From owner-proteins@net.bio.net Thu Aug 01 23:00:00 1996 Path: biosci!herts.ac.uk!P.Ofarrell From: P.Ofarrell@herts.ac.uk (Paul O'Farrell) Newsgroups: bionet.molbio.proteins Subject: Finding open reading frames Date: 2 Aug 1996 04:20:46 -0700 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 16 Sender: daemon@net.bio.net Distribution: world Message-ID: <7290.9608021123@altair.herts.ac.uk> NNTP-Posting-Host: net.bio.net "Does anyone know any site which analyses nucleotide sequences in all six reading frames, and identifies all ORF's ?" I dont know if this is exactly what your looking for, but the BLASTX programme Will compare all 6 reading frames of your sequence to the protein data bases for homology. It won't find orf's for you, but if you find good homology you'll know what reading frame to use. If you send a message to blast@ncbi.nlm.nih.gov with help as the body of the message they'll email you the full instructions. Paul From owner-proteins@net.bio.net Thu Aug 01 23:00:00 1996 Path: biosci!rutgers!uwm.edu!news-res.gsl.net!news.gsl.net!hunter.premier.net!news.cais.net!nntp04.primenet.com!news.shkoo.com!nntp.primenet.com!tank.news.pipex.net!pipex!oleane!in2p3.fr!swidir.switch.ch!01-newsfeed.univie.ac.at!Austria.EU.net!EU.net!Portugal.EU.net!news.rccn.net!scsing.switch.ch!swsbe6.switch.ch!surfnet.nl!news.wau.nl!usenet From: Pedro Silva Newsgroups: bionet.molbio.proteins Subject: Finding open reading frames Date: Fri, 02 Aug 1996 09:47:43 -0700 Organization: wau.nl Lines: 2 Message-ID: <3202312F.1A73@epr.bc.wau.nl> NNTP-Posting-Host: flex076.tran.wau.nl Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 2.02 (Win16; I) Does anyone know any site which analyses nucleotide sequences in all six reading frames, and identifies all ORF's ? From owner-proteins@net.bio.net Thu Aug 01 23:00:00 1996 Path: biosci!agate!news.ucdavis.edu!dcn67.dcn.davis.ca.us!user From: ijiwaru@wheel.dcn.davis.ca.us Newsgroups: bionet.molbio.proteins Subject: Re: ScFv-expression in pVIII??? Date: Fri, 02 Aug 1996 00:21:20 +0500 Organization: University of California, Davis Lines: 29 Message-ID: References: <31FF67DA.D63@mailserv.cuhk.hk> NNTP-Posting-Host: dcn67.dcn.davis.ca.us X-Newsreader: Value-Added NewsWatcher 2.0b27.1+ In article , ijiwaru@wheel.dcn.davis.ca.us wrote: > In article <31FF67DA.D63@mailserv.cuhk.hk>, b574713@mailserv.cuhk.hk wrote: > > > Dear phage-library workers' > > I would like to know whether ScFv-antibody normally expressed in > pIII protein can > > be expressed in pVIII protein. If not, are there any ways to increase > ScFv-antibody copy > > number per phage which is about 1-5 copies per phage in the case of pIII > protein? The > > problem I found is the ScFv-Ab phage derived from my hybridoma binds > only weakly to > > the antigen. > > You probably wouldn't have much success with displaying ScFvs on pVIII as [stuff deleted} I do have to correct myself, I do know of one successful display of something rather large in pVIII. Charles Craik has built phage with trypsin:pVIII incorporated into it and the trypsin is active. However, this is the only case I know of with something that large (~24kD). Good luck. > > Lyle Najita > Plant Pathology > University of California - Davis From owner-proteins@net.bio.net Thu Aug 01 23:00:00 1996 Path: biosci!bloom-beacon.mit.edu!spool.mu.edu!uwm.edu!news-res.gsl.net!news.gsl.net!usenet.eel.ufl.edu!tank.news.pipex.net!pipex!oleane!in2p3.fr!swidir.switch.ch!01-newsfeed.univie.ac.at!Austria.EU.net!EU.net!main.Germany.EU.net!fu-berlin.de!informatik.tu-muenchen.de!lrz-muenchen.de!uni-erlangen.de!winx03!wpxx02!not-for-mail From: krasel@wpxx02.toxi.uni-wuerzburg.de (Cornelius Krasel) Newsgroups: bionet.molbio.methds-reagnts,bionet.molbio.proteins Subject: Re: Qiagen DNA purification resin - what is it? Followup-To: bionet.molbio.methds-reagnts Date: 2 Aug 1996 08:24:17 GMT Organization: University of Wuerzburg, Germany Lines: 80 Message-ID: <4tsdvh$k7e@winx03.informatik.uni-wuerzburg.de> References: <4tlpbs$2hc4@news.doit.wisc.edu> <4tr6io$2l76@news.doit.wisc.edu> NNTP-Posting-Host: wpxx02.toxi.uni-wuerzburg.de X-Newsreader: TIN [UNIX 1.3 950824BETA PL0] Xref: biosci bionet.molbio.methds-reagnts:47539 bionet.molbio.proteins:8431 [fixed some attribution; followups to the cloners' group only] I wrote some BS in my last posting (seems my brain was on vacation while writing :-) I cancelled it. You will still find it in the archives :-( Dima Klenchin (klenchin@macc.wisc.edu) wrote: > In article , > gjenkins@facstaff.wisc.edu (Glenn Jenkins) wrote: > ->In article <4tlpbs$2hc4@news.doit.wisc.edu>, klenchin@macc.wisc.edu > ->(Dima Klenchin) wrote: > -> > ->> Just curious - what type of anion exchanger is it? > ->> QAE? And what is the support made of? Simply can't believe > ->> this is smth magic that qiagen developed. Must be some oldie > ->> for which a new application was found. > -> > ->My understanding is that it is has charged groups (ie. DEAE?) attached > ->to a silica matrix. These groups are attached in high density which allows > ->a greater salt gradient to be used to elute the DNA. Thus you will have > ->less overlap with proteins and RNA. > -> > ->I do not find a great advantage of their preps($$) except if I want > highly > ->pure DNA. Time wise is it not much of a savings either. One day when I > ->have time I want to do the basic miniprep and apply the Na Acetate/DNA > ->solution to a DEAE sepharose column to see what sticks. I suspect there > ->will not be much of a difference. > -> > ->Does anyone else have any ideas? > > Well, DEAE for *NA purification has been used *widely* in "dark ages" > (before all these fancy kit$). I'm still wondering what's that special > about Qiagen's stuff. And, alas, there is no information on _type_ of > exchanger in archives. Silica is the support? Great. That would mean that > the resin should not been used below pH ~ 6.0 Is this true? Some of the Quiagen blurb: ---start--- QIAGEN resin is a unique anion-exchanger with a hydrophilic surface modification that selectively separates nucleic acids from other substances, such as proteins, carbohydrates, metabolites, or dyes. The macroporous anion-exchanger, with a particle size of approx. 100 um, has a hydrophilic surface coating containing DEAE groups, creating an extremely high surface charge density. The large pore size, together with the high density of anion-exchange groups, provide the extraordinarily broad separation range that makes QIAGEN resin unique. The separation range of QIAGEN resin extends from 0.1 M to 1.6 M salt. Conventional anion-exchangers, based on cellulose, dextran or agarose, have separation ranges only up to 0.4 M salt. Binding and elution of all substances is thus limited to a narrow range of salt concentrations. As the elution peaks of proteins, RNA, and DNA overlap extensively with one another, a satisfactory separation cannot be achieved. QIAGEN resin will not function in the presence of anionic detergents such as SDS, or at a pH less than 4.0. ---end--- I don't think their support is silica: Qiagen has been fairly quick to sue competitors who use similar resins; however, they have apparently not been able to sue Macherey & Nagel, who sell a similar kit (called "Nucleobond") where the resin is derivatized silica. Also, the description of the resin (the "hydrophilic coating") does not speak for silica. The DNA from Qiagen columns in my experience is about as clean as DNA from CsCl preps (but contains less salt). It's much less hassle, however. Usual disclaimers apply. (I don't work for Qiagen. I don't get any profit from this posting. I don't ... :-) --Cornelius. -- /* Cornelius Krasel, U Wuerzburg, Dept. of Pharmacology, Versbacher Str. 9 */ /* D-97078 Wuerzburg, Germany email: phak004@rzbox.uni-wuerzburg.de SP3 */ /* "Science is the game we play with God to find out what His rules are." */ From owner-proteins@net.bio.net Thu Aug 01 23:00:00 1996 Path: biosci!rutgers!uwm.edu!spool.mu.edu!howland.reston.ans.net!nntp.coast.net!fu-berlin.de!informatik.tu-muenchen.de!lrz-muenchen.de!news From: diabetes@lrz.uni-muenchen.de (Wolfgang Schechinger) Newsgroups: bionet.molbio.methds-reagnts,bionet.molbio.proteins Subject: Q: Looking for GFP-cDNA Date: Fri, 02 Aug 1996 21:48:34 GMT Organization: Institute for Diabetes Research Lines: 17 Distribution: world Message-ID: <4ttt3g$7g@sparcserver.lrz-muenchen.de> NNTP-Posting-Host: dial146.ppp.lrz-muenchen.de X-Newsreader: Forte Free Agent 1.0.82 Xref: biosci bionet.molbio.methds-reagnts:47582 bionet.molbio.proteins:8434 Hi everyone out there. Actually, everything is in the subject line. I want to link GFP to a protein I plan to express in E. coli and in eucaryotic cells. If you could share afew nanograms of GFP-cDNA and some microliters of PCR-primers, please let me know. I would be very happy. Greetings from Munich! Wolfgang Schechinger Institute for Diabetes Research Kölner Platz 1 80804 Muenchen Germany Phone +49 (89) 30 79 31 24 Fax 30 81 73 3 E-mail: DIABETES@LRZ.UNI-MUENCHEN.DE From owner-proteins@net.bio.net Thu Aug 01 23:00:00 1996 Path: biosci!iata.csic.es!poseva From: poseva@iata.csic.es (Eva Dupille) Newsgroups: bionet.molbio.proteins Subject: (none) Date: 2 Aug 1996 06:42:36 -0700 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 2 Sender: daemon@net.bio.net Distribution: world Message-ID: <01BB8088.62C4E8C0@alzacarias.iata.csic.es> NNTP-Posting-Host: net.bio.net unsubscribe From owner-proteins@net.bio.net Fri Aug 02 23:00:00 1996 Path: biosci!agate!spool.mu.edu!uwm.edu!cs.utexas.edu!howland.reston.ans.net!vixen.cso.uiuc.edu!newsfeed.internetmci.com!cpk-news-hub1.bbnplanet.com!cpk-news-feed4.bbnplanet.com!maze.dpo.uab.edu!topaz.microbio.uab.edu!sai From: Arioch Newsgroups: bionet.molbio.proteins Subject: Discussion:questions about gel filtration... Date: Sat, 3 Aug 1996 21:03:51 -0500 Organization: University of Alabama at Birmingham Lines: 29 Message-ID: NNTP-Posting-Host: topaz.microbio.uab.edu Mime-Version: 1.0 Content-Type: TEXT/PLAIN; charset=US-ASCII hey all, ...cant seem to get a straight answer out of the other people in my lab, so i'll try here...ok..after having tried dialysis and the ultrafree concentration device and not having too much success with them, i've turned to gel filtration to (a).desalt my protein sample/buffer exchange into buffer of choice...; and (b) use gel filtration to separate out some of the proteins that are NOT too close apart in size...we are using the pharmacia gels in our lab; specifically the g-50 for my purposes of desalting. my question has to do with trying to give some good detail to my protocol...i already know about what length of column to pour, column packing and stuff like that. my question is after i have poured my column and packed it, how do i replace the buffer the gel is in (20% ethanol) with my eluent buffer (i.e. the final buffer i want my protein to be in)...do i just repeatedly wash the column with my eluent buffer? if so, then wont the salts in my eluent buffer get retarded in the gel matrix (i.e. the pores)?? wont this just result in the salts taken out of my eluent buffer? how, then, does this cause the column to be "equilibrated" with my eluent buffer? also, when i run my protein samples thru it, how does my protein then go thru its buffer exchange? i can understand that any salts that the current buffer the protein is in will get trapped and the protein will flow thru...wont it end up coming out in just water then? i would appreciate it if someone can clarify this for me...thx much...cheers... Arioch (sai@topaz.microbio.uab.edu) Graduate Student ( and Lord of Chaos...) Dept. of Biochemistry and Molecular Genetics... Milieu of Entropy (a.k.a University of Alabama at Birmingham...) From owner-proteins@net.bio.net Fri Aug 02 23:00:00 1996 Path: biosci!agate!spool.mu.edu!usenet.eel.ufl.edu!news.alt.net!news1.alt.net!news.exodus.net!www1.hlc.net!ppp3.dialup.peakaccess.net!user From: wdspiv@peakaccess.net (Warren D. Spivack) Newsgroups: bionet.molbio.proteins Subject: Cathepsin D Date: Mon, 29 Jul 1996 18:28:26 -0400 Organization: None Lines: 5 Message-ID: NNTP-Posting-Host: ppp3.dialup.peakaccess.net X-Newsreader: Yet Another NewsWatcher 2.0.1 Does anybody know the cleavage specificity for Cathepsin D? Need this info a.s.a.p. -- "When the bough breaks the cradle will fall... From owner-proteins@net.bio.net Fri Aug 02 23:00:00 1996 Path: biosci!agate!spool.mu.edu!uwm.edu!newsspool.doit.wisc.edu!news.doit.wisc.edu!klenchin From: klenchin@facstaff.wisc.edu (Dima Klenchin) Newsgroups: bionet.molbio.proteins Subject: Re: Discussion:questions about gel filtration... Date: Sun, 04 Aug 96 04:23:41 GMT Organization: UW-Madison Lines: 71 Message-ID: <4u154l$2kqg@news.doit.wisc.edu> References: NNTP-Posting-Host: f182-117.net.wisc.edu Mime-Version: 1.0 Content-Type: text/plain; charset=KOI8-R Content-Transfer-Encoding: 8bit X-Newsreader: News Xpress 2.0 Beta #2 X-No-Archive: Yes In article , Arioch wrote: #hey all, # ...cant seem to get a straight answer out of the other people in #my lab, so i'll try here...ok..after having tried dialysis and the #ultrafree concentration device and not having too much success with them, #i've turned to gel filtration to (a).desalt my protein sample/buffer #exchange into buffer of choice...; and (b) use gel filtration to separate #out some of the proteins that are NOT too close apart in size...we are #using the pharmacia gels in our lab; specifically the g-50 for my purposes #of desalting. For small volumes (> 200 ul) spin columns work best with no dilutions. Alternatively, you can try PD-10 columns from Pharmacia (<2.5 ml loading). In both cases, Sephadex G-25 is used and works better for desalting of protein solutions. # my question has to do with trying to give some good detail #to my protocol...i already know about what length of column to pour, #column packing and stuff like that. my question is after i have poured my #column and packed it, how do i replace the buffer the gel is in (20% #ethanol) with my eluent buffer (i.e. the final buffer i want my protein #to be in)...do i just repeatedly wash the column with my eluent buffer? Yes. At least 5 (better 10) column volumes. #if so, then wont the salts in my eluent buffer get retarded in the gel #matrix (i.e. the pores)?? wont this just result in the salts taken out of #my eluent buffer? how, then, does this cause the column to be #"equilibrated" with my eluent buffer? Everything gets retarded to some extent. That's why *many* volumes of equilibration. After a while, concentration of everything inside and outside beads becomes equal, i.e. equibrated. #also, when i run my protein samples #thru it, how does my protein then go thru its buffer exchange? i can #understand that any salts that the current buffer the protein is in will #get trapped and the protein will flow thru...wont it end up coming out in #just water then? No, it won't. It'll come out in the buffer with which the column was equilibrated AND eluted. To help you out with understanding how gel filtration works in it's extreme (and simplest) case of desalting works, I can give an example that illustrates the principle: imagine that you have a beaker full with protein in solution A and a sponge with pores that do not allow the protein to get into it. You want to change the solution to B. What you do is: 1. soak the sponge (it absorbes, say, 1/2 of total volume), remove it and add solution B to the beaker. Now you have 50% of each A,B). 