Re: [ccp4bb] how to create a figure of a blob
I would suggest the same Best __ Rafael Marques da Silva PhD Student – Structural Biology University of Leicester Mestrando em Física Biomolecular Universidade de São Paulo Bacharel em Ciências Biológicas Universidade Federal de São Carlos phone: +55 16 99766-0021 "A sorte acompanha uma mente bem treinada" De: CCP4 bulletin board em nome de Jon Cooper <488a26d62010-dmarc-requ...@jiscmail.ac.uk> Enviado: sexta-feira, 19 de julho de 2024 14:56 Para: CCP4BB@JISCMAIL.AC.UK Assunto: Re: [ccp4bb] how to create a figure of a blob I am afraid I don't have knowledge of pymol options but can't one make a separate pdb file with a dummy atom at the desired position so that the map is centred there and then turn that molecule/atom off for the rendering step with the main molecule left on. Probably I completely misunderstood. CueMol does this, too. Best wishes, Jon Cooper. jon.b.coo...@protonmail.com Sent from Proton Mail Android Original Message On 18/07/2024 15:33, Andrea Smith wrote: Hi, I have a “green blob” in my map and I want to create a picture of it. What is the best option to do this? Normally, to make figures of maps, I use pymol's "display ccp4 maps", but pymol shows maps around “site”. So if I don't fit anything into my blob, I can't create a “site” and display the map. Is printscreen from Coot my only option? Thank you for suggestions, Andrea To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=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] Unknown density
Dear John Ethanol would be my first guess, Best wishes __ Rafael Marques da Silva PhD Student – Structural Biology University of Leicester Mestrando em Física Biomolecular Universidade de São Paulo Bacharel em Ciências Biológicas Universidade Federal de São Carlos phone: +55 16 99766-0021 "A sorte acompanha uma mente bem treinada" De: CCP4 bulletin board em nome de John Smith Enviado: quinta-feira, 11 de julho de 2024 09:35 Para: CCP4BB@JISCMAIL.AC.UK Assunto: [ccp4bb] Unknown density Dear all! We have solved a structure at 2.2 A resolution. We see this interesting density near a Tyrosine. It is not fully aligned, like pi stacking. Otherwise, negatively charged or polar residues surround the smaller 'head'. Does anybody have any idea? Thanks for the help! Best John To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=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/
[ccp4bb] RES: [ccp4bb] message yesterday on the CCP4BB
It reminds me of that movie, the wishmaster, a thrash one. “Be careful about what you desire” Rafael Marques da Silva PhD Student – Structural Biology University of Leicester Mestre em Física Biomolecular Universidade de São Paulo Bacharel em Ciências Biológicas Universidade Federal de São Carlos phone: +55 16 99766-0021 "A sorte acompanha uma mente bem treinada" De: Tim Gruene<mailto:tim.gru...@univie.ac.at> Enviado:quarta-feira, 10 de julho de 2024 11:02 Para: CCP4BB@JISCMAIL.AC.UK<mailto:CCP4BB@JISCMAIL.AC.UK> Assunto: Re: [ccp4bb] message yesterday on the CCP4BB On Wed, 10 Jul 2024 08:46:06 + "Hough, Michael (DLSLtd,RAL,LSCI)" <69715b1ac6c0-dmarc-requ...@jiscmail.ac.uk> wrote: > or the magic porridge pot... > > -Original Message- > From: CCP4 bulletin board On Behalf Of Harry > Powell Sent: Wednesday, July 10, 2024 9:43 AM > To: CCP4BB@JISCMAIL.AC.UK > Subject: Re: [ccp4bb] message yesterday on the CCP4BB > > Hi > > Pandora's box comes to mind... It's called 'internet' Cheers, Tim -- -- Tim Gruene Head of the Core Facility Crystal Structure Analysis Faculty of Chemistry University of Vienna Phone: +43-1-4277-70202 https://ccsa.univie.ac.at GPG Key ID = A46BEE1A To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=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/ To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=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/
[ccp4bb] RES: [ccp4bb] Crystal optimization
Hi, It seems to my eyes that you have multiple nucleation events happening simultaneously. I would recommend you to try crystallization under 10 or 4 ºC Best of luck Rafael Marques da Silva PhD Student – Structural Biology University of Leicester Mestre em Física Biomolecular Universidade de São Paulo Bacharel em Ciências Biológicas Universidade Federal de São Carlos phone: +55 16 99766-0021 "A sorte acompanha uma mente bem treinada" De: CCP4 bulletin board em nome de 白雪慧 Enviado: Friday, May 31, 2024 3:32:52 AM Para: CCP4BB@JISCMAIL.AC.UK Assunto: [ccp4bb] Crystal optimization Thank you very much for your suggestions. I have a question. My crystal grows microcrystals under multiple conditions, as shown in the figure. After orthogonal optimization of the precipitant and pH, the crystal growth is still very small and difficult to obtain diffraction. What is the method to optimize and increase the crystal size in this situation? To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=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/
[ccp4bb] RES: [ccp4bb] What could these crystals be?
It seems to me that your drop has dried. By the pic, I would not say these are crystals Best Rafael Marques da Silva PhD Student – Structural Biology University of Leicester Mestre em Física Biomolecular Universidade de São Paulo Bacharel em Ciências Biológicas Universidade Federal de São Carlos phone: +55 16 99766-0021 "A sorte acompanha uma mente bem treinada" De: CCP4 bulletin board em nome de 张慧欣 Enviado: Monday, May 13, 2024 3:15:09 PM Para: CCP4BB@JISCMAIL.AC.UK Assunto: [ccp4bb] What could these crystals be? Hi all We have been trying with no success to crystalize a protein. Recently we got these strange shape "crystals". They are hard and flat . Any ideas as to what could cause this? The crystallization conditions is:0.06M Divalents 0.1M BICINE 0.1M Tris pH8.5 20% Ethylene glycol 10% PEG8000 To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=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] Fwd: AlphaFold3 Transparency and Reproducibility
I tried it yesterday and I was really shocked by how fast it is. When I was preparing to submit my second job, the first one was already finished, which made me think that I was definitely doing something wrong. Probably I was...Best wishes Rafael Marques On 13 May 2024 09:53, Harry Powell <193323b1e616-dmarc-requ...@jiscmail.ac.uk> wrote:Hi folks This arrived in my inbox this morning, and I believe that it may provoke some discussion… Wikipedia tells me: στέργει γὰρ οὐδεὶς ἄγγελον κακῶν ἐπῶν Best wishes! Harry > From: Stephanie Wankowicz > Sent: Saturday, May 11, 2024 3:31 PM > To: James Fraser ; Pedro Beltrao ; Benjamin Cravatt ; Roland Dunbrack ; Anthony Gitter ; Kresten Lindorff-Larsen ; Sergey Ovchinnikov ; Polizzi, Nicholas F. ; Brian Shoichet > Subject: AlphaFold3 Transparency and Reproducibility > > > This email from mullane.stepha...@gmail.com originates from outside Imperial. Do not click on links and attachments unless you recognise the sender. If you trust the sender, add them to your safe senders list to disable email stamping for this address. > > Hello, > > > As many of you, we were incredibly disappointed with the lack of code or even executables accompanying the publication of AlphaFold3 in Nature. AlphaFold3 was released without the means to test and use the software in a high-throughput manner. This does not align with the principles of scientific research, which rely on the ability of the community to evaluate, use, and build upon existing work. > > > > We have written a letter, which will be posted on Zenodo and submitted as a Letter to the Editor in the coming days. > > > > Please see the entire letter here. If you want to endorse this letter, please fill out your name, affiliation, and email in the form. > > > > Additionally, a PDF version of the letter can be found here . > > > > Thank you, > > > > Stephanie Wankowicz, UCSF > > Pedro Beltrao, ETH > Benjamin Cravatt, Scripps > Roland Dunbrack, FCCC > Anthony Gitter, UW Madison > Kresten Lindorff-Larsen, Copenhagen > Sergey Ovchinnikov, MIT > Nicholas Polizzi, DFCI/HMS > Brian Shoichet, UCSF > James Fraser, UCSF > > To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=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/ To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1
[ccp4bb] RES: [ccp4bb] Rwork and Rfree the same?
