Re: [ccp4bb] Looking for a student and/or help with finalizing new virus spike structures
I am willing to help for your work. Can we communicate for the issue throgh my e-mail fenghui...@163.com with fanfeng...@mail.tsinghua.edu.cn cced? Dr Fenghui Fan Replied Message | From | Grant Hansman| | Date | 07/09/2024 12:52 | | To | CCP4BB@JISCMAIL.AC.UK | | Cc | | | Subject | [ccp4bb] Looking for a student and/or help with finalizing new virus spike structures | Help needed ASAP. I am just setting up my new laboratory in Australia (alone at the moment) and already have at least 5 new X-ray structures that need checking and a bit more refinement. High resolution and should not be too much work. Therefore, I am looking for a student with experience and/or any help with finalizing the structures (will provide all details to those interested). Happy to add as a co-first author to any manuscripts, as all the wet lab, data collection, and refinements were done by myself ;) Cheers. 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/
Re: [ccp4bb] recombinant protein-based fluorophores
eGFP, for example, can be fused to the C- terminal of your target protein. Fluorescent proteins were usually highly soluble, especially if you select to express in insect cells, which has the tendency to express proteins in soluble states. Smith | | Smith Liu | | 邮箱:smith_liu...@163.com | 签名由 网易邮箱大师 定制 On 12/07/2021 07:21, samer halabi wrote: Dear All, Sorry for disturbing you all with my current inquiry. However, I am in the process of designing and expressing soluble proteins and I thought to ask for your valuable advice. 1-What would be your first choice (protein-based) fluorophore to link it to the soluble protein at its C-terminus? 2-Would this fluorophore be suitable for carrying later experiments in vivo and in vitro withstanding such conditions and still having detectable fluorescence signal (medium to high)? 3-Would this fluorophore of choice stand denaturing and renaturing conditions, as refolding the protein chimera (soluble protein + the linked fluorophore) from bacterial inclusion bodies that were dissolved in 8M Urea or 6M Guanidine-HCl? 4-Have you had success with producing APC or R-PE in such manner (secreting them from insect cells or refolding them from bacterial inclusion bodies)? Thank you in advance. Best regards, Samer 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] AW: Re: [ccp4bb] AW: Re: [ccp4bb] coordinate transformation
sorry, how i move the mtz into the transformed pdb for the question in my previous email? | | Smith Liu | | 邮箱:smith_liu...@163.com | 签名由 网易邮箱大师 定制 在2017年12月18日 23:37,Smith Liu 写道: thanks. i may mean something other. for example, if i rotate the pdb by 30 degree (or 29.5 degree), or i shift the pdb along x-axis by something for example 0.123*a, then how i move the mtz map correspondingly for the fitting of mtz into the transformed map? | | Smith Liu | | 邮箱:smith_liu...@163.com | 签名由 网易邮箱大师 定制 在2017年12月18日 21:58,herman.schreu...@sanofi.com 写道: Dear Smith, The map extends through the whole crystal. What happens is that the map is calculated around the atom you clicked on during centering. So by centering on your transformed pdb, you will sample the same map at a different position. Just load your transformed pdb and untransformed mtz and try. If the transformed pdb does not fit the map, something went wrong during the transformation of your pdb. If you have applied an origin shift (is not equal to applying a crystallographic symmetry operation), you have to recalculate the mtz, e.g. by running another round of refinement. I hope this is clear so. Herman Von: Smith Liu [mailto:smith_liu...@163.com] Gesendet: Montag, 18. Dezember 2017 14:52 An: Schreuder, Herman /DE Betreff: [EXTERNAL] Re: [ccp4bb] AW: Re: [ccp4bb] coordinate transformation you mean the mtz map will transform simutaneously? | | Smith Liu | | 邮箱:smith_liu...@163.com | 签名由 网易邮箱大师 定制 在2017年12月18日 21:26,herman.schreu...@sanofi.com写道: If you use coot with on the fly map calculation (e.g. you load an mtz and not a map file), you do not need to transform the map. Otherwise I would recommend to run one more round of refinement and produce a new map your usual way. This will also get rid of any rounding errors due to the transformation. Best, Herman Von: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] Im Auftrag von Smith Liu Gesendet: Montag, 18. Dezember 2017 14:16 An:CCP4BB@JISCMAIL.AC.UK Betreff: [EXTERNAL] Re: [ccp4bb] coordinate transformation Dear All, If I have a set of PDB with the corresponding density map, after I transform the PDB based on the suggestion of everybody, is any way to transform the map so that the map will be fit with the transformed PDB? Smith At 2017-12-18 18:39:34, "Eleanor Dodson" <176a9d5ebad7-dmarc-requ...@jiscmail.ac.uk> wrote: I showed you pdbset .. Find the centre of mass for your assembly. Move it where you will pdbset xyzin mow.pdb end Find CoM 0.7 1.3 -0.2 Hmm - a little thought - centre at 1 -1 0 say pdbset yzin now.pdb xyzout changed.pdb symgen x , y-2, z end New CoM 0.7 -0.7 -0.2 Eleanor On 18 December 2017 at 00:19, Edward A. Berry wrote: Neat idea! And do you have a 1-line command for setting all the coordinates to 1,1,1? or 0.1,0.1,0.1 if I still want it near the origin but biased toward the inside of the positive-going cell? eab On 12/14/2017 07:23 PM, James Holton wrote: What I usually do for this is make a copy of the PDB file and change all the atom x-y-z positions to "1.000". Then I use something like reforigin or my "origins.com" script to shift the original coordinates via allowed symmetry operations, origin shifts, or perhaps indexing ambiguities until it is as close as possible to the "reference", which is at 1,1,1. I use 1,1,1 instead of 0,0,0 because there are generally at least two symmetry-equivalent places that are equidistant from the origin. Declaring the reference to be a bit off-center breaks that ambiguity, and also biases the result toward having all-positive x,y,z values. In case it is interesting, my script is here: http://bl831.als.lbl.gov/~jamesh/scripts/origins.com You need to have the CCP4 suite set up for it to work. Run it with no arguments to get instructions. -James Holton MAD Scientist On 12/13/2017 5:50 AM, Kajander, Tommi A wrote: Hello, If someone could point this out would be very helpful... Wasnt there a simple script somewhere that would transfer coordinates close to origin - if they for some reason are not? Just cant find anything right away. Sure i have done this before... Thanks, Tommi
Re: [ccp4bb] AW: Re: [ccp4bb] AW: Re: [ccp4bb] coordinate transformation
thanks. i may mean something other. for example, if i rotate the pdb by 30 degree (or 29.5 degree), or i shift the pdb along x-axis by something for example 0.123*a, then how i move the mtz map correspondingly for the fitting of mtz into the transformed map? | | Smith Liu | | 邮箱:smith_liu...@163.com | 签名由 网易邮箱大师 定制 在2017年12月18日 21:58,herman.schreu...@sanofi.com 写道: Dear Smith, The map extends through the whole crystal. What happens is that the map is calculated around the atom you clicked on during centering. So by centering on your transformed pdb, you will sample the same map at a different position. Just load your transformed pdb and untransformed mtz and try. If the transformed pdb does not fit the map, something went wrong during the transformation of your pdb. If you have applied an origin shift (is not equal to applying a crystallographic symmetry operation), you have to recalculate the mtz, e.g. by running another round of refinement. I hope this is clear so. Herman Von: Smith Liu [mailto:smith_liu...@163.com] Gesendet: Montag, 18. Dezember 2017 14:52 An: Schreuder, Herman /DE Betreff: [EXTERNAL] Re: [ccp4bb] AW: Re: [ccp4bb] coordinate transformation you mean the mtz map will transform simutaneously? | | Smith Liu | | 邮箱:smith_liu...@163.com | 签名由 网易邮箱大师 定制 在2017年12月18日 21:26,herman.schreu...@sanofi.com写道: If you use coot with on the fly map calculation (e.g. you load an mtz and not a map file), you do not need to transform the map. Otherwise I would recommend to run one more round of refinement and produce a new map your usual way. This will also get rid of any rounding errors due to the transformation. Best, Herman Von: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] Im Auftrag von Smith Liu Gesendet: Montag, 18. Dezember 2017 14:16 An:CCP4BB@JISCMAIL.AC.UK Betreff: [EXTERNAL] Re: [ccp4bb] coordinate transformation Dear All, If I have a set of PDB with the corresponding density map, after I transform the PDB based on the suggestion of everybody, is any way to transform the map so that the map will be fit with the transformed PDB? Smith At 2017-12-18 18:39:34, "Eleanor Dodson" <176a9d5ebad7-dmarc-requ...