Re: [ccp4bb] high Rwork / Rfree after MR
Dear Shanti Pal Gangwar, based on the information you give, this does not sound like solved structure to me. By the way, the ACA conference at the end of May will feature a session data processing with the pros, where difficult projects will be analyzed, and hopefully structures like this will be solved. If you want your data to be analysed in that session, please send me an email - of course, the raw data need to be available. Although there is still a shortage of interesting projects for that session, unfortunately I cannot promise that any particular project will be accepted, nor that any particular structure can be solved - it's just on the basis of best effort. best, Kay On Tue, 4 Feb 2014 15:06:36 +0530, Shanti Pal Gangwar gangwar...@gmail.com wrote: Dear All I have solved one structure by MR. The data data quality was poor so the Rmerge was high. The resolution of the data is 3.3 Angstrom.The refinement statistics are also very poor Rw/Rf= 40/42 %. After many efforts we are not able to get reasonable Rw/Rf. My question is can it be claimed that we have solved the structure? Thanks in advance -- regards Shanti Pal Gangwar School of Life Sciences Jawaharlal Nehru University New Delhi-110067 India Email:gangwar...@gmail.com
[ccp4bb] Summer Student Placements @Diamond
Dear All, I would be grateful if you could bring the enclosed/linked opportunity to the attention of any suitable candidates in your institutions. Please also encourage them to look at all the categories e.g. the computing category is a bit of a misleading label (that might change soon). The Diamond Student Summer Placement scheme allows undergraduate students studying for a degree in Science, Engineering, Computing or Mathematics (and who expect to gain a first or upper-second class honours degree) to gain experience working within a scientific environment at Diamond. These 8-12 week placements are paid positions and will provide successful students with an opportunity to work on a research or development project within Diamond. http://www.diamond.ac.uk/Home/Jobs/placements/Current.html Sent on behalf of the placement organisers. Alun -- This e-mail and any attachments may contain confidential, copyright and or privileged material, and are for the use of the intended addressee only. If you are not the intended addressee or an authorised recipient of the addressee please notify us of receipt by returning the e-mail and do not use, copy, retain, distribute or disclose the information in or attached to the e-mail. Any opinions expressed within this e-mail are those of the individual and not necessarily of Diamond Light Source Ltd. Diamond Light Source Ltd. cannot guarantee that this e-mail or any attachments are free from viruses and we cannot accept liability for any damage which you may sustain as a result of software viruses which may be transmitted in or with the message. Diamond Light Source Limited (company no. 4375679). Registered in England and Wales with its registered office at Diamond House, Harwell Science and Innovation Campus, Didcot, Oxfordshire, OX11 0DE, United Kingdom
Re: [ccp4bb] off-topic: bug busting
Dear Phoebe, We have a constant systems homogenizer that we use routinely as a service for researchers here at UGA. It is really easy to use and gets up to high pressure (40k psi) so you can lyse plant cells or other difficult to lyse cell types. You just pour/pump your resuspended cells, as low as 25mL, into the inlet, setup parameters (usually once) and hit the start button. The lysed cells will exit the outlet line. We usually have 2 cups to catch the outlet and we recycle 3-6 times. The unit will automatically shutoff when it runs out of liquid. Below is a link to the website for the company with a video. For greater than 1L of cells we have a Niro-Soavi high pressure homogenizer (second link) that I recommend. http://www.constantsystems.com/products/cell%20disruption%20systems/ts%20series%20cabinet http://www.nirosoavi.com/products/Ariete_NS2006.asp On Tue, Feb 4, 2014 at 11:49 AM, Phoebe A. Rice pr...@uchicago.edu wrote: Some time ago, there was a nice discussion of cost-effective, wimpy protein-friendly ways to break open E. coli. We're thinking about replacing an aging sonicator. If people have a favorite gizmo, could they repeat that advice? thank you, Phoebe Rice ++ Phoebe A. Rice Dept. of Biochemistry Molecular Biology The University of Chicago 773 834 1723; pr...@uchicago.edu http://bmb.bsd.uchicago.edu/Faculty_and_Research/ http://www.rsc.org/shop/books/2008/9780854042722.asp
[ccp4bb] no link_id
I am asking this question on behalf of a friend: -Thanks! Hi All - I am working at small biotech. We have minimal budget for software, only CCP4 and Coot. We have a ligand that binds covalently to cysteine. I define the parameter file using Prodrg in CCP4 to get drug.cif file and then make a link in Coot between the Cys and the Drug. The problem is Refmac keeps complaining that there is no link_id for this bond: I am reading library. Please wait. mon_lib.cif WARNING : link: CYS--UNK is without link_id link will be created with covalent bond only. Bond= 1.610 WARNING : description of link:CYS-UNK is not in the dictionary link will be created with bond_length = 1.610 Where do I specify this link_id, and where can I find an example for the correct syntax? Is this done in mon_lib.cif or the drug.cif file?? The bond length, if I'm not mistaken should be ~1.82 A for distance between carbon and sulfur. Thanks for any help.
