[ccp4bb] Molecular Replacement
Hi All; I have a native data set of membrane protein at 3.8A. I nearly use all the options for Molrep. I would like to ask, is some body has some special strategy which can work for difficult Molrep Thanks you in Advance Bashir -- Muhammad Bashir Khan ** Department Of Biochemistry University of Alberta, Edmonton Canada
[ccp4bb] MR_ROSETTA
Hi All; Could some body explain why I am getting the error message when running MR-Rosetta. Message is " Sorry cannot locate a binary starting with mr_protocol.default' in the directory /home/phenix/rosetta_2014.34.57213_bundle/main/source/bin I just download it and locate it in the preferences, wizard and then path to Rosetta. Thanks Bashir -- Muhammad Bashir Khan ** Department of Biochemistry University of Alberta, Canada Canada
Re: [ccp4bb] HKL2000 Display
Hi All; Thanks everybody. It was wrong detector site file. Now its working. Thanks Bashir On Wed, December 3, 2014 17:19, David Schuller wrote: > Yes, it is probably the wrong detector type, or the wrong parameters, > perhaps binned vs. unbinned. > > > On 12/03/14 07:25, David Waterman wrote: >> Hi Muhammed, >> >> It looks a lot to me like denzo thinks your detector has a larger >> image size than the actual number of elements in the array. However, I >> don't use denzo so I'm not commenting from experience. >> >> Cheers >> >> -- David >> >> On 3 December 2014 at 04:09, Muhammed bashir Khan >> > <mailto:muhammad.bashir.k...@univie.ac.at>> wrote: >> >> Hi All; >> >> Could somebody explain why my Display image in HKL2000 look like >> that. >> Image attached. >> Thanks for help in advance. >> >> Bashir >> >> >> Department of Biochemistry >> University of Alberta >> Edmonton Canada >> >> >> > > > -- > === > All Things Serve the Beam > === > David J. Schuller > modern man in a post-modern world > MacCHESS, Cornell University > schul...@cornell.edu > > -- Muhammad Bashir Khan ** Department for Structural and Computational Biology Max F. Perutz Laboratories University of Vienna Campus Vienna Biocenter 5 A-1030 Vienna Austria Austria Phone: +43(1)427752224 Fax: +43(1)42779522
[ccp4bb] Membrane Protein Crystal Optimization
Hi all; I have crystal of membrane protein (110 kDa) in several different conditions. I optimized several of them and tested it for their diffraction. It does not goes behind 10A at CLS and SSRL. The protein is purified by two steps nickel and gel filtration. The crystal grow very fast and reach to maximum size of 500*70*70 in approximately 30hrs. I mostly tried with different temperatures as well as additives. I am trying with Micro Batch crystallization in order to slow down the crystal growth. Any new idea which I can try will be highly appreciated from your own experience Thanks... -- Muhammad Bashir Khan ** Department of Biochemistry University of Alberta Edmonton Canada
[ccp4bb] Structure Refinement
Dear All I have a data at 2.75A. I process it in Space group P3121, using HKL3000. Run a molrep,find three molecule in a unit cell. I am trying to refine it with phenix, the R and R-free stuck at 34 and 41 respectively. Crystal: The crystal seems multiple thin plates and I tried to freeze the possible single crystal (plate). Any suggestion would be highly appreciated!! Bashir Muhammad Bashir Khan ** Structural Genome Consortium (SGC) University of Toronto Canada
Re: [ccp4bb] Crystal Optimization
Dear All, Thanks alot for your valuable suggestion.I hope I will find out the solution now. As far as to giveup is out of question Thanks once agin Regards; Bashir On Wed, July 11, 2012 05:25, Tuhin Bhowmick wrote: > Dear Muhammad, > > I had a similar case, and the crystals could indeed be optimized. A few > things to check first, > > 1) How long does it take for the needles to appear? Sometimes, if the > protein is degraded/ cleaved over time, a small population (possibly a > fragment of the whole >protein) from the heterogeneous mix can give similar crystals. Like > Bryan had suggested, it is also very useful to check the mol wt. of the > crystallized species through >mass spect/ native page. But do make sure to give the crystals serial > washes, so the test accounts for crystallizexd species, not the ones from > surrounding condition. > > 2) If the crystals indeed contain the protein of interest, they can be > used > for various seeding methods. I've got results from both streak and micro > seeding. > > Best, > > Tuhin. > > Tuhin Bhowmick > > Department of Physics > > Indian Institute of Science > > Bangalore: 560012 > Email: tuhin.i...