Sorry for jumping into the post, but I would like the community’s opinion about completeness, once this topic was raised here. What could be considered reasonable? Recently I have seen a 65% completeness Crystal structure and, surprisingly, the electron density map was not that bad for a > 3.2 A structure. How such a nice map could have been calculated with such poor parameters? I could only think of anisotropy. Best Rafael Marques da Silva PhD Student – Structural Biology University of Leicester Mestre em Física Biomolecular Universidade de São Paulo Bacharel em Ciências Biológicas Universidade Federal de São Carlos phone: +55 16 99766-0021 "A sorte acompanha uma mente bem treinada" De: CCP4 bulletin board em nome de Paul Adams Enviado: Wednesday, February 28, 2024 2:58:16 PM Para: CCP4BB@JISCMAIL.AC.UK Assunto: Re: [ccp4bb] Rwork and Rfree the same? By setting wxc (weight of the X-ray term) to 0.1 there is good chance that the refinement is dominated by the geometry term and the model isn’t really seeing the effect of the X-ray data. I suspect this would result in R-factors that are similar. Why they are so low is less clear, but as others have pointed out 38% completeness is a problem. It would be good to check if that is 38% overall, or just very incomplete in the higher resolution shells. If it is complete at lower resolution you might be able to do something with the dataset, but not using the default parameterization in refinement programs - you’ll need to apply constraints and additional restraints if you can, and look at the weighting (by modifying wxc_scale, not wxc). There is a Phenix mailing list you might want to use as well (I assume you are using phenix.refine): https://phenix-online.org/mailman/listinfo/phenixbb On Feb 28, 2024, at 8:21 AM, Justin Cruite wrote: Thanks everyone, I agree, 18.4% Rwork and Rfree is too good to be true for a 3.4 Å dataset. The data was processed using autoProc and the staranisano mtz was used for MR. The completeness is only 38%. It could be that the Rfree and Rwork reflection sets are small because of this? What is the best way to check the number of reflections used for Rwork and Rfree? Is this dataset usable at all? Thanks! Justin On Wed, Feb 28, 2024 at 10:21 AM nicfoos mailto:nicf...@embl.fr>> wrote: Hello Justin, There is something weird in your results. You mention Rwork/Rfree of 0.1837. This means a pretty good refinement and also is very unusual to be obtain for a resolution of 3.37. Additionally you should not have Rfree = Rwork. I suspect something wrong with you Rfree reflections sets. What size is it ? Is your dataset complet ? How did you cut the res. ? I hope this may help you. Nicolas On 2024-02-28 16:10, Justin Cruite wrote: > Hi everyone, > > What does it mean if your Rwork and Rfree are exactly the same? > > I solved a 3.37 Å structure with Phaser-MR and immediately ran 10 > cycles of refinement with wxc = 0.1. Everything else at default. The > Rwork and Rfree are both 0.1837. Is this bad? > > Thank you! > > Justin > > - > > To unsubscribe from the CCP4BB list, click the following link: > https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 -- Paul Adams (he/him/his) Associate Laboratory Director for Biosciences, LBL (https://biosciences.lbl.gov) Principal Investigator, Computational Crystallography Initiative, LBL (http://cci.lbl.gov) Vice President for Technology, the Joint BioEnergy Institute (http://www.jbei.org) Principal Investigator, ALS-ENABLE, Advanced Light Source (http://als-enable.lbl.gov) Laboratory Research Manager, ENIGMA Science Focus Area (http://enigma.lbl.gov) Adjunct Professor, Department of Bioengineering, UC Berkeley (http://bioeng.berkeley.edu) Member of the Graduate Group in Comparative Biochemistry, UC Berkeley (http://compbiochem.berkeley.edu) Building 91, Room 410 Building 978, Room 4126 Tel: 1-510-486-4225 http://cci.lbl.gov/paul ORCID: -0001-9333-8219 Lawrence Berkeley Laboratory 1 Cyclotron Road BLDG 91R0183 Berkeley, CA 94720, USA. Executive Assistant: Michael Espinosa [ meespin...@lbl.gov ][ 1-510-333-6788 ] Phenix Consortium: Ashley Dawn [ ashleyd...@lbl.gov ][ 1-510-486-5455 ] -- To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 This message was issued to member
[ccp4bb] RES: [ccp4bb] Crystals with DNA
Hi Careina. The top two things that come to my mind is that you need to screen for different crystallization conditions and do some seeding. If your Crystal conditions does not diffract at all, probably it won’t. Have you tried to optimize your condintion changing the pH or precipitant concentration? Best wishes Rafael Marques da Silva PhD Student – Structural Biology University of Leicester Mestre em Física Biomolecular Universidade de São Paulo Bacharel em Ciências Biológicas Universidade Federal de São Carlos phone: +55 16 99766-0021 "A sorte acompanha uma mente bem treinada" De: CCP4 bulletin board em nome de careinaedgo...@yahoo.com <02531c126adf-dmarc-requ...@jiscmail.ac.uk> Enviado: Thursday, February 8, 2024 8:25:25 AM Para: CCP4BB@JISCMAIL.AC.UK Assunto: [ccp4bb] Crystals with DNA Hello all. I am struggling to get defracting crystals with a protein DNA complex. The crystals are plentiful but they do not diffract. I am going back to the grind stone and relookong at my DNA sequence. Is there any wisdom you could give me with regards to what works best with DNA in crystals? >From my reading it seems if the length is a multiple of 7 (for B DNA) and >blunt ended, it will stretch over the length of the crystal and improve >crystalisability. But if you want crystals that diffract better, you will need >to play with length and even making it only one base longer or shorter can >make a difference, even changing the morphology of the crystal? Longer is >better than shorter, and overhangs are good for improving diffraction? >Presumably because they stabilize contacts? It is expensive to synthesize a >while bunch of sequences so I need to be strategic in my choice. Would >appreciate any advice. Thank you Careina. To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=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/
[ccp4bb] RES: [ccp4bb] What could these crystals be?