[ccp4bb] CeBEM/CCP4 South American School in Macromolecular Crystallography - Final week for applications
Dear Colleagues, This is a reminder about the second CeBEM/CCP4 South American School in Macromolecular Crystallography. The closing date for applications is Monday the 10th of February. This year's school will take place at Sao Carlos University in Brazil. Full details including the program and application information can be found at the school website: http://www.ifsc.usp.br/mx2014 Title: Macromolecular Crystallography School 2014: From data processing to structure refinement and beyond Dates: April 8th to 16th 2014 Purpose of the course and intended audience: The course is intended mainly for graduate students, postdoctoral researchers and young scientists in the area of structural biology. The school is not meant as an introductory level course to protein crystallography. It is designed more for applicants with reasonable expertise in crystallography and experience with the CCP4 suite. The purpose of the school is to address specific problems that the applicants face while processing diffraction data and while solving and refining novel structures. Applicants with diffraction data (solved, partially solved or unsolved) will be given strong consideration, although these are not mandatory requirements. The workshop will cover many popular programs used for data processing and structure solution with the software developers available to help throughout the week. Programs covered will include: Mosflm, Aimless, XDS, Refmac, ARP/wARP, Phaser, Crank, ARCIMBOLDO, Coot, SHELXC/D/E, Balbes, MrBUMP, Buccaneer and many more. Grants will be available for traveling students needing assistance with funding. Best wishes, Eduardo, Garib, Richard and Ronan -- Scanned by iCritical.
Re: [ccp4bb] no link_id
Hi Ronnie - Refmac has done the job for you (or for your friend). The Refmac log file says that Refmac has created a file called .cif (the exact name varies). You should open up that file and inspect it; it will be Refmac's best guess about the geometry of that link. I usually specify only bond lengths and bond angles, leaving torsion angles undefined (i.e. deleting those restraints) unless I know it should be restrained to a certain angle. The next time you run Refmac, use the Library file input line in ccp4i to feed it this cif file; refinement should proceed OK after this. You can also append the contents of the cif file that Refmac made to your drug.cif file. Just cut and paste with a text editor. Don't forget that you need to give your ligand a real 3-letter code, not just UNK, before depositing it. So you might as well do it now. (Even if the structure will remain private, not deposited, I would still change the code.) - Matt On 2/5/14 10:59 AM, Ronnie wrote: I am asking this question on behalf of a friend: -Thanks! Hi All - I am working at small biotech. We have minimal budget for software, only CCP4 and Coot. We have a ligand that binds covalently to cysteine. I define the parameter file using Prodrg in CCP4 to get drug.cif file and then make a link in Coot between the Cys and the Drug. The problem is Refmac keeps complaining that there is no link_id for this bond: I am reading library. Please wait. mon_lib.cif WARNING : link: CYS--UNK is without link_id link will be created with covalent bond only. Bond= 1.610 WARNING : description of link:CYS-UNK is not in the dictionary link will be created with bond_length = 1.610 Where do I specify this link_id, and where can I find an example for the correct syntax? Is this done in mon_lib.cif or the drug.cif file?? The bond length, if I'm not mistaken should be ~1.82 A for distance between carbon and sulfur. Thanks for any help. -- Matthew Franklin, Ph. D. Senior Scientist New York Structural Biology Center 89 Convent Avenue, New York, NY 10027 (212) 939-0660 ext. 9374
Re: [ccp4bb] no link_id
If there is a covalent link between CYS and UNK then it is better to create that using jligand from ccp4. There are nice tutorials designed by Andrey Lebedev. Tutorials are on this site: http://www.ysbl.york.ac.uk/mxstat/JLigand/ Regards Garib On 5 Feb 2014, at 15:59, Ronnie ronnie...@yahoo.com wrote: I am asking this question on behalf of a friend: -Thanks! Hi All - I am working at small biotech. We have minimal budget for software, only CCP4 and Coot. We have a ligand that binds covalently to cysteine. I define the parameter file using Prodrg in CCP4 to get drug.cif file and then make a link in Coot between the Cys and the Drug. The problem is Refmac keeps complaining that there is no link_id for this bond: I am reading library. Please wait. mon_lib.cif WARNING : link: CYS--UNK is without link_id link will be created with covalent bond only. Bond= 1.610 WARNING : description of link:CYS-UNK is not in the dictionary link will be created with bond_length = 1.610 Where do I specify this link_id, and where can I find an example for the correct syntax? Is this done in mon_lib.cif or the drug.cif file?? The bond length, if I'm not mistaken should be ~1.82 A for distance between carbon and sulfur. Thanks for any help. Dr Garib N Murshudov MRC-LMB Francis Crick Avenue Cambridge CB2 0QH UK Web http://www.mrc-lmb.cam.ac.uk, http://www2.mrc-lmb.cam.ac.uk/groups/murshudov/
