[ccp4bb] Two Positions (Staff Scientist & Technician) in New York City
Hi Everyone, We have two open positions in the Bhabha/Ekiert labs at NYU School of Medicine in New York City (http://bhabhaekiertlab.org). Our labs use structure-driven approaches to study bacteria, parasites, and host-pathogen interactions. In addition to these potential research areas, the candidate will also participate in structure determination efforts as part of a collaborative project with the lab of David Baker (HHMI/University of Washington; https://www.bakerlab.org). 1) a PhD-level staff scientist position for a structural biologist 2) a BS/MS-level position for technician interested in structural biology (ideal for a recent grad looking to gain more experience before applying to graduate school). Please see the links before below for more information about the positions and how to apply. https://jobrxiv.org/job/nyu-school-of-medicine-27778-staff-scientist-structural-biology/ https://jobrxiv.org/job/nyu-school-of-medicine-27778-research-technician-structural-biology/ Best, Damian - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - Damian Ekiert Assistant Professor Skirball Institute | NYU School of Medicine Depts. Cell Biology and Microbiology damian.eki...@ekiertlab.org www.bhabhaekiertlab.org To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1 This message was issued to members of www.jiscmail.ac.uk/CCP4BB, a mailing list hosted by www.jiscmail.ac.uk, terms & conditions are available at https://www.jiscmail.ac.uk/policyandsecurity/
Re: [ccp4bb] Phaser Warning Message
Hi Silvia, Looking at the end of the log file, it looks like you have a very clear solution and are ready to go! This message means that a better solution from the Fast Translation Function was excluded because it failed the packing test (too many clashes with neighboring molecules). Sometimes this will prevent you from getting a usable solution altogether, and this message would clue you into that. In such a situation, you could prune flexible/non-conserved loops from your protein and try MR again with the truncated model. But in this case, your still managed to get a very clear solution, so I think you are ready to look at your maps and begin rebuilding and refinement. Good luck! Best, Damian > On Jun 16, 2020, at 4:47 PM, Napolitano Silvia > wrote: > > > Dear ccp44 Bulletin Board, > > I am trying to solve a structure by molecular replacement. > I am doing that by using PhaserMR (simple one-component interface). > > I tried out setting up different parameters (e.g.changing # of components, > #search, different space group search settings...), no matter what I try I > always get this warning message: > > -- > Advisory: The top solution from a FTF did not pack > -- > > -- > Advisory: The top solution from a TF rescoring did not pack > > (please, see log file attached). > > What does this Warning mean? I tried to look it up online but, unfortunately, > I could not find the answer I was looking for. > > Please, let me know if you need more detailed information. > > Many thanks in advance for your help and your time! > > Best Regards > > Silvia > > Silvia Napolitano > ETH Zurich > Institute of Molecular Biology and Biophysics > Otto-Stern-Weg 5, HPK E14 > CH–8093 Zurich > Phone: +41 44 6332482 > > To unsubscribe from the CCP4BB list, click the following link: > https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1 > To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1 This message was issued to members of www.jiscmail.ac.uk/CCP4BB, a mailing list hosted by www.jiscmail.ac.uk, terms & conditions are available at https://www.jiscmail.ac.uk/policyandsecurity/
Re: [ccp4bb] Crystal Structures as Snapshots