@jiscmail.ac.uk> wrote: I showed you pdbset .. Find the centre of mass for your assembly. Move it where you will pdbset xyzin mow.pdb end Find CoM 0.7 1.3 -0.2 Hmm - a little thought - centre at 1 -1 0 say pdbset yzin now.pdb xyzout changed.pdb symgen x , y-2, z end New CoM 0.7 -0.7 -0.2 Eleanor On 18 December 2017 at 00:19, Edward A. Berry wrote: Neat idea! And do you have a 1-line command for setting all the coordinates to 1,1,1? or 0.1,0.1,0.1 if I still want it near the origin but biased toward the inside of the positive-going cell? eab On 12/14/2017 07:23 PM, James Holton wrote: What I usually do for this is make a copy of the PDB file and change all the atom x-y-z positions to "1.000". Then I use something like reforigin or my "origins.com" script to shift the original coordinates via allowed symmetry operations, origin shifts, or perhaps indexing ambiguities until it is as close as possible to the "reference", which is at 1,1,1. I use 1,1,1 instead of 0,0,0 because there are generally at least two symmetry-equivalent places that are equidistant from the origin. Declaring the reference to be a bit off-center breaks that ambiguity, and also biases the result toward having all-positive x,y,z values. In case it is interesting, my script is here: http://bl831.als.lbl.gov/~jamesh/scripts/origins.com You need to have the CCP4 suite set up for it to work. Run it with no arguments to get instructions. -James Holton MAD Scientist On 12/13/2017 5:50 AM, Kajander, Tommi A wrote: Hello, If someone could point this out would be very helpful... Wasnt there a simple script somewhere that would transfer coordinates close to origin - if they for some reason are not? Just cant find anything right away. Sure i have done this before... Thanks, Tommi
Re: [ccp4bb] coordinate transformation
Dear All, If I have a set of PDB with the corresponding density map, after I transform the PDB based on the suggestion of everybody, is any way to transform the map so that the map will be fit with the transformed PDB? Smith At 2017-12-18 18:39:34, "Eleanor Dodson" <176a9d5ebad7-dmarc-requ...@jiscmail.ac.uk> wrote: I showed you pdbset .. Find the centre of mass for your assembly. Move it where you will pdbset xyzin mow.pdb end Find CoM 0.7 1.3 -0.2 Hmm - a little thought - centre at 1 -1 0 say pdbset yzin now.pdb xyzout changed.pdb symgen x , y-2, z end New CoM 0.7 -0.7 -0.2 Eleanor On 18 December 2017 at 00:19, Edward A. Berry wrote: Neat idea! And do you have a 1-line command for setting all the coordinates to 1,1,1? or 0.1,0.1,0.1 if I still want it near the origin but biased toward the inside of the positive-going cell? eab On 12/14/2017 07:23 PM, James Holton wrote: What I usually do for this is make a copy of the PDB file and change all the atom x-y-z positions to "1.000". Then I use something like reforigin or my "origins.com" script to shift the original coordinates via allowed symmetry operations, origin shifts, or perhaps indexing ambiguities until it is as close as possible to the "reference", which is at 1,1,1. I use 1,1,1 instead of 0,0,0 because there are generally at least two symmetry-equivalent places that are equidistant from the origin. Declaring the reference to be a bit off-center breaks that ambiguity, and also biases the result toward having all-positive x,y,z values. In case it is interesting, my script is here: http://bl831.als.lbl.gov/~jamesh/scripts/origins.com You need to have the CCP4 suite set up for it to work. Run it with no arguments to get instructions. -James Holton MAD Scientist On 12/13/2017 5:50 AM, Kajander, Tommi A wrote: Hello, If someone could point this out would be very helpful... Wasnt there a simple script somewhere that would transfer coordinates close to origin - if they for some reason are not? Just cant find anything right away. Sure i have done this before... Thanks, Tommi
Re: [ccp4bb] secondary structure prediction
yes | | Smith Liu | | 邮箱:smith_liu...@163.com | 签名由 网易邮箱大师 定制 在2017年12月07日 09:11,zheng zhou 写道: Thanks for many advices. I was not clear in the previous email. I know the close homologous protein (20% identity, total 500aa), but the fragment hits (30~40aa, identity 40~50%) are from other proteins in PDB. I am trying to see whether the fragments from non-homologous proteins may help the secondary structure prediction. Best, Z On Wed, Dec 6, 2017 at 10:14 PM, zheng zhou wrote: > Dear CCP4 community, > > Sorry for the off-topic question. I am trying to design constructs for > structure studies. It only has a homolog structure in PDB with > sequence identity ~20%. When I blast against PDB sequence, there are > quite a few motif hits (30~40aa, identity 40~50%). Any prediction > tools utilize this information? > > Thanks for your advice in advance. > > Best, > > Zheng
Re: [ccp4bb] secondary structure prediction
Rosetta | | Smith Liu | | 邮箱:smith_liu...@163.com | 签名由 网易邮箱大师 定制 在2017年12月06日 22:14,zheng zhou 写道: Dear CCP4 community, Sorry for the off-topic question. I am trying to design constructs for structure studies. It only has a homolog structure in PDB with sequence identity ~20%. When I blast against PDB sequence, there are quite a few motif hits (30~40aa, identity 40~50%). Any prediction tools utilize this information? Thanks for your advice in advance. Best, Zheng
Re: [ccp4bb] script for shape complementarity
how about modify your selected part of protein into two molecules | | Smith Liu | | 邮箱:smith_liu...@163.com | 签名由 网易邮箱大师 定制 在2017年12月03日 16:21,joy yang 写道: Dear All, can anyone shed a clue on how to define the suface area used in ccp4 sc (shape complementarity) script? The script that I could find on internet is as following: sc XYZIN a22.pdb SURFIN1 A.grasp_surf SURFIN2 B.grasp_surf \ SURFOUT1 A.sc_surf SURFOUT2 B.sc_surf <
Re: [ccp4bb] RMSD calculation for large assemblies
how about mean square deviation of the rmsd of each c alpha? | | Smith Liu | | 邮箱:smith_liu...@163.com | 签名由 网易邮箱大师 定制 在2017年12月02日 22:48,Reza Khayat 写道: Hi, I'm analyzing the RMSD between 60 subunits of a virus. Can someone identify a program that can generate a spread for the RMSD between equivalent C-alpha atoms? For example, the C-alpha atom for amino acid 39 may have RMSD values from 0.1 to 1.5. Coot does a nice job of automatically detecting and calculating RMSD, but I'd like to have the spread for each atom in the final graph that Coot generates. Thank you. Best wishes, Reza Reza Khayat, PhD Assistant Professor City College of New York Department of Chemistry New York, NY 10031
Re: [ccp4bb] Vacuum pump in cold room
use the brand of vacuum pump without oil 发自网易邮箱大师 在2017年10月29日 01:17,Denis Rousseau 写道: Does anyone have experience with a vacuum pump in cold room? We have been told they may not start because of the viscosity of the oil? Thanks Denis Rousseau From: CCP4 bulletin board [CCP4BB@JISCMAIL.AC.UK] on behalf of Schulz, Eike-Christian [eike.sch...@mpsd.mpg.de] Sent: Friday, October 27, 2017 4:31 PM To:CCP4BB@JISCMAIL.AC.UK Subject: [ccp4bb] using CC1/2 to define resolution limit in Xscale Dear all, I would like to compare > 15 datasets and would like to use a common CC1/2 value as an objective criterion to determine the resolution cut-off. All data were integrated in XDS. Is there a convenient way to apply this in XSCALE or in any of its alternatives? With best regards, Eike
Re: [ccp4bb] OFF topic (Protein degrades during Dailysis)
suppose protease is the issue, then avoid overnight dialysis 发自网易邮箱大师 在2017年10月26日 22:18,Anamika Singh 写道: Dear All, I am purifying the His-tagged proteins ( 21Kda) using buffer 20mM Tris, 150mM Nacl, and 5mM MgCl2 with 500mM imidazole. Earlier I was facing the precipitation problem during dialysis but with the addition of 50mM L-Arg somehow managed to overcome the precipitation issue. But this time I have seen after overnight dialysis there are degradation products on SDS page. I have used protease inhibitor (PMSF) in my sonication buffer. Please suggest me to overcome this problem. Thanks -- Anamika
