From owner-proteins@net.bio.net Tue Oct 01 23:00:00 1996 Path: biosci!daresbury!not-for-mail From: ditlev@kemi.aau.dk (Ditlev Brodersen) Newsgroups: bionet.molbio.proteins Subject: Re: $$MAKE MONEY FAST$$ READ ME NOW!! Date: 2 Oct 1996 19:50:49 +0100 Lines: 24 Sender: lpddist@mserv1.dl.ac.uk Distribution: bionet Message-ID: <52udi9$t1g@mserv1.dl.ac.uk> X-Sender: ditlev@beauty.kemi.aau.dk Original-To: proteins@dl.ac.uk GOT YOUR MONEY BUT LOST YOUR MORAL !!!!!!! I most strongly urge you NOT to use serious and specific news-group lists like this for such junk. No one wants to listen to it anyway !!!! If you really had earned that amount of money, why would you want to tell the whole world about it ??? I certainly hope everyone reading this is clever enough not to spend money on such a silly thing. All intelligent people know that you can never make something out of nothing - the same goes for money. -- DB ------------------------------------------------------------------- Ditlev Brodersen Phone: +45 8942 3871 Dept. of Struct. and Mol. Biology Fax: +45 8619 6199 c/o Dept. of Chemistry Email: ditlev@kemi.aau.dk Langelandsgade 140 ditlev@beauty.kemi.aau.dk DK-8000 Aarhus C Denmark WWW:http://kaktus.kemi.aau.dk/~ditlev From owner-proteins@net.bio.net Tue Oct 01 23:00:00 1996 Path: biosci!agate!newsgate.duke.edu!news-feed-1.peachnet.edu!usenet.eel.ufl.edu!arclight.uoregon.edu!enews.sgi.com!news.sgi.com!www.nntp.primenet.com!nntp.primenet.com!news.mindspring.com!uunet!news-in2.uu.net!rcogate.rco.qc.ca!altitude!usenet From: "Achim Recktenwald, PhD" Newsgroups: bionet.molbio.proteins,sci.bio Subject: Re: proteins -please help! Date: Wed, 02 Oct 1996 08:02:03 -0500 Organization: IBEX Technologies, Inc., Biochemistry, 5485 Pare, Montreal, PQ, H4P 1P7, Canada Lines: 24 Message-ID: <325267CB.1E14@ibex.ca> References: <269370222961424ntc@ukonline.co.uk> 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 3.0 (Macintosh; I; PPC) To: trystandragon@ukonline.co.uk paul wrote: > > I am a college student and am writing a project on proteins. I need any > material, preferably in text form, on proteins and what they are, what > they do and how. Anything would be useful, but a general introduction > into proteins would be best. I have looked, but could not find much > material on the 'net, however my access time is severely limited. Site > urls would be useful, a private e-mail with the text in it would be > even better, as this would be quickest, and my access time is the > important factor. > > Thanks in advance for any help > > Paul > > -------------------------------------------- > trystandragon@ukonline.co.uk > -DFE-UDIC- Go into any university library and you can bury yourself in books about proteins. Achim From owner-proteins@net.bio.net Tue Oct 01 23:00:00 1996 Path: biosci!bcm.tmc.edu!news.msfc.nasa.gov!newsfeed.internetmci.com!howland.erols.net!EU.net!Norway.EU.net!sn.no!nntp.uio.no!nntp-oslo.UNINETT.no!nntp-trd.UNINETT.no!daresbury!is.bbsrc.ac.uk!news From: Mick Partis Newsgroups: bionet.molbio.proteins Subject: Re: Elution of protein from dry SDS_PAGE Date: 2 Oct 1996 11:45:56 GMT Organization: Horticulture Research International Lines: 27 Message-ID: <52tklk$ut8@is.bbsrc.ac.uk> References: <4104vsav-3009962009010001@mac118.envir.ulaval.ca> NNTP-Posting-Host: pc0233.hriw.bbsrc.ac.uk Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 1.1N (Windows; I; 16bit) 4104vsav@vm1.ulaval.ca (Savita Visal) wrote: >Dear Friends, >I want to sequence protein of interest. I have my protein of interest as a >single band on SDS_PAGE. I've dried this gel. I would like sequence this >protein..the problem is elution from the SDS-PAGE. I have read so many >protocols and at this stage I'm totally confused ..which protocol do I >use? MW of my protein is 55kDa. Could anyone share their experiences and >guide me to which is the best protocol I could use? >Thanks > >Savita >4104vsav@vm1.ulaval.ca I hope that you are not thinking of eluting the protein into solution as implied in your subject heading. If so, you will lose protein during elution, and may get modification of your protein. Much better to rehydrate your gel and blot your protein onto PDVF as described in Ron Tate's posting. For transfer buffer, I use a CAPS buffer rather than Tris-glycine to avoid a mammoth glycine peak in the first cycle. Mick -- mick.partis@hri.ac.uk Horticulture Research International http://www.geocities.com/CapeCanaveral/1957/ From owner-proteins@net.bio.net Tue Oct 01 23:00:00 1996 Path: biosci!daresbury!not-for-mail From: ditlev@kemi.aau.dk (Ditlev Brodersen) Newsgroups: bionet.molbio.proteins Subject: Re: protein-bound Zn measurement needed Date: 2 Oct 1996 16:53:00 +0100 Lines: 28 Sender: lpddist@mserv1.dl.ac.uk Distribution: bionet Message-ID: <52u34s$gvl@mserv1.dl.ac.uk> X-Sender: ditlev@beauty.kemi.aau.dk Original-To: proteins@dl.ac.uk >Does anyone konw of a lab which can identify protein-bound metals using >ug quantities of sample? > >==================================================== >* John H. Collins, Ph.D. * >* Medical Biotechnology Center * >* University of Maryland Biotechnology Institute * >* 108 N. Greene Street * >* Baltimore, MD 21201-1503 * >* USA * >* e-mail: collins@umbi.umd.edu * >==================================================== Why don't you somehow dissociate the metalion from the protein and run an atom absorbance spectroscopy - they can determine metals down to ppm/ppb which should suffice ------------------------------------------------------------------- Ditlev Brodersen Phone: +45 8942 3871 Dept. of Struct. and Mol. Biology Fax: +45 8619 6199 c/o Dept. of Chemistry Email: ditlev@kemi.aau.dk Langelandsgade 140 ditlev@beauty.kemi.aau.dk DK-8000 Aarhus C Denmark WWW:http://kaktus.kemi.aau.dk/~ditlev From owner-proteins@net.bio.net Tue Oct 01 23:00:00 1996 Path: biosci!faseb.org!cpk-news-feed1.bbnplanet.com!cpk-news-hub1.bbnplanet.com!www.nntp.primenet.com!nntp.primenet.com!howland.erols.net!EU.net!usenet2.news.uk.psi.net!uknet!usenet1.news.uk.psi.net!uknet!uknet!strath-cs!str-ccsun!usenet From: linda brooks Newsgroups: bionet.molbio.proteins Subject: Protein L Date: 1 Oct 1996 17:09:47 GMT Organization: university of strathclde Lines: 9 Message-ID: <52rj8r$9q4@rockall.cc.strath.ac.uk> NNTP-Posting-Host: oscar.imm.strath.ac.uk Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 1.12(Macintosh; I; PPC) X-URL: news:bionet.molbio.proteins Hello, I wish to use protein L to create an immunoaffinity column for purification of recombinant antibody variable light chains - does anyone know of a commercial supplier of protein L? Thanks Linda From owner-proteins@net.bio.net Tue Oct 01 23:00:00 1996 Path: biosci!agate!howland.erols.net!news-peer.gsl.net!news.gsl.net!nntp.coast.net!oleane!in2p3.fr!swidir.switch.ch!scsing.switch.ch!rzunews.unizh.ch!NewsWatcher!user From: zjons@vetbio.unizh.ch (Zophonias O. Jonsson) Newsgroups: bionet.molbio.proteins Subject: Re: protein-bound Zn measurement needed Date: Wed, 02 Oct 1996 22:08:59 +0200 Organization: Universitat Zurich-Irchel Lines: 29 Distribution: bionet Message-ID: References: <52u34s$gvl@mserv1.dl.ac.uk> NNTP-Posting-Host: 130.60.120.18 In article <52u34s$gvl@mserv1.dl.ac.uk>, ditlev@kemi.aau.dk (Ditlev Brodersen) wrote: > >Does anyone konw of a lab which can identify protein-bound metals using > >ug quantities of sample? > > > > Why don't you somehow dissociate the metalion from the protein and run an > atom absorbance