Hi all. Just wanted to add a bit of doubt to the topic. Is it possible that they are salt crystals? I don’t know, but some spots are too intense and far from each other to me. But maybe I just need better glasses. Best wishes Rafael Marques da Silva PhD Student – Structural Biology University of Leicester Mestre em Física Biomolecular Universidade de São Paulo Bacharel em Ciências Biológicas Universidade Federal de São Carlos phone: +55 16 99766-0021 "A sorte acompanha uma mente bem treinada" De: careinaedgo...@yahoo.com<mailto:02531c126adf-dmarc-requ...@jiscmail.ac.uk> Enviado:sexta-feira, 24 de novembro de 2023 09:00 Para: CCP4BB@JISCMAIL.AC.UK<mailto:CCP4BB@JISCMAIL.AC.UK> Assunto: Re: [ccp4bb] What could these crystals be? I wanted to thank everyone for their suggestions and ideas regarding the eyeball shaped crystals that I got at the beginning of the month. I can confirm that these crystals do indeed contain protein and DNA which is good news. I have tried a number of different buffers and salts since then. I have also tried seeding but nothing I have tried has changed the morphology of the crystals. They remain thin, flat and eyeball/pumpkin seed shaped. It is not difficult to make the crystals, they form quite easily under quite a few conditions. The difficulty is in the diffraction. By changing the buffer conditions we can now see some very weak diffraction (previously there was no diffraction at all). Are there any suggestions as to how to improve diffraction of these crystals? I did try different cryoprotectants, parabar, glycerol, PEG but no difference. I think perhaps the problem is heterogeneity considering my sample contains both protein and DNA. Any suggestions or thoughts are welcome. Thank you Careina On Wednesday, November 8, 2023 at 06:18:46 PM GMT+2, careinaedgo...@yahoo.com <02531c126adf-dmarc-requ...@jiscmail.ac.uk> wrote: The reservior solution is 0.2 M NaCl2, 0.1 M HEPES pH 7.5, and 25% 3350 PEG Protein buffer is 300 mM NaCl, 20 mM HEPES pH 7.5, 3mM TCEP The drop contains 1.5ul protein-DNA complex and 1.5 ul reservior solution On Wednesday, November 8, 2023 at 05:12:25 PM GMT+2, stephen.c...@rc-harwell.ac.uk wrote: Hi Careina, Without knowing what's in your protein buffer or crystallisation condition it's hard to comment. Best wishes, Steve Dr Stephen Carr Research Complex at Harwell (RCaH) Rutherford Appleton Laboratory Harwell Oxford Didcot Oxon OX11 0FA United Kingdom Email stephen.c...@rc-harwell.ac.uk tel 01235 567717 From: CCP4 bulletin board on behalf of careinaedgo...@yahoo.com <02531c126adf-dmarc-requ...@jiscmail.ac.uk> Sent: 08 November 2023 15:00 To: ccp4bb Subject: [ccp4bb] What could these crystals be? Hi all We have been trying with no success to crystalize a protein. Recently we got these strange shape "crystals". They are hard and flat but they do not diffract at all. Any ideas as to what could cause this? Careina To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 This email and any attachments may contain confidential, copyrightand or privileged material, and are for the use of the intended addressee only. If you are not the intended addressee or an authorized 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 this email. Any views or opinions presented are solely those of the author and do not necessarily represent those of the Research Complex at Harwell. There is no guarantee that this email 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. We use an electronic filing system. Please send electronic versions of documents, unless paper is specifically requested. This email may have a protective marking, for an explanation, please see: http://www.mrc.ac.uk/About/informationandstandards/documentmarking/index.htm. To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=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/
[ccp4bb] RES: [ccp4bb] low resolution data refinement
Hi, As the others have already mentioned, it is hard to get any decent MR solution at this resolution. The best thing to do would be to set up more crystal plates and harvest crystals growing at different conditions. You also have not mentioned where you diffracted your crystals. If you did it using an in-house system you should consider a synchrotron light source. Best wishes Rafael Marques da Silva PhD Student – Structural Biology University of Leicester Mestre em Física Biomolecular Universidade de São Paulo Bacharel em Ciências Biológicas Universidade Federal de São Carlos phone: +55 16 99766-0021 "A sorte acompanha uma mente bem treinada" De: Paul Emsley<mailto:pems...@mrc-lmb.cam.ac.uk> Enviado:terça-feira, 21 de novembro de 2023 12:08 Para: CCP4BB@JISCMAIL.AC.UK<mailto:CCP4BB@JISCMAIL.AC.UK> Assunto: Re: [ccp4bb] low resolution data refinement On 21/11/2023 12:02, Yahui Liu wrote: > > Dear all, > I got a protein crystal dataset of 4.3 A and would like to some the > structure with MR. > Now I am suffering with the refinement. Just for the record, then intention of refinement is not to make you suffer. Paul. To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 This message was issued to members of www.jiscmail.ac.uk/CCP4BB<http://www.jiscmail.ac.uk/CCP4BB>, a mailing list hosted by www.jiscmail.ac.uk<http://www.jiscmail.ac.uk>, terms & conditions are available at https://www.jiscmail.ac.uk/policyandsecurity/ To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=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/
[ccp4bb] RES: [ccp4bb] What could these crystals be?
Hi Careina Initially I was sure they were protein crystals, principally because you only have NaCl, Hepes and TCEP in your buffer. And then you said they are hard. If you try to press them against the bottom of the well, they should easily break if they are made of proteins. Once they did not diffract, I think it is something you should try with the remaining ones. Alternatively, if you have a source of UV light coupled to a camera, they should “light up”. I wish you the best of luck. Rafael Marques da Silva PhD Student – Structural Biology University of Leicester Mestre em Física Biomolecular Universidade de São Paulo Bacharel em Ciências Biológicas Universidade Federal de São Carlos phone: +55 16 99766-0021 "A sorte acompanha uma mente bem treinada" De: Jurgen Bosch<mailto:jxb...@case.edu> Enviado:quarta-feira, 8 de novembro de 2023 17:08 Para: CCP4BB@JISCMAIL.AC.UK<mailto:CCP4BB@JISCMAIL.AC.UK> Assunto: Re: [ccp4bb] What could these crystals be? You could also run them on a gel if you don’t want to shot with canons on small birds. Jürgen On Nov 8, 2023, at 11:33 AM, M T wrote: Dear Careina, If you have easy access to mass spectrometry, you can try to fish/rince your « pumpkin seeds » and send them to mass to try to identify what is inside to see if it needs optimization or not. Best. Le 8 nov. 2023 à 16:03, 02531c126adf-dmarc-requ...@jiscmail.ac.uk a écrit : Hi all We have been trying with no success to crystalize a protein. Recently we got these strange shape "crystals". They are hard and flat but they do not diffract at all. Any ideas as to what could cause this? Careina To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=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/
[ccp4bb] RES: [ccp4bb] About model building
Hi Sam. If you still have any of your crystals or any protein solution left in the well you harvested your crystals, I would run a MS/MS with them. Next step would be to run AF with your known chain A and your best Mass Spec hit (s), and use the resulting model for MR. Good luck Rafael Marques da Silva PhD Student – Structural Biology University of Leicester Mestre em Física Biomolecular Universidade de São Paulo Bacharel em Ciências Biológicas Universidade Federal de São Carlos phone: +55 16 99766-0021 "A sorte acompanha uma mente bem treinada" De: CCP4 bulletin board em nome de Boaz Shaanan Enviado: Monday, November 6, 2023 2:41:43 PM Para: CCP4BB@JISCMAIL.AC.UK Assunto: Re: [ccp4bb] About model building Hi, If you still have crystals left, you could soak crystals with KI3 and collect data at Cu wavelength for SAD phasing, which could help you to resolve the missing piece. Maybe. Cheers, Boaz Boaz Shaanan, Ph.D. Dept. of Life Sciences Ben Gurion University Beer Sheva, Israel On Nov 4, 2023 10:04, Sam Tang wrote: Dear community, I am solving the structure of a complex between proteins A and B, where A is a protein with known homologs and B is a novel protein isolated from plant. The diffraction data was at 1.9 Ang collected in-house, indexed to P321. Using A as the search model, we have got a reasonable solution where, after one round of refinement, the A chain fits the map pretty well. What's left was to extend the termini and fit a few rotamers. For protein B (B chain) I have tried the web version of ARP/wARP but the outcome was not really good. The model was not successfully built as indicated by low model completeness and score. The tricky thing may be that we do not have the complete sequence information of this protein B in-hand. (The other way round, we more or less wish to rely on the high resolution data to confirm its sequence.) What approach would you then recommend to build the B chain in this scenario? Thanks in advance and best regards, Sam To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=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/
[ccp4bb] [OFF TOPIC] On Ni-NTA resin TEV cleavage
Hi everyone, I have been looking on this bb and other websites as well but I could not find a veredict. We are suspecting that when I elute my sample from my Ni-NTA column, the imidazole concentration (250 mM) is making it to precipitate. Once my sample has a cleavable TEV site, I was planning to incubate my loaded resin overnight with TEV and get my sample back simply using my lysis buffer. And here lies the problem. Most of the TEVs are kept in EDTA and DTT and I wonder if they are essential for its protease activity or if I could use another reducing agent more compatible with my resin (or maybe do not add both). I saw that someone did not have EDTA and used b-mercap. instead of DTT. May I have your comments if you guys already faced a similar situation? Best wishes __ Rafael Marques da Silva PhD Student – Structural Biology University of Leicester Mestrando em Física Biomolecular Universidade de São Paulo Bacharel em Ciências Biológicas Universidade Federal de São Carlos phone: +55 16 99766-0021 "A sorte acompanha uma mente bem treinada" To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=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/
[ccp4bb] Updated pbd metrics and other small things
Hi folks, I was wondering if someone has the link of that page where we could check the graphs “resolution vs R-value” and “r-value vs r-free”. The ones that I found are outdated by let’s say 20 years. Off topic. If you guys have some examples of diffraction patterns of Neutron and Electron scattering, could you please send me? I am preparing a presentation and a single google search has been really misleading. Best wishes Rafael Marques da Silva PhD Student – Structural Biology University of Leicester Mestre em Física Biomolecular Universidade de São Paulo Bacharel em Ciências Biológicas Universidade Federal de São Carlos phone: +55 16 99766-0021 "A sorte acompanha uma mente bem treinada" To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=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/
[ccp4bb] RES: [ccp4bb] Is this Spam? This looks like Spam.