@gmail.com > > On Wed, Jul 11, 2012 at 12:34 AM, Bernhard Rupp (Hofkristallrat a.D.) < > hofkristall...@gmail.com> wrote: > >> > Always give up. >> >> ** ** >> >> ...definitely not the kind of guy I want to sit up front in an >> airliner…** >> ** >> >> ** ** >> >> Best regards, BR >> >> - >> >> Bernhard Rupp, ATP-B737, CFII-MEI >> >> Vienna Air International >> >> Professional Aviation Services >> >> 001 (925) 209-7429 >> >> +43 (676) 571-0536 >> >> b...@vienna-air.com >> >> b...@ruppweb.org >> >> http://www.vienna-air.com/ >> >> - >> >> It is not your aptitude but your attitude >> >> that determines your altitude. (or your crystals) >> >> - >> >> ** ** >> >> ** ** >> >> ** ** >> >> From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of >> yybbll >> >> Sent: Tuesday, July 10, 2012 11:58 AM >> >> To: CCP4BB@JISCMAIL.AC.UK >> >> Subject: Re: [ccp4bb] Crystal Optimization >> >> ** ** >> >> Hi, >> >> >> >> In my experience, it is very very very difficult to optimize this needle >> like crystal. Always give up. >> >> >> >> Good luck! >> >> >> >> >> >> Dear All; >> >> Could somebody give a nice suggestion how the following type crystal >> could >> >> >> be optimized, I almost tried everything. >> >> Crystal Image is attached >> >> Crystal condition: 20% w/v PEG3350 and 200mM NaCl. >> >> Thanks in advance >> >> Bashir >> >> -- >> >> Muhammad Bashir Khan >> >> ** >> >> Structural Genome Consortium (SGC). University of Toronto >> >> Toronto, Canada >> > -- Muhammad Bashir Khan ** Department for Structural and Computational Biology Max F. Perutz Laboratories University of Vienna Campus Vienna Biocenter 5 A-1030 Vienna Austria Austria Phone: +43(1)427752224 Fax: +43(1)42779522
[ccp4bb] Structure Refinment problem
Dear all; I have a structure at 3.3A resolution of about 140kDa protein containing eight domains, in tetragonal space group.I also have the structure of most of the individual domains.I almost refined the structure in all the possible space groups, the best space group at the moment are the P42 with the R/Rfree of 32 and 38%. Could somebody suggest what else I should try to to get better R/Rfree values. I am using Phenix for refinment. Regarding the protein it contain quite flexible domains. Thanks in advance for your suggestions. Regards, Bashir -- Muhammad Bashir Khan ** Department for Structural and Computational Biology Max F. Perutz Laboratories University of Vienna Campus Vienna Biocenter 5 A-1030 Vienna Austria Austria Phone: +43(1)427752224 Fax: +43(1)42779522
Re: [ccp4bb] cryo protection
Hi Len; I was having exactly the same problem with my crystals, but when we grow the crystals in presence of increasing concentration of Glycerol and MPD starting from 0.5 to 10%. The crystal doesn't appear after 3% of Glycerol or MPD but the one which appear in 2.5 to 3 % were much resistant to cracking than the original crystals. Good luck Bashir On Wed, October 26, 2011 18:46, Leonard Thomas wrote: > Hi All, > > I have run into a very sensitive crystals system when it comes to cryo > protecting them. I have run through the usual suspects and trays are > going to be setup with a cryo protectant as part of crystallization > cocktail. The one problem that seems to be occurring is that the > crystals crack as soon as they are transfered out of the original > drop. I am running out of ideas and really would love some new ones. > > Thanks in advance. > > Len > > Leonard Thomas Ph.D. > Macromolecular Crystallography Laboratory Manager > University of Oklahoma > Department of Chemistry and Biochemistry > Stephenson Life Sciences Research Center > 101 Stephenson Parkway > Norman, OK 73019-5251 > > lmtho...@ou.edu > http://barlywine.chem.ou.edu > Office: (405)325-1126 > Lab: (405)325-7571 > -- Muhammad Bashir Khan ** Department for Structural and Computational Biology Max F. Perutz Laboratories University of Vienna Campus Vienna Biocenter 5 A-1030 Vienna Austria Austria Phone: +43(1)427752224 Fax: +43(1)42779522