2. sqeeze the sponge, soak again, and fill with B again (easy to see that now you have 75% B and 25% A). Repeat 1, 2 until you have only acceptable traces of A in a beaker full with protein dissolved now in B. That's how column gel filtration works but instead of repeating the same procedure with many sponges (or sqeezing one repeatedly), you pack them in a huge pile and force the solution to go through them sequentially (= column). - Dima From owner-proteins@net.bio.net Fri Aug 02 23:00:00 1996 Path: biosci!agate!nature.berkeley.edu!lhom From: lhom@nature.berkeley.edu (Louis Hom) Newsgroups: bionet.molbio.proteins,bionet.molbio.methds-reagnts Subject: Continuous gel ref Date: 4 Aug 1996 05:20:50 GMT Organization: University of California, Berkeley Lines: 7 Message-ID: <4u1bvi$ah4@agate.berkeley.edu> NNTP-Posting-Host: nature.berkeley.edu Xref: biosci bionet.molbio.proteins:8437 bionet.molbio.methds-reagnts:47602 Can someone point me to a good reference on continuous buffer systems for PAGE? TIA. -- _______________________________________________________________________________ Lou Hom >K '93 "Putney is confusing obscenity lhom@nature.berkeley.edu with originality." http://www.ocf.berkeley.edu/~lhom -- Myron X in "Putney Swope" From owner-proteins@net.bio.net Sat Aug 03 23:00:00 1996 Newsgroups: ebi.announce,bionet.general,bionet.molbio.embldatabank,bionet.molbio.proteins,bionet.molecules.peptides,embnet.general Path: biosci!daresbury!hgmp.mrc.ac.uk!ebi.ac.uk!saturn.ebi.ac.uk!apweiler From: apweiler@saturn.ebi.ac.uk (Rolf Apweiler) Subject: ANNOUNCEMENT: Second Beta release of TREMBL, a supplement to SWISS-PROT Sender: news@ebi.ac.uk (Mr news) Message-ID: Date: Thu, 1 Aug 1996 15:20:34 GMT Lines: 81 Reply-To: apweiler@saturn.ebi.ac.uk (Rolf Apweiler) Organization: European Bioinformatics Institute (EMBL) - UK X-Newsreader: mxrn 6.18-31 Keywords: Protein database Xref: biosci bionet.general:22893 bionet.molbio.embldatabank:666 bionet.molbio.proteins:8439 bionet.molecules.peptides:393 Second Beta release of TREMBL, a protein sequence database supplementing the SWISS-PROT Protein Sequence Data Bank INTRODUCTION ============ TREMBL is a protein sequence database supplementing the SWISS-PROT Protein Sequence Data Bank. TREMBL contains the translations of all coding sequences (CDS) present in the EMBL Nucleotide Sequence Database not integrated in SWISS-PROT. At the moment we treat TREMBL as an independent dataset in SWISS-PROT format, but shortly TREMBL will become a part of SWISS-PROT. SECOND BETA RELEASE OF TREMBL ============================= This TREMBL release is created from the EMBL Nucleotide Sequence Database release 47 and contains 107'065 sequence entries, comprising 28'667'743 amino acids. TREMBL is split in two main sections; SP-TREMBL and REM-TREMBL: SP-TREMBL (SWISS-PROT TREMBL) contains the entries (89'883) which should be incorporated into SWISS-PROT. SP-TREMBL is partially redundant against SWISS-PROT, since approximately 40'000 SP-TREMBL entries are only additional sequence reports of proteins already in SWISS-PROT. We will try to merge these sequence reports as fast as possible with the already existing SWISS-PROT entries for these proteins, so as to make SWISS-PROT and TREMBL completely nonredundant. SP-TREMBL is organized in subsections: fun.dat (Fungi): 4939 entries hum.dat (Human): 5985 entries inv.dat (Invertebrates): 11476 entries mam.dat (Other Mammals): 2096 entries mhc.dat (MHC proteins): 2221 entries org.dat (Organelles): 6466 entries phg.dat (Bacteriophages): 928 entries Pln.dat (Plants): 5790 entries pro.dat (Prokaryotes): 17393 entries rod.dat (Rodents): 5473 entries unc.dat (Unclassified): 243 entries vrl.dat (Viruses): 24595 entries vrt.dat (Other Vertebrates): 2278 entries REM-TREMBL (REMaining TREMBL) contains the entries (17'182) that we do not want to include in SWISS-PROT. ACCESS/DATA DISTRIBUTION ======================== FTP server: ftp.ebi.ac.uk/pub/databases/trembl TREMBL is also available on the SWISS-PROT CD-ROM. TREMBL HAS BEEN PREPARED BY: ============================ Rolf Apweiler, Alain Gateau, Vivien Junker, Fiona Lang, Claire O'Donovan, and Nicoletta Mitaritonna at the EMBL Outstation - European Bioinformatics Institute (EBI) in Hinxton, UK; Amos Bairoch at the Medical Biochemistry Department of the University of Geneva. ======================================================================= Rolf Apweiler | EMBL Outstation | Email:apweiler@ebi.ac.uk European Bioinformatics Institute (EBI) | URL: http://www.ebi.ac.uk Wellcome Trust Genome Campus, Hinxton | Tel: +44 (1223) 494435 Cambridge CB10 1SD, UK | Fax: +44 (1223) 494968 ======================================================================== From owner-proteins@net.bio.net Sun Aug 04 23:00:00 1996 Path: biosci!daresbury!lyra.csx.cam.ac.uk!news From: Paul Digard Newsgroups: bionet.molbio.proteins,bionet.molbio.methds-reagnts Subject: Re: Continuous gel ref Date: Mon, 05 Aug 1996 17:46:40 +0000 Organization: University of Cambridge, England Lines: 17 Message-ID: <32063380.5132@mole.bio.cam.ac.uk> References: <4u1bvi$ah4@agate.berkeley.edu> NNTP-Posting-Host: mac08.vrl.path.cam.ac.uk Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 2.01 (Macintosh; I; 68K) To: Louis Hom Xref: biosci bionet.molbio.proteins:8441 bionet.molbio.methds-reagnts:47635 Louis Hom wrote: > > Can someone point me to a good reference on continuous buffer systems for > PAGE? TIA. > -- McClellan, T. (1982). Electrophoresis buffers for polyacrylamide gels at various pH. Anal. Biochem. 126, 94-99. This might be helpfull? Paul Digard Division of Virology Department of Pathology University of Cambridge From owner-proteins@net.bio.net Sun Aug 04 23:00:00 1996 Path: biosci!biosci!not-for-mail From: jmerelo@kal-el.ugr.es (J.J. Merelo Guervos) Newsgroups: bionet.software,bionet.molbio.proteins,comp.ai.neural-nets Subject: [Web]: protein 2ary structure from CD spectra using NNs Date: 5 Aug 1996 15:34:54 -0700 Organization: GeNeura Team Lines: 48 Sender: daemon@net.bio.net Distribution: world Message-ID: <51raprl7l6.fsf@kal-el.ugr.es> NNTP-Posting-Host: net.bio.net Xref: biosci bionet.software:16239 bionet.molbio.proteins:8444 comp.ai.neural-nets:23410 Dear usenetters: We announce the establishment of a web server for prediction of protein secondary structure percentages from UV circular dichroism spectra. You only need to submit 41 CD values ranging from 200 nm to 240 nm (given in deg cm^2 dmol^-1 multiplied by 0.001) and the server gives back the estimated percentages of helix, beta and rest of secondary structure of your protein plus an estimation of the accuracy of the prediction. The prediction is done using a Kohonen neural network with a 2-dimensional output layer. The algorithm has been published [1, 2] and possitively referenced in a recent review [3]. The program itself (k2d) can be retrieved by anonymous ftp (see the k2d home page for details). The http address of the k2d server is: http://kal-el.ugr.es/k2d/spectra.html The home page of the k2d program is at: http://kal-el.ugr.es/k2d/k2d.html or at http://www.embl-heidelberg.de/~andrade/k2d.html As the server just began we would be very grateful in receiving your feedback about it. J.J. Merelo M.A. Andrade References [1] Merelo, J.J., M.A. Andrade, A. Prieto and F. Moran. 1994. Neurocomputing. 6, 443-454 [2] Andrade, M.A., P. Chacon, J.J. Merelo and F. Moran. 1993. Prot. Eng. 6, 383-390 [3] Greenfield, N.J. 1996. Anal. Biochem., 235, 1-10 -- JJ Merelo | http://kal-el.ugr.es/htbin/jj-plex Grupo Geneura ---- Univ. Granada | http://kal-el.ugr.es/ From owner-proteins@net.bio.net Sun Aug 04 23:00:00 1996 Path: biosci!rutgers!uwm.edu!news-res.gsl.net!news.gsl.net!usenet.eel.ufl.edu!warwick!news.nott.ac.uk!NewsWatcher!user From: kevin.bailey@nottingham.ac.uk () Newsgroups: bionet.molbio.proteins Subject: Re: Cathepsin D Followup-To: bionet.molbio.proteins Date: Mon, 05 Aug 1996 12:31:16 +0100 Organization: Cripps Computing Centre, The University of Nottingham Lines: 12 Message-ID: References: NNTP-Posting-Host: wpbjwk1.biochem.nottingham.ac.uk In article , wdspiv@peakaccess.net (Warren D. Spivack) wrote: > Does anybody know the cleavage specificity for Cathepsin D? > Need this info a.s.a.p. Don't know if this is much use but I think cathepsins sequencially remove dipeptides from the N-terminal end of proteins, but I'm not sure how specific they are, I think that an N-terminal lys or arg will block cathepsin C from removing dipeptides. The only ref I know of for cath.D is Barrett in Biochemical Journal vol 117 p.601 (but it's an old ref, may be a lot more info known these days). From owner-proteins@net.bio.net Sun Aug 04 23:00:00 1996 Path: biosci!daresbury!bioftp.unibas.ch!ubaclu.unibas.ch!ubaclu.unibas.ch!nntp Newsgroups: bionet.molbio.proteins Subject: NMR post-doc positions -> Biozentrum, U. Basel, Switzerland Message-ID: <32066C01.167E@ubaclu.unibas.ch> From: andrei Date: Mon, 05 Aug 1996 23:47:45 +0200 Nntp-Posting-Host: nmr.bioz.unibas.ch X-Mailer: Mozilla 3.0b5aGold (X11; I; IRIX 5.3 IP17) MIME-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit Lines: 62 Applications are invited for two Biomolecular-NMR post doctoral positions in the Department of Structural Biology of the Biozentrum, University of Basel - Switzerland. Persons who have received a Ph.D. within the last 6 months, or who expect to receive their Ph.D. within 3 months are strongly preferred. One position is for a person with a technical NMR background. The applicant should have strong computational skills, and extensive experience in programming multidimensional heteronuclear NMR pulse sequences in a UNIX environment. The second position is for an NMR spectroscopist with a molecular biology background. Applicants should ideally have hands-on experience with PCR, mutagenesis, T7 expression systems, 13C/15N isotopic labeling, and protein purification. In addition, outstanding candidates with a primary interest in protein folding and original ideas on how to make progress with THE PROBLEM are also strongly encouraged to apply. Our group's current research interests include: 1) Folding and dynamics of non-homologous proteins that share an OB-fold structural motif. 2) Protein engineering. 3) Isotopically labeled peptides for NMR studies of peptide-protein complexes. 