The same happened to me. I had to subscribe again cause I was not receiving ccp4 mails anymore. Best Rafael Marques da Silva PhD Student – Structural Biology University of Leicester Mestre em Física Biomolecular Universidade de São Paulo Bacharel em Ciências Biológicas Universidade Federal de São Carlos phone: +55 16 99766-0021 "A sorte acompanha uma mente bem treinada" De: Goldman, Adrian<mailto:adrian.gold...@helsinki.fi> Enviado:quinta-feira, 12 de janeiro de 2023 06:34 Para: CCP4BB@JISCMAIL.AC.UK<mailto:CCP4BB@JISCMAIL.AC.UK> Assunto: Re: [ccp4bb] Is this Spam? This looks like Spam. Me too. My uni says that a Microsoft set of addresses ended up on a spam blacklist, which is why this happened. I suspect your institutions are also using office365 outlook (also known as crapmail) as mail server and client As roberto, I complain all the time to my uni about their mail and it. Probably as roberto, it has no effect whatsoever. Adrian Sent from my iPhone On 11 Jan 2023, at 23:13, Becker, Stefan wrote: I also received this kind of message. Is it definitely spam? I waited for 24 hours and noticed that no messages appeared under CCP4BB. Then I re-registered with CCP4BB. From that moment I received messages again. Best, Stefan Becker Am 10.01.2023 um 15:23 schrieb Bernhard Rupp mailto:hofkristall...@gmail.com>>: I got unsubscribed from ccp4em yesterday with a similar message for b...@ruppweb.org<mailto:b...@ruppweb.org>. That one has a minor certificate problem that however does not affect other email recepients. For some reason, the ssl certificate for https://ruppweb.org/ is valid, but the mailserver still seems to have an issue. May be your IT can check yours Best br On Tue, Jan 10, 2023, 02:57 David Briggs mailto:david.bri...@crick.ac.uk>> wrote: I received this odd-looking message from the board address. Did anyone else get anything similar? [cid:b20d59ac-fc29-4cbb-b7f0-65ef44f06b97] I'm sharing a screenshot because I'm not: (a) going to click those links (b) get others to click those links. Can the board admin confirm if this is legitimate or not? D -- Dr David C. Briggs CSci MRSB Principal Laboratory Research Scientist Signalling and Structural Biology Lab The Francis Crick Institute London, UK == about.me/david_briggs<http://about.me/david_briggs> The Francis Crick Institute Limited is a registered charity in England and Wales no. 1140062 and a company registered in England and Wales no. 06885462, with its registered office at 1 Midland Road London NW1 1AT To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=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/
[ccp4bb] RES: [ccp4bb] Fwd: Re: [ccp4bb] Calculate charge on protein surface
Thank you very much, guys. What I wanted actually was a software that gives the formal charge looking at different axis of my model instead of particular residues. Then, placing the center of my model at coordinates 0,0,0, I would be able to see the sum up of charges at +/- x, +/-y and +/-z. I will take a look on the sites and in chimera. Best wishes Rafael Marques da Silva PhD Student – Structural Biology University of Leicester Mestre em Física Biomolecular Universidade de São Paulo Bacharel em Ciências Biológicas Universidade Federal de São Carlos phone: +55 16 99766-0021 "A sorte acompanha uma mente bem treinada" De: Jon Cooper<mailto:488a26d62010-dmarc-requ...@jiscmail.ac.uk> Enviado:quinta-feira, 29 de dezembro de 2022 13:20 Para: CCP4BB@JISCMAIL.AC.UK<mailto:CCP4BB@JISCMAIL.AC.UK> Assunto: Re: [ccp4bb] Fwd: Re: [ccp4bb] Calculate charge on protein surface Dear Boaz I think your original answer must have been steganographic ;-0 Maybe white text on a white background ;-? Anyway, I can see it below now that I have started typing this reply, and advice is excellent. My answer to Rafael's question would be to look at apbs: https://www.poissonboltzmann.org/ This gives you a map of the potential. It's a while since I dabbled with it, but I remember using it and it was all pretty workable as a standalone program, even for an electrostatics non-specialist. You might need to do some scripting to sum up charges, etc, though. Good luck. Jon.C. Sent from ProtonMail mobile Original Message On 28 Dec 2022, 15:57, Boaz Shaanan < bshaa...@bgu.ac.il> wrote: Boaz Shaanan, Ph.D. Dept. of Life Sciences Ben Gurion University Beer Sheva, Israel -- Forwarded message -- From: בעז שאנן Date: Dec 28, 2022 17:13 Subject: Re: [ccp4bb] Calculate charge on protein surface To: Rafael Marques Cc: Hi In ucsf-chimera you can "walk" with the cursor on the surface and it'll show the actual porential value at each point in the command line. You have to enable this option in the surface drawing panel. I've used this option quite often. There may be a similar option in chimerax by now. My 2p. Cheers, Boaz Boaz Shaanan, Ph.D. Dept. of Life Sciences Ben Gurion University Beer Sheva, Israel On Dec 28, 2022 17:03, Rafael Marques wrote: Hello guys. I hope you are having a great break and still eating the leftovers of your Xmas turkey. I wonder if there is any software that could provide me numbers regarding the formal charge of different sides of my structure. Although I can clearly see that one face of it is pretty red, I would like to know “how much” red it is. Best wishes Rafael Marques da Silva PhD Student – Structural Biology University of Leicester Mestre em Física Biomolecular Universidade de São Paulo Bacharel em Ciências Biológicas Universidade Federal de São Carlos phone: +55 16 99766-0021 "A sorte acompanha uma mente bem treinada" To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=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/
[ccp4bb] Calculate charge on protein surface
Hello guys. I hope you are having a great break and still eating the leftovers of your Xmas turkey. I wonder if there is any software that could provide me numbers regarding the formal charge of different sides of my structure. Although I can clearly see that one face of it is pretty red, I would like to know “how much” red it is. Best wishes Rafael Marques da Silva PhD Student – Structural Biology University of Leicester Mestre em Física Biomolecular Universidade de São Paulo Bacharel em Ciências Biológicas Universidade Federal de São Carlos phone: +55 16 99766-0021 "A sorte acompanha uma mente bem treinada" To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=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/
[ccp4bb] RES: [ccp4bb] Multiplicity is more than 20
Problematic is low multiplicity, not high, I’d say... Best Rafael Marques da Silva Mestre em Física Biomolecular Universidade de São Paulo Bacharel em Ciências Biológicas Universidade Federal de São Carlos phone: +55 16 99766-0021 "A sorte acompanha uma mente bem treinada" De: Winter, Graeme (DLSLtd,RAL,LSCI)<mailto:6a19cead4548-dmarc-requ...@jiscmail.ac.uk> Enviado:segunda-feira, 19 de setembro de 2022 16:27 Para: CCP4BB@JISCMAIL.AC.UK<mailto:CCP4BB@JISCMAIL.AC.UK> Assunto: Re: [ccp4bb] Multiplicity is more than 20 Hi Prasun You just measured the reflections a lot. 33 is fine. More likely even with cubic space groups I’d love to know where you got the idea of the limit from. I have data sets with more than 100 fold multiplicity The R conversation I’ll leave to others Best wishes Graeme On 19 Sep 2022, at 20:32, Prasun Kumar wrote: Hi All: I have collected a dataset for a crystal of a 30 residues long helical peptide that makes a trimer in the solution. I also solved the structure to get a trimer. My issues start when I start preparing for a deposition. Details about the data: space group: I 21 3 Resolution: 1.6 Current Rfree/ Rwork: 0.21/0.19 Problems: According to Aimless, Multiplicity: 33.9, and I understand that the value should be less than or equal to 20. Does it mean that I have a lot of random noise or ice rings or something similar? For the inner shell, R work is also higher than R free. Please guide me in solving the above issue. Thank You Prasun To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 -- 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 To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=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/
[ccp4bb] RES: [ccp4bb] Off-topic question related to ITC binding studies