Re: [ccp4bb] off-topic: bug busting
Interesting 'off-topic' thread. I'm a rather long-time user of Microfluidics cell disruptors (for E. coli or P. pastoris or S. ceresvisiae), and a shared-use M110-P Plug and Play is used by most of the membrane-heads at my place. I've generally been happy (or at least not unhappy) with it. However, we've been having some QC issues with a membrane protein that we're making in S. ceresvisiae (Sc), and I'm having some concerns about sample heating. Can anyone comment on Microfludics vs Avestin vs Constant Systems vs Retsch vs whatever-else for cracking Sc cells? These days, we're working up ~300-400g of paste at a time. Thank you very much! -MW Michael C. Wiener, Ph.D. Professor Department of Molecular Physiology and Biological Physics University of Virginia PO Box 800886 Charlottesville, VA 22908-0886 434-243-2731 434-982-1616 (FAX) On Wed, 5 Feb 2014 00:34:11 -0500 Anirban Banerjee ani...@gmail.com wrote: I will be curious to know about people's experiences with membrane proteins and lysing yeast cells with the Microfluidizer and how that compares with using a Retsch Miller, i.e. grinding in a liquid nitrogen cooled stainless steel chamber and plunging in liquid nitrogen in between grinding cycles. I am worried that the Microfluidizer is not as mild w.r.t. heating as they claim it to be. That would, of course, perfectly qualify as my OCD. Any insights will be really appreciated. Thanks, Anirban On Tue, Feb 4, 2014 at 11:58 AM, Matthew Franklin mfrank...@nysbc.org wrote: Hi Phoebe - Cost-effective may not be the applicable word here, but the Microfluidizer works very well: http://www.microfluidicscorp.com/index.php?option=com_contentview=articleid=19Itemid=76 This gadget runs on house compressed air (don't try to use a compressed air tank - you'll empty it in minutes). It's a bit noisy, but so is a sonicator. The Microfluidizer really shines with large volumes of lysate - like 1 L and up. If you're only processing 100-200 mL at a time, I think sonication is the best way to go. Hope that helps, Matt On 2/4/14 11:49 AM, Phoebe A. Rice wrote: Some time ago, there was a nice discussion of cost-effective, wimpy protein-friendly ways to break open E. coli. We're thinking about replacing an aging sonicator. If people have a favorite gizmo, could they repeat that advice? thank you, Phoebe Rice ++ Phoebe A. Rice Dept. of Biochemistry Molecular Biology The University of Chicago 773 834 1723; pr...@uchicago.edu http://bmb.bsd.uchicago.edu/Faculty_and_Research/ http://www.rsc.org/shop/books/2008/9780854042722.asp -- Matthew Franklin, Ph. D. Senior Scientist New York Structural Biology Center 89 Convent Avenue, New York, NY 10027 (212) 939-0660 ext. 9374
Re: [ccp4bb] off-topic: bug busting
If heat is a concern, you can place your Emulsiflex into a bin with lots of ice. We have the version with cooling after lysis, but if you are paranoid you can cool the whole thing. It has the footprint of a 15” MacBook Pro but it’s way more expensive than that :-) Jürgen .. Jürgen Bosch Johns Hopkins University Bloomberg School of Public Health Department of Biochemistry Molecular Biology Johns Hopkins Malaria Research Institute 615 North Wolfe Street, W8708 Baltimore, MD 21205 Office: +1-410-614-4742tel:%2B1-410-614-4742 Lab: +1-410-614-4894tel:%2B1-410-614-4894 Fax: +1-410-955-2926tel:%2B1-410-955-2926 http://lupo.jhsph.edu On Feb 5, 2014, at 11:49 AM, Michael C. Wiener mwie...@virginia.edumailto:mwie...@virginia.edu wrote: Interesting 'off-topic' thread. I'm a rather long-time user of Microfluidics cell disruptors (for E. coli or P. pastoris or S. ceresvisiae), and a shared-use M110-P Plug and Play is used by most of the membrane-heads at my place. I've generally been happy (or at least not unhappy) with it. However, we've been having some QC issues with a membrane protein that we're making in S. ceresvisiae (Sc), and I'm having some concerns about sample heating. Can anyone comment on Microfludics vs Avestin vs Constant Systems vs Retsch vs whatever-else for cracking Sc cells? These days, we're working up ~300-400g of paste at a time. Thank you very much! -MW Michael C. Wiener, Ph.D. Professor Department of Molecular Physiology and Biological Physics University of Virginia PO Box 800886 Charlottesville, VA 22908-0886 434-243-2731 434-982-1616 (FAX) On Wed, 5 Feb 2014 00:34:11 -0500 Anirban Banerjee ani...