4) NMR structure determination. NMR experiments will be carried out on a dedicated 600 MHz Varian Unity-Plus spectrometer, on which our group (3 NMR users) has 30 % measurement time. This instrument is equipped with pulse field gradients, triple resonance probes, and 4 channels for multidimensional heteronuclear experiments. An SGI Crimson, and INDY computer are available for NMR data processing and analysis. Successful candidate should be highly motivated, and should have a track-record of original publications. Interested and qualified applicants should send a C.V. as well as the names and addresses of 3 referees familiar with the applicant's work to: Dr. Andrei T. Alexandrescu Biozentrum University of Basel Klingelbergstrasse 70 Basel/Switzerland CH-4056 Tel: 41-61-267-2091 Fax: 41-61-267-2109 email: alexandrescu@ubaclu.unibas.ch Use of Fax or e-mail is appreciated, and will expedite responses to inquiries. From owner-proteins@net.bio.net Sun Aug 04 23:00:00 1996 Path: biosci!NANOTHINC.COM!tess From: tess@NANOTHINC.COM (Tess Mercer) Newsgroups: bionet.molbio.proteins Subject: Nanobiology Call for Papers Date: 5 Aug 1996 14:25:40 -0700 Organization: Nanothinc Lines: 57 Sender: daemon@net.bio.net Distribution: world Message-ID: <32060527.42B8@nanothinc.com> Reply-To: tess@nanothinc.com NNTP-Posting-Host: net.bio.net Topical Overview: Call for Papers - Nanobiology: Research, Theory, and Current Applications Nanotechnology, as a general topic, covers a very wide array of different disciplines and areas of research. However, in the realm of biological and biomolecular applications and developments, there are a number of current examples, including some commercial applications, which are reshaping the approach to which previously difficult, or even "unsolvable" procedures can now be resolved, or greatly improved upon. It is specifically in the realm of thoeretical, near term, or currently available technologies, and how they are affected by application of "nanobiological" processes, that will be reviewed in this section of the journal. By definition, a nanobiological process could be described, in this context, as any process which utilizes the contrived manipulation of individual biomolecules or biomolecular structures to fullfill a desired task, create an object, a material or application of a material, or as an intermediate step in fullfilling a multistaged series of assembly or construction tasks involving biomolecular materials. This definition could also be expanded to include the methodologies of visualiztion or detection of nanoscale activities in a biomolecular environment, or in the development of instrumentation and processes related to these tasks. Also, there are other tangential aspects of nanobiological processes which are also relevant, and deserve attention. Particularly in the areas of potential nanoscale computing or information processing applications which may utilize or incorporate biological components or processes, there has been considerable interest and research ativities. Please let us know if you are interested. Information about our company, Nanothinc, can be found at http://www.nanothinc.com. Also, I have additional Notes for Contributors which I can fax to your attention. Sincerely, Maritess T. Mercer Director of Administration Project Manager Nanothinc 1797 Union Street San Francisco, CA 94123 Tel: (415) 202-9969, Fax: (415) 202-9975 URL: http://www.nanothinc.com Email: tess@nanothinc.com From owner-proteins@net.bio.net Sun Aug 04 23:00:00 1996 Path: biosci!bcm.tmc.edu!news.msfc.nasa.gov!newsfeed.internetmci.com!in2.uu.net!news.biu.ac.il!news.huji.ac.il!avdat3.bgu.ac.il!usenet From: irisp@bgumail.bgu.ac.il (Iris Porat) Newsgroups: bionet.molbio.proteins Subject: Participation on confrence "Thermophiles '96', sep, 96, Athens Date: Mon, 05 Aug 1996 20:00:02 GMT Organization: Ben Gurion University Lines: 4 Message-ID: <32065204.4499656@news.bgu.ac.il> NNTP-Posting-Host: bgumail.bgu.ac.il X-Newsreader: Forte Agent .99c/16.141 I will participate on the conference Thermophiles '96. I will rent a room for 13$ for night. The hostel is located about 10 miles from Athens (15 minute by car). I will rent a car, for 20$ for day. I am looking for a travel companion. From owner-proteins@net.bio.net Sun Aug 04 23:00:00 1996 Path: biosci!biosci!not-for-mail From: BIOSCI Administrator Newsgroups: bionet.molbio.proteins Subject: Research Diagnostics, Inc. sponsors BIOSCI! Date: 5 Aug 1996 17:40:14 -0700 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 47 Sender: biohelp@net.bio.net Distribution: world Message-ID: <4u649e$nl0@net.bio.net> NNTP-Posting-Host: net.bio.net We are pleased to welcome a new sponsor of the BIOSCI project Research Diagnostics, Inc., distributor of fine immunochemical products Research Diagnostics is sponsoring the CELL-BIOLOGY, DIAGNOSTICS, IMMUNOLOGY, and PROTEIN-ANALYSIS groups. You can view their ads on those groups by visiting our WWW archives at http://www.bio.net/archives.html. Please thank them for their support of BIOSCI if you contact them. This is our complete list of sponsors: Molecular Dynamics, Inc., a leading producer of innovative instrumentation systems for the life sciences Knight-Ridder Information, Inc., a major provider of electronic information to business, research, and scientific professionals BIOSIS, publisher of Biological Abstracts and Zoological Record CLONTECH Laboratories, Inc., a leading manufacturer of innovative reagents and kits for the life sciences The UnCover Company, an internationally-known firm that offers article delivery and current issue alerting from 17,000 periodicals. LI-COR, Inc., manufacturer of automated infrared DNA sequencing and genetic analysis systems. The Nest Group, Inc. a value-added distributor of HPLC columns, DNA kits, and electrophoresis gels for biomolecule separation. Sun Microsystems, Inc., a leading provider of solutions for open network computing environments. Academic Press, a leading scientific publisher for more than 50 years. QIAGEN, a leading provider of innovative, user-friendly technologies for nucleic acid and protein purification. Trends in Biochemical Sciences, the leading monthly magazine in biochemisty and molecular biology. Research Diagnostics, Inc., distributor of fine immunochemical products The assistance of these organizations will allow us to continue to provide our service to you beyond the end of our grant this year. Please thank our sponsors for their support of BIOSCI in your communications with them. Sincerely, Dave Kristofferson BIOSCI/bionet Manager biosci-help@net.bio.net From owner-proteins@net.bio.net Mon Aug 05 23:00:00 1996 Path: biosci!bcm.tmc.edu!cs.utexas.edu!swrinde!tank.news.pipex.net!pipex!oleane!in2p3.fr!swidir.switch.ch!scsing.switch.ch!swsbe6.switch.ch!surfnet.nl!highway.leidenuniv.nl!ruly46!mirihyel From: Peter Hohenstein Newsgroups: bionet.molbio.proteins Subject: custom made knock out mice Date: Mon, 5 Aug 1996 11:11:44 +0200 Organization: Leiden University, The Netherlands Lines: 19 Message-ID: NNTP-Posting-Host: ruly46.medfac.leidenuniv.nl Mime-Version: 1.0 Content-Type: TEXT/PLAIN; charset=US-ASCII X-Sender: mirihyel@ruly46 Hello, I'm looking for companies who make mice from (alreaddy targeted) ES cells. Does anybody have information on this? Thanks in advance. Peter Hohenstein Dept. Human Genetics University of Leiden The Netherlands From owner-proteins@net.bio.net Mon Aug 05 23:00:00 1996 Path: biosci!rutgers!uwm.edu!chi-news.cic.net!cs.utexas.edu!geraldo.cc.utexas.edu!usenet From: "Brett S. Phinney" Newsgroups: bionet.molbio.proteins Subject: Protein precipitation and detergent Date: 6 Aug 1996 13:38:20 GMT Organization: University of Texas Lines: 11 Message-ID: <01bb839c$899e2440$6a715380@UT.cc.utexas.edu> NNTP-Posting-Host: slip-c-10.ots.utexas.edu X-Newsreader: Microsoft Internet News 4.70.1132 I know this topic was just discussed, but I was curious if chloroform methanol precipitation of protein will remove detergents such as SDS or Triton X, has anyone had any experience with this. Thanks a lot -- Brett Phinney Cell Research Institute University of Texas, Austin From owner-proteins@net.bio.net Mon Aug 05 23:00:00 1996 Path: biosci!biosci!not-for-mail From: a.c.noorlandt@chem.ruu.nl (Albert Noorlandt) Newsgroups: bionet.biophysics,bionet.cellbiol,bionet.immunology,bionet.metabolic-reg,bionet.molbio.ageing,bionet.molbio.methds-reagnts,bionet.molbio.proteins,bionet.molbio.proteins.7tms_r,alt.med,alt.pharm,sci.bio.misc,sci.chem,sci.chem.analytical,sci.med,sci.misc,sci.techniques Subject: 1997 FEBS Meeting on Cell Signalling Mechanisms Date: 6 Aug 1996 17:24:42 -0700 Organization: CBLE Lines: 29 Sender: biohelp@net.bio.net Distribution: world Message-ID: NNTP-Posting-Host: net.bio.net Xref: biosci bionet.biophysics:2198 bionet.cellbiol:5225 bionet.immunology:9491 bionet.metabolic-reg:805 bionet.molbio.ageing:2916 bionet.molbio.methds-reagnts:47700 bionet.molbio.proteins:8451 bionet.molbio.proteins.7tms_r:804 sci.bio.misc:4117 sci.chem:61653 sci.chem.analytical:5118 sci.med:133749 sci.misc:12601 FEBS Special Meeting on Cell Signalling Mechanisms, From membrane to nucleus Amsterdam, The Netherlands, June 29­July 3, 1997 Everything you want to know about this meeting on www-page: http://www.cble.chem.ruu.nl/febs97/index.html The meeting will cover the following topics: € Tyrosine kinases € MAP-, Ser/Thr-kinases € G-coupled receptors € Lymphokine receptors € Ras, Rac, Rho-proteins € Ca2+, cGMP, NO € Lipid signalling € Steroid receptors € Transcription factors € Cell cycle, Apoptosis For further information contact: Secretariat FEBS Special Meeting 1997 Lidy Groot Congress Events P.O. Box 83005 1080 AA Amsterdam The Netherlands Tel. +31.20.679 3218 Fax: +31.20.675 8236 Email: lidy.groot@inter.nl.net From owner-proteins@net.bio.net Mon Aug 05 23:00:00 1996 Path: biosci!MANI.CBS.UMN.EDU!npv From: npv@MANI.CBS.UMN.EDU (Nora Plesofsky-Vig) Newsgroups: bionet.molbio.proteins Subject: re:protein precipitation Date: 6 Aug 1996 11:21:22 -0700 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 6 Sender: daemon@net.bio.net Distribution: world Message-ID: <9608061821.AA00675@mani.cbs.umn.edu> Reply-To: nora@biosci.cbs.umn.edu NNTP-Posting-Host: net.bio.net I have used ethanol (4 vol) to precipitate proteins away from Triton X-100 (with 1 ethanol wash) for running the proteins in SDS PAGE. It works well enough for this purpose, but I don't know how much Triton is left behind. Nora Plesofsky-Vig From owner-proteins@net.bio.net Mon Aug 05 23:00:00 1996 Path: biosci!bcm.tmc.edu!news.uth.tmc.edu!exrady335.mdacc.tmc.edu!ylek From: ylek@utmdacc.uth.tmc.edu (David Voehringer) Newsgroups: bionet.molbio.proteins Subject: Cytochrome C antibody Date: Tue, 6 Aug 1996 14:46:09 Organization: University of Texas Health Science Center Lines: 7 Message-ID: NNTP-Posting-Host: exrady335.mdacc.tmc.edu Mime-Version: 1.0 Content-Type: text/plain;charset=US-ASCII Content-Transfer-Encoding: 7bit X-Newsreader: Trumpet for Windows [Version 1.0 Rev A] Does anyone know where I could find a anti-mouse Cytochrome C antibody?? David W. Voehringer Department of Experimental Radiotherapy Univ. of Texas M.D. Anderson Cancer Center 1515 Holcombe Bvld. Houston, Tx. 77030 (713) 792-3797 From owner-proteins@net.bio.net Tue Aug 06 23:00:00 1996 Path: biosci!bcm.tmc.edu!pendragon!news.msfc.nasa.gov!newsfeed.internetmci.com!in3.uu.net!newsxfer2.itd.umich.edu!agate!hpg30a.csc.cuhk.hk!news.cuhk.edu.hk!news.hklink.net!icon1.intercon.net!usenet From: TPM Newsgroups: bionet.molbio.proteins Subject: chemifluorescence vs chemiluminescence Date: Wed, 07 Aug 1996 18:18:43 -0700 Organization: CTS Lines: 10 Message-ID: <32094073.169F@icon.intercon.net> Reply-To: ctsngai@icon.intercon.net NNTP-Posting-Host: anp5.intercon.net Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 3.0b6 (Win95; I; 16bit) Hi Everybody, Does anyone compare the methods "Chemifluorescence" and "Chemiluminescence"? What are the pros and cons of these methods? The experience from all the existing user is most welcomed. Thanks. TPM From owner-proteins@net.bio.net Tue Aug 06 23:00:00 1996 Path: biosci!agate!spool.mu.edu!uwm.edu!cs.utexas.edu!howland.erols.net!surfnet.nl!highway.leidenuniv.nl!ruly46!mirihyel From: Peter Hohenstein Newsgroups: bionet.molbio.proteins Subject: glycine strand in tagged protein Date: Wed, 7 Aug 1996 17:02:56 +0200 Organization: Leiden University, The Netherlands Lines: 24 Message-ID: NNTP-Posting-Host: ruly46.medfac.leidenuniv.nl Mime-Version: 1.0 Content-Type: TEXT/PLAIN; charset=US-ASCII X-Sender: mirihyel@ruly46 Hello, I'm working on the expression of a cDNA in 3T3 and I intend to tag this protein C terminally with the HA tag. Somebody told me about putting several glycine residues between the protein of interest and the tag, the idea being that glycine is the smallest amino acid and therefor has the best 'moving' capacity. In this way the tag would be maximum available for the antibodies. However, the person who told me this had no idea where he had it from, and I can't find any references on it. So my questions are: does anybody have experience with this, and if so, how many glycines should I put between protein and HA-tag? Any information on this would be appreciated! Peter Hohenstein Dept. Human Genetics University of Leiden The Netherlands From owner-proteins@net.bio.net Tue Aug 06 23:00:00 1996 Newsgroups: bionet.molbio.proteins Path: biosci!agate!spool.mu.edu!usenet.eel.ufl.edu!warwick!yama.mcc.ac.uk!liv!woodie From: woodie@liverpool.ac.uk (Dr I.P. Woodrow) Subject: Re: chemifluorescence vs chemiluminescence Message-ID: Sender: news@liverpool.ac.uk (News System) Nntp-Posting-Host: ash-22.liv.ac.uk Organization: The University of Liverpool References: <32094073.169F@icon.intercon.net> Date: Wed, 7 Aug 1996 15:08:41 GMT Lines: 43 In article <32094073.169F@icon.intercon.net>, TPM writes: > Hi Everybody, > > Does anyone compare the methods "Chemifluorescence" and > "Chemiluminescence"? What are the pros and cons of these methods? The > experience from all the existing user is most welcomed. > > Thanks. > > > TPM The major difference is that fluorescence is due to absorbed light being re-emitted at a lower wavewlength (and doesn't need the "chemi" as it naturally occurs in chemicals anyway) , whilst chemiluminsecence is the production of photons of light throught chemical reactions. This means that you have to "excite" fluorescent molecules with light to see them, whilst chemiluminsecence occurs only when the relevant chemicals are mixed (the most commonly encountered lab example of this is in luciferase from fireflies). With fluorescence you often get some background interference from the incident (excitation) light as there is often some overlap in the wavelength peaks. It is very sensitive technique all the same. Fluorescent dyes are tailored