I am not a proper chemist, but I remember that during my college classes my professor emphasized that ^x is a measured value and not really related to stoichiometry. If someone has something to add… Best Rafael Marques da Silva Mestre em Física Biomolecular Universidade de São Paulo Bacharel em Ciências Biológicas Universidade Federal de São Carlos phone: +55 16 99766-0021 "A sorte acompanha uma mente bem treinada" De: ABHISHEK SUMAN<mailto:2fccd9428006-dmarc-requ...@jiscmail.ac.uk> Enviado:terça-feira, 16 de agosto de 2022 14:41 Para: CCP4BB@JISCMAIL.AC.UK<mailto:CCP4BB@JISCMAIL.AC.UK> Assunto: [ccp4bb] Off-topic question related to ITC binding studies Hello everyone! Hope this email finds you well. I have an off-topic question regarding ITC binding studies, which was asked by a reviewer. We performed an ITC binding study (using Affinity ITC, TA Instruments) to evaluate protein-DNA interaction which resulted in a perfect sigmoidal curve. We used the ‘one set of sites’ binding algorithm (“independent” model) for curve fitting and to calculate binding and thermodynamic parameters. The study suggested two copies of the protein binding to a single duplex DNA, i.e., the stoichiometry of protein:DNA is 2:1 (N=2). The ITC calculated the KD (equilibrium molar dissociation constant) in μM (micromolar). But the reviewer is asking to report the KD in (micromolar)^2 instead of micromolar mentioning that the binding reaction is 2A + B <-> (A2)B and the complex is (A2)B and not AB. Though we're trying to explain to the reviewer that we couldn't find any software that can compute the KD in (micromolar)^2 for the stoichiometry of 2 but he is not agreeing to it. We have used the NanoAnalyze software from the TA instrument. This software does not have a model to measure the KD in (micromolar)^2. I would be grateful if you could help me to resolve this problem or at least let me know what explanation might be appropriate to answer the reviewer’s concern that it’s a general practice to report the KD in the Molar irrespective of stoichiometry. Thanks in advance. Regards Abhishek Dr. Abhishek Suman Ph.D (Structural Biology) Indian Institute of Technology Hyderabad Kandi 502 284 Sangareddy Telangana INDIA Contact: +91 91002 74548, +91 80843 11898 Email: abhisheks.i...@gmail.com<mailto:abhisheks.i...@gmail.com> P Please don't print this e-mail or attachment unless necessary. Preserve trees on the planet. Disclaimer:- This footer text is to convey that this email is sent by one of the users of IITH. So, do not mark it as SPAM. To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=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/
[ccp4bb] RES: [ccp4bb] Coot 1
If Paul had said that there is also a Windows version I would be sure that it was April fools’ joke. I can’t wait to use it 😊 Best Rafael Marques da Silva Mestrando em Física Biomolecular Universidade de São Paulo Bacharel em Ciências Biológicas Universidade Federal de São Carlos phone: +55 16 99766-0021 "A sorte acompanha uma mente bem treinada" De: Jurgen Bosch<mailto:jxb...@case.edu> Enviado:sexta-feira, 1 de abril de 2022 19:59 Para: CCP4BB@JISCMAIL.AC.UK<mailto:CCP4BB@JISCMAIL.AC.UK> Assunto: Re: [ccp4bb] Coot 1 Yay!! I can now blend it with blender. Jürgen __ Jürgen Bosch, PhD, MBA Center for Global Health & Diseases Case Western Reserve University https://www.linkedin.com/in/jubosch/ CEO & Co-Founder at InterRayBio, LLC Johns Hopkins University Bloomberg School of Public Health Department of Biochemistry & Molecular Biology On Apr 1, 2022, at 18:46, Paul Emsley mailto:pems...@mrc-lmb.cam.ac.uk>> wrote: That, for the record, is more or less what Ralf said 18 years ago. On 01/04/2022 23:38, Pavel Afonine wrote: It's April 1st today, isn't it? -;) On Fri, Apr 1, 2022 at 3:15 AM Paul Emsley mailto:pems...@mrc-lmb.cam.ac.uk>> wrote: Coot 1 18 years after the release of Coot 0 it's time that I actually released Coot 1. To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=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/
[ccp4bb] Postdoc position at USP (São Carlos, Brazil)
The Sérgio Mascarenhas Biophysics and Structural Biology Group, from IFSC/USP, is looking for exceptionally motivated and innovative candidates to carry out a postdoctoral fellowship in the Thematic Project “Septin filaments: structure, polymerization and action in pathologies” to develop a project using Cryo-EM as the main technique. In addition, negative staining electron microscopy, MALS and SAXS can also be used to determine the oligomerization state, shape and homogeneity of septin complexes. Candidates must have a PhD degree obtained no more than 7 years ago and experience in protein purification. Experience in Structural Biology and Cryo-EM are desirable, but not mandatory. This opportunity is open to applicants of all nationalities. Interested candidates should send, by February 25th, to the project coordinator, Richard C. Garratt (rich...@ifsc.usp.br ), the following documents for registration: – A letter presenting your interest; – A short CV, presenting academic background, publications, and information that proves your scientific experience; - Letter of recommendation. Best Regards Rafael Marques da Silva Mestrando em Física Biomolecular Universidade de São Paulo Bacharel em Ciências Biológicas Universidade Federal de São Carlos phone: +55 16 99766-0021 "A sorte acompanha uma mente bem treinada" To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=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/
[ccp4bb] RES: [ccp4bb] 60 kDa contamination in Rosetta cells
I do agree with the others about what they said previously and I would not bother about the ~60kDa protein. I suppose the tag is in the 15kDa one by the amount of protein you have in the gel. I think you might be facing two different “problems”. One of your proteins (or the complex) is precipitating during SEC. Do you centrifuge your samples before running SEC? Do you keep them in 4ºC? What concentration do you get after your first elution? Your sample may look very clean in solution but maybe this is not really real. The other “problem” is that maybe your proteins are forming bigger oligomers in solution. You can try native gel to find this out but firstly I would use another SEC column, a bigger one. Superdex 200 10/300 is an analytical column and should not be used for purification itself (although I have done this many times). I would try maybe S200 16/60. Best Rafael Marques da Silva Mestrando em Física Biomolecular Universidade de São Paulo Bacharel em Ciências Biológicas Universidade Federal de São Carlos phone: +55 16 99766-0021 "A sorte acompanha uma mente bem treinada" De: Dilip Badgujar<mailto:dilip@gmail.com> Enviado:terça-feira, 13 de julho de 2021 11:23 Para: CCP4BB@JISCMAIL.AC.UK<mailto:CCP4BB@JISCMAIL.AC.UK> Assunto: Re: [ccp4bb] 60 kDa contamination in Rosetta cells Hello, Please find the attached gel picture for the reference and some additional information. Lysis buffer-50 mM Tris-pH 8.0, 300 mM NaCl, 1 mM DTT. 1X protease inhibitor cocktail. SEC column – Superdex 200. 16/300 One of the proteins is 37 kDa while the other one is15 kDa. I do not think that they are making that strong heterodimer. The total molecular of the complex would be ~ 52 kDa but now it is around 60 kDa. I am currently using lower temperature (16 ᵒC) but can try around 12 with low IPTG conc. Regards Dilip Badgujar On Mon, Jul 12, 2021 at 11:56 PM zaigham khan mailto:mahmood.zaig...@gmail.com>> wrote: Hey Dilip, There are many reasons for this observation. Rafael is right, please do share the image of the gel. Also what are the exact sizes of the two proteins that you co-expressed? I have observed the heterodimeric and pentameric proteins on SDS-PAGE albeit the presence of DTT in the sample buffer, and despite boiling of the samples. Could this be that 60 KD is actually the hetero-dimer? One can perform western blotting, followed by the use of anti-polyhistidine and anti-Streptavidin antibodies on separate blots to confirm the suspected bands. Likewise SEC followed by WB may confirm the identity of the eluted proteins in different fractions after size exclusion chromatography. You may also cleave the tags, and then see the magic! Tom has correctly pointed out that induction at lower temperature is best achieved upon incubation of culture so that the temperature is dropped before the induction. Bon Voyage! -Z Zaigham M Khan, PhD Associate Scientist Icahn School of Medicine at Mount Sinai Department of Oncological Sciences 1470 Madison Avenue New York United States On Mon, Jul 12, 2021 at 2:45 AM Dilip Badgujar mailto:dilip@gmail.com>> wrote: Greetings, everyone I am trying to co-express two mammalian proteins (less than 50 kDa MW) in Rosetta cells but getting a contaminating band around 60 kDa. One of the construct is in pET28a with His tag while the other is in pET21c with Strep tag and I am adding all the three selection markers during growth of pre-culture and during induction. Initially, cells were grown at 37 °C till OD reaches to 0.6 then induced with 0.5mM IPTG and incubated at 16°C ON. When I do purification using Streptactin resin; I can see proteins of my interest bound to the resin along with contaminating protein at 60 kDa. I have tried performing size exclusion as a follow up step but they are co-eluting in void volume. I have also tried to wash with MgCL2-ATP solution but it co-elution with contaminant. I am looking for valuable suggestions to avoid the contamination during or after expression. Thanks in advance. Regards Dilip To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 -- Dilip C. Badgujar, (PhD) Post Doctoral Fellow, IIT Bomaby To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=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/
