumar atulsingh21...@gmail.com wrote: Dear all, I have solved the structure of my protein by molecular replacement at 2.9A, with Rfactor and Rfree 18 and 22 respectively. Overall everything seems fine, its a two domain protein NTD and CTD, the NTD have high average B factor compared to CTD. A helix of NTD seems to be disordered, I tried different geometric weights but the refined structure does not seem to follow proper geometry for this helix. The B-factor of this helix is very high compared to overall B factor for NTD and omit map shows only some partial density in this region( off course not conclusive). All the homologous structure have helix in this region although with high B-factor. Should I submit the current pdb or need more refinement? thanks and regards Atul Kumar
[ccp4bb] PISA interface question
Hello everybody, I am trying to evaluate solvatation energy of my protein and. Unfortunately the protein highly glycosilated and it seems that PISA does not take into account hydrogen bonds between mannose from one molecule and symmetry related molecule. Is there any way to tell PISA to use ligands in calculations of the assembly solvatation energy. -- Eugene Osipov Junior Research Scientist Laboratory of Enzyme Engineering A.N. Bach Institute of Biochemistry Russian Academy of Sciences Leninsky pr. 33, 119071 Moscow, Russia e-mail: e.m.osi...@gmail.com
[ccp4bb] A faculty position in the Department of Biophysics, NIMHANS, Bangalore, India
Dear all, Opening for Associate/Assistant Professor Position in the Department of Biophysics, National Institute of Mental Health and Neuro Sciences (NIMHANS), Bangalore, INDIA. Research experience: preference to Neuroscience/membrane proteins. Here is the link to the advertisement: http://www.nimhans.kar.nic.in/notifications.htm The last date for receipt of Application: *05 Jun 2013* Please forward to interested candidates. Thanks. -B. Padmanabhan * B. Padmanabhan, Ph. D Additional Professor Department of Biophysics NIMHANS Bangalore - 560029 INDIA
[ccp4bb] Postdoctoral Position available at Brown University, Providence, RI
The Page Laboratory at Brown University has an opening for a highly motivated postdoctoral research scientist to undertake structural analysis of signaling complexes, with a focus on the interactions between phosphatases, kinases and their regulatory proteins. Most studies typically employ a combination of x-ray crystallography, NMR spectroscopy and SAXS. Applicants with a strong background in biochemical techniques and structure determination are encouraged to apply. The Page laboratory is housed in a 105,000 sq. ft. research laboratory in Providence, RI. The laboratories are open-lab-space facilities and allow for close interactions with the adjunct structural biology and molecular biology groups at Brown University. Recent upgrades to the Brown University structural biology infrastructure include new X-ray crystallography instrumentation (Rigaku FR-E+ SuperBright with VariMax-HF ArcSec optics, a Saturn 944 CCD detector and ACTOR robotic crystal mounting system), a BIOSAXS-1000 small angle X-ray scattering instrument, and the addition of an 850 MHz NMR spectrometer. Providence, RI, is located on the coast between Boston (45 min.) and New York City (3 hours). Starting date, salary and project details are negotiable. Please e-mail a Cover Letter and CV to Rebecca Page (rebecca_p...@brown.edu), Brown University, Box G-E4, Providence, RI 02912.