Re: [ccp4bb] Off topic: denaturing urea gels
your enzyme cannot give definite fragment. thus smear 发自网易邮箱大师 在2017年09月30日 19:28,Mohammad Khan 写道: Dear all, I am working with an exonuclease and I run the digested DNA on a 8Murea-20%acrylamide gel in TBE buffer. I use the Mini-Protean BioRad system and cast gels of about 8.6x6.5 cm dimensions with 1.5 mm thickness. I use a 15 well comb. I run my gels at 70 V for as long as 4 hours till my undigested DNA reaches half the gel distance. I use 20-30 nt long susbtrates. I am mostly not able to get distinct bands of the digested products but rather get a smear. Is there any way to make sure that I get distinct digested products rather than a smear? I am looking forward for suggestions from all! Thank you. Ciao!
Re: [ccp4bb] Biotin binding protein
if necessary, phage or yeast display to screen 发自网易邮箱大师 在2017年09月24日 07:05,Reza Khayat 写道: Hi, Sorry for the non crystallography question. Is anyone aware of monomeric proteins (<20kDa) that can bind to biotin? We need this for a couple of different projects where biotin covalently modifies a ligand of interest. We'd like to complex the biotin to a protein Thanks. Best wishes, Reza Reza Khayat, PhD Assistant Professor City College of New York Department of Chemistry New York, NY 10031
Re: [ccp4bb] Difficult purification with imac columns
change buffer,or use different resin matrix from another compny 发自网易邮箱大师 在2017年09月15日 19:53,Narayanan Ramasubbu 写道: Hi. We are working on a periplasmic protein that breaks naked glycans in peptidoglycans. There is truncated structure available but our target is the full length protein. The difficulty us that it strongly binds to the resin with or without his.tag. Changing the resin to acrylamide did not help. Has anyone come across similar problem and how was it resolved. The pdb structure is the catalytic domain and mussing a region that, in my opinion, binds to the resin. Thank you in advance Sent from my iPhone
Re: [ccp4bb] Fast searching of articles related to your PubMed indexed paper
can you updated the server so that the most recent articles can be found? 发自网易邮箱大师 在2017年07月25日 11:49,Yaoqi Zhou 写道: Fast searching of articles related to your PubMed indexed paper Are you spending too much time searching the literature to prepare your grant proposals, manuscripts and keeping track of research happening in your field? We have built a literature-based search engine (http://pubmed.ict.griffith.edu.au/) which is powered by a combination of state-of-the-art methods to locate relevant articles to your published PubMed-indexed papers within a few seconds. While providing you the ability to locate relevant articles, we are seeking your feedback regarding the performance of these algorithms. By annotating recommended articles as relevant, somewhat relevant, or irrelevant, you are participating in a community-wide effort of establishing a gold-standard benchmark for relevant literature search. The accuracy of this benchmark is ensured by your domain knowledge and experience as your judgements will be based in the context of your own article. With your expertise, the entire process will take only a few minutes to complete (1 paper, 6 recommendations). Upon completion, this community-wide dataset will be available to freely download and redistribute, encouraging development of next-generation literature searching and retrieval methodologies, something we all desperately need. Due to current data availability, this relevant-paper search is limited to papers indexed by PubMed between January 1, 2007 to December 31, 2016 (the end of last year). Click http://pubmed.ict.griffith.edu.au/ to start. As this is a product in development, any constructive comments and suggestions will be truly appreciated. If you like this idea of community-wide benchmark for literature recommendation, please forward this email to your colleagues in the scientific community. Initial feedbacks to our server. “The process was certainly fast!”, “Although the articles found in the search were not necessarily exactly relevant, many are articles that I don't think I would have come across easily with a traditional search”, “Very nice work. I think there can be a longer list” (limited to 30 currently). Thank you! Yaoqi Professor Yaoqi Zhou | Research Leader Institute for Glycomics Griffith University | Gold Coast campus | QLD 4222 | Institute for Glycomics (G24) Room 2.10 T +61 7 5552 8228 | F +61 7 5552 9040 | email yaoqi.z...@griffith.edu.au http://sparks-lab.org (Group webpage) http://griffith.edu.au/institute-glycomics (Institute Webpage) On 25 Jul 2017, at 9:01 AM, CCP4BB automatic digest system wrote: There are 14 messages totaling 9878 lines in this issue. Topics of the day: 1. Buccaneer places residues in different asymmetric units (3) 2. Primer design (7) 3. PhD position at University of Oslo 4. Postdoctoral position at Boston Children’s Hospital 5. About weighting factor settings in new ccp4i2 (2) -- Date:Mon, 24 Jul 2017 16:56:51 +0800 From:Lingxiao Zeng Subject: Buccaneer places residues in different asymmetric units Dear All, I tried to use buccaneer to build a model. The starting model is a partial model, after model building the Rwork and Rfree are reasonable but buccaneer places residues in different asymmetric units and the model looks really weird. Is there any way to build the model into the same ASU or put different parts together after model building? Thanks! Best, Alice -- Lingxiao Zeng PhD candidate School of Biomedical Sciences The University of Hong Kong -- Date:Mon, 24 Jul 2017 10:35:17 +0100 From:Jon Agirre Subject: Re: Buccaneer places residues in different asymmetric units Dear Alice, there's an option in both ccp4i and ccp4i2 interfaces that lets you tell Buccaneer that you want to build the new model in the same place as the partial model supplied - see my screenshots for reference. It might not be on by default and perhaps it should be. Please be aware that most newer developments and improvements will be put on the ccp4i2 interface to Buccaneer - it would be helpful if you could have a go and let us know what you think! Hope this helps, Jon On 24 July 2017 at 09:56, Lingxiao Zeng wrote: Dear All, I tried to use buccaneer to build a model. The starting model is a partial model, after model building the Rwork and Rfree are reasonable but buccaneer places residues in different asymmetric units and the model looks really weird. Is there any way to build the model into the same ASU or put different parts together after model building? Thanks! Best, Alice -- Lingxiao Zeng PhD candidate School of Biomedical Sciences The University of Hong Kong -- Dr Jon Agirre York Structural Biology Laboratory / Department of Chemistry University of York, Heslington, YO10 5DD, York, England http://www.york.ac.uk/chemist
Re: [ccp4bb] Off topic: Flourescence anisotropy measurement
May i ask, whether the fluoresnce anisotropy method was reliable enough to determine the stoichiometry of a protein complex? 发自网易邮箱大师 在2017年07月22日 03:44,Phoebe A. Rice 写道: You might also be getting aggregation. If you do an old-fashioned EMSA ("gel shift") assay, does it hang up in the well? ++ Phoebe A. Rice Dept. of Biochemistry & Molecular Biology The University of Chicago pr...@uchicago.edu From: CCP4 bulletin board [CCP4BB@JISCMAIL.AC.UK] on behalf of Morgan Milton [eilise.mil...@gmail.com] Sent: Friday, July 21, 2017 9:44 AM To:CCP4BB@JISCMAIL.AC.UK Subject: Re: [ccp4bb] Off topic: Flourescence anisotropy measurement Completely agree, you need a higher DNA concentration. We have had luck with 10 nM DNA. Also, bubbles have a HUGE impact on how the fluorescent signal is measured. Make sure you spin your plates down (assuming you are using them) to remove any bubbles. We just had an undergrad read his anisotropy assay and the data looked horrible. He then realized he had not spun his plate down, after doing so the data was much more consistent. Are you doing technical replicates? We do at least triplicates per plate. All the best, Morgan Morgan E. Milton, PhD On Fri, Jul 21, 2017 at 9:48 AM, Opher Gileadi wrote: FP is the ratio between two fluorescence measurements; if the fluorescence signal is too low, you will still get a ratio but it will be essentially noise. Try to perform the measurements at 10-50 nM DNA. If your binding affinity is in the low nM range, you may have to use other methods to measure KD.