@gmail.commailto:ani...@gmail.com wrote: I will be curious to know about people's experiences with membrane proteins and lysing yeast cells with the Microfluidizer and how that compares with using a Retsch Miller, i.e. grinding in a liquid nitrogen cooled stainless steel chamber and plunging in liquid nitrogen in between grinding cycles. I am worried that the Microfluidizer is not as mild w.r.t. heating as they claim it to be. That would, of course, perfectly qualify as my OCD. Any insights will be really appreciated. Thanks, Anirban On Tue, Feb 4, 2014 at 11:58 AM, Matthew Franklin mfrank...@nysbc.orgmailto:mfrank...@nysbc.org wrote: Hi Phoebe - Cost-effective may not be the applicable word here, but the Microfluidizer works very well: http://www.microfluidicscorp.com/index.php?option=com_contentview=articleid=19Itemid=76 This gadget runs on house compressed air (don't try to use a compressed air tank - you'll empty it in minutes). It's a bit noisy, but so is a sonicator. The Microfluidizer really shines with large volumes of lysate - like 1 L and up. If you're only processing 100-200 mL at a time, I think sonication is the best way to go. Hope that helps, Matt On 2/4/14 11:49 AM, Phoebe A. Rice wrote: Some time ago, there was a nice discussion of cost-effective, wimpy protein-friendly ways to break open E. coli. We're thinking about replacing an aging sonicator. If people have a favorite gizmo, could they repeat that advice? thank you, Phoebe Rice ++ Phoebe A. Rice Dept. of Biochemistry Molecular Biology The University of Chicago 773 834 1723; pr...@uchicago.edu http://bmb.bsd.uchicago.edu/Faculty_and_Research/ http://www.rsc.org/shop/books/2008/9780854042722.asp -- Matthew Franklin, Ph. D. Senior Scientist New York Structural Biology Center 89 Convent Avenue, New York, NY 10027 (212) 939-0660 ext. 9374
[ccp4bb] Computational Crystallography Newsletter - Volume 5, Number 1
Dear Colleagues, I am pleased to announce the publication of the latest issue of the Computational Crystallography Newsletter: http://www.phenix-online.org/newsletter/ A listing of the articles and short communications is given below. Please note that the newsletter accepts articles of a general nature of interest to all crystallographers. Please send any articles to me at nwmoria...@lbl.gov noting that there is a Word Template on the website to streamline production. Articles FEL Detectors and ImageCIF Quantum Mechanics-based Refinement in Phenix/DivCon Short communications Short Communications Phenix tools for interpretation of BIOMT and MTRIX records of PDB files Coping with BIG DATA image formats: integration of CBF, NeXus and HDF5 Fitting tips #7 - Getting the Pucker Right in RNA Structures Cheers Nigel -- Nigel W. Moriarty Building 64R0246B, Physical Biosciences Division Lawrence Berkeley National Laboratory Berkeley, CA 94720-8235 Phone : 510-486-5709 Email : nwmoria...@lbl.gov Fax : 510-486-5909 Web : CCI.LBL.gov
[ccp4bb] Method to calculate/measure the mean solvent electron density calculated from the crystallization conditions?
Dear All, Is there a way or program that is good at providing a solid estimate of F000 using the standard mean protein electron density of 0.433 e/A**3 and a mean solvent electron density calculated from the crystallization conditions? I am wondering how the latter might affect F000 if the conditions are a bit different then the values assumed in most programs. I am assuming from these two values I can calculate a pretty good estimate of F000 using: F000 = mean_electron_density*Vcell(e-), where mean_electron_density=fraction_protein*0.433 e/A**3 + fraction_solvent*mean_solvent_electron_density(e/A**3). In short, how can one calculate the average solvent density instead of using the value assumed in many programs of 0.35? I know that 4M ammonium sulphate is 0.41 and pure water is 0.33 but not sure how these values are measured (or calculated). Thanks! Joe __ Joseph P. Noel, Ph.D. Arthur and Julie Woodrow Chair Investigator, Howard Hughes Medical Institute Professor, The Jack H. Skirball Center for Chemical Biology and Proteomics The Salk Institute for Biological Studies 10010 North Torrey Pines Road La Jolla, CA 92037 USA Phone: (858) 453-4100 extension 1442 Cell: (858) 349-4700 Fax: (858) 597-0855 E-mail: n...@salk.edu Publications Citations: http://scholar.google.com/citations?user=xiL1lscJ Homepage Salk: http://www.salk.edu/faculty/noel.html Homepage HHMI: http://hhmi.org/research/investigators/noel.html __