Along the lines of Roger's second point, there was a very nice paper a few years back that found very good agreement between the conformational ensemble sampled by ubiquitin in solution (by NMR) with the ensemble of conformations observed in a large number of crystal structures: Lange OF, Lakomek NA, Farès C, Schröder GF, Walter KF, Becker S, Meiler J, Grubmüller H, Griesinger C, de Groot BL. Recognition dynamics up to microseconds revealed from an RDC-derived ubiquitin ensemble in solution. Science. 2008 Jun 13;320(5882):1471-5. PubMed PMID: 18556554. Best, Damian Ekiert On Feb 10, 2012, at 12:50 PM, Roger Rowlett wrote: I believe the most justifiable assumption one can make is that crystal structures are likely to represent the least soluble conformations of a protein under the conditions of crystallization (which might be a broad range of conditions, including physiological). This can be quite vexing if you are studying an allosteric protein and one of the two conformations is typically much less soluble than the other. BTDT. I'm sure others have had the same experience. Having said that, the solvent content of protein crystals (which is close to that of cellular conditions), the observation of enzymatic activity in many protein crystals, and the *general* concordance of XRD and NMR structures of proteins (when both have been determined) leads one to believe that XRD structures are likely representative of physiologically relevant conformations. Cheers, ___ Roger S. Rowlett Gordon Dorothy Kline Professor Department of Chemistry Colgate University 13 Oak Drive Hamilton, NY 13346 tel: (315)-228-7245 ofc: (315)-228-7395 fax: (315)-228-7935 email: rrowl...@colgate.edu On 2/10/2012 3:34 PM, Nat Echols wrote: On Fri, Feb 10, 2012 at 12:29 PM, James Stroud xtald...@gmail.com wrote: How could they not be snapshots of conformations adopted in solution? Packing billions of copies of an irregularly-shaped protein into a compact lattice and freezing it to 100K isn't necessarily representative of solution, especially when your solution contains non-physiological amounts of salt and various organics (and possibly non-physiological pH too). -Nat
Re: [ccp4bb] Coot pudding? (a.k.a N-linked carbohydrate addition)
For an ideal glycan, you could used a model from a high resolution structure, or something that has been energy minimized, etc. Mostly I find this helps in getting the sugars in about the right place (keeping bond lengths and angles reasonable). I perhaps could stand to fiddle more and maybe I'll look through the updated documentation. Last I tried, the Asn-NAG1 linkage wasn't enforced, with would allow the whole glycan to slide down into the protein, or off into space. I am typically building relatively small glycans (2-5 residues) into ~3A data, so the density itself doesn't keep things in place very well. Regarding BMA vs MAN: When I have tried to used BMA in REFMAC, it doesn't seem to recognize it and requires a library file. But if you use MAN, it adds a MODRES record to the header, enforcing beta-mannose geometry. Not sure if this is just a REFMAC version issue or what. Best, Damian Paul Emsley wrote: If I can chip into this somewhat sacrilegiously-named thread 1) I *would* use real-space refinement :), specifically Sphere Refinement. You can dial down the density weight if needed, of course. 2) the documentation on refining carbohydrates in Coot has recently been updated http://www.biop.ox.ac.uk/coot/doc/coot/Refining-Carbohydrates.html 3) Coot does not (yet) correct chiral centre inversions in glycosidic linkages on refinement 4) or delete the O1s :) Paul. Robbie Joosten wrote: Dear Steve, I would also use Damian's approach, but the sequence of the core should be NAG-NAG-BMA-(MAN)2. This is improtant because the correct stereochemistry restraints for beta-mannose can only be applied when you call the residue BMA. Building carbohydrates also comes with special validation requirements. PDB-care and CARP are both very usefull. Unfortunately, the service is currently down (http://www.dkfz.de/spec/glycosciences.de). Just make sure the links between your carbs are correct and, please, remove the O1 atoms when needed ;) Cheers, Robbie Joosten Date: Mon, 21 Dec 2009 17:48:31 -0800 From: dceki...@scripps.edu Subject: Re: [ccp4bb] Coot pudding? (a.k.a N-linked carbohydrate addition) To: CCP4BB@JISCMAIL.AC.UK Steve, My general strategy is to start with an ideal glycan (an Asn linked to NAG-NAG-(MAN)3 ) and superimpose the Asn on the residue from my protein. Then you can move the whole glycan as a rigid body until the Asn and first NAG are roughly positioned. Then you can tweak any sugars further out on the chain to get them to fit. Unless you have really great density, usually it is best to avoid real pace refine zone. Better to fit the sugars using the manual rigid body fitting tools, do the best you can, then REFMAC usually brings them in OK. I have some models that I could send you if you need them. Best, Damian Ekiert Soisson, Stephen M wrote: Hi everyone- I was searching for some information on what might be the best way to add N-linked sugars in coot, and Google has let me down. Searching adding sugars in coot returns a very nice recipe for Coot Pudding. ***_Recipe for_/ Coot//__/_ Pudding - American_/ Coots/* It has plenty of fat,/ sugar/, and starch, and probably some calcium from the milk.