[ccp4bb] 回复:[ccp4bb] suggestions are welcome
please characterize your a and b to see they really what they are 发自网易邮箱大师 在2017年07月18日 10:25,高艺娜 写道: Hi all , It has been reported the Negative stain EM of a protein A-B complex, but according to my gel filtration results (I purified A and B respectively for incubation) , I found that A could not bind to B, of course I tried different buffer condition with various pH value, even the binding condition only had 50 mm Kcl. Do you have any suggestion or methods that I can try to get the protein A-B complex? Any suggestion is welcome, Thank you all , Best,
Re: [ccp4bb] long loop
Fab fragment binding towards long loop 在2017年07月04日 23:22,dongxiaofei 写道: Dear ALL, I want to make a protein crystal,but there is a long loop between domains of protein , which contains two small domains owning about 40 amino acids respectively and a loop about 70 amino acids. Loop is so long and flexible ,but I don't want to delete some fragments,because it may be important for protein's function of a histone demethylase. Besides, the surface charge of protein is whole negative . I have tried a long time but it is hard to me to get crystal. Would be very grateful for any advice! Thanks Dong Xiao
Re: [ccp4bb] Separating Monomers and Dimers
it was not stable for frozen storage. if necessary,using protein fresh without frozen 发自网易邮箱大师 在2017年06月27日 20:22,jai mohan 写道: Dear all, I am working on a Red Fluorescent Protein (His-Tag) molecular weight around 27kDa. After purification I ran a SDS page, the band at 27kDa confirms the monomer. The protein was stored at -20C, a week later again I ran a gel, this time I saw another new band between 50-60kDa, it confirms the protein solution contains both monomers and dimers. I would like to know, what is the best way to separate the monomers and dimers? One of my colleague advice me to go for sucrose gradient centrifugation and size exclusion chromatography. However, I seek all your valuable suggestions and advice. With best regards Dr. S.M.Jaimohan
[ccp4bb] Fw:[ccp4bb] on Cell & Symmetry in coot
Dear All, I mean if the radius set in the Coot "Cell Symmetry" was too small, not enough monomers (less than 6) can be displayed to show the "continuous helix with a six-fold screw axis". If the radius was too large, as for the "continuous helix with a six-fold screw axis" can be regarded as a "rod", too large radius will lead to show several rods in one window. But with the Coot window, it cannot distinguish which monomer was from which rod. Thus I cannot identify 6 monomers forming the single rod, i.e., a "continuous helix with a six-fold screw axis". Can anyone explain in this situation how can I identify the 6 monomers in the Coot "Cell and SYmmetry" windows forming a single "continuous helix with a six-fold screw axis"? Smith Forwarding messages From: "Smith Lee" <0459ef8548d5-dmarc-requ...@jiscmail.ac.uk> Date: 2016-12-09 18:12:20 To: CCP4BB@JISCMAIL.AC.UK Subject: [ccp4bb] on Cell & Symmetry in coot Dear All, There is a pdb, once opended in coot, it was a monomer (space group P 65 2 2 ). But in the correspondence paper, it writes, "the subunits form a continuous helix with a six-fold screw axis". I have tried to view with Coot the "six-fold screw axis" formed by 6 monomers. But in the "Cell & Symmetry" in Coot, if the radius is small, 6 monomers cannot be shown. If I increase the radius, more than 6 monomers would occur in the window, and it can hardly distinguish the 6 monomers forming the "six-fold screw axis". In this situation, will you please let me know how to use Coot to identify the 6 monomers forming the "six-fold screw axis"? In addition, suppose 6 monomers forming the "six-fold screw axis" have been identified in Coot, in order to save the pdb of each monomer, I need to click each monomer in mouse, then by "Save symmetry coordinates" to save the pdb of each monomer, right? I am looking forward to getting your reply. Smith
[ccp4bb] on R-free label
Dear All, Will you please tell me how to know whether my mtz file has the R-free label or not? SMith
Re: [ccp4bb] Rfree below Rwork
If both the PDB and mtz for the pdb have been assigned to P1 space group for some reason, can this lead to Rwork higher than Rfree during refinement? If after converting my PDB and mtz to P1 space group, and I have forgotten what is the original space group for my PDB and mtz before conversion to P1 space group, is any method which can recover the original space group for my PDBand mtz, so that in the following refine Rwork would be lower than Rfree? Smith At 2015-07-01 01:55:22, "Eleanor Dodson" wrote: I suppose if I was the referee for this structure and your FreeR is so close to the Rfactor I would ask you to ensure you had the right space group - is the 6 fold NCS actually 2 fold NCS with a crystallographic 3 fold.. Cases occur where R32 is indexed as C2.. Certainly if the Rfree set is assigned randomly to reflections which are symmetry equivalents then you see this phenomena of Rfree = Rfactor Eleanor On 30 June 2015 at 18:26, Gerard Bricogne wrote: Dear Wolfram, I have a perhaps optimistic view of the effect of high-order NCS on Rfree, in the sense that I don't view it as a "problem". People have agonised to extreme degrees over the "difficulty" of choosing a free set of reflections that would produce the expected gap between Rwork and Rfree, and some of the conclusions were that you would need to hide almost half of your data in some cases! I think it is best to remember that the idea of cross-validation by Rfree is to prevent overfitting, i.e. ending up with a model that fits the amplitudes too well compared to how well it determines the phases. In the case of high-order NCS (in your case, the U/V ratio that the old papers on NCS identified as the key quantity to measure the phasing power of NCS would be less than 0.1!) the phases and the amplitudes are so tightly coupled that it is simply impossible to fit the amplitudes without delivering phases of an equally good quality. In other words there is no overfitting problem (provided you do have good and complete data) and the difference between Rfree and Rwork is simply within the bounds of the statistical spread of Rfree depending on the free set chosen. You are lucky to have 6-fold NCS, so don't let any reviewer convince you that it is a curse, and make you suffer for it :-) . With best wishes, Gerard. -- On Tue, Jun 30, 2015 at 12:58:44PM -0400, wtempel wrote: > Hello, > my question concerns refinement of a structure with 6-fold NCS (local > automatic restraints in REFMAC) against 2.8 A data. The size of my free set > is 1172 selected in thin resolution shells (SFTOOLS) and corresponding to > 4.3 % of reflections. > A refmac run of 10 cycles of TLS and 10 cycles of CGMAT starts out at > Rfree/Rcryst 0.271/0.272. After the 10th TLS cycle I have 0.227/0.224. Yes, > Rfree < Rcryst. At the end of CGMAT I have 0.2072/0.2071. > I understand that NCS stresses the independence assumption of the free set. > Am I correct in believing that Rfree *may* be smaller than Rcryst even in > the absence of a major mistake? My hope is that the combined wisdom of > ccp4bb followers can point out my possible mistake, suggest tests that I > may perform to avoid them and, possibly, arguments in defense of a > crystallographic model with Rfree < Rcryst. > Many thanks, > Wolfram Tempel -- === * * * Gerard Bricogne g...@globalphasing.com * * * * Global Phasing Ltd. * * Sheraton House, Castle Park Tel: +44-(0)1223-353033 * * Cambridge CB3 0AX, UK Fax: +44-(0)1223-366889 * * * ===
[ccp4bb] on mosaicity estimation by iMosflm
Dear All, When I do mosaicity estimation by iMOSFLM, it shows, "The mosaicity estimation has not worked for some reason. Message from Mosflm is - Unable to estimate mosaicity automatically from this image-determine a value visually. You should enter an estimated value to replace 0.05". First I have input a lot of images, I think " Unable to estimate mosaicity automatically from this image-determine a value visually" should be " Unable to estimate mosaicity automatically from theseimages-determine a value visually". Secondly, will you please tell me how can I determine a value visually? And in which step I can get a reliable mosaicity estimation? I am looking forward to getting a reply from you. Smithy.