Re: [ccp4bb] disordered helix
Sometimes a floppy bit of a protein is even more floppy in a particular crystal form. Your maps do not appear to support your model of a helix in this location. I would not build it unless maps based on later refinement show something reasonable in the omit map. (Of course if you leave out the helix, all your maps will be omit maps.) It is quite common to submit models to the PDB that do not contain all of the amino acids expected based on the sequence. If you can't see where the chain goes you certainly can't be expected to build it. Dale Tronrud On 05/13/2013 04:23 AM, atul kumar wrote: I have attached the map and omit map(after deleting helix) images. 2Fo-Fc(1 sigma) Fo-Fc(3sigma) On 5/13/13, Eleanor Dodson eleanor.dod...@york.ac.uk wrote: Hard to say without seeing the maps and experimenting. My first check would be to set the NTD occupancies to 0.0 - refine the CTD alone, then look at the maps in COOT. Or maybe let an automatic modelling building program such as Buccaneer try to rebuild the NTD section, with starting phases from the CTD. Eleanor On 13 May 2013 09:04, atul kumar atulsingh21...@gmail.com wrote: Dear all, I have solved the structure of my protein by molecular replacement at 2.9A, with Rfactor and Rfree 18 and 22 respectively. Overall everything seems fine, its a two domain protein NTD and CTD, the NTD have high average B factor compared to CTD. A helix of NTD seems to be disordered, I tried different geometric weights but the refined structure does not seem to follow proper geometry for this helix. The B-factor of this helix is very high compared to overall B factor for NTD and omit map shows only some partial density in this region( off course not conclusive). All the homologous structure have helix in this region although with high B-factor. Should I submit the current pdb or need more refinement? thanks and regards Atul Kumar
[ccp4bb] Overide refmac restraints
H ithere, I am trying to refine a covalently bound ligand to my protein and I am having trouble with the restraints. I have generated the cif file and link within Jligand and this is reasonable. However it appears that REFMAC is overriding these and fitting the ligand to the density. I have added a keyword text file with externtal restraints such as : exte angle first chain A resi 1 atom S12 second chain A resi 1 atom C10 third chain A resi 2 atom N value 120 sigma 3.0 The resulting structures has incorrect angles which do not match the external restraints or cif file. Am I missing something? Any help muchly appreciated. (I am using the most upto date version of the CCP4 package on a PC) Joel _ 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
Re: [ccp4bb] Overide refmac restraints
Hi Joel IF you use external restraints then you need to add type 0 at the end. Then refmac will assume that you want to override standard restraints. I.e. exte angle first chain A resi 1 atom S12 second chain A resi 1 atom C10 third chain A resi 2 atom N value 120 sigma 3.0 type 0 It is very strange that refmac is overriding definitions from your cif file. Would it be possible to have a look your cif file and how use it in refmac. For example some portion of your protein with cif file might be helpful. Regards Garib On 13 May 2013, at 22:45, Joel Tyndall wrote: H ithere, I am trying to refine a covalently bound ligand to my protein and I am having trouble with the restraints. I have generated the cif file and link within Jligand and this is reasonable. However it appears that REFMAC is overriding these and fitting the ligand to the density. I have added a keyword text file with externtal restraints such as : exte angle first chain A resi 1 atom S12 second chain A resi 1 atom C10 third chain A resi 2 atom N value 120 sigma 3.0 The resulting structures has incorrect angles which do not match the external restraints or cif file. Am I missing something? Any help muchly appreciated. (I am using the most upto date version of the CCP4 package on a PC) Dr Garib N Murshudov Group Leader, MRC Laboratory of Molecular Biology Francis Crick Avenue Cambridge Biomedical Campus Cambridge CB2 0QH UK Email: ga...@mrc-lmb.cam.ac.uk Web http://www.mrc-lmb.cam.ac.uk, http://www2.mrc-lmb.cam.ac.uk/groups/murshudov/
Re: [ccp4bb] PISA interface question
On 05/13/2013 06:34 PM, Evgeny Osipov wrote: Hello everybody, I am trying to evaluate solvatation energy of my protein and. Unfortunately the protein highly glycosilated and it seems that PISA does not take into account hydrogen bonds between mannose from one molecule and symmetry related molecule. Is there any way to tell PISA to use ligands in calculations of the assembly solvatation energy. Isn't it a job for APBS? http://www.poissonboltzmann.org/apbs
[ccp4bb] Fwd: bioassay g-csf
-- Forwarded message -- From: megha goyal mgbio...@gmail.com Date: Mon, May 6, 2013 at 1:16 PM Subject: bioassay g-csf To: CCP4BB@jiscmail.ac.uk we perform bioassay of g-csf using m-nfs-60 cell lines but we do not get reproducible results. some time a sample fails and on repeating the assay for same sample it passes. we do not get proper gradation too. can someone please provide me a protocol for g-csf bioassay that has been successfully used. please help me.
[ccp4bb] reference for true multiplicity?
Hi, I'm meant to know this but I'm blanking, so I'll crowdsource instead: Anybody know a (the) reference where it was showed that the best SAD data is obtained by collecting multiple revolutions at different crystal offsets (kappa settings)? It's axiomatic now (I hope!), but I remember seeing someone actually show this. I thought Sheldrick early tweens, but PubMed is not being useful. (Oh dear, this will unleash references from the 60s, won't it.) phx