Re: [ccp4bb] off-topic: bug busting
We always have used ice with the Microfluidics disruptor. We also, on occasion, monitor the temperature of the process stream. I'm not convinced, at all, that there is actually any problem with heating, but I thought it reasonable to put the query out there. Thanks to all who emailed me. -MW On Wed, 5 Feb 2014 12:07:18 -0500 Bosch, Juergen jubo...@jhsph.edu wrote: If heat is a concern, you can place your Emulsiflex into a bin with lots of ice. We have the version with cooling after lysis, but if you are paranoid you can cool the whole thing. It has the footprint of a 15” MacBook Pro but it’s way more expensive than that :-) Jürgen .. Jürgen Bosch Johns Hopkins University Bloomberg School of Public Health Department of Biochemistry Molecular Biology Johns Hopkins Malaria Research Institute 615 North Wolfe Street, W8708 Baltimore, MD 21205 Office: +1-410-614-4742tel:%2B1-410-614-4742 Lab: +1-410-614-4894tel:%2B1-410-614-4894 Fax: +1-410-955-2926tel:%2B1-410-955-2926 http://lupo.jhsph.edu On Feb 5, 2014, at 11:49 AM, Michael C. Wiener mwie...@virginia.edumailto:mwie...@virginia.edu wrote: Interesting 'off-topic' thread. I'm a rather long-time user of Microfluidics cell disruptors (for E. coli or P. pastoris or S. ceresvisiae), and a shared-use M110-P Plug and Play is used by most of the membrane-heads at my place. I've generally been happy (or at least not unhappy) with it. However, we've been having some QC issues with a membrane protein that we're making in S. ceresvisiae (Sc), and I'm having some concerns about sample heating. Can anyone comment on Microfludics vs Avestin vs Constant Systems vs Retsch vs whatever-else for cracking Sc cells? These days, we're working up ~300-400g of paste at a time. Thank you very much! -MW Michael C. Wiener, Ph.D. Professor Department of Molecular Physiology and Biological Physics University of Virginia PO Box 800886 Charlottesville, VA 22908-0886 434-243-2731 434-982-1616 (FAX) On Wed, 5 Feb 2014 00:34:11 -0500 Anirban Banerjee ani...@gmail.commailto:ani...@gmail.com wrote: I will be curious to know about people's experiences with membrane proteins and lysing yeast cells with the Microfluidizer and how that compares with using a Retsch Miller, i.e. grinding in a liquid nitrogen cooled stainless steel chamber and plunging in liquid nitrogen in between grinding cycles. I am worried that the Microfluidizer is not as mild w.r.t. heating as they claim it to be. That would, of course, perfectly qualify as my OCD. Any insights will be really appreciated. Thanks, Anirban On Tue, Feb 4, 2014 at 11:58 AM, Matthew Franklin mfrank...@nysbc.orgmailto:mfrank...@nysbc.org wrote: Hi Phoebe - Cost-effective may not be the applicable word here, but the Microfluidizer works very well: http://www.microfluidicscorp.com/index.php?option=com_contentview=articleid=19Itemid=76 This gadget runs on house compressed air (don't try to use a compressed air tank - you'll empty it in minutes). It's a bit noisy, but so is a sonicator. The Microfluidizer really shines with large volumes of lysate - like 1 L and up. If you're only processing 100-200 mL at a time, I think sonication is the best way to go. Hope that helps, Matt On 2/4/14 11:49 AM, Phoebe A. Rice wrote: Some time ago, there was a nice discussion of cost-effective, wimpy protein-friendly ways to break open E. coli. We're thinking about replacing an aging sonicator. If people have a favorite gizmo, could they repeat that advice? thank you, Phoebe Rice ++ Phoebe A. Rice Dept. of Biochemistry Molecular Biology The University of Chicago 773 834 1723; pr...@uchicago.edu http://bmb.bsd.uchicago.edu/Faculty_and_Research/ http://www.rsc.org/shop/books/2008/9780854042722.asp -- Matthew Franklin, Ph. D. Senior Scientist New York Structural Biology Center 89 Convent Avenue, New York, NY 10027 (212) 939-0660 ext. 9374
[ccp4bb] N-linked glycans mediate inter-molecular protein-protein interactions
Dear colleagues, I am looking for structures of protein complexes in which N-linked glycans mediate inter-molecular protein-protein interactions. Can anyone point me in the right direction? Thanks! Tianyu Gmail via foxmail
Re: [ccp4bb] Method to calculate/measure the mean solvent electron density calculated from the crystallization conditions?