[ccp4bb] on secondary structure restraint
Dear All, Even with the Coot secondary structure (for example helix) restraint selected and by this way we keep the secondary structure, I find the protein secondary structure formed in this way was not so typpical, for example, not all CO ( i) and NH (i+4) forms H-bonds, and there are H-bonds formed by CO ( i) and NH (i+3). I checks some RCSB PDBs, and find this kind of untypical H-bonds for the Helix backbone were not rare. Do we accept this kind of untypical H-bonds containing PDB as normal, or do we think we need further refine the structure based on the map? Smith
[ccp4bb] on resolution and explaination of the intersubunit bonds
Dear All, In order to acceptably explain the salt bridges, hydrophobic interactions and H-bonds among subunits in the crystal structure of a protein complex, is there a threshold resolution of the crystal, for example, if the crystal is poorer than 4A or 5A, the crystal structure solved cannot be used to acceptably explain the intersubunit interactions at the non-covalen bond level? Smith
[ccp4bb] on b-factor
Dear All, In the PDB file, the b-factors were only determined by the quality of the map, is this view right or not? Smith
[ccp4bb] HIS related crystallography issue
Dear All, Suppose my protein has 4 same subunits (not exactly 4 subunits, in order to explain things clear, it contains not less than 2 identical subunitss), each subunit has the potential H-bond involving N of HIS. I have run the optimization by both PDB_REDO and the Phenix refine with the phenix flips checked. I have checkecd the PDB_REDO refine results and the Phenix refine results, the answer is, suppose in PDB_REDO it tells us subunit A has that specific H-bond formed, Phenix refine rells us that subunit B has that specific H-bond formed. In fact, supposing it has 4 subunits, the symmetry determines that the 4 subunits should be identical, and they should all contains the specific H-bonds, or all do not contains the specific H-bonds. Any more suggestions welcome. Smith At 2015-05-20 22:55:13, "Robbie Joosten" wrote: Hi Smith, Just to contrast Pavel's phenix plug. PDB_REDO does HQN-flips automatically based on WHAT _CHECK results and refines your model in Refmac. Cheers, Robbie Sent with my Windows Phone Van: Robbie Joosten Verzonden: 20-5-2015 14:30 Aan: Smith Liu; CCP4BB@JISCMAIL.AC.UK Onderwerp: RE: [ccp4bb] HIS related crystallography issue You can typically assign the histidine orientation based on analysis of the hydrogen bond network. In ambiguous cases you might have to look a few residues deep. WHAT_CHECK does this for you by a global optimization of the hydrogen bond network. Cheers, Robbie Sent with my Windows Phone Van: Smith Liu Verzonden: 20-5-2015 14:12 Aan: CCP4BB@JISCMAIL.AC.UK Onderwerp: [ccp4bb] HIS related crystallography issue Dear All, Suppose the protein crystal resolution is about 2-3A, then in the map it should be rather difficult to distinguish the C and N in the sidechain of HIS. In this way we may regard the sidechain of HIS is flippable. But suppose in one flipped conformation of the HIS, the free N in the sidechain of HIS can form H-bond with the neighbour H of the OH of the Thr, in the other flipped conformation of the HIS, the free N in the sidechain of HIS cannot form H-bond with the neighbour H of the OH of the Thr (caused by distance issue). Suppose whether the H-bond forms between the free N in the sidechain of HIS and the neighbour H of the OH of the Thr was very important biologically, in this situation how can we distinguish whether the H-bond forms? Smith
[ccp4bb] HIS related crystallography issue
Dear All, Suppose the protein crystal resolution is about 2-3A, then in the map it should be rather difficult to distinguish the C and N in the sidechain of HIS. In this way we may regard the sidechain of HIS is flippable. But suppose in one flipped conformation of the HIS, the free N in the sidechain of HIS can form H-bond with the neighbour H of the OH of the Thr, in the other flipped conformation of the HIS, the free N in the sidechain of HIS cannot form H-bond with the neighbour H of the OH of the Thr (caused by distance issue). Suppose whether the H-bond forms between the free N in the sidechain of HIS and the neighbour H of the OH of the Thr was very important biologically, in this situation how can we distinguish whether the H-bond forms? Smith
[ccp4bb] how to actively assign H to N of His
Dear All, After CCP4 or phenix refinement, I find the H position of N in specific His residues needs to be regularized based on the function of that His. Will you please advise on how to actively assign H to one or two of the 2 N in the His residue in the PDB file? Or do we have a server, which can help us to assign H to the N of the His in the PDB based on the function of the His, or based on our intention which N we need to have H in the His? In addition, after H regularuzed in the His in the PDB, do you think whether further CCP4 or phenix refinement is needed? Smith
[ccp4bb] on the contour level in Coot
Dear All, For the contour level in the Properties in the Dssplay of the Display Manager in Coot, the contour level should be exacly the sigma value we see in the published crystallography paper, am I right? But I have paid attention to a paper which has a sigma level of 6-7 for its densty. Will you please explain why the sigma can be so high and how can we make it so high? Smith
[ccp4bb] on space group
Dear All, Alhough there are on-line explainations on the space group, I found it was difficult to fully understand. Here we take P 1 21 1 as an exmaple, will you please explain to me with easy language what each number indicates?Or do we have a on-line server which can demonstrate the meaning of each number? How can we get out crystal arrangement with repeating the unit cell in the format of space group? Smith
[ccp4bb] on the contour level in Coot
If for both a mtz density and mrc map I set the contour level as 0.15, does the 015 has the comparable significance for the mtz density and mrc map? Smith
Re: [ccp4bb] self-rotation in the absence of NCS