to a range of uses, so you should have no problem finding what you need. I have less experience with chemiluminescence. You are certainly limited by the applications as luminescent reactions are pretty rare if memory serves. Havbing said this it shouldbe more sensitive than fluorecence as there is no incident radiation (there are likelty to be other practical considerations to this as there always are!) Fluorescence is usually measured on a fluorimeter (oddly enough!), with the fluorescence measured usualy at 90 deg to the incident light to minimise interference from this. Luminescence can be measured on a scintilation counter (scintillation is avery similar phenomenon). Also flashier (no pun intended!) fluorimters can measure luminescence. Hope this is some help! Iain From owner-proteins@net.bio.net Tue Aug 06 23:00:00 1996 Path: biosci!agate!newsxfer2.itd.umich.edu!howland.erols.net!howland.reston.ans.net!vixen.cso.uiuc.edu!uwm.edu!news-res.gsl.net!news.gsl.net!hunter.premier.net!news.cais.net!nntp.uio.no!news.kth.se!news From: Arne Elofsson Newsgroups: bionet.molbio.proteins,bionet.molec-model Subject: Prediction servers and predictions Date: 07 Aug 1996 11:38:25 +0200 Lines: 30 Message-ID: NNTP-Posting-Host: rune.biokemi.su.se Mime-Version: 1.0 Content-Type: text/plain; charset=iso-8859-1 Content-Transfer-Encoding: 8bit X-Newsreader: Gnus v5.2.37/XEmacs 19.14 Xref: biosci bionet.molbio.proteins:8453 bionet.molec-model:1100 Dear netters I will teach a course in "Theoretical DNA and Protein analysis" and as more and more of the tools get available over the WWW I will focus on using the WWW to analyze the sequence/protein you are interested in. The students will work on a small set of selected sequence and use the WWW to try to find out as much as possible about these proteins. In a way it is similar to what Tim Hubbard and Anna Tramontano did but on a much smaller scale. We will use this course as a test of different servers that are available over the net. A summary evaluating the different servers will be published on the net. SO I would rally appreciate 1) Any server that you want the students to use and test. (If you have a small, easy to use and fast program that you want ous to test we can provide a WWW interface for the server) 2) Any comprehensive list of servers and others related links. 3) Any protein sequence that you want the students to work on. thank you a lot arne -- ----------------------------------------------------------------- From: Arne Elofsson Email: arne@rune.biokemi.su.se Tel:+46(0)8-161553 WWW: http://www.biokemi.su.se/~arne/ From owner-proteins@net.bio.net Tue Aug 06 23:00:00 1996 Path: biosci!agate!newsxfer2.itd.umich.edu!newsfeed.internetmci.com!in2.uu.net!EU.net!main.Germany.EU.net!fu-berlin.de!informatik.tu-muenchen.de!lrz-muenchen.de!uni-regensburg.de!news.uni-stuttgart.de!news.urz.uni-heidelberg.de!phoenix.mgen.uni-heidelberg.de!chw From: Christoph H. Winter Newsgroups: bionet.molbio.proteins Subject: Large MW Standard f. SDSPAGE Date: 7 Aug 1996 08:25:27 GMT Organization: Molecular Genetics Lines: 11 Distribution: world Message-ID: <4u9jtn$blt@sun0.urz.uni-heidelberg.de> NNTP-Posting-Host: phoenix.mgen.uni-heidelberg.de Mime-Version: 1.0 Content-Type: text/plain; charset=ISO-8859-1 Content-Transfer-Encoding: 8bit X-Newsreader: Nuntius 2.0.4_68K X-XXMessage-ID: X-XXDate: Wed, 7 Aug 1996 08:25:27 GMT I need a super-high-marker protein standard for SDS-PAGE (200-300kd). Does anybody know (available) proteins which work in this range to supplement a normal high marker? Thank you in advance Christoph --- < mailto:chw@sirius.mgen.uni-heidelberg.de > From owner-proteins@net.bio.net Tue Aug 06 23:00:00 1996 Path: biosci!bcm.tmc.edu!cs.utexas.edu!howland.erols.net!vixen.cso.uiuc.edu!newsfeed.internetmci.com!in3.uu.net!gwu.edu!mnrao From: mnrao@gwis2.circ.gwu.edu (Manjunath N.Rao) Newsgroups: bionet.molbio.proteins Subject: Protein binding to its ligand Date: 7 Aug 1996 19:55:17 GMT Organization: The George Washington University, Washington DC Lines: 19 Message-ID: <4uasb5$au5@cronkite.seas.gwu.edu> NNTP-Posting-Host: 128.164.127.252 X-Newsreader: TIN [version 1.2 PL2] Hello Everyone, I am experiencing a strange problem in studying the ligand binding property of a protein isolated recently. This protein binds a hydrophobic lipid ligand ( a small moleculae with a mol wt. of 600). I have conducted one experiment wherein I have incubated increasing amounts of the radioactive ligand alone and radioactive ligand plus 20 fold excess of cold ligand with the protein concentration also increasing proportionately. I expected to find out the specific binding as well as protein to ligand (p:l) binding ratio. However, I now find that at lower protein concentration the ratio of p:l is very high (1:16) and as protein concentration increases the p:l ratio decreases and almost reaches 1:1. I would like to know whether anyone of you have heard of or experienced such a property in a ligand binding protein. I appreciate any help in this regard. Thanks in advance MNR. -- ******************************* ************************* From owner-proteins@net.bio.net Tue Aug 06 23:00:00 1996 Path: biosci!agate!howland.reston.ans.net!vixen.cso.uiuc.edu!newsfeed.internetmci.com!in2.uu.net!rcogate.rco.qc.ca!altitude!usenet From: Achim Recktenwald Newsgroups: bionet.molbio.proteins Subject: Re: Discussion:questions about gel filtration... Date: Wed, 07 Aug 1996 08:15:34 -0500 Organization: IBEX Technologies, Inc., Biochemistry, 5485 Pare, Montreal, PQ, H4P 1P7, Canada Lines: 25 Message-ID: <320896F6.4BA@ibex.ca> References: NNTP-Posting-Host: ibex.hip.cam.org Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 2.02 (Macintosh; I; PPC) To: Arioch Arioch wrote: > [snip] > my question is after i have poured my > column and packed it, how do i replace the buffer the gel is in (20% > ethanol) with my eluent buffer (i.e. the final buffer i want my protein > to be in)...do i just repeatedly wash the column with my eluent buffer? With gel filtration, esp. these Sephadex gels, it is a very bad move to introduce ethanol. I usually use a Buchner funnel or a glass-frit to get my gel into the buffer i want and then make my column. But before I pour the column, I degass the gel very thoroughly. This is a very important step. If you have your gel already in the column but in etanol, you will never succeed in removing all the air-bubbles in those pores deep in the gel. But these pores are the most important for the efficiency of the gel filtratrion. I would recommend to unpack your column, get the gel into the right buffer, degass it, and pour it again. After that I would use ~2CV to wash the column with your running buffer. If you want to keep the column for further runs, store it in PBS with some azide. Before using it again you should wash it with ~10CV of buffer. Achim From owner-proteins@net.bio.net Tue Aug 06 23:00:00 1996 Path: biosci!CSHL.ORG!leemor From: leemor@CSHL.ORG (Leemor Joshua-Tor) Newsgroups: bionet.molbio.proteins Subject: postdoctoral position at Cold Spring Harbor Date: 7 Aug 1996 06:21:57 -0700 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 49 Sender: daemon@net.bio.net Distribution: world Message-ID: NNTP-Posting-Host: net.bio.net POSTDOCTORAL POSITION in STRUCTURAL BIOLOGY at COLD SPRING HARBOR A postdoctoral position is available at Cold Spring Harbor Laboratory in either of two areas: 1. Structural and biochemical studies of a unique DNA-binding protease. Gal6 (yeast bleomycin hydrolase) is a conserved protease that limits the use of the anti-cancer drug bleomycin. We are combining structural and biochemical information of the protein, its complexes and various mutants, along with yeast molecular biology and genetics which is done in collaboration with Stephen Johnston at Southwestern Medical Center at Dallas, to study its functions and conserved cellular role. See: L. Joshua-Tor, H. E. Xu, S. A. Johnston and D. C. Rees, Crystal Structure of a Conserved Protease That Binds DNA: The Bleomycin Hydrolase, Gal6, Science, 269, 945-950 (1995). 2. Structural studies of complexes of eukaryotic transcription factors. We are combining structural studies with genetic information to study the molecular interactions important for transcriptional activation in eukaryotes. Our X-ray crystallography facilities include an R-axis detector equipped with an Oxford cryosystem, several graphics workstations and have access to the nearby National Synchrotron Light Source (NSLS) at Brookhaven. We also have state-of-the-art facilities for carrying out biochemical and molecular biology studies. Cold Spring Harbor provides a unique stimulating environment for rich scientific interactions. The positions are available for highly motivated applicants with a Ph. D. and experience in either: (a) protein crystallography or a related field; or (b) those with experience in biochemistry or molecular biology and a keen interest in protein crystallography and structural biology. Please send CV, list of publications, a summary of research experience and interests, and names and numbers of three references to Dr. Leemor Joshua-Tor, Cold Spring Harbor Laboratory, P. O. Box 100, Cold Spring Harbor, NY 11724. Tel. (516) 367 8821. For additional information please call or send e-mail to leemor@cshl.org ******************************************************************* Leemor Joshua-Tor, Ph.D. Cold Spring Harbor Laboratory Tel. (516) 367 8821 1 Bungtown Road Fax (516) 367 8873 Cold Spring Harbor, NY 11724 e-mail: leemor@cshl.org ******************************************************************* From owner-proteins@net.bio.net Wed Aug 07 23:00:00 1996 Path: biosci!agate!newsxfer2.itd.umich.edu!portc01.blue.aol.com!news-res.gsl.net!news.gsl.net!usenet.eel.ufl.edu!warwick!leicester!usenet From: "Dr E. Buxbaum" Newsgroups: bionet.molbio.proteins Subject: Re: Protein precipitation and detergent Date: 8 Aug 1996 09:42:52 GMT Organization: University of Leicester, UK (PCFS User) Lines: 3 Message-ID: <4uccqs$39f@falcon.le.ac.uk> References: <01bb839c$899e2440$6a715380@UT.cc.utexas.edu> NNTP-Posting-Host: pc47.msb.le.ac.uk Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 1.22 (Windows; I; 16bit) It does. The problem usually is to get the protein back into solution afterwards. From owner-proteins@net.bio.net Wed Aug 07 23:00:00 1996 Path: biosci!agate!spool.mu.edu!usenet.eel.ufl.edu!usenet.cis.ufl.edu!usenet.ufl.edu!cybernet.cse.fau.edu!usenet From: Dr Peter Lutz Newsgroups: bionet.molbio.proteins Subject: HELP protein analysis Date: Wed, 07 Aug 1996 13:39:51 -0400 Organization: Florida Atlantic University Lines: 14 Message-ID: <3208D4E7.165F@acc.fau.edu> NNTP-Posting-Host: bsmac03.sci.fau.edu Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 2.0 (Macintosh; I; PPC) I am trying to measure monoamine concentration in brain tissue and report it in ng/mg protein. I am using an extraction procedure to quantify the monoamines. Does anyone know of a method or a source where I can find a mthod to help with this procedure? You can reach me either at the above e-mail address or at dbrager@gate.net Thanks for all your help, Darrin Brager Florida Atlantic University From owner-proteins@net.bio.net Wed Aug 07 23:00:00 1996 Path: biosci!agate!newsgate.duke.edu!news-feed-1.peachnet.edu!usenet.eel.ufl.edu!warwick!bham!usenet From: T.M.Franklin@bham.ac.uk (Tanya) Newsgroups: bionet.molbio.proteins Subject: Histone 1 Ab Date: 8 Aug 1996 14:17:07 GMT Organization: The University of Birmingham, UK Lines: 9 Message-ID: <4ucst3$9bd@sun4.bham.ac.uk> NNTP-Posting-Host: bcs124.bham.ac.uk X-Newsreader: WinVN version 0.80 Dear All, Is there anyone out there who has an antibody raised to histone 1, preferably the concerved region. I have a protein which I suspect might be a histone and I need to be able to confirm or disprove the posibility of its being a Histone 1. I have sent the protein for amino acid analysis but would like to be able to provide further prof. Thanks in advance, Tan. From owner-proteins@net.bio.net Wed Aug 07 23:00:00 1996 Path: biosci!agate!howland.erols.net!newsserver.jvnc.net!crick.bms.com!usenet From: "Patricia W. Cash" Newsgroups: bionet.molbio.proteins Subject: Re: glycine strand in tagged protein Date: Thu, 08 Aug 1996 08:16:13 -0700 Organization: Bristol-Myers Squibb Lines: 17 Message-ID: <320A04BD.457@bms.com> References: NNTP-Posting-Host: 140.176.114.56 Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 2.02 (Win16; I) Peter Hohenstein wrote: > > Hello, > > I'm working on the expression of a cDNA in 3T3 and I intend to tag this > protein C terminally with the HA tag. Somebody told me about putting > several glycine residues between the protein of interest and the tag, Peter, We've used a spacer amino acid such as amino caproic acid to separate peptides or proteins from tags. The amino caproic acid is convenient because it's added in one step and it seemed to give enough distance so as not to interfere with the binding of the protein. Pat From owner-proteins@net.bio.net Wed Aug 07 23:00:00 1996 Path: biosci!agate!howland.erols.net!vixen.cso.uiuc.edu!newsfeed.internetmci.com!in3.uu.net!newsfeed.pitt.edu!newsflash.concordia.ca!news.mcgill.ca!VM1.MCGILL.CA From: Susan James Newsgroups: bionet.molbio.proteins Subject: Re: Tryptic digest problems Date: 08 AUG 96 16:43:56 EST Organization: McGill University Lines: 35 Sender: usenet@MUSICA.MCGILL.CA Message-ID: <08AUG96.18070864.0093@VM1.MCGILL.CA> References: <01bb7e20$2afd09a0$5d715380@UT.cc.utexas.edu> NNTP-Posting-Host: vm1.mcgill.ca In article tedmunds@world.std.com (Tim Edmunds) writes: >In article <01bb7e20$2afd09a0$5d715380@UT.cc.utexas.edu>, "Brett S. >Phinney" wrote: > >> I was wondering if anybody had any experience doing tryptic digests using >> SDS-PAGE and reversed phase HPLC to separate their protein fragments. I'm >> trying to digest 2 proteins, and collect and sequence the fragments, the >> problem is that when I digest the protein and run it on an SDS-PAGE gel I >> see what I consider is a good digestion if not complete, i.e. the two >> places where the proteins should be are now gone. But when I run the same >> sample on an HPLC using a 2 mm C-18 column, I see 1 big peek and a few >> smaller ones. The big peek is about 50-100 times bigger then the smaller >> ones. Could it be possible that all of my fragments are co-eluting, or I'm >> not getting a good digestion. If I'm not getting a good digestion, then why >> do I not see any of my original protein, or at least very large fragments, >> when I run them on a gel. I should get about 30 fragments after a trypsin >> digestion. > >You do not say if you reduced and alkylated the protein prior to digestion >or if you are running reduced SDS gels. If the tryptic digest was done on >native protein then the disulfide bonds would remain intact during HPLC >giving rise to fewer and larger peptides than you would see on a reducing >SDS gel. I would suggest reducton and alkylation prior to digestion or >following digstion but prior to HPLC depending on what your objective is. > >Tim Edmunds >. >. I have done many tryptic digests on peptides. One problem with reducing and alkylating them first (at least with vinyl pyridine) is that the trypsin will often cleave at the site of the vinyl-pyridine, giving rise to way too many fragments that are hard to interpret. We do the digest first and the alkylation after. Sue From owner-proteins@net.bio.net Wed Aug 07 23:00:00 1996 Path: biosci!agate!ihnp4.ucsd.edu!gondor!newsfeeder.sdsu.edu!news.sgi.com!swrinde!cs.utexas.edu!howland.erols.net!vixen.cso.uiuc.edu!newsfeed.internetmci.com!in3.uu.net!EU.net!usenet2.news.uk.psi.net!uknet!usenet1.news.uk.psi.net!uknet!uknet!strath-cs!queens-belfast.ac.uk!queens-belfast.ac.uk!nntp Newsgroups: bionet.molbio.proteins Subject: Recombinant ICE and Apopain/CPP32 Message-ID: <320A05DF.78D2@queens-belfast.ac.uk> From: Andrew Wallace Date: Thu, 08 Aug 1996 16:21:03 +0100 Nntp-Posting-Host: awall.bc.qub.ac.uk X-Mailer: Mozilla 2.0 (Macintosh; I; PPC) MIME-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit Lines: 12 Does anyone know where I can get some recombinant ICE and apopain/CPP32? I am in contact with a company called Upstate Biotechnology which sells His6-tagged apopain but they are out of it at the moment. Are there any other apoptosis-associated aspases currently available? Thanks, Andrew -- ====================================================================== Andrew Wallace,Ph.D., Queen's University Belfast, N. Ireland (UK) a.wallace@qub.ac.uk http://www.qub.ac.uk/b&bchem/awpage/wallace.htm ====================================================================== From owner-proteins@net.bio.net Wed Aug 07 23:00:00 1996 Path: biosci!ihnp4.ucsd.edu!gondor!newsfeeder.sdsu.edu!news.sgi.com!swrinde!cs.utexas.edu!howland.erols.net!vixen.cso.uiuc.edu!newsfeed.internetmci.com!in3.uu.net!EU.net!usenet2.news.uk.psi.net!uknet!usenet1.news.uk.psi.net!uknet!uknet!strath-cs!queens-belfast.ac.uk!queens-belfast.ac.uk!nntp Newsgroups: bionet.molbio.proteins Subject: Re: Cathepsin D Message-ID: <320A055A.4C44@queens-belfast.ac.uk> From: Andrew Wallace Date: Thu, 08 Aug 1996 16:18:50 +0100 References: Nntp-Posting-Host: awall.bc.qub.ac.uk X-Mailer: Mozilla 2.0 (Macintosh; I; PPC) MIME-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit Lines: 15 According to the Benyon and Bond book on proteolytic enzymes, the specificity of Cathepsin D is similar to that of pepsin, i.e. -P1-P1'- (P1 = non-specific but aromatic and other hydrophobic residues preferred, especially Phe, Leu; P1, P1' cannot be Val, Ala, Gly). Reference: Takayuki, T. and Tang, J. (1980) Methods Enzym. v.80, p.565. Andrew -- ====================================================================== Andrew Wallace,Ph.D., Queen's University Belfast, N. Ireland (UK) a.wallace@qub.ac.uk http://www.qub.ac.uk/b&bchem/awpage/wallace.htm ====================================================================== From owner-proteins@net.bio.net Wed Aug 07 23:00:00 1996 Path: biosci!agate!newsgate.duke.edu!solaris.cc.vt.edu!newsfeed.internetmci.com!in2.uu.net!01-newsfeed.univie.ac.at!03-newsfeed.univie.ac.at!news.univie.ac.at!emb1.bcc.univie.ac.at!rc150295 From: Christian Radauer Newsgroups: bionet.molbio.proteins Subject: Re: solubilisation for 2-D gels Date: Thu, 8 Aug 1996 21:13:43 +0200 Organization: Vienna University, Austria Lines: 55 Message-ID: References: <9607230031.AA21598@maths.marc.cri.nz> NNTP-Posting-Host: emb1.bcc.univie.ac.at Mime-Version: 1.0 Content-Type: TEXT/PLAIN; charset=US-ASCII In-Reply-To: <9607230031.AA21598@maths.marc.cri.nz> On 22 Jul 1996, Laurence Barker wrote: > Subject: solubilisation for 2-D gels > > Hello, > I am attempting to run membrane proteins on a 2-D gel, but am > experiencing no separation in the IEF first dimension. I have tried using > 0.5% Triton X-100 in the solubilisation buffer, and have now been > informed that minimal SDS/DTT might help also. Does anyone know about the > use of chloryl hydrate to help in solubilisation/inhibition of > aggregation, if that is what is happening ? > Any help glady accepted, > > Laurence Barker. > > The Horticultural and Food Research Institute of New Zealand, > Mt Albert Research Centre, > Auckland, > New Zealand > l.barker@auckland.ac.nz Hi Laurence, I have no experience with membrane proteins, but I don't have problems separating clear and DNA-free solutions using the following solutions: Sample solution: 9 M Urea, 4% CHAPS, 0.5% SDS, 2% 2-Mercaptoethanol, 2% Ampholytes Gel solution: 3.9% Acrylamide, 0.1% PDA, 1.5% CHAPS, 0.5% NP-40, 8.5 M Urea, 2% Ampholytes The dipolar detergent CHAPS is said to be quite efffective in solubilizing proteins, small concentrations of SDS should enhance the resolution. The addition of reducing agents (2-ME or DTT) has no influence on protein solubilisation but disturbs your pH-Gradient above pH 7.5, where the thiol groups dissociate, so use it only if you are not intersted in basic proteins. Hope that helps, Christian ------------------------------------------------------- Christian Radauer Institute of General and Experimental Pathology University of Vienna AKH 3Q Phone: +43 (1) 40400 5116 Waehringer Guertel 18-20 FAX: +43 (1) 40400 5130 A-1090 Wien Austria e-mail: rc150295@emb1.bcc.univie.ac.at ------------------------------------------------------- From owner-proteins@net.bio.net Wed Aug 07 23:00:00 1996 Path: biosci!JASON.UTHCT.EDU!SHAUN From: SHAUN@JASON.UTHCT.EDU ("Shaun D. Black") Newsgroups: bionet.molbio.proteins Subject: Sequencing Proteins from Westerns of Urea-treated samples Date: 8 Aug 1996 11:05:52 -0700 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 28 Sender: daemon@net.bio.net Distribution: world Message-ID: <960808130444.6777@jason.uthct.edu> NNTP-Posting-Host: net.bio.net Hi All, Recently, we have had trouble sequencing bands were cut from Western blots of samples that have been extracted with 3.5 M urea . We know that the sequencer is working correctly from known controls run before and after (at the same pmol level). Briefly, samples in 0.1 M Tris pH 7.4 were treated with 3.5 M urea for 15 min at RT. Urea is ultrapure molecular-biology grade used without further purification. Samples are then centrifuged and supernatant and pellet fractions are separated. Fractions are then diluted about 1:1 vol/vol with SDS-PAGE gel loading buffer and submitted to SDS-PAGE with a Laemmli system. Samples were boiled for 2 min before loading. Then, when gels had been run, they were soaked briefly in blotting buffer (Towbin with a smidge of SDS), blotted to PVDF, and then stained with amido black. Bands were excised with a razor blade, and bands were submitted to sequence analysis with an ABI gas-phase instrument. No peaks above 1-2 pmol were seen at any cycle (with multiple bands from the blot). Any idea why? Could small amounts of cyanate from the urea be blocking our proteins, even in the presence of 0.1 M Tris as a scavenger? Any ideas how we can get around this? Thanks, in advance! Best regards, Shaun =-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-= = Shaun D. Black, PhD | Internet address: shaun@jason.uthct.edu = = Dept. of Biochemistry | University of Texas Health Center, at Tyler = = World Wide Web: http://pegasus.uthct.edu/UTHCT-Home/Welcome.html = =-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-= From owner-proteins@net.bio.net Thu Aug 08 23:00:00 1996 Path: biosci!agate!spool.mu.edu!usenet.eel.ufl.edu!news-res.gsl.net!news.gsl.net!news.mathworks.com!newsfeed.internetmci.com!in2.uu.net!news.compuserve.com!news.production.compuserve.com!news From: Pier Carlo Montecucchi <100143.765@CompuServe.COM> Newsgroups: bionet.molbio.proteins,bionet.molecules.peptides,sci.bio.technology,sci.chem Subject: Snail toxins block pain Date: 9 Aug 1996 10:23:56 GMT Organization: HI-TECH INDUSTRY CONSULTANTS Lines: 5 Distribution: inet Message-ID: <4uf3js$6ui$1@mhade.production.compuserve.com> Xref: biosci bionet.molbio.proteins:8469 bionet.molecules.peptides:403 sci.bio.technology:5890 sci.chem:61860 Does some people know something about omega toxin developed at Neurex Corp.? Regards Pier Carlo Montecucchi E mail 100143,765@compuserve.com From owner-proteins@net.bio.net Thu Aug 08 23:00:00 1996 Path: biosci!agate!howland.erols.net!vixen.cso.uiuc.edu!newsfeed.internetmci.com!info.ucla.edu!unixg.ubc.ca!van-bc!n1van.istar!van.istar!west.istar!ott.istar!istar.net!tor.istar!east.istar!newsjunkie.ans.net!newsfeeds.ans.net!cronkite.d48.lilly.com!newsmgr From: "David B. Flora" Newsgroups: bionet.molbio.proteins Subject: Re: Sequencing Proteins from Westerns of Urea-treated samples Date: Fri, 09 Aug 1996 15:44:15 +0000 Organization: Lilly Research Laboratories Lines: 13 Message-ID: <320B5CCF.2DC5@lilly.com> References: <960808130444.6777@jason.uthct.edu> NNTP-Posting-Host: tray.d50.lilly.com Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 2.02 (Macintosh; I; PPC) Shaun D. Black wrote: > > Any idea why? Could small amounts of cyanate from the urea be blocking > our proteins, even in the presence of 0.1 M Tris as a scavenger? Any ideas > how we can get around this? Thanks, in advance! Best regards, Shaun Hello, It sounds like the urea you are using may be blocking your N-termini...you could try passing your urea solution over a mixed bed ion-exchange resin such as Amberlite Monobed (made by Rohm and Haas). Dave From owner-proteins@net.bio.net Thu Aug 08 23:00:00 1996 Path: biosci!agate!usenet From: Zia Newsgroups: bionet.molbio.proteins Subject: Sequencing of small peptides Date: Fri, 09 Aug 1996 13:38:15 -0700 Organization: University of California, Berkeley Lines: 19 Message-ID: <320BA1B7.3FF@uclink4.berkeley.edu> Reply-To: ziadn@uclink4.berkeley.edu NNTP-Posting-Host: ziadn.hip.berkeley.edu Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 3.0b5aGold (Win16; I) Hi! In our laboratory we got some custom peptides syntesized by company. We had asked to bitonylate peptides at NH2 terminal. When peptides came some peptides related to one virus are labelled Biot-Spacer A.A.-Amino Acids of our sequence-OH while dor other virus the peptides are labelled Biot-Spacer A.A.