[ccp4bb] RES: [ccp4bb] 60 kDa contamination in Rosetta cells
Hello, Dilip. You have not told us about your buffer composition. Also, it would be great if you could provide us a pic of your gels. I would suggest to increase the salt concentration (~300 to 500 mM NaCl) to check if by doing this the interaction between contaminant and your samples stops. Are your proteins monomers in solution? If you are not sure, try a bigger SEC column. Regards Rafael Marques da Silva Mestrando em Física Biomolecular Universidade de São Paulo Bacharel em Ciências Biológicas Universidade Federal de São Carlos phone: +55 16 99766-0021 "A sorte acompanha uma mente bem treinada" De: Dilip Badgujar<mailto:dilip@gmail.com> Enviado:segunda-feira, 12 de julho de 2021 03:45 Para: CCP4BB@JISCMAIL.AC.UK<mailto:CCP4BB@JISCMAIL.AC.UK> Assunto: [ccp4bb] 60 kDa contamination in Rosetta cells Greetings, everyone I am trying to co-express two mammalian proteins (less than 50 kDa MW) in Rosetta cells but getting a contaminating band around 60 kDa. One of the construct is in pET28a with His tag while the other is in pET21c with Strep tag and I am adding all the three selection markers during growth of pre-culture and during induction. Initially, cells were grown at 37 °C till OD reaches to 0.6 then induced with 0.5mM IPTG and incubated at 16°C ON. When I do purification using Streptactin resin; I can see proteins of my interest bound to the resin along with contaminating protein at 60 kDa. I have tried performing size exclusion as a follow up step but they are co-eluting in void volume. I have also tried to wash with MgCL2-ATP solution but it co-elution with contaminant. I am looking for valuable suggestions to avoid the contamination during or after expression. Thanks in advance. Regards Dilip To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=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/
[ccp4bb] RES: [ccp4bb] Micro/Macro crystal seeding experience
Thank you so much guys for all the suggestions. I will definitely try some of them. Just telling you a bit more about my case, especially the average size crystal: - I screened for different protein concentration (15, 10 and 5 mg/mL), different precipitant/protein ratio (2:1, 1:1 and 1:2) and also different room temperatures (4, 10 and 20ºC). I also tried different commercial kits (Morpheus, BCS, SG1, PACT and INDEX) - I tried co crystallization with some of its bindings too (GTP, GDP and GTP-y)...no improvement - As cryo protector I used glycerol, PEG200 and PEG400 (no difference concerning diffraction) - I screened manually a range of pH and PEG percentage from my best condition (the biggest crystals), even changed the volume of the drops and the reservoir solution (sometimes I didn’t even add it) - the crystals are rod-like...pointless tells me they are P63 2 2 and I couldn’t get another space group till now. And yes, there is a lot of solvent (~79%) and I believe that this is behind with this protein crystal don’t diffract beyond the necessary. Thank you very much again, guys Best regards Rafael Marques da Silva Mestrando em Física Biomolecular Universidade de São Paulo Bacharel em Ciências Biológicas Universidade Federal de São Carlos phone: +55 16 99766-0021 "A sorte acompanha uma mente bem treinada" De: Andrew Purkiss<mailto:a.purk...@mail.cryst.bbk.ac.uk> Enviado:sexta-feira, 18 de dezembro de 2020 12:50 Para: Rafael Marques<mailto:rafael_mmsi...@hotmail.com>; CCP4BB@JISCMAIL.AC.UK<mailto:CCP4BB@JISCMAIL.AC.UK> Assunto: Re: [ccp4bb] Micro/Macro crystal seeding experience We recently had a projects where removing the HIS-tag changed an unsolvable crystal form into one where Molecular Replacement worked immediately (and with an NMR ensemble as the search model to boot). In this case the HIS-tag was about 5% of the total domain mass and the tags were probably tangled up in the centre of the unit cell. Along with all the other suggestions, I would also put a word in for crystallisation at a lower/different temperature. We've had several projects where this has improved diffraction from worse than 4 Ang. to better than 2 Ang. or where different forms have grown. You can try all the other suggestions in the cold room as well as well as your normal condition. I've been trying to get all our users to screen new crystal forms (both new projects and new conditions in existing projects) in-situ, before trying cryo-cooled data collection. This gives a baseline before opening the drops for cryo-protection and cryo-cooling and may also give some guidelines for dehydration etc. Good luck. Andy Purkiss On Thu, 2020-12-17 at 21:30 +, Rafael Marques wrote: > Hello all, how are you doing? > > I have been working with two different proteins for 3 years. I was > able to purify them by IMAC and SEC, no problems at all, and I have > got some crystals of both. However, the best data I have got until > now is around 4A. The diffraction experiment was carried out several > times at DLS. > > The crystals for each protein are different. One is really small > whereas the other one is a regular size (Images below). Some people > have told me to try microseeding and I did this, but with no success > at all. > > So I am asking here if there is someone experienced in this kind of > witchcraft that could give me a hand and also answer me these two > things: > > Have you ever had good size crystals that didn’t diffract beyond 4A > and after trying seeding you have got very good diffraction data (in > these case with no change concerning the crystal size)? > > What is the best protocol that you have tried to increase the size of > your crystals and make them diffract better? > > Many thanks in advance. > > Best > > > > > Rafael Marques da Silva > Mestrando em Física Biomolecular > Universidade de São Paulo > > Bacharel em Ciências Biológicas > Universidade Federal de São Carlos > > phone: +55 16 99766-0021 > >"A sorte acompanha uma mente bem treinada" > > > > To unsubscribe from the CCP4BB list, click the following link: > https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 -- Andrew Purkiss Structural Biology STP The Francis Crick Institute To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=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/
[ccp4bb] CCP4 tutorials (files)
Hello folks. During the quarantine I am trying to use some softwares on CCP4 that I have never used before and also to learn how to solve structures differently than the usual MR (for me). However, the ccp4 tutorial website is down and error 404 is everywhere. Does anybody know a link where I can find files for doing SAD, MAD, MR, SIRAS and so on? Many thanks in advance __ Rafael Marques da Silva Mestrando em Física Biomolecular Universidade de São Paulo Bacharel em Ciências Biológicas Universidade Federal de São Carlos phone: +55 16 99766-0021 "A sorte acompanha uma mente bem treinada" To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=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/
[ccp4bb] RES: [ccp4bb] RES: [ccp4bb] Looking for suggestions with protein expression
Thank you for pointing me this out. I don’t know why I said BL21...maybe in the rush to say to use Rosetta for protein expression. We use Dh5a for plasmid replication (although I have already used BL21 for plasmid purification and it worked nicely). My bad. Rafael Marques da Silva Mestrando em Física Biomolecular Universidade de São Paulo Bacharel em Ciências Biológicas Universidade Federal de São Carlos phone: +55 16 99766-0021 "A sorte acompanha uma mente bem treinada" De: bogba...@yahoo.co.uk<mailto:bogba...@yahoo.co.uk> Enviado:sexta-feira, 10 de julho de 2020 00:30 Para: Rafael Marques<mailto:rafael_mmsi...@hotmail.com> Cc:CCP4BB@JISCMAIL.AC.UK<mailto:CCP4BB@JISCMAIL.AC.UK> Assunto: Re: [ccp4bb] RES: [ccp4bb] Looking for suggestions with protein expression Re: "In our lab we generally use BL21 for plasmid replication." I am interested because on the occasions that I prepared plasmids in BL21 (usually by mistake), they were never any good for cloning or sequencing. I always had to transform them back into a cloning strain and do another plasmid prep! Probably my incompetence! Jon Cooper On 9 Jul 2020 21:44, Rafael Marques wrote: Hi Umar, I must say that it would be better use as an expression system Rosetta DE3 or Rosetta-gami instead of BL21. In our lab we generally use BL21 for plasmid replication. Also, there are a few things that you could try before trying another construction. 