I recently solved a 4-identical chain protein structure. First I got it from initial model with NCS, good enough. Then from the initial model I process it without NCS, quality better than solved with NCS. Smith At 2015-04-30 02:07:31, "Eleanor Dodson" wrote: Hmm - I think these peaks MUST be related to some internal symmetry in the 1200 aa solved structure. Is there some arrangement of helices or other features which are replicated in another part of the structure? A phi value of ~ 19 degrees can't be explained by a different related space group I don't think Eleanor On 29 April 2015 at 15:56, Ian Tickle wrote: Peer, you didn't say which program you are using for this? Polarrfn or Molrep? Do you get the same results with both programs? Also did you try sharpening the Fs with Ecalc and/or using all your data? In my experience sharpening works better with self- than with cross-rotation functions because the differences between NCS-related monomers are usually much less those than between non-isomorphous ones. Cheers -- Ian On 29 April 2015 at 15:00, Peer Mittl wrote: Zbyszek Otwinowski and Fred Vellieux suggested to run the self-rotation on Fcalcs. This suggestion "solved" the problem, since there are similar peaks on the kappa=180° planes as well. However, I wasn't able to get rid of those peaks by playing around with resolution and integration radius. I must say that I am surprized, because - as Eleanor pointed out - I also expected to find peaks on the kappa=180° planes only in case of P6522 symmetry. Anyway, this experience reminds me to run some simple tests beforehand. -Peer On 29.04.2015 15:31, Eleanor Dodson wrote: Well - PG P6/mmm (possible SG P6522) will have peaks at kappa = 180 omega = 90 phi = 0 30 60 etc.. But if there is only one molecule / asymm unit there cant be an extra 2-fold. How big are the relative domains? Your interesting domain couldnt just be cleaved off could it? Eleanor On 29 April 2015 at 12:59, Peer Mittl mailto:mi...@bioc.uzh.ch>> wrote: We are working with a multi-domain protein crystallized in SG P6_5 with one molecule per asymmetric unit. The structure was refined at 2.00 A resolution with reasonable R-factors but unfortunately the domain we are most interested in seems to be disordered. Interestingly, the self-rotation function shows peaks on the kappa=180° plane (omega=90°, phi=19° (and every 30°)), with more than 50% origin peak height. Therefore, we are wondering if perhaps the space group assignment might be sub-optimal. Any explanations how these self-rotation peaks could occur and how we could extract meaningful information to resolve the disordered domain are welcome. Best regards, Peer P.S. Some additional information: pointless suggests SG P6_5, the data doesn't seem to be twinned (L-test), the refined part of the structure has no "internal symmetry" and refinement in P1 doesn't reveal the "lost" domain.
[ccp4bb] on the electron microscopy function of refmac5
Dear All, The CCP4 document says refmac can process electron microscopy map for refinement. But I cannot localize that function in the refmac5 of the CCP4. Will you please advise how to have the refmac process the electron microscopy map? Smith
Re: [ccp4bb] on NCS restraint
Dear Jurgen, My understanding is that NCS restraint can significantly enhance the speed of calculation, but considering the subunits even with the eactly same sequence may not be identical, to have NCS restraint may be not necessary or may be not good for the refinement, am I right? Smith At 2015-04-17 09:09:05, "Jurgen Bosch" wrote: yes. Have two sets of NCS operators one that describe the four subunits and one describing the two subunits. If during the refinement of your structure you should find out that the subunits are not identical to each other you can relax the NCS weights. Jürgen .. Jürgen Bosch Johns Hopkins University Bloomberg School of Public Health Department of Biochemistry & Molecular Biology Johns Hopkins Malaria Research Institute 615 North Wolfe Street, W8708 Baltimore, MD 21205 Office: +1-410-614-4742 Lab: +1-410-614-4894 Fax: +1-410-955-2926 http://lupo.jhsph.edu On Apr 16, 2015, at 9:02 PM, Smith Lee <0459ef8548d5-dmarc-requ...@jiscmail.ac.uk> wrote: Dear All, If a protein contains 6 subunits, 4 subunits from the same sequence (subunit A, B, C, D all from the same sequence), each of the 2 other subunits from 2 diffrent sequences (subunit E from the second sequence, subunit F from the third sequence), in this situation should I use NCS restraint or not? If my protein contains 2 subunits, both of the 2 subunits composed of the eaxctly same sequence, however supposing the 2 subunits have a little diffrent conformation, in this situation should we use NCS retraint or not? Smith
Re: [ccp4bb] Problem in optimization
Additive screening At 2015-04-11 18:00:21, "高艺娜" wrote: >Dear all >I have tried a variety of methods on regular optimization of the crystal ,but >all have failed,the 17 kDa protein crystal still have many nucleation and poor >diffraction also poor resolution(the best 3.5-4.0Å) >The crystallization conditions are: >0.1 M Tris pH-6.5, >0.2 M Sodium acetate >1.8 M Ammonium sulphate > >Could some one suggest me how to trouble shoot this problem? > >Best wishes,
[ccp4bb] on building a helix fragment by coot
Dear All, Caused by poor resolution, I can only build a series of Ala into my helix density (for sure the density is for helix). Then I tried to use the dock sequence in the extension of the Coot to change to my target sequence. Coot says it was not confidence on the alignment, thus it did not change the Alas into my target sequence. In the "Molecule to be sequenced" in the coot dock sequence, if we paste the sequence, show we still need to input the pir file? For the sequenc we paste, it should be my target sequence rather than the series of Alas , right? Why in this graphical interface there are "Sequence closest fragment" and "Sequnece all fragments!"? I ask these questions in order to avoid my failling to do something which Coot can do. I am looking forward to getting your reply. Smith Forwarding messages From: "Smith Liu" Date: 2015-04-01 12:58:01 To: lists...@jiscmail.ac.uk,"CCP4BB@JISCMAIL.AC.UK" Subject: on building a helix fragment by coot Dear All, If I need to build a fragment of helix to a fragment of density by coot, should I use baton build method or should I use place Helix here function? I have tried both, but I find it is difficult to place the sidechains to the map mosition which looks like where shold be occupied by the sidechains. I am looking forward to getting your message on how to correctly buld the helix. Best regards. Smith
Re: [ccp4bb] on building a helix fragment by coot
Dear Rhys, My difficulty is, suppose I have 20 residues (just an example), the first 8 sidechains fit the dnsity, the last 8 fit the density, however for the middle 4 residues, the map can hold 5 sidechains or hold 3 sidechains, not exactly 4 sidechains. This is caused by the poor resolution. But my boss needs me to get a good fit. Can you give more advices? Smith At 2015-04-01 13:01:38, "RHYS GRINTER" wrote: >Hi Smith, > >What resolution are you working at and what kind of phases do you have >(experimental?, good/bad?)? >I've had the most success using the place helix function, then trimming it >back so it fits the denisty. Once the helix is place you can try rigid body >fitting it so it is orientated correctly for adding sidechains. >If your resolution and or phases are poor however it can be difficult to fit >fragments into density. > >Good luck, > >Rhys > >From: CCP4 bulletin board [CCP4BB@JISCMAIL.AC.UK] On Behalf Of Smith Liu >[smith_liu...@163.com] >Sent: 01 April 2015 05:58 >To: CCP4BB@JISCMAIL.AC.UK >Subject: [ccp4bb] on building a helix fragment by coot > >Dear All, > >If I need to build a fragment of helix to a fragment of density by coot, >should I use baton build method or should I use place Helix here function? I >have tried both, but I find it is difficult to place the sidechains to the map >mosition which looks like where shold be occupied by the sidechains. > >I am looking forward to getting your message on how to correctly buld the >helix. > >Best regards. > >Smith > >