-Amino Acids of our sequence-NH2. I feel that company might have done some mistake either in labelling or actually synthesizing the peptide in reverse sequence. The sequence we supplied is about 15-20 A.A. long and spacer is about 5-6 A.A. long. I am interested to sequence these peptides. Can any one tell me how easy/hard it may be to sequence these peptides? We do not have capability to sequence the peptides, so is it very expensive to get it sequenced commercially or can any lab help to sequence those peptides as a help to me? I do not need to sequence all of them. May be one or two peptides of each virus should be enough to sequence to figure out whether there is any mistake in synthesizing the peptides. Thanks, Zia. From owner-proteins@net.bio.net Thu Aug 08 23:00:00 1996 Path: biosci!agate!spool.mu.edu!uwm.edu!chi-news.cic.net!News1.mcs.net!van-bc!unixg.ubc.ca!lpss From: lpss@unixg.ubc.ca (Alex Chang) Newsgroups: bionet.molbio.proteins Subject: BioRad PROTEIN A BINDING BUFFER Date: 9 Aug 1996 19:01:33 GMT Organization: University of British Columbia, Vancouver, B.C., Canada Lines: 13 Message-ID: <4ug1ud$qra@nntp.ucs.ubc.ca> NNTP-Posting-Host: netinfo.ubc.ca X-Newsreader: TIN [version 1.2 PL2] I am using BioRad's Protein A column to purify some antibody. They have a binding buffer (protein A MAPS II Binding Buffer) comes with the column. I am running out of the buffer now and it is very expensive to buy a new batch. Anyone knows the recepe of the buffer please email me. -- Alex Chang Pathology University of British Columbia achang@hivnet.ubc.ca From owner-proteins@net.bio.net Thu Aug 08 23:00:00 1996 Path: biosci!agate!howland.erols.net!vixen.cso.uiuc.edu!newsfeed.internetmci.com!in3.uu.net!ott.istar!istar.net!tor.istar!east.istar!news.nstn.ca!newsflash.concordia.ca!news.mcgill.ca!VM1.MCGILL.CA From: Susan James Newsgroups: bionet.molbio.proteins Subject: Re: Histone 1 Ab Date: 09 AUG 96 14:34:23 EST Organization: McGill University Lines: 24 Sender: usenet@MUSICA.MCGILL.CA Message-ID: <09AUG96.15739001.0074@VM1.MCGILL.CA> References: <4ucst3$9bd@sun4.bham.ac.uk> NNTP-Posting-Host: vm1.mcgill.ca In article kevin.bailey@nottingham.ac.uk () writes: >In article <4ucst3$9bd@sun4.bham.ac.uk>, T.M.Franklin@bham.ac.uk (Tanya) >wrote: > >> Dear All, >> >> Is there anyone out there who has an antibody raised to histone 1, preferably >> the concerved region. I have a protein which I suspect might be a histone and >> I need to be able to confirm or disprove the posibility of its being a Histone 1. >> I have sent the protein for amino acid analysis but would like to be able to >> provide further prof. >> >> Thanks in advance, Tan. > > You could also consider digesting the protein and sequencing a fragment. >A clear sequence would identify it as histone, though I appreciate there >are regions common to more than one class of histones. >. >. You might also try to have and mass done of your sample. Either ionspray or Maldi should give you an exact mass with relatively little material. Less than 1nmole should be enough. Susan From owner-proteins@net.bio.net Thu Aug 08 23:00:00 1996 Path: biosci!agate!howland.erols.net!vixen.cso.uiuc.edu!newsfeed.internetmci.com!usenet.eel.ufl.edu!warwick!news.nott.ac.uk!NewsWatcher!user From: kevin.bailey@nottingham.ac.uk () Newsgroups: bionet.molbio.proteins Subject: Re: Histone 1 Ab Followup-To: bionet.molbio.proteins Date: Fri, 09 Aug 1996 13:30:43 +0100 Organization: Cripps Computing Centre, The University of Nottingham Lines: 16 Message-ID: References: <4ucst3$9bd@sun4.bham.ac.uk> NNTP-Posting-Host: wpbjwk1.biochem.nottingham.ac.uk In article <4ucst3$9bd@sun4.bham.ac.uk>, T.M.Franklin@bham.ac.uk (Tanya) wrote: > Dear All, > > Is there anyone out there who has an antibody raised to histone 1, preferably > the concerved region. I have a protein which I suspect might be a histone and > I need to be able to confirm or disprove the posibility of its being a Histone 1. > I have sent the protein for amino acid analysis but would like to be able to > provide further prof. > > Thanks in advance, Tan. You could also consider digesting the protein and sequencing a fragment. A clear sequence would identify it as histone, though I appreciate there are regions common to more than one class of histones. From owner-proteins@net.bio.net Thu Aug 08 23:00:00 1996 Path: biosci!agate!howland.erols.net!vixen.cso.uiuc.edu!newsfeed.internetmci.com!in3.uu.net!ott.istar!istar.net!van.istar!west.istar!n1van.istar!van-bc!unixg.ubc.ca!news.bc.net!uvaix3e1.comp.UVic.CA!usenet From: olafson@uvic.ca (Robert Olafson) Newsgroups: bionet.molbio.proteins Subject: Re: Sequencing Proteins from Westerns of Urea-treated samples Date: Fri, 09 Aug 1996 23:10:56 GMT Organization: University of Victoria Lines: 56 Message-ID: <4ugge1$1g2g@uvaix3e1.comp.UVic.CA> References: <960808130444.6777@jason.uthct.edu> NNTP-Posting-Host: masamune.bioc.uvic.ca X-Newsreader: Forte Free Agent 1.0.82 SHAUN@JASON.UTHCT.EDU ("Shaun D. Black") wrote: >Hi All, > Recently, we have had trouble sequencing bands were cut from Western >blots of samples that have been extracted with 3.5 M urea . We know >that the sequencer is working correctly from known controls run before and >after (at the same pmol level). > Briefly, samples in 0.1 M Tris pH 7.4 were treated with 3.5 M urea for >15 min at RT. Urea is ultrapure molecular-biology grade used without further >purification. Samples are then centrifuged and supernatant and pellet >fractions are separated. Fractions are then diluted about 1:1 vol/vol >with SDS-PAGE gel loading buffer and submitted to SDS-PAGE with a Laemmli >system. Samples were boiled for 2 min before loading. Then, when gels had >been run, they were soaked briefly in blotting buffer (Towbin with a >smidge of SDS), blotted to PVDF, and then stained with amido black. Bands >were excised with a razor blade, and bands were submitted to sequence analysis >with an ABI gas-phase instrument. No peaks above 1-2 pmol were seen at >any cycle (with multiple bands from the blot). > Any idea why? Could small amounts of cyanate from the urea be blocking >our proteins, even in the presence of 0.1 M Tris as a scavenger? Any ideas >how we can get around this? Thanks, in advance! Best regards, Shaun > =-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-= > = Shaun D. Black, PhD | Internet address: shaun@jason.uthct.edu = > = Dept. of Biochemistry | University of Texas Health Center, at Tyler = > = World Wide Web: http://pegasus.uthct.edu/UTHCT-Home/Welcome.html = > =-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-= Hi Shaun, Cyanate from urea is notorious for blocking N-terminals in neutral to basic pH. The best way I know of to guard against this is to acidify your urea stock. In acidic donditions (pH 6.0) the cyanates are destroyed. So make sure it is acidified prior to using it. The formation of these compounds is time dependant so the shorter your protein is exposed to urea in a non acidic environment, the better. Since you are loading the samples onto a gel immediately, I don't believe you will have a problem. However, if you still don't get any sequence, take the PVDF out of the sequencer and subject it to amino acid analysis. The binding of stain to protein is always sample dependant, and you may not have as much material as you think you do. I should tell you that there is an alternative method for cleaning up your urea, and that is to store it over few mL of mixed bed ion exchange resin which will suck up all the "nasties" as my boss likes to call them. I think Amberlite resin is the one that has been used in this lab in the past. I've never done this, but my supervisor assures me that it works. He's from the old school, so the older the procedure, the better it works. Good luck, Dustin Lippert From owner-proteins@net.bio.net Thu Aug 08 23:00:00 1996 Path: biosci!agate!howland.erols.net!vixen.cso.uiuc.edu!newsfeed.internetmci.com!info.ucla.edu!unixg.ubc.ca!van-bc!n1van.istar!van.istar!west.istar!ott.istar!istar.net!tor.istar!east.istar!newsjunkie.ans.net!newsfeeds.ans.net!cronkite.d48.lilly.com!newsmgr From: "David B. Flora" Newsgroups: bionet.molbio.proteins Subject: Re: Sequencing Proteins from Westerns of Urea-treated samples Date: Fri, 09 Aug 1996 15:46:42 +0000 Organization: Lilly Research Laboratories Lines: 13 Message-ID: <320B5D62.74F8@lilly.com> References: <960808130444.6777@jason.uthct.edu> NNTP-Posting-Host: tray.d50.lilly.com Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 2.02 (Macintosh; I; PPC) Shaun D. Black wrote: > > Any idea why? Could small amounts of cyanate from the urea be blocking > our proteins, even in the presence of 0.1 M Tris as a scavenger? Any ideas > how we can get around this? Thanks, in advance! Best regards, Shaun Hello, It sounds like the urea you are using may be blocking your N-termini...you could try passing your urea solution over a mixed bed ion-exchange resin such as Amberlite Monobed (made by Rohm and Haas). Dave From owner-proteins@net.bio.net Thu Aug 08 23:00:00 1996 Path: biosci!agate!howland.erols.net!vixen.cso.uiuc.edu!newsfeed.internetmci.com!in2.uu.net!EU.net!usenet2.news.uk.psi.net!uknet!usenet1.news.uk.psi.net!uknet!uknet!yama.mcc.ac.uk!news.salford.ac.uk!aber!bath.ac.uk!dcl-cs!strath-cs!st-and!gatty!njc From: Nicholas James Cole Newsgroups: bionet.molbio.proteins Subject: phosphorylation Date: Fri, 9 Aug 1996 17:04:03 +0100 Organization: St. Andrews University Lines: 4 Message-ID: NNTP-Posting-Host: gatty.st-and.ac.uk Mime-Version: 1.0 Content-Type: TEXT/PLAIN; charset=US-ASCII NNTP-Posting-User: njc X-Sender: njc@gatty hello can anyone out there suggest a method to analyse proteins (Muscle) in such a way that the same sample is first analysed as normal and then analysed fully phosphorylated and then dephosphorylated...using gel electrophoresis? many thanks for any advice, From owner-proteins@net.bio.net Thu Aug 08 23:00:00 1996 Path: biosci!agate!howland.erols.net!vixen.cso.uiuc.edu!newsfeed.internetmci.com!info.ucla.edu!unixg.ubc.ca!news.bc.net!uvaix3e1.comp.UVic.CA!usenet From: olafson@uvic.ca (Robert Olafson) Newsgroups: bionet.molbio.proteins Subject: Re: Sequencing Proteins from Westerns of Urea-treated samples Date: Fri, 09 Aug 1996 23:32:42 GMT Organization: University of Victoria Lines: 19 Message-ID: <4ughmq$2nb4@uvaix3e1.comp.UVic.CA> References: <960808130444.6777@jason.uthct.edu> NNTP-Posting-Host: masamune.bioc.uvic.ca X-Newsreader: Forte Free Agent 1.0.82 Our resident protein sequencing expert had some further thoughts on your situation. She has apparently never seen a successful sequence of an amido black stained protein, however, this probably is the nature of the protein and not the stain. Part of the problem might be the staining solution. I'm not sure what the conditions of staining are, but if your amido black is in acetic acid, the acid may be acetylating your N-terminal. This commonly occurs when people use standard protocols for Coomassie staining, which are all done in 5-10% acetic acid. Most of our samples are stained with Coomassie in 40% methanol and destained with 50% methanol. You might consider this as an alternative. The only other possibility that we can think of is that your protein is naturally blocked, in which case you're hooped...hope that's not your problem. again, good luck Dustin Lippert From owner-proteins@net.bio.net Fri Aug 09 23:00:00 1996 Path: biosci!agate!howland.erols.net!cs.utexas.edu!swrinde!news.sgi.com!news.msfc.nasa.gov!newsfeed.internetmci.com!in3.uu.net!news00.sunet.se!sunic!news99.sunet.se!news.uni-c.dk!news From: jmollerup@aki.ku.dk (Jens Mollerup) Newsgroups: bionet.molbio.proteins Subject: Re: antibody to alkaline phosphatase (E.coli) Date: Fri, 09 Aug 1996 10:52:29 GMT Organization: News Server at UNI-C, Danish Computing Centre for Research and Education. Lines: 19 Message-ID: <4uf4j1$ef8@news.uni-c.dk> References: <4t49dj$c5h@cc-server9.massey.ac.nz> NNTP-Posting-Host: pc152.aki.ku.dk X-Newsreader: Forte Free Agent 1.0.82 Richard Johnson wrote: >Does anyone know of a good source for anti-alkaline phosphatase antibody? >thanks >Richard DAKO A/S has a rabbit anti-calf alkaline phosphatase antibody (code no. Z0271), that probably could be delivered to you by MED-BIO Enterprises Ltd. in New Zealand Good Luck Jens Mollerup Univ. Copenhagen From owner-proteins@net.bio.net Fri Aug 09 23:00:00 1996 Path: biosci!agate!howland.erols.net!vixen.cso.uiuc.edu!newsfeed.internetmci.com!in2.uu.net!server-b.cs.interbusiness.it!jussieu.fr!univ-lr.fr!usenet From: jlopes@ketch.univ-lr.fr (Joao Lopes) Newsgroups: bionet.molbio.proteins Subject: Cutinase Date: 10 Aug 1996 18:01:46 GMT Organization: Universite de La Rochelle Lines: 15 Message-ID: <4uiiqa$b1n@hpuniv.univ-lr.fr> NNTP-Posting-Host: bio.univ-lr.fr Keywords: Cutinase, Enzyme, Chromatography, Purification I'm trying to purify Cutinase by ion exchange chromatography from a mix of proteins ( about 50 % cutinase ). Until now, two collums were used ( cation and anion exchangers ), but in my lab we've optimized a method using only a cation exchanger. The problem of the purified enzyme Cutinase is that it aggregates and so we loose some activity. Methods used for breaking aggregates are well knowed but all methods I've try gave week results. Sonication, pH changes, additives are an exemple. Dilution raises activity but then I have a biggest problem. If someone can give an opinion, I'll be grateful. joao.lopes@bio.univ-lr.fr From owner-proteins@net.bio.net Fri Aug 09 23:00:00 1996 Path: biosci!agate!howland.erols.net!vixen.cso.uiuc.edu!newsfeed.internetmci.com!info.ucla.edu!nnrp.info.ucla.edu!unixg.ubc.ca!van-bc!n1van.istar!van.istar!west.istar!ott.istar!istar.net!tor.istar!east.istar!newsjunkie.ans.net!newsfeeds.ans.net!news-m01.ny.us.ibm.net!news.biu.ac.il!news.huji.ac.il!avdat3.bgu.ac.il!usenet From: irisp@bgumail.bgu.ac.il (Iris Porat) Newsgroups: bionet.molbio.proteins Subject: Lolking for PCR program Date: Sat, 10 Aug 1996 14:45:01 GMT Organization: Ben Gurion University Lines: 6 Message-ID: <320c9fcf.697550@news.bgu.ac.il> NNTP-Posting-Host: bgumail.bgu.ac.il X-Newsreader: Forte Agent .99c/16.141 I loking for PCR program, for PC, something similar to the program Amplify that exist for Machintosh. Thank you, Iris Porat Ben Gurion University Israel From owner-proteins@net.bio.net Fri Aug 09 23:00:00 1996 Path: biosci!CC.CSIC.ES!ciblr1a From: ciblr1a@CC.CSIC.ES (Gloria) Newsgroups: bionet.molbio.proteins Subject: Help with dimers Date: 10 Aug 1996 13:12:15 -0700 Organization: cib Lines: 637 Sender: daemon@net.bio.net Distribution: world Message-ID: <01I84KI1PWSI008CDH@pinarvms.csic.es> NNTP-Posting-Host: net.bio.net Hello! We have a little problem with our protein. When we analyse the cellular extracts by by Western blotting we detect two bands: one correspondig wito the protein and the other with an apparent molecular weight according with a dimer. The problem appears when we try to disgregate the dimer with DTT 0.1 M, Urea+SDS+heat that it results imposible. Could someone tell us how to get it? ormay be everything is an artefact. Thanks. Gloria Gonzale> PROTEIN-ANALYSIS/bionet.molbio.proteins Newsgroup Archive > > -------------------------------------------------------------------- > Please visit our sponsors' WWW pages and tell them you saw their ad > on BIOSCI. > > [Research Diagnostics, Inc.] [The Nest Group, Inc.] > > See us for antibodies, antigens... RPC, HILIC, IEX & HIC Chromatography > > -------------------------------------------------------------------- > Post a new article to PROTEIN-ANALYSIS/bionet.molbio.proteins via > the BIOSCI mail-Usenet gateway. 