* ...* The/ coots/ will not tolerate/ adding/ eggs in any form, so this is an egg* ...* ///_www.beaky_//_*coot*.com/pudding.html_/// / -/ _Similar_ // I did not know that coots had such an aversion to eggs. :) Anyway, would anyone have any top tips on adding N-linked sugars using coot? I can import the NAG monomers, but linking them up to the protein seems non-trivial Many thanks in advance, Steve Notice: This e-mail message, together with any attachments, contains information of Merck Co., Inc. (One Merck Drive, Whitehouse Station, New Jersey, USA 08889), and/or its affiliates Direct contact information for affiliates is available at http://www.merck.com/contact/contacts.html) that may be confidential, proprietary copyrighted and/or legally privileged. It is intended solely for the use of the individual or entity named on this message. If you are not the intended recipient, and have received this message in error, please notify us immediately by reply e-mail and then delete it from your system.
Re: [ccp4bb] Coot pudding? (a.k.a N-linked carbohydrate addition)
Pierre, For what its worth, I just tried using Real Space Refine Zone on something I am working on now (2.8A resolution, in coot 0.5.2, build 1691). With the LINKR record generated by REFMAC5, my glycan slithers down into the protein density. Similarly, using Regularize Zone does not enforce the Asn-NAG linkage. I also tried replacing the R from LINKR with a space, but it doesn't make a difference. My records read: 1 2 3 4 5 6 7 8 12345678901234567890123456789012345678901234567890123456789012345678901234567890 LINKRC1 NAG A 401 ND2 ASN A 21NAG-ASN LINK C1 NAG A 401 ND2 ASN A 21NAG-ASN However, I checked the page that Paul mentioned in an earlier email, and it states: LINK and LNKR cards are not used to determine the geometry of the restraints. If other people don't have this problem, I would love to know how to get this to work because, admittedly, placing everything using other tools is a pain. Best, Damian Pierre Rizkallah wrote: Hi Damian, I have used both Coot and Refmac for glycan modelling/refinement in the past, last time was 2 or 3 years ago. They both behaved correctly for me. To get the correct behaviour, you must have the correct LINK cards in the header. At the time, the Chemical ID BMA did not exist in the PDB. There was only MAN. Presumably, the PDB had originally decided that the saccahride linkage is a bit like the peptide bond, where it can be one sort or another, so you have to specify it explicitly. But convenience, or avoiding confusion, must have been the reason for the recent creation of BMA. If Coot/Refmac do not have it in their library, well you can add it easily into the correct folder on your disk, by editing the MAN-b-D.cif entry and calling it BMA.cif. Garib told me, when I was grappling with this problem, that Refmac works out from the coordinates which form of the saccahride linkage it is and imposes the correct restraints. Admittedly, there must be room for confusion if modelling is not accurate enough. So BMA is better all round. The branching of the glycan tree does have a good template in PDB entry 1LGB, first deposited in 1994. It has been reviewed to include BMA earlier this year. The branching pattern is correct, so care must be taken to enter the correct numbers in the LINK cards if you are to reproduce a similar branching pattern in your glycan. Incidentally, if you search the PDB for chemical id BMA, there are apparently 36 entries. 1LGB was not among them, and I don't know why. I hope this is useful. Good Luck. Pierre Rizkallah Dr. Pierre Rizkallah, Senior Lecturer in Structural Biology, WHRI, School of Medicine, Cardiff University, Heath Park, CF14 4XN email: rizkall...