[ccp4bb] on building a helix fragment by coot
Dear All, If I need to build a fragment of helix to a fragment of density by coot, should I use baton build method or should I use place Helix here function? I have tried both, but I find it is difficult to place the sidechains to the map mosition which looks like where shold be occupied by the sidechains. I am looking forward to getting your message on how to correctly buld the helix. Best regards. Smith
[ccp4bb] on refinement by coot
Dear All, Suppose there is a 10-residue helix, there are 2 of the 10 Calphas do not position so well so the sidechains of the 2 Calphas do not align with the density. Is any Coot commands which can refine the 2 Calpha positions and the corresponding sidechains? By the way, if I use Wincoot, will you let me know how to use Wincoot by inputing command? Smith
[ccp4bb] on Baton build
Dear All, In coot when I try to build a peptide by baton method, I find the starting point of the baton starts from somewhere in the sidechain rather than from the Calpha position. In addition, when I try to changing the starting residue by Baton-build params, the starting point of the Baton does not change residue at all. Will you please tell me how to setting the issue? Smith
[ccp4bb] on openning the PDB file and the mtz file by Coot
Dear All, When we by Coot open the PDB fle and the mtz file from the same refinement, the protein backbone (and the sidechains) and the electron density always fit automatically. Will you please tell me the mechanism of the Coot how the PDB file automatically fit the mtz file in its graphical window? Suppose I have a PDB file and a mtz file (PDB file from protein A, mtz file from protein B, which is a homology protein of protein A) which are not from the same refinment (thus not fit automatically in Coot), will you please tell me what modification I should make on the files in order to have the Coot to fit the protein bakbone (and the sidechains) and the electron density? I am looking forward to getting your reply. Smith
Re: [ccp4bb] how to recover my data
Dear All, For the corrupted mtz file, if we open it by notepad, it would be messy code or unreadble code. In addition, not all the mtz files in the damaged hardware were corrupted, some mts files are still readable by Coot. Do you have any suggestions to correct the unreadbale mtz files to readable? Smith At 2015-03-07 22:51:12, "Zhijie Li" wrote: Smith, As Ian said, mtzdump can give you some clues, but will not solve your problem. The file that mtzdump could not read apparently has changed length due to some sort of insertion. The other file that mtzdump could process has the correct length and the stats look reasonable, so it might be OK, although we still can not rule out the possibility that a few numbers might have been changed in the dataset. Have you tried to do anything with the latter file, such as a few rounds of refinement? What did you see? If you send me the two mtz I can take a look and make some guesses. If the problem was as simple as introduction of some 0D 0A insertion/deletion it can be fixed. If you can consult the technician who recovered the data and tell us what software he used it would be helpful. Zhijie From:Smith Liu Sent: Saturday, March 07, 2015 9:24 AM To:CCP4BB@JISCMAIL.AC.UK Subject: Re: [ccp4bb] how to recover my data Thanks Ian. I got it from the bin folder you mentioned. A moment ago I have failed to localize it from the CCP4 graphical interface. Smith At 2015-03-07 21:13:58, "Ian Tickle" wrote: Hi Smith Not sure what you mean, the current version of CCP4 (as have all previous versions) certainly does contain mtzdump: > which mtzdump /software/CCP4-6.5.0/ccp4-6.5/bin/mtzdump The "limited output" is intentional: mtzdump by default only lists the first 10 reflexions, and you have exactly 10 in your list. If you want more then supply an NREF value, e.g. echo nref 9 |mtzdump HKLIN my.mtz |less But mtzdump (or any MTZ utility for that matter) is not going to help you with a corrupted file, since the MTZ read routines assume that the file has a strictly defined format which has most likely been modified when it was transferred from the damaged disk, and is therefore now unreadable by any program that assumes the defined format. What does the technician who transferred the data have to say about it? He is the only person who can possibly tell you how the file has been corrupted. What you need is a hex dump of the file (e.g. using 'hexdump') and someone who understands a) the correct format and b) exactly how it was corrupted. Then it may be possible to write a bespoke program tor recover your data. But that is not going to be easy: it will require someone with some expertise in programming. Attempting to read the file with random programs has even less chance of working than I have of winning the lottery! As others have suggested there are much easier ways of recovering your data (such as reprocessing the images). Cheers -- Ian On 7 March 2015 at 06:29, Smith Liu wrote: Dear All, The current version of CCP4 does not contain mtzdump. I use UCLA MBI — mtzdump to process it. For a mtz file, the output says there was no valid reflection data. For another mtz file, it only gave about 50 lines information (not a new mtz file). As for the original mtz is very large, I think at least some message in the mtz file has been missing or has not been recovered by UCLA MBI — mtzdump. Please feel free for further advise. Smith Follwing: output from the UCLAMBI-mtzdump mtzdump - dump data from an MTZ reflection data file. A CCP4 program Your mtzdump Results Cell Dimensions : (obsolete - refer to dataset cell dimensions above) 130.3260 130.3260 360.3400 90. 90. 90. * Resolution Range : 0.30.22676 (180.170 - 2.100 A ) * Sort Order : 1 2 3 0 0 * Space group = 'I 4 2 2' (number 97) | | | 1 ASC 0 62 0 100.00 32.5 32.5 180.17 2.10 H H | | | 2 NONE 0 43 0 100.00 13.4 13.4 180.17 2.10 H K | | | 3 NONE 0 171 0 100.00 64.4 64.4 180.17 2.10 H L | | | 4 NONE0.019.0 0 100.00 9.51 9.51 180.17 2.10 I FreeR_flag | | | 5 NONE 65.8466807.5 47890 47.08 14985.84 14985.84 41.21 2.10 J IMEAN | | | 6 NONE 22.7 37273.6 47890 47.08 1161.73 1161.73 41.21 2.10 Q SIGIMEAN | | | 7 NONE 65.8466807.5 50197 44.54 15601.05 15601.05 41.21 2.10 K I(+) | | | 8 NONE 36.9 37273.6 50197 44.54 1723.38 1723.38 41.21 2.10 M SIGI(+) | | | 9 NONE 65.8466807.5 53778 40.58 16626.28 16626.28 41.21 2.10 K I(-) | | | 10 NONE 36.9 37273.6 53778 40.58 1568.40 1568.40 41.21 2.10 M SIGI(-) | | | 11 NONE 67.3 6770.8 47890 47.08 1006.23 1006.23 41.21 2.10 F F | | | 12 NONE5.6 446.6 47890 47.