Starting with the density (or specific volume) of water, you can obtain rho(water)=0.33 e-/A^3. It's a nifty little bit of arithmetic and dimensional analysis. -MW Michael C. Wiener, Ph.D. Professor Department of Molecular Physiology and Biological Physics University of Virginia PO Box 800886 Charlottesville, VA 22908-0886 434-243-2731 434-982-1616 (FAX) On Wed, 5 Feb 2014 10:39:57 -0800 Joseph Noel n...@salk.edu wrote: Dear All, Is there a way or program that is good at providing a solid estimate of F000 using the standard mean protein electron density of 0.433 e/A**3 and a mean solvent electron density calculated from the crystallization conditions? I am wondering how the latter might affect F000 if the conditions are a bit different then the values assumed in most programs. I am assuming from these two values I can calculate a pretty good estimate of F000 using: F000 = mean_electron_density*Vcell(e-), where mean_electron_density=fraction_protein*0.433 e/A**3 + fraction_solvent*mean_solvent_electron_density(e/A**3). In short, how can one calculate the average solvent density instead of using the value assumed in many programs of 0.35? I know that 4M ammonium sulphate is 0.41 and pure water is 0.33 but not sure how these values are measured (or calculated). Thanks! Joe __ Joseph P. Noel, Ph.D. Arthur and Julie Woodrow Chair Investigator, Howard Hughes Medical Institute Professor, The Jack H. Skirball Center for Chemical Biology and Proteomics The Salk Institute for Biological Studies 10010 North Torrey Pines Road La Jolla, CA 92037 USA Phone: (858) 453-4100 extension 1442 Cell: (858) 349-4700 Fax: (858) 597-0855 E-mail: n...@salk.edu Publications Citations: http://scholar.google.com/citations?user=xiL1lscJ Homepage Salk: http://www.salk.edu/faculty/noel.html Homepage HHMI: http://hhmi.org/research/investigators/noel.html __
Re: [ccp4bb] N-linked glycans mediate inter-molecular protein-protein interactions
Dear Tianyu, I have found the recent review by: Nagae and Yamaguchi in Int. J. Mol. Sci. 2012, 13, 8398-8429; doi:10.3390/ijms13078398 quite a good starting point. best Savvas On 05 Feb 2014, at 19:53, Gmail adams.wan...@gmail.com wrote: Dear colleagues, I am looking for structures of protein complexes in which N-linked glycans mediate inter-molecular protein-protein interactions. Can anyone point me in the right direction? Thanks! Tianyu Gmail via foxmail
Re: [ccp4bb] no link_id
Just to add to Matthew's good advice, UNK is the standard residue name for unidentified amino acids. Using it for your compound may cause conflicts in other programs such as COOT. Cheers, Robbie Sent from my Windows Phone Van: Matthew Franklin Verzonden: 5-2-2014 17:50 Aan: CCP4BB@JISCMAIL.AC.UK Onderwerp: Re: [ccp4bb] no link_id Hi Ronnie - Refmac has done the job for you (or for your friend). The Refmac log file says that Refmac has created a file called .cif (the exact name varies). You should open up that file and inspect it; it will be Refmac's best guess about the geometry of that link. I usually specify only bond lengths and bond angles, leaving torsion angles undefined (i.e. deleting those restraints) unless I know it should be restrained to a certain angle. The next time you run Refmac, use the Library file input line in ccp4i to feed it this cif file; refinement should proceed OK after this. You can also append the contents of the cif file that Refmac made to your drug.cif file. Just cut and paste with a text editor. Don't forget that you need to give your ligand a real 3-letter code, not just UNK, before depositing it. So you might as well do it now. (Even if the structure will remain private, not deposited, I would still change the code.) - Matt On 2/5/14 10:59 AM, Ronnie wrote: I am asking this question on behalf of a friend: -Thanks! Hi All - I am working at small biotech. We have minimal budget for software, only CCP4 and Coot. We have a ligand that binds covalently to cysteine. I define the parameter file using Prodrg in CCP4 to get drug.cif file and then make a link in Coot between the Cys and the Drug. The problem is Refmac keeps complaining that there is no link_id for this bond: I am reading library. Please wait. mon_lib.cif WARNING : link: CYS--UNK is without link_id link will be created with covalent bond only. Bond= 1.610 WARNING : description of link:CYS-UNK is not in the dictionary link will be created with bond_length = 1.610 Where do I specify this link_id, and where can I find an example for the correct syntax? Is this done in mon_lib.cif or the drug.cif file?? The bond length, if I'm not mistaken should be ~1.82 A for distance between carbon and sulfur. Thanks for any help. -- Matthew Franklin, Ph. D. Senior Scientist New York Structural Biology Center 89 Convent Avenue, New York, NY 10027 (212) 939-0660 ext. 9374
Re: [ccp4bb] off-topic: bug busting