@cf.ac.uk phone + 44 29 2074 2248 Damian Ekiert dceki...@scripps.edu 22/12/09 5:46 PM For an ideal glycan, you could used a model from a high resolution structure, or something that has been energy minimized, etc. Mostly I find this helps in getting the sugars in about the right place (keeping bond lengths and angles reasonable). I perhaps could stand to fiddle more and maybe I'll look through the updated documentation. Last I tried, the Asn-NAG1 linkage wasn't enforced, with would allow the whole glycan to slide down into the protein, or off into space. I am typically building relatively small glycans (2-5 residues) into ~3A data, so the density itself doesn't keep things in place very well. Regarding BMA vs MAN: When I have tried to used BMA in REFMAC, it doesn't seem to recognize it and requires a library file. But if you use MAN, it adds a MODRES record to the header, enforcing beta-mannose geometry. Not sure if this is just a REFMAC version issue or what. Best, Damian Paul Emsley wrote: If I can chip into this somewhat sacrilegiously-named thread 1) I *would* use real-space refinement :), specifically Sphere Refinement. You can dial down the density weight if needed, of course. 2) the documentation on refining carbohydrates in Coot has recently been updated http://www.biop.ox.ac.uk/coot/doc/coot/Refining-Carbohydrates.html 3) Coot does not (yet) correct chiral centre inversions in glycosidic linkages on refinement 4) or delete the O1s :) Paul. Robbie Joosten wrote: Dear Steve, I would also use Damian's approach, but the sequence of the core should be NAG-NAG-BMA-(MAN)2. This is improtant because the correct stereochemistry restraints for beta-mannose can only be applied when you call the residue BMA. Building carbohydrates also comes with special validation requirements. PDB-care and CARP are both very usefull. Unfortunately, the service is currently down (http://www.dkfz.de/spec/glycosciences.de). Just make sure the links between your carbs are correct and, please, remove the O1
Re: [ccp4bb] Coot pudding? (a.k.a N-linked carbohydrate addition)
Steve, My general strategy is to start with an ideal glycan (an Asn linked to NAG-NAG-(MAN)3 ) and superimpose the Asn on the residue from my protein. Then you can move the whole glycan as a rigid body until the Asn and first NAG are roughly positioned. Then you can tweak any sugars further out on the chain to get them to fit. Unless you have really great density, usually it is best to avoid real pace refine zone. Better to fit the sugars using the manual rigid body fitting tools, do the best you can, then REFMAC usually brings them in OK. I have some models that I could send you if you need them. Best, Damian Ekiert Soisson, Stephen M wrote: Hi everyone- I was searching for some information on what might be the best way to add N-linked sugars in coot, and Google has let me down. Searching adding sugars in coot returns a very nice recipe for Coot Pudding. ***_Recipe for_/ Coot//__/_ Pudding - American_/ Coots/* http://www.beakycoot.com/pudding.html It has plenty of fat,/ sugar/, and starch, and probably some calcium from the milk.* ...* The/ coots/ will not tolerate/ adding/ eggs in any form, so this is an egg* ...* ///_www.beaky_//_*coot*.com/pudding.html_/// file://www.beakycoot.com/pudding.html/ -/ _Similar_ file:///search?hl=enq=related:www.beakycoot.com/pudding.html+adding+sugars+in+cootsa=Xei=r6YhS-T1F4PQlAeP2ZT8CQved=0CAgQHzAA// I did not know that coots had such an aversion to eggs. :) Anyway, would anyone have any top tips on adding N-linked sugars using coot? I can import the NAG monomers, but linking them up to the protein seems non-trivial Many thanks in advance, Steve Notice: This e-mail message, together with any attachments, contains information of Merck Co., Inc. (One Merck Drive, Whitehouse Station, New Jersey, USA 08889), and/or its affiliates Direct contact information for affiliates is available at http://www.merck.com/contact/contacts.html) that may be confidential, proprietary copyrighted and/or legally privileged. It is intended solely for the use of the individual or entity named on this message. If you are not the intended recipient, and have received this message in error, please notify us immediately by reply e-mail and then delete it from your system.