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RPC, HILIC, IEX & HIC Chromatography > > -------------------------------------------------------------------- > Post a new article to PROTEIN-ANALYSIS/bionet.molbio.proteins via > the BIOSCI mail-Usenet gateway. (Requires that your browser be able > to handle mailto URLs.) > -------------------------------------------------------------------- > Read the most recent PROTEIN-ANALYSIS/bionet.molbio.proteins > articles (Aug 96) > -------------------------------------------------------------------- > Back to the BIOSCI Home Page > -------------------------------------------------------------------- > The archive is broken down by month (MM) and year (YY) > The articles are stored in directories named YYMM > > -------------------------------------------------------------------- > The archives are indexed using freeWAIS. To search for a keyword or > phrase occurring in the PROTEIN-ANALYSIS/bionet.molbio.proteins > newsgroup, enter your query below. freeWAIS supports the Boolean > operators "and" and "not" and the truncation operator *. If multiple > search terms are entered without boolean operators, messages > containing any one or more of the terms are returned, i.e., an "or" > search is the default. For example > > PCR and sequenc* not DNA > > will find messages that contain the text "PCR" and any words > starting with "sequenc" (sequence, sequencing, etc.) but exclude > messages containing the text "DNA." Terms do not occur in any > specific order in the message. > > Search for: > > Maximum hits returned: > > > > -------------------------------------------------------------------- > > [ ] Name Last modified Size Description > -------------------------------------------------------------------- > > [DIR] Parent Directory 10-Aug-96 12:08 - > > [DIR] 8912/ 13-Nov-95 01:42 - > > [DIR] 9001/ 13-Nov-95 01:42 - > > [DIR] 9002/ 13-Nov-95 01:42 - > > [DIR] 9003/ 13-Nov-95 01:42 - > > [DIR] 9004/ 13-Nov-95 01:42 - > > [DIR] 9005/ 13-Nov-95 01:42 - > > [DIR] 9006/ 13-Nov-95 01:42 - > > [DIR] 9007/ 13-Nov-95 01:42 - > > [DIR] 9008/ 13-Nov-95 01:42 - > > [DIR] 9009/ 13-Nov-95 01:42 - > > [DIR] 9010/ 13-Nov-95 01:43 - > > [DIR] 9011/ 13-Nov-95 01:43 - > > [DIR] 9012/ 13-Nov-95 01:43 - > > [DIR] 9101/ 13-Nov-95 01:43 - > > [DIR] 9102/ 13-Nov-95 01:43 - > > [DIR] 9103/ 13-Nov-95 01:43 - > > [DIR] 9104/ 13-Nov-95 01:43 - > > [DIR] 9105/ 13-Nov-95 01:43 - > > [DIR] 9106/ 13-Nov-95 01:43 - > > [DIR] 9107/ 13-Nov-95 01:43 - > > [DIR] 9108/ 13-Nov-95 01:43 - > > [DIR] 9109/ 13-Nov-95 01:43 - > > [DIR] 9110/ 13-Nov-95 01:43 - > > [DIR] 9111/ 13-Nov-95 01:43 - > > [DIR] 9112/ 13-Nov-95 01:43 - > > [DIR] 9201/ 13-Nov-95 01:43 - > > [DIR] 9202/ 13-Nov-95 01:43 - > > [DIR] 9203/ 13-Nov-95 01:43 - > > [DIR] 9204/ 13-Nov-95 01:43 - > > [DIR] 9205/ 13-Nov-95 01:43 - > > [DIR] 9206/ 13-Nov-95 01:43 - > > [DIR] 9207/ 13-Nov-95 01:43 - > > [DIR] 9208/ 13-Nov-95 01:44 - > > [DIR] 9209/ 13-Nov-95 01:44 - > > [DIR] 9210/ 13-Nov-95 01:44 - > > [DIR] 9211/ 13-Nov-95 01:44 - > > [DIR] 9212/ 13-Nov-95 01:44 - > > [DIR] 9301/ 13-Nov-95 01:44 - > > [DIR] 9302/ 13-Nov-95 01:44 - > > [DIR] 9303/ 13-Nov-95 01:44 - > > [DIR] 9304/ 13-Nov-95 01:44 - > > [DIR] 9305/ 13-Nov-95 01:44 - > > [DIR] 9306/ 13-Nov-95 01:44 - > > [DIR] 9307/ 13-Nov-95 01:44 - > > [DIR] 9308/ 13-Nov-95 01:45 - > > [DIR] 9309/ 13-Nov-95 01:45 - > > [DIR] 9310/ 13-Nov-95 01:45 - > > [DIR] 9311/ 13-Nov-95 01:45 - > > [DIR] 9312/ 13-Nov-95 01:46 - > > [DIR] 9403/ 13-Nov-95 01:46 - > > [DIR] 9404/ 13-Nov-95 01:47 - > > [DIR] 9405/ 13-Nov-95 01:47 - > > [DIR] 9406/ 13-Nov-95 01:47 - > > [DIR] 9407/ 13-Nov-95 01:48 - > > [DIR] 9408/ 13-Nov-95 01:48 - > > [DIR] 9409/ 13-Nov-95 01:49 - > > [DIR] 9410/ 13-Nov-95 01:49 - > > [DIR] 9411/ 13-Nov-95 01:49 - > > [DIR] 9412/ 13-Nov-95 01:50 - > > [DIR] 9501/ 13-Nov-95 01:50 - > > [DIR] 9502/ 13-Nov-95 01:51 - > > [DIR] 9503/ 13-Nov-95 01:51 - > > [DIR] 9504/ 13-Nov-95 01:52 - > > [DIR] 9505/ 13-Nov-95 01:53 - > > [DIR] 9506/ 13-Nov-95 01:53 - > > [DIR] 9507/ 13-Nov-95 01:54 - > > [DIR] 9508/ 13-Nov-95 01:54 - > > [DIR] 9509/ 13-Nov-95 01:55 - > > [DIR] 9510/ 13-Nov-95 01:56 - > > [DIR] 9511/ 27-Dec-95 19:27 - > > [DIR] 9512/ 31-Dec-95 18:25 - > > [DIR] 9601/ 31-Jan-96 15:49 - > > [DIR] 9602/ 29-Feb-96 21:40 - > > [DIR] 9603/ 31-Mar-96 19:13 - > > [DIR] 9604/ 30-Apr-96 23:01 - > > [DIR] 9605/ 31-May-96 21:26 - > > [DIR] 9606/ 30-Jun-96 19:42 - > > [DIR] 9607/ 31-Jul-96 18:57 - > > [DIR] 9608/ 10-Aug-96 11:26 - z From owner-proteins@net.bio.net Fri Aug 09 23:00:00 1996 Path: biosci!agate!howland.erols.net!cs.utexas.edu!swrinde!news.sgi.com!news.msfc.nasa.gov!newsfeed.internetmci.com!info.ucla.edu!nnrp.info.ucla.edu!wlheye.jsei.ucla.edu!hassane From: hassane@wlheye.jsei.ucla.edu (hassane mchaourab) Newsgroups: bionet.molbio.proteins Subject: Beckman HPLC:system gold nouveau Date: 10 Aug 1996 21:34:09 GMT Organization: Jules Stein Eye Institute, UCLA Lines: 7 Message-ID: <4uiv8h$19a8@uni.library.ucla.edu> NNTP-Posting-Host: wlheye.jsei.ucla.edu Hi, I am setting up a new lab. I am considering for HPLC the beckman system gold noveau(Plastic Pumps). Since I have never used this equipment and could not find many users in my department I am looking for advice. Thanks From owner-proteins@net.bio.net Sat Aug 10 23:00:00 1996 Message-ID: <320EC071.1E80@oa.net> Date: Mon, 12 Aug 1996 00:26:09 -0500 From: Charles X-Mailer: Mozilla 2.0 (Macintosh; I; PPC) MIME-Version: 1.0 Newsgroups: bionet.cellbiol,bionet.immunology,bionet.general,bionet.molbio.proteins Subject: Re: Eukaryotic Promoter Database References: Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit NNTP-Posting-Host: cewhiteh.virtual.oa.net Lines: 4 Path: biosci!rutgers!gatech!arclight.uoregon.edu!usenet.eel.ufl.edu!news-res.gsl.net!news.gsl.net!swrinde!cs.utexas.edu!howland.erols.net!news.sprintlink.net!news-stk-200.sprintlink.net!news.sprintlink.net!news-pen-14.sprintlink.net!newshost.oa.net! Xref: biosci bionet.cellbiol:5245 bionet.immunology:9521 bionet.general:22945 bionet.molbio.proteins:8485 Dear Brent: The Signal Scan data base can be used over the Web: http://bimas.dcrt.nih.gov/molbio/signal/ Good Luck...... From owner-proteins@net.bio.net Sun Aug 11 23:00:00 1996 Path: biosci!rutgers!gatech!news.jsums.edu!news.uoregon.edu!newsfeed.orst.edu!newshub.tc.umn.edu!newsstand.tc.umn.edu!dialup-4-6.gw.umn.edu!user From: bilha001@gold.tc.umn.edu (Brent) Newsgroups: bionet.cellbiol,bionet.immunology,bionet.general,bionet.molbio.proteins Subject: Eukaryotic Promoter Database Date: Sun, 11 Aug 1996 15:31:02 -0600 Organization: J&B INC. Lines: 7 Message-ID: NNTP-Posting-Host: dialup-4-6.gw.umn.edu Xref: biosci bionet.cellbiol:5246 bionet.immunology:9522 bionet.general:22946 bionet.molbio.proteins:8486 I have heard of a searchable database that will allow me to search promoters for potential target genes based on the consensus binding site of a DNA binding protein. Can somebody direct me to this database? Thanks, BRENT From owner-proteins@net.bio.net Sun Aug 11 23:00:00 1996 Message-ID: <320EC272.599D@oa.net> Date: Mon, 12 Aug 1996 00:34:42 -0500 From: Charles X-Mailer: Mozilla 2.0 (Macintosh; I; PPC) MIME-Version: 1.0 Newsgroups: bionet.cellbiol,bionet.immunology,bionet.general,bionet.molbio.proteins To: Brent Subject: Re: Eukaryotic Promoter Database References: Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit NNTP-Posting-Host: cewhiteh.virtual.oa.net Lines: 4 Path: biosci!rutgers!uwm.edu!cs.utexas.edu!news.sprintlink.net!news-stk-200.sprintlink.net!news.sprintlink.net!news-pen-14.sprintlink.net!newshost.oa.net! Xref: biosci bionet.cellbiol:5247 bionet.immunology:9524 bionet.general:22947 bionet.molbio.proteins:8488 Brent: You can use the Signal Scan Program at this Web site: http://bimas.dcrt.nih.gov/molbio/signal/ Good Luck...... From owner-proteins@net.bio.net Sun Aug 11 23:00:00 1996 Path: biosci!rutgers!news.sgi.com!news.msfc.nasa.gov!newsfeed.internetmci.com!newsxfer2.itd.umich.edu!portc01.blue.aol.com!newsstand.cit.cornell.edu!newstand.syr.edu!usenet From: dslomczy@mailbox.syr.edu Newsgroups: bionet.molbio.proteins Subject: Re: Beckman HPLC:system gold nouveau Date: 11 Aug 1996 19:11:36 GMT Organization: Syracuse University, Syracuse, NY (USA) Lines: 16 Message-ID: <4ulb98$449@newstand.syr.edu> References: <4uiv8h$19a8@uni.library.ucla.edu> NNTP-Posting-Host: sudial-69.syr.edu X-Newsreader: News for Windows NT X1.0-74 >I am setting up a new lab. I am considering for HPLC the beckman >system gold noveau(Plastic Pumps). Since I have never used this >equipment and could not find many users in my department I am looking >for advice. >Thanks What are going to use the equipment for? Will it be for Protein purification or analytical methods. First ask this question then look for the equipment which is best suited for your needs. There are many excellent manufacturers of HPLC equipment (HP, Waters, LKB-Pharmacia, Isco, and others). Look at all the equipment and then decide - it can be an expensive mistake. I hope this helps David Slomczynski dslomczy@mailbox.syr.edu From owner-proteins@net.bio.net Sun Aug 11 23:00:00 1996 Path: biosci!rutgers!uwm.edu!cs.utexas.edu!howland.erols.net!surfnet.nl!news.wau.nl!usenet From: William Kloek Newsgroups: bionet.molbio.proteins Subject: Program for schematic representation proteins Date: 12 Aug 1996 09:10:40 GMT Organization: Wageningen Agricultural University Lines: 14 Message-ID: <4umseg$or4@Trex.IenD.wau.nl> NNTP-Posting-Host: flex060.biot.wau.nl Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 1.22 (Windows; I; 16bit) HI, I am looking for a program that can make from an aminoacid sequence of a protein that characterizes the protein with a straight line. On this line information should be given by means of markers concerning hydrophobic residues, Pro residues and possibilities for S-S bridges.It should also be possible to indicate the charge by a vertical line up -or down, depending on the charge being positive or negative and the length should indicate the magnitude of the charge. Does anybody know if such a program is available in the public domain. Please also E-mail, Thanks From owner-proteins@net.bio.net Sun Aug 11 23:00:00 1996 Path: biosci!rutgers!uwm.edu!news-res.gsl.net!news.gsl.net!usenet.eel.ufl.edu!warwick!bham!usenet From: altabios@bham.ac.uk (John E. Fox) Newsgroups: bionet.molbio.proteins Subject: Re: Sequencing of small peptides Date: 12 Aug 1996 12:39:59 GMT Organization: Alta Bioscience Lines: 22 Message-ID: <4un8mv$grl@sun4.bham.ac.uk> References: <320BA1B7.3FF@uclink4.berkeley.edu> NNTP-Posting-Host: bcs118.bham.ac.uk X-Newsreader: WinVN 0.90.6 In article <320BA1B7.3FF@uclink4.berkeley.edu>, Zia says: > >Hi! In our laboratory we got some custom peptides syntesized by company. >We had asked to bitonylate peptides at NH2 terminal. When peptides came >some peptides related to one virus are labelled Biot-Spacer A.A.-Amino >Acids of our sequence-OH while dor other virus the peptides are labelled Dear Zia, Looks to me as though some of the peptides have free acid groups at the C-terminal and written as sequence - OH The others labelled - NH2 will have amide C-terminal. By any chance do the NH2 peptides have proline as their C-amino acid? That would explain the choice of amide. John Fox ****************************************************************** Alta BioScience Email: altabios@bham.ac.uk School of Biochemistry Phone: 0121-414-5450 The University of Birmingham Fax: 0121-414-3376 Edgbaston, BIRMINGHAM, B15 2TT, UK From owner-proteins@net.bio.net Sun Aug 11 23:00:00 1996 Path: biosci!GUITAR.ROCKEFELLER.EDU!ilya From: ilya@GUITAR.ROCKEFELLER.EDU (Ilya Vakser) Newsgroups: bionet.molbio.proteins Subject: Announcement - protein docking program GRAMM Date: 12 Aug 1996 15:30:16 -0700 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 92 Sender: daemon@net.bio.net Distribution: world Message-ID: <320FB1B6.167E@guitar.rockefeller.edu> NNTP-Posting-Host: net.bio.net ----------- GRAMM v1.03 ----------- This note is to announce the availability of GRAMM (Global RAnge Molecular Matching) software. GRAMM is a program for protein docking. To predict the structure of a complex, it requires only the atomic coordinates of the two molecules (no information about the binding sites is needed). The program performs an exhaustive 6-dimensional search through the relative translations and rotations of the molecules. The molecular pairs may be: two proteins, a protein and a smaller compound, two transmembrane (TM) helices, etc. GRAMM may be used for high- resolution molecules, for inaccurate structures (where only the gross structural features are known), in cases of large conformational changes, etc. The Global Range Molecular Matching (GRAMM) methodology is an empirical approach to smoothing the intermolecular energy function by changing the range of the atom-atom potentials. The technique allows to locate the area of the global minimum of intermolecular energy for structures of different accuracy. The quality of the prediction depends on the accuracy of the structures. Thus, the docking of high-resolution structures with small conformational changes yields an accurate prediction, while the docking of ultralow-resolution structures will give only the gross features of the complex. _________________ I am making GRAMM publicly available following a number