Dear all, Our lab is predominantly interested in membrane proteins and currently uses a Constant Systems device for E coli hooked up to a cooling unit which pumps 4C water around the outside of the main chamber and keeps the sample cool even when using large volumes. This works really well for E. coli but we are expressing/studying more and more (mycobacterial) proteins in M. smegmatis these days and have found that our French press is still far superior to the Constant Systems device. The French press is aging though and I was interested if anyone else has experience with lysing M. smeg and if so which system/device they would recommend? Cheers Adam Dr Adam Heikal Department of Microbiology and Immunology University of Otago +64 (3) 471-6464 adam.hei...@otago.ac.nz http://micro.otago.ac.nz/our-people/adamheikal On 6/02/14 5:49 AM, Michael C. Wiener mwie...@virginia.edu wrote: Interesting 'off-topic' thread. I'm a rather long-time user of Microfluidics cell disruptors (for E. coli or P. pastoris or S. ceresvisiae), and a shared-use M110-P Plug and Play is used by most of the membrane-heads at my place. I've generally been happy (or at least not unhappy) with it. However, we've been having some QC issues with a membrane protein that we're making in S. ceresvisiae (Sc), and I'm having some concerns about sample heating. Can anyone comment on Microfludics vs Avestin vs Constant Systems vs Retsch vs whatever-else for cracking Sc cells? These days, we're working up ~300-400g of paste at a time. Thank you very much! -MW Michael C. Wiener, Ph.D. Professor Department of Molecular Physiology and Biological Physics University of Virginia PO Box 800886 Charlottesville, VA 22908-0886 434-243-2731 434-982-1616 (FAX) On Wed, 5 Feb 2014 00:34:11 -0500 Anirban Banerjee ani...@gmail.com wrote: I will be curious to know about people's experiences with membrane proteins and lysing yeast cells with the Microfluidizer and how that compares with using a Retsch Miller, i.e. grinding in a liquid nitrogen cooled stainless steel chamber and plunging in liquid nitrogen in between grinding cycles. I am worried that the Microfluidizer is not as mild w.r.t. heating as they claim it to be. That would, of course, perfectly qualify as my OCD. Any insights will be really appreciated. Thanks, Anirban On Tue, Feb 4, 2014 at 11:58 AM, Matthew Franklin mfrank...@nysbc.org wrote: Hi Phoebe - Cost-effective may not be the applicable word here, but the Microfluidizer works very well: http://www.microfluidicscorp.com/index.php?option=com_contentview=artic leid=19Itemid=76 This gadget runs on house compressed air (don't try to use a compressed air tank - you'll empty it in minutes). It's a bit noisy, but so is a sonicator. The Microfluidizer really shines with large volumes of lysate - like 1 L and up. If you're only processing 100-200 mL at a time, I think sonication is the best way to go. Hope that helps, Matt On 2/4/14 11:49 AM, Phoebe A. Rice wrote: Some time ago, there was a nice discussion of cost-effective, wimpy protein-friendly ways to break open E. coli. We're thinking about replacing an aging sonicator. If people have a favorite gizmo, could they repeat that advice? thank you, Phoebe Rice ++ Phoebe A. Rice Dept. of Biochemistry Molecular Biology The University of Chicago 773 834 1723; pr...@uchicago.edu http://bmb.bsd.uchicago.edu/Faculty_and_Research/ http://www.rsc.org/shop/books/2008/9780854042722.asp -- Matthew Franklin, Ph. D. Senior Scientist New York Structural Biology Center 89 Convent Avenue, New York, NY 10027 (212) 939-0660 ext. 9374
Re: [ccp4bb] no link_id
UNL is for unknown ligand, while UNX is for unknown atom/ion. And as Robbie said, UNK implies an amino acid of unknown identity. http://www.wwpdb.org/documentation/format32/sect4.html On 2/5/14, 1:38 PM, Robbie Joosten wrote: Just to add to Matthew's good advice, UNK is the standard residue name for unidentified amino acids. Using it for your compound may cause conflicts in other programs such as COOT. Cheers, Robbie Sent from my Windows Phone Van: Matthew Franklin Verzonden: 5-2-2014 17:50 Aan: CCP4BB@JISCMAIL.AC.UK Onderwerp: Re: [ccp4bb] no link_id Hi Ronnie - Refmac has done the job for you (or for your friend). The Refmac log file says that Refmac has created a file called .cif (the exact name varies). You should open up that file and inspect it; it will be Refmac's best guess about the geometry of that link. I usually specify only bond lengths and bond angles, leaving torsion angles undefined (i.e. deleting those restraints) unless I know it should be restrained to a certain angle. The next time you run Refmac, use the Library file input line in ccp4i to feed it this cif file; refinement should proceed OK after this. You can also append the contents of the cif file that Refmac made to your drug.cif file. Just cut and paste with a text editor. Don't forget that you need to give your ligand a real 3-letter code, not just UNK, before depositing it. So you might as well do it now. (Even if the structure will remain private, not deposited, I would still change the code.) - Matt On 2/5/14 10:59 AM, Ronnie wrote: I am asking this question on behalf of a friend: -Thanks! Hi All - I am working at small biotech. We have minimal budget for software, only CCP4 and Coot. We have a ligand that binds covalently to cysteine. I define the parameter file using Prodrg in CCP4 to get drug.cif file and then make a link in Coot between the Cys and the Drug. The problem is Refmac keeps complaining that there is no link_id for this bond: I am reading library. Please wait. mon_lib.cif WARNING : link: CYS--UNK is without link_id link will be created with covalent bond only. Bond= 1.610 WARNING : description of link:CYS-UNK is not in the dictionary link will be created with bond_length = 1.610 Where do I specify this link_id, and where can I find an example for the correct syntax? Is this done in mon_lib.cif or the drug.cif file?? The bond length, if I'm not mistaken should be ~1.82 A for distance between carbon and sulfur. Thanks for any help. -- Matthew Franklin, Ph. D. Senior Scientist New York Structural Biology Center 89 Convent Avenue, New York, NY 10027 (212) 939-0660 ext. 9374
[ccp4bb] Position in Chemical and systems biology in Oslo
Dear CCP4bb please forward to candidates that might be interested http://uio.easycruit.com/vacancy/1125923/70931?iso=gb best Preben -- J. Preben Morth, Ph.D Group Leader Membrane Transport Group Nordic EMBL Partnership Centre for Molecular Medicine Norway (NCMM) University of Oslo P.O.Box 1137 Blindern 0318 Oslo, Norway Tel: +47 2284 0794
[ccp4bb] Murnau conference on structural biology, Sept 10-13
Dear colleagues, We'd like to bring to your attention this year's 5th Murnau conference on structural biology. This biennial structural biology meeting takes place in Murnau, a picturesque village at one of the lakes south from Munich, Germany. The meeting this year will be held from Sept 10th to 13th and will have a focus on signal transduction, but it will also cover sessions on broader structural subjects - please visit the meeting web site for further information: http://www.murnauconference.de/2014/index.html http://www.murnauconference.de/2014/index.html We'd be happy to welcome many of you in September! With best wishes, Clemens Steegborn Oliver Einsle -- Don't miss the 5th Murnau conference on structural biology - Focus topic: Signal transduction September 10-13, 2014 Murnau am Staffelsee, Germany http://www.murnauconference.de/2014/index.html --- Prof. Dr. Clemens Steegborn University of Bayreuth Dept. Biochemistry, NW III Universitaetsstr. 30 95447 Bayreuth, Germany phone: ++49 0921 / 55 - 7831 fax: ++49 0921 / 55 - 7832 email: mailto:clemens.steegb...@uni-bayreuth.de clemens.steegb...@uni-bayreuth.de web: http://www.biochemie.uni-bayreuth.de/ www.biochemie.uni-bayreuth.de
[ccp4bb] Glycosylation in Gram-positive bacteria such as bacillus and paenibacillus
Colleagues, Many bacilli are known to have S-layer proteins outlining their surface and such proteins are known to be glycosylated. I have a few questions below: 1) What's the signature motif of such glycosylation and what pathway is involved? 2) Do you know any experts in this area, esp. in Paenibacillus? Thanks, Yong-Fu Li Elanco Animal Health
[ccp4bb] American Crystallographic Association - invitation to submit an abstract
We invite you to submit an abstract to the Wavelengths and Particles as Tools in Structural Analyses session scheduled May 26, 2014, Monday morning during the ACA meeting in Albuquerque. The focus will be on new applications and methods in X-ray and/or neutron crystallography that explore the use of non-conventional wavelengths or particle properties, or the wave-particle duality of photons and neutrons for structural analyses. Confirmed speakers are: John Helliwell, Dean Myles, B.- C. Wang, Wayne Hendrickson. Two abstracts will be chosen for oral presentation. Abstracts should be submitted by Friday, February 7. The organizers, B. C. Wang and Vivian Stojanoff
[ccp4bb] ACA 2014, Albuquerque: invitation to submit abstracts to Opportunities in structural biology with new light sources session
On behalf of Soichi Wakatsuki: Dear all We would like to invite you submit abstract to the structural biology session: Opportunities in structural biology with new light sources scheduled from 1:30PM to 5:00PM on May 28, 2014 as part of the American Crystallography Association meeting in Albuquerque. Our session will discuss recent results and future developments in structural biology using new light sources, in particular, but not limited to, XFELs. Three to four abstracts will be selected for aural presentations. Invited speakers include Vadim Cherezoff (Scripps Institute), Sebastien Boutet (LCLS, SLAC), Richard Neutze (University of Gothenburg), and Brenda Hogue (Arizona State University). The deadline for abstract submission has been extended to Feb 7th: http://www.amercrystalassn.org/2014-homepage Hope to see you in Albuquerque. Soichi Wakatsuki SLAC, Photon Science Stanford School of Medicine, Structural Biology Department Eaton Lattman Hauptman-Woodward Institute University of Buffalo SUNY