1. Lower the temperature during the expression. 2. Try to use a different range of pH in your buffer. Maybe you could add a bit of NaCl (150 – 300 mM) and/or glycerol (2 – 10%) 3. I must say that I have already obtained very different results using Co or Ni columns for IMAC. You could take a look at this. Regards ______ Rafael Marques da Silva Mestrando em Física Biomolecular Universidade de São Paulo Bacharel em Ciências Biológicas Universidade Federal de São Carlos phone: +55 16 99766-0021 "A sorte acompanha uma mente bem treinada" De: Lau Kelvin<mailto:kelvin@epfl.ch> Enviado:quarta-feira, 8 de julho de 2020 16:22 Para: CCP4BB@JISCMAIL.AC.UK<mailto:CCP4BB@JISCMAIL.AC.UK> Assunto: Re: [ccp4bb] Looking for suggestions with protein expression Hello Umar, I would not pin down your difficulties solely due to an Fe-S proteins. I have produced some with no fusion partners and they work wonderfully. They were expressed in an aerobic environment and then reduced in an anaerobic one before usage in reactions. 1) On the Fe-S side, there are plasmids you can co-transform to increase Fe-S production. This plasmid pH151 has the synthetic genes necessary for Fe-S formation. https://www.jbc.org/content/279/33/34721.abstract 2) On the general protein side, have you hhpred your protein? Different constructs (not just tags), temperature? Strain? Media? 3) For these proteins we typically use His Excel (or Protein Ark Ni2+ Advance) that is resistant to most chelators since more often than not, they contain other metals and can snatch Ni2+ from normal Ni-NTA resins. Strep tags also work well. On Jun 27, 2020, at 9:14 AM, Umar Farook mailto:umarfaroo...@gmail.com>> wrote: Dear All, Sorry for an offtopic question, your suggestions are highly appreciated. We have been working on iron sulfur cluster binding protein, which is usually expressed as a nice soluble protein expressed in BL21 cells but aggregated in the affinity column itself and unable to recover from it. We had made n number of truncations and fused to soluble tags such as MBP, but always ended up in large aggregates. Anyone has experience in working with iron-sulfur cluster binding protein before, please let us know the critical steps in purification of such proteins, whether you have completely done the expression, purification and crystallization in anaerobic conditions? or else changing the expression system to eukaryotic system such as Baculo or HEK 293T would help? Please share your valuable experience, thank you. -- Best Regards, Umar Farook To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 This message was issued to members of www.jiscmail.ac.uk/CCP4BB, a mail
[ccp4bb] RES: [ccp4bb] Looking for suggestions with protein expression
Hi Umar, I must say that it would be better use as an expression system Rosetta DE3 or Rosetta-gami instead of BL21. In our lab we generally use BL21 for plasmid replication. Also, there are a few things that you could try before trying another construction. 1. Lower the temperature during the expression. 2. Try to use a different range of pH in your buffer. Maybe you could add a bit of NaCl (150 – 300 mM) and/or glycerol (2 – 10%) 3. I must say that I have already obtained very different results using Co or Ni columns for IMAC. You could take a look at this. Regards __ Rafael Marques da Silva Mestrando em Física Biomolecular Universidade de São Paulo Bacharel em Ciências Biológicas Universidade Federal de São Carlos phone: +55 16 99766-0021 "A sorte acompanha uma mente bem treinada" De: Lau Kelvin<mailto:kelvin@epfl.ch> Enviado:quarta-feira, 8 de julho de 2020 16:22 Para: CCP4BB@JISCMAIL.AC.UK<mailto:CCP4BB@JISCMAIL.AC.UK> Assunto: Re: [ccp4bb] Looking for suggestions with protein expression Hello Umar, I would not pin down your difficulties solely due to an Fe-S proteins. I have produced some with no fusion partners and they work wonderfully. They were expressed in an aerobic environment and then reduced in an anaerobic one before usage in reactions. 1) On the Fe-S side, there are plasmids you can co-transform to increase Fe-S production. This plasmid pH151 has the synthetic genes necessary for Fe-S formation. https://www.jbc.org/content/279/33/34721.abstract 2) On the general protein side, have you hhpred your protein? Different constructs (not just tags), temperature? Strain? Media? 3) For these proteins we typically use His Excel (or Protein Ark Ni2+ Advance) that is resistant to most chelators since more often than not, they contain other metals and can snatch Ni2+ from normal Ni-NTA resins. Strep tags also work well. On Jun 27, 2020, at 9:14 AM, Umar Farook mailto:umarfaroo...@gmail.com>> wrote: Dear All, Sorry for an offtopic question, your suggestions are highly appreciated. We have been working on iron sulfur cluster binding protein, which is usually expressed as a nice soluble protein expressed in BL21 cells but aggregated in the affinity column itself and unable to recover from it. We had made n number of truncations and fused to soluble tags such as MBP, but always ended up in large aggregates. Anyone has experience in working with iron-sulfur cluster binding protein before, please let us know the critical steps in purification of such proteins, whether you have completely done the expression, purification and crystallization in anaerobic conditions? or else changing the expression system to eukaryotic system such as Baculo or HEK 293T would help? Please share your valuable experience, thank you. -- Best Regards, Umar Farook To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=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/
[ccp4bb] [OFF TOPIC] Cell biology bb
Hi Folks, how are you doing? Hope everyone is safe! I was wondering if there is a bulletin board email group like this one concerning Cell Biology, where we could find not only discussions about related topics but also post-doc opportunities. I would be very glad if you guys know any and could share it with me. Kind Regards __ Rafael Marques da Silva Mestrando em Física Biomolecular Universidade de São Paulo Bacharel em Ciências Biológicas Universidade Federal de São Carlos phone: +55 16 99766-0021 "A sorte acompanha uma mente bem treinada" To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=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/
[ccp4bb] RES: [ccp4bb] how to get protein crystal
Dear Amala, The first thing I would try is to run your protein sequence on this server: http://xtalpred.godziklab.org/XtalPred-cgi/xtal.pl. This is a webserver that will compare you aa sequence to other previously crystallized proteins. The result will also suggest which parts of your protein should be removed (if there are disordered regions). Have in mind that a bad score does NOT mean that your protein will not crystallize. Protein crystallization is not the rule, it is the exception. You must be aware that some proteins will never give you crystals, no matter how much you try. And unfortunately, you can only know it after trying. As people already said, always use fresh protein in your screenings and, if possible, purify it by SEC to ensure that you have homogeinity in your sample. If your protein is very soluble, I would even increase the protein concentratrion (in my group we have got better diffracting crystals using 25mg/mL) Good luck Rafael Marques da Silva Mestrando em Física Biomolecular Universidade de São Paulo Bacharel em Ciências Biológicas Universidade Federal de São Carlos phone: +55 16 99766-0021 "A sorte acompanha uma mente bem treinada" De: amala mathimaran<mailto:amalat...@gmail.com> Enviado:sexta-feira, 27 de dezembro de 2019 10:31 Para: CCP4BB@JISCMAIL.AC.UK<mailto:CCP4BB@JISCMAIL.AC.UK> Assunto: [ccp4bb] how to get protein crystal Dear all, Can you suggest me how to get protein crystal??? I purified my target protein and concentrated to 12mg/ml (pI 6.04). Final purified protein contains 50mM HEPES pH 8.0 and 50mM NaCl. Then the initial screening was done using hanging drop method but no crystal. So 2mM NADP cofactor was added again screen with Hampton (PEG ion, PEG RX, crystal screen, Index) and Molecular dimensions conditions etc. I got precipitate like formation the image was attached below. From this formation what I can do… mean while I increase the protein concentration and did screening for that selected conditions again I got same kind of formations. I am new to protein crystallography kindly suggesting me. And how much concentration is suitable for protein crystallization?? How to find which concentration is enough for our target protein crystallization?? Thanks in advance To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB&A=1 To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB&A=1
[ccp4bb] RES: [ccp4bb] Xray-dataset usable despite low completeness ?