of requests from different labs. I would like to make it clear, however, that both the methodology and the program, at present, are in the process of active development and validation, especially in the area of the low-resolution docking, and have to be viewed like that. The program is free. However, I would expect proper references. I will also appreciate bug reports. To get GRAMM, send the request to ilya@guitar.rockefeller.edu. The GRAMM site on the Web is http://guitar.rockefeller.edu/. GRAMM was created during my stay in the Weizmann Institute (1991-1993), in the Washington University (1993-1995), and in the Rockefeller University (1995-1996). I deeply appreciate the assistance of my colleagues in all these institutions (especially, Profs. Ephraim Katchalski-Katzir, Garland Marshall, and Andrej Sali). Platforms Presently, GRAMM is compiled on SGI R4000, SGI R4400, SGI R8000, and SGI R10000 Unix workstations. In the near future I will expand this list, so check the GRAMM site for the updates. Interestingly, GRAMM also works on a PC platform under Windows95 (the performance on P5-120 with 16 MB RAM is only two times slower than on SGI 250 MHZ Indigo2 R4400). Papers on GRAMM methodology * E. Katchalski-Katzir, I. Shariv, M. Eisenstein, A. A. Friesem, C. Aflalo, I. A. Vakser, 1992, Molecular surface recognition: Determination of geometric fit between proteins and their ligands by correlation techniques, Proc. Natl. Acad. Sci. USA, 89, 2195-2199. (Basic algorithm of protein recognition by correlation technique with Fast Fourier transform. High-resolution 'geometric' docking). * I. A. Vakser, C. Aflalo, 1994, Hydrophobic docking: A proposed enhancement to molecular recognition techniques, Proteins, 20, 320-329. (High-resolution 'hydrophobic' docking). * I. A. Vakser, G. V. Nikiforovich, 1995a, Protein docking in the absence of detailed molecular structures, in: Methods in Protein Structure Analysis (M. Z. Atassi & E. Appella, eds.), Plenum Press, New York, pp. 505-514. * I. A. Vakser, 1995b, Protein docking for low-resolution structures, Protein Eng., 8, 371- 377. ('Low-resolution' protein docking). * I. A. Vakser, 1996a, Long-distance potentials: An approach to the multiple-minima problem in ligand-receptor interaction, Protein Eng., 9, 37-41. (Interpretation of the low-resolution docking in terms of energy potentials). * I. A. Vakser, 1996b, Low-resolution docking: Prediction of complexes for underdetermined structures, Biopolymers, in press (39, No.3). (Validation of the low-resolution docking). * I. A. Vakser, 1996c, Main-chain complementarity in protein-protein recognition, Protein Eng., in press (9, No.7). (Docking of Ca structures). ------------------------------------------------------------------ Ilya A. Vakser The Rockefeller University, Box 270 1230 York Avenue, New York, NY 10021 Phone: (212) 327-7206; Fax: (212) 327-7540 Email: ilya@guitar.rockefeller.edu ------------------------------------------------------------------ From owner-proteins@net.bio.net Sun Aug 11 23:00:00 1996 Message-ID: <320EC2A4.4AE3@oa.net> Date: Mon, 12 Aug 1996 00:35:32 -0500 From: Charles X-Mailer: Mozilla 2.0 (Macintosh; I; PPC) MIME-Version: 1.0 Newsgroups: bionet.cellbiol,bionet.immunology,bionet.general,bionet.molbio.proteins Subject: Re: Eukaryotic Promoter Database References: Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit NNTP-Posting-Host: cewhiteh.virtual.oa.net Lines: 8 Path: biosci!rutgers!uwm.edu!cs.utexas.edu!news.sprintlink.net!news-stk-200.sprintlink.net!news.sprintlink.net!news-pen-14.sprintlink.net!newshost.oa.net! Xref: biosci bionet.cellbiol:5248 bionet.immunology:9525 bionet.general:22948 bionet.molbio.proteins:8489 Brent wrote: > > I have heard of a searchable database that will allow me to search > promoters for potential target genes based on the consensus binding site > of a DNA binding protein. Can somebody direct me to this database? > > Thanks, > BRENT From owner-proteins@net.bio.net Sun Aug 11 23:00:00 1996 Path: biosci!rutgers!uwm.edu!vixen.cso.uiuc.edu!newsfeed.internetmci.com!in2.uu.net!EU.net!usenet2.news.uk.psi.net!uknet!usenet1.news.uk.psi.net!uknet!uknet!lyra.csx.cam.ac.uk!daresbury!bioftp.unibas.ch!news.vub.ac.be!news.belnet.be!swsbe6.switch.ch!scsing.switch.ch!rzunews.unizh.ch!NewsWatcher!user From: maga@vetbio.unizh.ch (Giovanni Maga) Newsgroups: bionet.molbio.proteins Subject: Re: phosphorylation Date: Mon, 12 Aug 1996 17:37:43 +0200 Organization: University of Zurich Irchel- Biochemistry Lines: 19 Message-ID: References: NNTP-Posting-Host: 130.60.120.18 In article , Nicholas James Cole wrote: > hello can anyone out there suggest a method to analyse proteins (Muscle) > in such a way that the same sample is first analysed as normal and then > analysed fully phosphorylated and then dephosphorylated...using gel > electrophoresis? many thanks for any advice, Know it sounds obvious but...what about start with a certain amount (i.e. 1 ug) of your protein in 70 ul (just to say), take out 20 ul and load the gel (unmodified sample), add the kinase, make the reaction take out 20 ul, load (phosphorylated sample), add the phosphatase, make the reaction, take 20 ul and load (dephosphorylated sample). This if you worry about the reproducibility. If you do not, the easiest thing is to set up four parallel reactions (unmodified sample incubated alone and plus phosphatase as negative controls; sample incubated with the kinase, sample incubated with kinase and then with phosphatase). Don't know...doesn't it sound good enough to you? Giovanni. From owner-proteins@net.bio.net Sun Aug 11 23:00:00 1996 Path: biosci!rutgers!uwm.edu!news-res.gsl.net!news.gsl.net!nntp.coast.net!news.dacom.co.kr!usenet.seri.re.kr!news.postech.ac.kr!usenet.kornet.nm.kr!agate!newsgate.duke.edu!godot.cc.duq.edu!newsfeed.pitt.edu!gatech!EU.net!main.Germany.EU.net!fu-berlin.de!news.th-darmstadt.de!tutor.oc.chemie.th-darmstadt.de!martin From: martin@tutor.oc.chemie.th-darmstadt.de (Martin Kroeker) Newsgroups: bionet.molbio.proteins Subject: Re: inhibitor modelling based on 3D structures Date: 12 Aug 1996 15:53:53 GMT Organization: Technische Hochschule Darmstadt Lines: 18 Distribution: world Message-ID: <4unk2h$ods@rs18.hrz.th-darmstadt.de> References: <320F4683.F84@pclsp2.kuicr.kyoto-u.ac.jp> NNTP-Posting-Host: tutor.oc.chemie.th-darmstadt.de X-Newsreader: TIN [version 1.2 PL2] mihara (mihara@PCLSP2.KUICR.KYOTO-U.AC.JP) wrote: : Dear Sir, : We are looking for an inhibitor for L-haloacid dehalogenase, i.e., compound(s) : structurally similar to L-2-chloropropionic acid. We heard that through Cambridge : Structural Database(CSD), a lot of compounds understood by X-ray analysis might have : such information but we can not get access to this database. Could you tell us how to : access to CSD or introduce to us candidate compounds? Thank you in advance. : Li and Nishihara The CSD is distributed as a single CD-ROM containing crystallographic data of approx. 140000 compounds. Please check out http://www.ccdc.cam.ac.uk for more details and postal addresses of their regional contacts. Hope this helps, Martin -- Dr.-Ing. Martin Kroeker Inst. f. Organ. Chemie martin@oc2.oc.chemie.th-darmstadt.de Univ. (TH) Darmstadt db7p@hrzpub.th-darmstadt.de Germany From owner-proteins@net.bio.net Sun Aug 11 23:00:00 1996 Path: biosci!rutgers!uwm.edu!newsfeed.internetmci.com!newsxfer2.itd.umich.edu!agate!newsgate.duke.edu!news-server.ncren.net!concert!newz.oit.unc.edu!usenet From: "Helen R. Mott" Newsgroups: bionet.molbio.proteins Subject: GMPPCP Date: Mon, 12 Aug 1996 11:41:39 -0400 Organization: University of North Carolina Lines: 14 Message-ID: <320F50B3.41C6@med.unc.edu> NNTP-Posting-Host: indy2.med.unc.edu Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 2.0 (X11; I; IRIX 5.3 IP22) Does anyone happen to know of a supplier of the non-hydrolysable GTP analog GMPPCP within the USA? We used to get supplies from Boehringer, Fluka or Calbiochem but they all seem to have stopped making it! Thanks in advance Helen Mott ----------------------------------------------------------------- Dr Helen R. Mott Department of Biochemistry and Biophysics hrm@med.unc.edu University of North Carolina, CB #7260 Chapel Hill, NC 27599 Fax(919) 966 2852 Ph:(919) 966 6781 ----------------------------------------------------------------- From owner-proteins@net.bio.net Sun Aug 11 23:00:00 1996 Path: biosci!PCLSP2.KUICR.KYOTO-U.AC.JP!mihara From: mihara@PCLSP2.KUICR.KYOTO-U.AC.JP (mihara) Newsgroups: bionet.molbio.proteins Subject: inhibitor modelling based on 3D structures Date: 12 Aug 1996 06:54:50 -0700 Organization: Institute for Chemical Research ,Kyoto University Lines: 7 Sender: daemon@net.bio.net Distribution: world Message-ID: <320F4683.F84@pclsp2.kuicr.kyoto-u.ac.jp> Reply-To: mihara@pclsp2.kuicr.kyoto-u.ac.jp NNTP-Posting-Host: net.bio.net Dear Sir, We are looking for an inhibitor for L-haloacid dehalogenase, i.e., compound(s) structurally similar to L-2-chloropropionic acid. We heard that through Cambridge Structural Database(CSD), a lot of compounds understood by X-ray analysis might have such information but we can not get access to this database. Could you tell us how to access to CSD or introduce to us candidate compounds? Thank you in advance. Li and Nishihara From owner-proteins@net.bio.net Sun Aug 11 23:00:00 1996 Path: biosci!rutgers!uwm.edu!spool.mu.edu!munnari.OZ.AU!news.ecn.uoknor.edu!solace!eru.mt.luth.se!bloom-beacon.mit.edu!news.mathworks.com!enews.sgi.com!news.sgi.com!cgl!picasso.ucsf.edu!smiller From: smiller@picasso.ucsf.edu (Stephen Miller) Newsgroups: bionet.molbio.proteins,bionet.molecules.peptides,sci.bio.technology,sci.chem Subject: Re: Snail toxins block pain Date: 12 Aug 96 20:20:20 GMT Organization: UCSF Computer Graphics Lab Lines: 14 Distribution: inet Message-ID: References: <4uf3js$6ui$1@mhade.production.compuserve.com> NNTP-Posting-Host: picasso-yp.ucsf.edu Xref: biosci bionet.molbio.proteins:8496 bionet.molecules.peptides:405 sci.bio.technology:5911 sci.chem:61984 Pier Carlo Montecucchi <100143.765@CompuServe.COM> writes: >Does some people know something about omega toxin developed at >Neurex Corp.? Look up papers by Baldomero Olivera in Medline or CC. Neurex also publishes some papers, usually including J. Ramachandran among the authors. There's another competing group in Tokyo at the Mitsubishi Institute of (something). The omega-conotoxins are neuronal calcium-channel blockers, and are all rigid peptides with THREE disulfide bonds. Impressive conformational restriction! Steve. From owner-proteins@net.bio.net Mon Aug 12 23:00:00 1996 Path: biosci!rutgers!csn!nntp-xfer-1.csn.net!magnus.acs.ohio-state.edu!math.ohio-state.edu!howland.erols.net!agate!spool.mu.edu!munnari.OZ.AU!news.unimelb.EDU.AU!murphy From: rm@clyde.its.unimelb.edu.au (Roger ) Newsgroups: bionet.molbio.proteins,bionet.molecules.peptides,sci.bio.technology,sci.chem Subject: Re: Snail toxins block pain Date: Wed, 14 Aug 96 03:47:51 GMT Organization: University of Melbourne Lines: 23 Distribution: inet Message-ID: <4urloq$5at@news.unimelb.EDU.AU> References: <4uf3js$6ui$1@mhade.production.compuserve.com> NNTP-Posting-Host: murphy.pharmacology.unimelb.edu.au X-Newsreader: News Xpress 2.0 Beta #0 Xref: biosci bionet.molbio.proteins:8504 bionet.molecules.peptides:407 sci.bio.technology:5920 sci.chem:62010 In article , smiller@picasso.ucsf.edu (Stephen Miller) wrote: >Pier Carlo Montecucchi <100143.765@CompuServe.COM> writes: > >>Does some people know something about omega toxin developed at >>Neurex Corp.? > >Look up papers by Baldomero Olivera in Medline or CC. Neurex also publishes >some papers, usually including J. Ramachandran among the authors. There's >another competing group in Tokyo at the Mitsubishi Institute of (something). >The omega-conotoxins are neuronal calcium-channel blockers, and are all >rigid peptides with THREE disulfide bonds. Impressive conformational >restriction! > >Steve. > Neurex have apparently used omega-conotoxin MVIIA injected intrathecally for chronic pain relief. Otherwise, I agree with Steve - look up either Olivera's papers or K. Sato's papers. Cheers, Roger Murphy From owner-proteins@net.bio.net Mon Aug 12 23:00:00 1996 Path: biosci!rutgers!csn!nntp-xfer-1.csn.net!carbon!night.primate.wisc.edu!ames!enews.sgi.com!news.sgi.com!news.cs.indiana.edu!purdue!mozo.cc.purdue.edu!news From: Vadim Beilinson Newsgroups: bionet.molbio.proteins Subject: info on polyclonal antibodies against soybean BBI needed Date: 13 Aug 1996 18:48:18 GMT Organization: Purdue University Lines: 5 Message-ID: <4uqili$qui@mozo.cc.purdue.edu> NNTP-Posting-Host: soypr2.agry.purdue.edu Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 1.2N (Windows; I; 16bit) Dear Researcher, would you please provide me with information on a source for polyclonal antibodies against soybean Bowman-Birk inhibitor (BBI). Thank you very much for your time. Vadim. From owner-proteins@net.bio.net Mon Aug 12 23:00:00 1996 Path: biosci!rutgers!uwm.edu!lll-winken.llnl.gov!enews.sgi.com!news.sgi.com!swrinde!howland.erols.net!agate!newsgate.duke.edu!godot.cc.duq.edu!usenet From: "Frank R. Gorga" Newsgroups: bionet.molbio.proteins Subject: Re: Specificity of NEM toward cysteine residues Date: Tue, 13 Aug 1996 13:43:24 -0700 Organization: Duquesne University Lines: 35 Message-ID: <3210E8EC.6EE0@next.duq.edu> References: NNTP-Posting-Host: phosphorus.chemistry.duq.edu Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 2.0 (Win16; I) Dominique DELFORGE wrote: > > My questions are: Does someone know something about the c