Re: [ccp4bb] how to recover my data
Thanks Ian. I got it from the bin folder you mentioned. A moment ago I have failed to localize it from the CCP4 graphical interface. Smith At 2015-03-07 21:13:58, "Ian Tickle" wrote: Hi Smith Not sure what you mean, the current version of CCP4 (as have all previous versions) certainly does contain mtzdump: > which mtzdump /software/CCP4-6.5.0/ccp4-6.5/bin/mtzdump The "limited output" is intentional: mtzdump by default only lists the first 10 reflexions, and you have exactly 10 in your list. If you want more then supply an NREF value, e.g. echo nref 9 |mtzdump HKLIN my.mtz |less But mtzdump (or any MTZ utility for that matter) is not going to help you with a corrupted file, since the MTZ read routines assume that the file has a strictly defined format which has most likely been modified when it was transferred from the damaged disk, and is therefore now unreadable by any program that assumes the defined format. What does the technician who transferred the data have to say about it? He is the only person who can possibly tell you how the file has been corrupted. What you need is a hex dump of the file (e.g. using 'hexdump') and someone who understands a) the correct format and b) exactly how it was corrupted. Then it may be possible to write a bespoke program tor recover your data. But that is not going to be easy: it will require someone with some expertise in programming. Attempting to read the file with random programs has even less chance of working than I have of winning the lottery! As others have suggested there are much easier ways of recovering your data (such as reprocessing the images). Cheers -- Ian On 7 March 2015 at 06:29, Smith Liu wrote: Dear All, The current version of CCP4 does not contain mtzdump. I use UCLA MBI — mtzdump to process it. For a mtz file, the output says there was no valid reflection data. For another mtz file, it only gave about 50 lines information (not a new mtz file). As for the original mtz is very large, I think at least some message in the mtz file has been missing or has not been recovered by UCLA MBI — mtzdump. Please feel free for further advise. Smith Follwing: output from the UCLAMBI-mtzdump mtzdump - dump data from an MTZ reflection data file. A CCP4 program Your mtzdump Results Cell Dimensions : (obsolete - refer to dataset cell dimensions above) 130.3260 130.3260 360.3400 90. 90. 90. * Resolution Range : 0.30.22676 (180.170 - 2.100 A ) * Sort Order : 1 2 3 0 0 * Space group = 'I 4 2 2' (number 97) | | | 1 ASC 0 62 0 100.00 32.5 32.5 180.17 2.10 H H | | | 2 NONE 0 43 0 100.00 13.4 13.4 180.17 2.10 H K | | | 3 NONE 0 171 0 100.00 64.4 64.4 180.17 2.10 H L | | | 4 NONE0.019.0 0 100.00 9.51 9.51 180.17 2.10 I FreeR_flag | | | 5 NONE 65.8466807.5 47890 47.08 14985.84 14985.84 41.21 2.10 J IMEAN | | | 6 NONE 22.7 37273.6 47890 47.08 1161.73 1161.73 41.21 2.10 Q SIGIMEAN | | | 7 NONE 65.8466807.5 50197 44.54 15601.05 15601.05 41.21 2.10 K I(+) | | | 8 NONE 36.9 37273.6 50197 44.54 1723.38 1723.38 41.21 2.10 M SIGI(+) | | | 9 NONE 65.8466807.5 53778 40.58 16626.28 16626.28 41.21 2.10 K I(-) | | | 10 NONE 36.9 37273.6 53778 40.58 1568.40 1568.40 41.21 2.10 M SIGI(-) | | | 11 NONE 67.3 6770.8 47890 47.08 1006.23 1006.23 41.21 2.10 F F | | | 12 NONE5.6 446.6 47890 47.0876.6976.69 41.21 2.10 Q SIGF | | | 13 NONE -514.8 537.4 56085 38.03 0.2187.57 41.21 2.10 D DANO | | | 14 NONE0.0 380.1 56085 38.0391.6391.63 41.21 2.10 Q SIGDANO | | | 15 NONE 67.3 6770.8 50197 44.54 1032.56 1032.56 41.21 2.10 G F(+) | | | 16 NONE8.2 446.6 50197 44.5490.1390.13 41.21 2.10 L SIGF(+) | | | 17 NONE 67.3 6770.8 53778 40.58 1075.89 1075.89 41.21 2.10 G F(-) | | | 18 NONE7.6 446.6 53778 40.5878.6778.67 41.21 2.10 L SIGF(-) | | | 19 NONE 0 2 47890 47.08 0.4 0.4 41.21 2.10 Y ISYM | | | | | LIST OF REPLECTIONS === 0 0 28.00 ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? 0 0 4 10.00 ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? 0 0 69.00 ? ? ? ? ? ? ? ? ? ? ? ? ?
Re: [ccp4bb] how to recover my data
Dear All, The current version of CCP4 does not contain mtzdump. I use UCLA MBI — mtzdump to process it. For a mtz file, the output says there was no valid reflection data. For another mtz file, it only gave about 50 lines information (not a new mtz file). As for the original mtz is very large, I think at least some message in the mtz file has been missing or has not been recovered by UCLA MBI — mtzdump. Please feel free for further advise. Smith Follwing: output from the UCLAMBI-mtzdump mtzdump - dump data from an MTZ reflection data file. A CCP4 program Your mtzdump Results Cell Dimensions : (obsolete - refer to dataset cell dimensions above) 130.3260 130.3260 360.3400 90. 90. 90. * Resolution Range : 0.30.22676 (180.170 - 2.100 A ) * Sort Order : 1 2 3 0 0 * Space group = 'I 4 2 2' (number 97) | | | 1 ASC 0 62 0 100.00 32.5 32.5 180.17 2.10 H H | | | 2 NONE 0 43 0 100.00 13.4 13.4 180.17 2.10 H K | | | 3 NONE 0 171 0 100.00 64.4 64.4 180.17 2.10 H L | | | 4 NONE0.019.0 0 100.00 9.51 9.51 180.17 2.10 I FreeR_flag | | | 5 NONE 65.8466807.5 47890 47.08 14985.84 14985.84 41.21 2.10 J IMEAN | | | 6 NONE 22.7 37273.6 47890 47.08 1161.73 1161.73 41.21 2.10 Q SIGIMEAN | | | 7 NONE 65.8466807.5 50197 44.54 15601.05 15601.05 41.21 2.10 K I(+) | | | 8 NONE 36.9 37273.6 50197 44.54 1723.38 1723.38 41.21 2.10 M SIGI(+) | | | 9 NONE 65.8466807.5 53778 40.58 16626.28 16626.28 41.21 2.10 K I(-) | | | 10 NONE 36.9 37273.6 53778 40.58 1568.40 1568.40 41.21 2.10 M SIGI(-) | | | 11 NONE 67.3 6770.8 47890 47.08 1006.23 1006.23 41.21 2.10 F F | | | 12 NONE5.6 446.6 47890 47.0876.6976.69 41.21 2.10 Q SIGF | | | 13 NONE -514.8 537.4 56085 38.03 0.2187.57 41.21 2.10 D DANO | | | 14 NONE0.0 380.1 56085 38.0391.6391.63 41.21 2.10 Q SIGDANO | | | 15 NONE 67.3 6770.8 50197 44.54 1032.56 1032.56 41.21 2.10 G F(+) | | | 16 NONE8.2 446.6 50197 44.5490.1390.13 41.21 2.10 L SIGF(+) | | | 17 NONE 67.3 6770.8 53778 40.58 1075.89 1075.89 41.21 2.10 G F(-) | | | 18 NONE7.6 446.6 53778 40.5878.6778.67 41.21 2.10 L SIGF(-) | | | 19 NONE 0 2 47890 47.08 0.4 0.4 41.21 2.10 Y ISYM | | | | | LIST OF REPLECTIONS === 0 0 28.00 ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? 0 0 4 10.00 ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? 0 0 69.00 ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? 0 0 8 18.00 ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? 0 0 103.00 ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? 0 0 12 17.00 3090.57268.91 3090.57268.91 3090.57 268.91554.30 24.36 0.00 0.00554.30 24.36554.30 24.36 1.00 0 0 149.00 23242.80 1362.28 23242.80 1362.28 23242.80 1362.28 1522.33 44.83 0.00 0.00 1522.33 44.83 1522.33 44.83 1.00 0 0 165.00 29139.17 1697.95 29139.17 1697.95 29139.17 1697.95 1704.50 49.90 0.00 0.00 1704.50 49.90 1704.50 49.90 1.00 0 0 18 12.00478.99 57.42478.99 57.42478.99 57.42217.65 13.29 0.00 0.00217.65 13.29217.65 13.29 1.00 0 0 203.00 113229.90 6443.99 113229.90 6443.99 113229.90 6443.99 3358.69 96.10 0.00 0.00 3358.69 96.10 3358.69 96.10 1.00 end, thus very limited information output At 2015-03-06 15:56:19, "Tim Gruene" wrote: >-BEGIN PGP SIGNED MESSAGE- >Hash: SHA1 > >Dear Smith, > >an mtz-file is a binary file and none of the programs you list is >suitable to open it in a meaningful manner. > >Why don't you follow the advice you were alre