spectroscopy - they can determine metals down to ppm/ppb > which should suffice Why would you have to dissociate the metal from the protein for atomic absorbtion? There will probably not be very much left of the protein in the flame or in the carbon arc anyway by the time you vaporise the metal. Or am I missing something obvious?? As to the sensitivity I agree, if the protein in question has reasonable affinity for the metal. I actually tried to detect Zn in this way once, but didn't find any, probably because the protein didn't bind Zink. At least it was an easy experiment! Zophonias _____________________________________________________________________ Zophonias O. Jonsson Institut fur Veterinarbiochemie Tel: (41-1)-257-54-75 Universitat Zurich-Irchel Fax: (41-1)-362-05-01 Winterthurerstrasse 190 CH-8057 Zurich Switzerland _____________________________________________________________________ From owner-proteins@net.bio.net Tue Oct 01 23:00:00 1996 Path: biosci!agate!howland.erols.net!cs.utexas.edu!swrinde!news.sgi.com!enews.sgi.com!decwrl!tribune.usask.ca!skyfox.usask.ca!yangj From: yangj@skyfox.usask.ca Newsgroups: bionet.molbio.proteins Subject: RE: $$MAKE MONEY FAST$$ READ ME NOW!! Date: 2 OCT 96 12:42:53 GMT Organization: University of Saskatchewan Lines: 1 Message-ID: <2OCT96.12425369@skyfox.usask.ca> References: NNTP-Posting-Host: sask.usask.ca Really? Then stay away from this newsgroup. You stupid jerk. From owner-proteins@net.bio.net Tue Oct 01 23:00:00 1996 Path: biosci!faseb.org!cpk-news-feed1.bbnplanet.com!cpk-news-hub1.bbnplanet.com!www.nntp.primenet.com!nntp.primenet.com!news.sgi.com!news-out.microserve.net!news-in.microserve.net!news.supernet.net!news.thenet.net!uunet!in3.uu.net!caen!umass.edu!nic.umass.edu!austen.oit.umass.edu!JSUMNER From: JSUMNER@austen.oit.umass.edu (Jason C Sumner) Newsgroups: bionet.molbio.proteins Subject: chaperone molecules Date: 3 Oct 1996 03:35:26 GMT Organization: University of Massachusetts, Amherst Lines: 9 Message-ID: <52vc9u$djq@nic.umass.edu> NNTP-Posting-Host: austen.oit.umass.edu X-Newsreader: TIN [version 1.2 PL2v [VAX/VMS]] I'm considering doing a research paper on the repairing ability of chaperone melecules (and possibly how the chaperone molecules find their own structures in the first place). Any interesting journal articles, web pages, information, etc would be greatly apprecialted. Jason Sumner jsumner@student.umass.edu From owner-proteins@net.bio.net Tue Oct 01 23:00:00 1996 Path: biosci!SEMOVM.SEMO.EDU!C478SCB From: C478SCB@SEMOVM.SEMO.EDU (Bilbrey Robert) Newsgroups: bionet.molbio.proteins Subject: Re[2]: protein-bound Zn measurement needed Date: 2 Oct 1996 14:00:10 -0700 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 43 Sender: daemon@net.bio.net Distribution: world Message-ID: <02OCT96.16809753.0119.MUSIC@SEMOVM> NNTP-Posting-Host: net.bio.net Why would the metal ions need to be removed from the protein sample? It doesn't seem to me that inclusion of protein in a sample would necessarily interfere with identification of metalic ligands. You should be able to figure out what cations are required for enzyme activity, if the protein in question is an enzyme, by treatment with a chelating agent then adding back individual ionic species and seeing which ones reactivate the zyme. Robert Bilbrey c478scb@semovm.semo.edu Dept of Biology Southeast Missouri State Umiversity >>Does anyone konw of a lab which can identify protein-bound metals using >>ug quantities of sample? >> >>==================================================== >>* John H. Collins, Ph.D. * >>* Medical Biotechnology Center * >>* University of Maryland Biotechnology Institute * >>* 108 N. Greene Street * >>* Baltimore, MD 21201-1503 * >>* USA * >>* e-mail: collins@umbi.umd.edu * >>==================================================== > >Why don't you somehow dissociate the metalion from the protein and run an >atom absorbance spectroscopy - they can determine metals down to ppm/ppb >which should suffice > >------------------------------------------------------------------- >Ditlev Brodersen Phone: +45 8942 3871 >Dept. of Struct. and Mol. Biology Fax: +45 8619 6199 >c/o Dept. of Chemistry Email: ditlev@kemi.aau.dk >Langelandsgade 140 ditlev@beauty.kemi.aau.dk >DK-8000 Aarhus C >Denmark > >WWW:http://kaktus.kemi.aau.dk/~ditlev > > > From owner-proteins@net.bio.net Tue Oct 01 23:00:00 1996 Path: biosci!bcm.tmc.edu!news.msfc.nasa.gov!news.sgi.com!howland.erols.net!EU.net!usenet2.news.uk.psi.net!uknet!usenet1.news.uk.psi.net!uknet!uknet!strath-cs!str-ccsun!usenet From: linda brooks Newsgroups: bionet.molbio.proteins Subject: Protein L Date: 1 Oct 1996 17:09:19 GMT Organization: university of strathclde Lines: 9 Message-ID: <52rj7v$93h@rockall.cc.strath.ac.uk> NNTP-Posting-Host: oscar.imm.strath.ac.uk Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 1.12(Macintosh; I; PPC) X-URL: news:bionet.molbio.proteins Hello, I wish to use protein L to create an immunoaffinity column for purification of recombinant antibody variable light chains - does anyone know of a commercial supplier of protein L? Thanks Linda From owner-proteins@net.bio.net Tue Oct 01 23:00:00 1996 Path: biosci!agate!usenet From: Matthias Dreyer Newsgroups: bionet.molbio.proteins Subject: Re: SDS-page Date: 3 Oct 1996 03:58:17 GMT Organization: Data Communication and Newtorking Services Lines: 28 Message-ID: <52vdkp$smg@agate.berkeley.edu> References: <324BA74D.6AD91F0E@csc.fi> <52mfon$46h@winx03.informatik.uni-wuerzburg.de> NNTP-Posting-Host: sb6.cchem.berkeley.edu Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 1.1N (X11; I; IRIX 5.3 IP12) X-URL: news:52mfon$46h@winx03.informatik.uni-wuerzburg.de >Jussi Kankare (Jussi.Kankare@csc.fi) wrote: >> I'm trying to crystallize a protein and would therefore like >> to have my sample as homogeneous as possible. However when the >> purity of the sample is checked with SDS-page the gel shows a >> major and small minor band just beneath the major one. I have >> not purified the sample myself but the persons who have done >> the work claim that the sample is pure but the protein just has >> the property that it shows two bands in SDS-page ?!? I would naturally >> think that this means that the sample is NOT homogeneous. What could >> cause this sort behaviour of a protein or is there simply a need for >> repurifying the protein. I observed an additional band on SDS-PAGs that resultet from boiling the samples. The band did not appear when I simply incubated the sample for a long time in the sample buffer at room temperature. -- --------------------------------------------------------- Matthias Dreyer ____________ Department of Chemistry \ / Calvin Lab \ / University of California \ / Berkeley, CA 94720-5230 \ / USA \ / e-mail: dreyer@lcbvax.cchem.berkeley.edu \/ --------------------------------------------------------- From owner-proteins@net.bio.net Wed Oct 02 23:00:00 1996 Path: biosci!daresbury!is.bbsrc.ac.uk!news From: Mick Partis Newsgroups: bionet.molbio.proteins Subject: Re: chaperone molecules Date: 3 Oct 1996 11:06:04 GMT Organization: Horticulture Research International Lines: 21 Message-ID: <5306ms$3mr@is.bbsrc.ac.uk> References: <52vc9u$djq@nic.umass.edu> NNTP-Posting-Host: pc0233.hriw.bbsrc.ac.uk Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 1.1N (Windows; I; 16bit) JSUMNER@austen.oit.umass.edu (Jason C Sumner) wrote: > >I'm considering doing a research paper on the repairing ability of >chaperone melecules (and possibly how the chaperone molecules >find their own structures in the first place). Any interesting journal >articles, web pages, information, etc would be greatly apprecialted. > >Jason Sumner >jsumner@student.umass.edu > You should find lots of info and links on the chaperonin page http://bioc09.uthscsa.edu/~seale/Chap/chap.html Mick -- mick.partis@hri.ac.uk Horticulture Research International http://www.geocities.com/CapeCanaveral/1957/ From owner-proteins@net.bio.net Wed Oct 02 23:00:00 1996 Path: biosci!daresbury!sunsite.doc.ic.ac.uk!lyra.csx.cam.ac.uk!macr3-20.welc.cam.ac.uk!user From: whc@mole.bio.cam.ac.uk (W.H. Colledge) Newsgroups: bionet.molbio.proteins Subject: Wanted:peptide biotinylation or fluorocinylation method Date: Thu, 03 Oct 1996 20:02:05 +0000 Organization: University of Cambridge Lines: 9 Message-ID: NNTP-Posting-Host: macr3-20.welc.cam.ac.uk Does anyone have a simple method for labelling peptides with biotin or fluorocine? How can the labelled peptide be purified from the reaction products? -- Dr.W.H.Colledge Physiological Laboratory University of Cambridge Cambridge CB2 3EG From owner-proteins@net.bio.net Wed Oct 02 23:00:00 1996 Path: biosci!SCRIPPS.EDU!zhluo From: zhluo@SCRIPPS.EDU (Jean Luo) Newsgroups: bionet.molbio.proteins Subject: EGFR's intrinsic kinase activity and Serine/Threonine phosphorylation Date: 3 Oct 1996 14:03:45 -0700 Organization: The Scripps Research Institute Lines: 29 Sender: daemon@net.bio.net Distribution: world Message-ID: <32542A84.2BF5@scripps.edu> Reply-To: zhluo@scripps.edu NNTP-Posting-Host: net.bio.net Hello, everybody, Epidermal Growth Factor (EGF) and its receptor (EGFR) have been hot research projects for years. Dr. Stanley Cohen from Vanderbilt Univ. even got his Nobel prize on that. However, I could not even understand some basic biochemical characteristics of EGFR by reading their papers. I would appreciate any comments or inputs. My questions are: 1) Is the intrinsic kinase activity of EGFR needed for ligand EGF induced tyrosine kinase activity? 2) If yes, is serine / threonine phosphorylation required for this intrinsic kinase activity? 3) What are other structural motifs of the active EGFR cytoplasmic domain (such as dimerization)? 4) EGFR cytoplasmic domain (EGFRc) expressed in insect cells is active. However, bacterial expressed EGFRc (soluble) is inactive. What is mssing in EGFRc expressed from bacteria? Please email your suggestions / answers to zhluo@scripps.edu Thanks! Jean From owner-proteins@net.bio.net Wed Oct 02 23:00:00 1996 Path: biosci!bcm.tmc.edu!cs.utexas.edu!swrinde!news-peer.gsl.net!news.gsl.net!portc01.blue.aol.com!newstf01.news.aol.com!newsbf02.news.aol.com!not-for-mail From: blackorb@aol.com (Black Orb) Newsgroups: bionet.molbio.proteins Subject: Question in organismal biology Date: 4 Oct 1996 01:05:34 -0400 Organization: America Online, Inc. (1-800-827-6364) Lines: 7 Sender: root@newsbf02.news.aol.com Message-ID: <5325uu$s16@newsbf02.news.aol.com> Reply-To: blackorb@aol.com (Black Orb) NNTP-Posting-Host: newsbf02.mail.aol.com I need some help with this question I got from my biology professor: - Why do ribosomes appear to be the first membrane bound organelle that developed in procaryotic cells? I don't quite know how to begin, but any help would be appreciated!! From owner-proteins@net.bio.net Wed Oct 02 23:00:00 1996 Path: biosci!agate!spool.mu.edu!howland.erols.net!EU.net!usenet2.news.uk.psi.net!uknet!usenet1.news.uk.psi.net!uknet!uknet!lyra.csx.cam.ac.uk!macr3-20.welc.cam.ac.uk!user From: whc@mole.bio.cam.ac.uk (W.H. Colledge) Newsgroups: bionet.molbio.proteins Subject: Cell marking Date: Thu, 03 Oct 1996 19:52:40 +0000 Organization: University of Cambridge