Lol __ Rafael Marques da Silva Mestrando em Física Biomolecular Universidade de São Paulo Bacharel em Ciências Biológicas Universidade Federal de São Carlos phone: +55 16 99766-0021 "A sorte acompanha uma mente bem treinada" De: CCP4 bulletin board em nome de Bernhard Rupp Enviado: Friday, November 29, 2019 1:18:41 AM Para: CCP4BB@JISCMAIL.AC.UK Assunto: Re: [ccp4bb] Xray-dataset usable despite low completeness ? It has come to our attention that on this bulletin board insensitive and hurtful comments have been made towards animals with disability. Particularly concerning is the display of white privilege and racial bias towards a minority individual given that the turkey is also referred to in German as 'Indian'. In view of this non-inclusive and divisive display of unwokeness, the faculty Bias Response Team will contact you shortly and allow you to present your self-critique. We want this board to remain a safe zone inclusive of all animals, complete or not. Stuffed, BR -Original Message- From: CCP4 bulletin board On Behalf Of Jurgen Bosch Sent: Thursday, November 28, 2019 13:51 To: CCP4BB@JISCMAIL.AC.UK Subject: Re: [ccp4bb] Xray-dataset usable despite low completeness ? Think of completeness with an analogy to turkey. Say you happen to find a one-legged turkey (incomplete by conventional standard) you could still stuff it and put it in the oven and enjoy 93% of the turkey. The 7% missing, who cares? Other than I like both legs of the turkey :-) Happy Thanksgiving everyone Jürgen P.S. back to my wine & ducks to be roasted. @BR, mit Rotkraut & Kartoffelknödel > On Nov 28, 2019, at 4:38 PM, Bernhard Rupp wrote: > > Sorry for being late on this thread - > > but the completeness myth is one of these conventional wisdoms I am > seriously questioning and completeness as a global statistic is almost > uninformative, short of telling you 'fewer than all recordable reflections up > to the reported (likely isotropic) resolution limit given whatever (likely > isotropic) cutoff was applied'. Sounds not very clear to me. > > Kay mentioned already that any information is better than no > information, with the caveat that you cannot expect map quality (being > an upper limit for model quality - not going into precision vs > accuracy issue here) corresponding to the highest resolution reported, which > is in reality frequently anisotropic (but not reported or reflected > adequately in the PDB reports). > We posted some remarks to this effect recently, pointing out that > highly incomplete and anisotropic data can still yield limited but > useful information as long as your claim remains correspondingly > modest. Section 3.4 in > http://journals.iucr.org/d/issues/2019/12/00/di5032/index.html > > Having said that, while random incompleteness is not problematic, > systematic reciprocal space incompleteness leads to corresponding > systematic real space effects on the map, the simplest being > anisotropic data reflecting anisotropic reciprocal map resolution. > This is different for example when wedges are missing or absence of serial > extinctions makes space group determination more challenging (although we are > almost in the age where 'record 360 deg of data and try every SG' works). > James Holton has video examples for incompleteness effects and some images > are also in my book. > https://bl831.als.lbl.gov/~jamesh/movies/ > > Cheers & Happy Thanksgiving, BR > > PS: A systemic rant regarding data quality representation can be found > here > https://www.cell.com/structure/fulltext/S0969-2126(18)30138-2 > > -- > Bernhard Rupp > http://www.hofkristallamt.org/ > b...@hofkristallamt.org > -- > All models are wrong > but some are useful. > -- > > -Original Message- > From: CCP4 bulletin board On Behalf Of Kay > Diederichs > Sent: Monday, November 25, 2019 08:07 > To: CCP4BB@JISCMAIL.AC.UK > Subject: Re: [ccp4bb] Xray-dataset usable despite low completeness ? > > Dear Matthias, > > Of course, high completeness is better than low completeness. > But as long as your low resolution is pretty much complete, there is no such > thing as "too low completeness" at high resolution. Each reflection adds > information to the map, and serves as a restraint in refinement. > > best, > Kay > > > On Mon, 25 Nov 2019 14:11:52 +0100, Matthias Oebbeke > wrote: > >> Dear ccp4 Bulletin Board, >> >> I collect
[ccp4bb] RES: [ccp4bb] Problem in real space - please sign & invite other scientists to sign this letter
Dear Daniel.
I really understand your point of view and I must say that I agree with it, at
certain extent. Science is all about evidence and not what we (sometimes) want
to see or want to believe. You are completely right when you say that, inside
academia, evidence should be always discussed by those who have deep knowledge
about the subject. Good science is based on good standards, and good standards
are exactly what make our science so robust.
While I must say that it bothers me when someone who is not an expert in "my"
field criticizes my job, I recognize that what makes what we do different than
any religion is exactly the possibility of being criticized, both by those
related to the field and those who might have a good point of view.
Also, we know the rules inside the academia, how to analyze data and we know
too that, in general, the population does not understand at all the meaning and
the methods inside science. If we try to explain any scientific data to these
people using our methods and standards it will be a huge failure (I can't even
change the mind of my nephews, which after a NatGeo show started to believe in
mermaids). When we deal with a non-academic public, we must emphasize how
important is to make science, the good things that came from it and, most
important, we need to be understood.
In fact, there is some discussion if the global warming is only natural or if
it has been happening because the human activity. However, it is pretty clear
that mankind is affecting the weather by the increase of carbon dioxide in the
atmosphere and the large impact caused by cattle. A five minutes search on
Google can give you back several papers, but I am going to present you only
this one, concerned in devaluation.
HTTPS://www.scientificamerican.com/article/co2-emissions-reached-an-all-time-high-in-2018/
As scientists, we are the ones who might have no knowledge in the field, but
the ones able to point out a critical view. When we refuse to talk to the
population cause we are not experts is that field, someone else takes our
place. And generally, they attack science and scientists, giving simple answers
for complex problems. That is because they know what the population wants to
hear. That is how Trump has become the USA president and Billboards did the
same here in Brazil.
When we refuse to look for information, refuse to spread it to the general
population and hide inside the academia, we must be aware that someone, for
sure, is taking our place. That is exactly why currently so many people believe
that the Earth is flat and that vaccines provoke diseases. That is also the
meaning why Greta Thunders, a sixteen years old teenager activist , is better
known than the experts in the field. If we want to be believed we must take
back our place. To hind is not an option.
Regards
Rafael Marques da Silva
Mestrando em Física Biomolecular
Universidade de São Paulo
Bacharel em Ciências Biológicas
Universidade Federal de São Carlos
phone: +55 16 99766-0021
"A sorte acompanha uma mente bem treinada"
De: CCP4 bulletin board em nome de Daniel M. Himmel,
Ph. D.
Enviado: Tuesday, August 20, 2019 10:23:05 PM
Para: CCP4BB@JISCMAIL.AC.UK
Assunto: Re: [ccp4bb] Problem in real space - please sign & invite other
scientists to sign this letter
Dear colleagues,
Since when does being a structural biologist make us experts in climatology,
and isn't it a breach of basic ethical practice and professionalism as
scientists
to sign on as authors to an article for which we have neither contributed
research nor intellectual content of the manuscript? Are we now going against
the standard to which the editorial policies of leading reputable biological
journals normally hold us as authors? And doesn't it hurt the credibility
of a serious scientific article, its authors, and the journal in which it
appears
if biologists with no expertise in earth science/astrophysics appear
without humility as authors to such an article?
Are you not embarrassed to put your name to an article that uses physical
sciences data as a platform for preaching about religion, politics, and economic
theory ("...social and economic justice for all...")?
Does it not upset you when someone unfamiliar with structural biology draws
firm conclusions that heavily depend on the part of a structural model that has
high
B-factors? So why are you unconcerned that you may be guilty of an analogous
error when, as structural biologists, you put your name to a controversial
interpretation
of selected earth science data? See, for example,
https://blogs.agu.org/geospace/2017/02/24/living-warm-peak-ice-ages/ about the
ways
climate data can be misinterpreted by choosing too tight a time interval, and
lets stick to
structural biology and allied sciences in the C