[ccp4bb] XSCALE!!! ERROR
Dear All; I am trying to to xsale several data sets together, but it gives an error of !!! ERROR !!! INSUFFICIENT NUMBER OF COMMON STRONG REFLECTIONS. PROGRAM IS UNABLE TO PRODUCE A SCALED DATA SET. resolution of data sets is ranged from 3.25 to 5A Any suggestion will be highly encouraged. Thanks in adv. Bashir -- Muhammad Bashir Khan ** Department for Structural and Computational Biology Max F. Perutz Laboratories University of Vienna Campus Vienna Biocenter 5 A-1030 Vienna Austria Austria Phone: +43(1)427752224 Fax: +43(1)42779522
Re: [ccp4bb] glycerol
Hi Ray; In case of my protein 5%-10% of glycerol help to increase the solubility both of the soluble domain as well as the trans-membrane domain,improve crystal quality dramatically and even prevent radiation damage to some extent. Best of luck Bashir On Thu, March 10, 2011 23:04, Ray Brown wrote: > Hi all, > > I was intrigued by the recent question of whether glycerol had any adverse > effects on the final purity of protein isolated by chromatography. > Glycerol > certainly helps to solubilize some proteins. Does anyone know of any > negative > effects of glycerol in protein purification, on protein crystal quality or > use in cryocrystallography and on X-ray diffraction results? > > Cheers. > > Ray Brown -- Muhammad Bashir Khan ** Department for Structural and Computational Biology Max F. Perutz Laboratories University of Vienna Campus Vienna Biocenter 5 A-1030 Vienna Austria Austria Phone: +43(1)427752224 Fax: +43(1)42779522
Re: [ccp4bb] Structure based and motif based sequence alignment
Dear Fred; Thanks for the reply! extending at the c-terminus means that this protein only N-terminus structure has been solved and I want to include the residues as well in the sequence alignment which are not in the structure. Regards; Bashir On Fri, November 26, 2010 13:38, Vellieux Frederic wrote: > Muhammed bashir Khan wrote: >> Dear All; >> >> I have structures of two protein one full-length while the other >> truncated >> at the c-terminus(one from prokaryote while the other from eukaryotes). >> Now I want to do the sequence alignment of these two proteins from all >> species in such a way that the structure based sequence remain constant >> while extending the sequence only at the c-terminus. Remember the >> structure are known only for the two proteins. >> >> Any suggestion will be highly appreciated! >> >> Regards and have a nice weekend. >> >> Bashir >> > Hi there, > > Ages ago, for this type of work (fine-tuning sequence alignments), I was > loading pre-aligned (or not pre-aligned) sequences and was editing the > alignment "by hand" using a sequence alignment editor. This editor was > working on VAX/VMS systems, which no-one uses anymore (I haven't touched > VMS in many many years). > > So I had a look at what sequence alignment editors are available today > using google, and I came across this: Jalview (http://www.jalview.org ). > Unfortunately, it seems it does not wish to install on my Linux box so I > don't know if the software does what you want it to do. And it is not > clear to me exactly what you mean by "extending the sequence only at the > c-terminus". > > Fred. > > -- Muhammad Bashir Khan ** Department for Structural and Computational Biology Max F. Perutz Laboratories University of Vienna Campus Vienna Biocenter 5 A-1030 Vienna Austria Austria Phone: +43(1)427752224 Fax: +43(1)42779522
[ccp4bb] Structure based and motif based sequence alignment
Dear All; I have structures of two protein one full-length while the other truncated at the c-terminus(one from prokaryote while the other from eukaryotes). Now I want to do the sequence alignment of these two proteins from all species in such a way that the structure based sequence remain constant while extending the sequence only at the c-terminus. Remember the structure are known only for the two proteins. Any suggestion will be highly appreciated! Regards and have a nice weekend. Bashir
Re: [ccp4bb] per-residue RMSD calculation for homologous structure
Dear Dhiraj; Please try the following web site http://www.cgl.ucsf.edu/home/meng/grpmt/structalign.html Here you will find a number of option for structure base sequence alignment no matter what is the similarity of your structures. Regarding the RMSD of every residues you can find this option by using the Dalilite server. Good luck Bashir On Fri, November 19, 2010 23:55, Srivastava, Dhiraj (MU-Student) wrote: > Hi All >does anyone know any software that can calculate and print out RMSD > of every residue (c alpha will be good) for homologous structures > which has only 30-40 % sequence similarity? I looked on the web but > all the software that I found require the sequence to be the same > for both structure. > > Thank you > > Dhiraj > -- Muhammad Bashir Khan ** Department for Structural and Computational Biology Max F. Perutz Laboratories University of Vienna Campus Vienna Biocenter 5 A-1030 Vienna Austria Austria Phone: +43(1)427752224 Fax: +43(1)42779522
[ccp4bb] Structure Based Sequence Alignment
Dear All; Can some body tell me a website for structure based sequence alignment, which can also pin point the similar and identical residues in different colors. regards Bashir -- Muhammad Bashir Khan Department for Biomolecular Structural Chemistry Max F. Perutz Laboratories University of Vienna Campus Vienna Biocenter 5 A-1030 Vienna Austria Phone: +43(1)427752224 Fax: +43(1)42779522
[ccp4bb] Solvent accessible regions of the channel (Pore)
Dear all; Can anybody help me how I can make the solvent accessible surface(inside) of the channel(pore)by pymol, or by any other programme. Thanks in Advance -- Muhammad Bashir Khan Department for Biomolecular Structural Chemistry Max F. Perutz Laboratories University of Vienna Campus Vienna Biocenter 5 A-1030 Vienna Austria Phone: +43(1)427752224 Fax: +43(1)42779522
[ccp4bb] Crystals Soaking in Divalent metals salts.
Dear all; I am working with protein which are supposed to be bound with Magnesium and Cobalt metals.I have now its crystals which are diffracting quite nicely.I want to know the Magnesium and Cobalt binding residues in the target protein. When I add magnesium or Cobalt salts to protein solution even 1mM about 50 percent of the protein precipitated out. When I soaked the crystals in about 10mM of the metals salt it lose diffraction. I varied the soaking time and the metal salt concentration,and even the types of metals salts as well but its seems that it does n,t work. Any idea would be highly appreciated. Bashir -- Muhammad Bashir Khan Department for Biomolecular Structural Chemistry Max F. Perutz Laboratories University of Vienna Campus Vienna Biocenter 5 A-1030 Vienna Austria Phone: +43(1)427752224 Fax: +43(1)42779522
[ccp4bb] R merge and R init
Dear All; I have a crystal structure collected on in house X-ray facility from Bruker using Xprep. I submitted the paper but the reviewer ask for the R merge. As I can't access to the computer at the moment its crashed out. But I have the prp file which have the R init values. My question is!!! 1)Can I use the R init value instead of Rmerge? 2)If yes how it has been calculated, I mean the mathematical formula so that I can write below the table. Thank you very much for the help in advance Sincerely Bashir -- Muhammad Bashir Khan Department for Biomolecular Structural Chemistry Max F. Perutz Laboratories University of Vienna Campus Vienna Biocenter 5 A-1030 Vienna Austria Phone: +43(1)427752224 Fax: +43(1)42779522
[ccp4bb] Molecular Replacement
Dear All; We have solved a crystal structrure of protein at 1.8 A. I have now another crystal of the same protein in aother unit cell,for the new crystal type resolution is 3.6 A but when I use our structure as a seach model It does't give any solution. Any suggestion would be highly appreciated. -- Muhammad Bashir Khan Department for Biomolecular Structural Chemistry Max F. Perutz Laboratories University of Vienna Campus Vienna Biocenter 5 A-1030 Vienna Austria Phone: +43(1)427752224 Fax: +43(1)42779522