Lines: 8 Message-ID: NNTP-Posting-Host: macr3-20.welc.cam.ac.uk Does anyone have a method for labelling (non-radioactive) or marking non-dividing cells that will persists for the lifetime of the cell? -- Dr.W.H.Colledge Physiological Laboratory University of Cambridge Cambridge CB2 3EG From owner-proteins@net.bio.net Wed Oct 02 23:00:00 1996 Path: biosci!daresbury!not-for-mail From: l.kempster@ic.ac.uk (L.Kempster) Newsgroups: bionet.molbio.proteins Subject: Please help! Date: 3 Oct 1996 18:21:57 +0100 Lines: 11 Sender: lpddist@mserv1.dl.ac.uk Distribution: bionet Message-ID: <530snl$n4k@mserv1.dl.ac.uk> X-Sender: lkhl@tolstoy.nhli.ic.ac.uk Original-To: proteins@dl.ac.uk Hi Does anyone know of the existance of a mammalian REVERSIBLE translation elongation inhibitor? Thanks in advance, Lee Kempster Imperial College. From owner-proteins@net.bio.net Thu Oct 03 23:00:00 1996 Path: biosci!agate!howland.erols.net!www.nntp.primenet.com!nntp.primenet.com!hunter.premier.net!news.cais.net!nntp.uio.no!nntp-oslo.UNINETT.no!nntp-trd.UNINETT.no!daresbury!is.bbsrc.ac.uk!news From: dunnsm@bbsrc.ac.uk (steve dunn) Newsgroups: bionet.molbio.proteins Subject: selenomethionine media... Date: 4 Oct 1996 08:21:51 GMT Organization: Institute of Arable Crops Research, Rothamsted Lines: 11 Message-ID: <532hev$7i6@is.bbsrc.ac.uk> NNTP-Posting-Host: pc773.res.bbsrc.ac.uk X-Newsreader: WinVN 0.92.6+ Dear Netters, I'm looking to produce a selenomethionine derivative of an over-expressed recombinant protein from an E.coli met auxotroph for MAD phasing (crystals!!). I understand that I grow the cells in liquid minimal media (M9 glucose) + amino acids + vits. I don't however, have a specific recipe that informs me as to the concentrations of the added amino acids (incuding selenomet.) and vitamins to include in the media. Can anybody help?? Many thanks in advance, Steve dunnsm@bbsrc.ac.uk or please post replies to board. From owner-proteins@net.bio.net Thu Oct 03 23:00:00 1996 Path: biosci!agate!spool.mu.edu!uwm.edu!nntp.primenet.com!news-peer.gsl.net!news.gsl.net!news-stock.gsl.net!news.gsl.net!snunews.snu.ac.kr!usenet From: "Lee, Ji Hyun" Newsgroups: bionet.molbio.proteins Subject: Best Condition for CNBr cleavage of protein? Date: Fri, 04 Oct 1996 22:54:53 +0900 Organization: College of Pharmacy, Seoul National University Lines: 15 Message-ID: <3255172D.3CF6@plaza.snu.ac.kr> Reply-To: newera@plaza.snu.ac.kr NNTP-Posting-Host: 147.46.8.60 Mime-Version: 1.0 Content-Type: text/plain; charset=iso-8859-2 Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 3.0 (Win95; I) I'm trying to cleave a peptide from its linker with CNBr and the linker has methionine residue in its c-terminal. According my books such as Guide to protein purification and Short protocols in molecular biology, the reaction is carried in 70% formic acid and excess CNBr. Where can I get detailed information about the reaction such as a paper, book or WWW site etc. When I observed SDS-PAGE of sample after overnight incubation, I came to see that only 50 % of all peptides are cleaved by now. my protein is as follows Linker-Met-Peptide ^ cleave here Thanks. From owner-proteins@net.bio.net Thu Oct 03 23:00:00 1996 Path: biosci!agate!spool.mu.edu!howland.erols.net!newsfeed.internetmci.com!btnet!netcom.net.uk!sunsite.doc.ic.ac.uk!charlie.lif.icnet.uk!europa.lif.icnet.uk!sternber From: sternber@europa.lif.icnet.uk (Michael Sternberg) Newsgroups: bionet.molbio.proteins Subject: ANNOUNCE - PROT SEC STR PREDN WWW SERVER Date: 4 Oct 1996 14:53:30 GMT Organization: Imperial Cancer Research Fund Lines: 61 Message-ID: <5338da$e9t@charlie.lif.icnet.uk> NNTP-Posting-Host: europa.lif.icnet.uk X-Newsreader: TIN [version 1.2 PL2] From rd_king@icrf.icnet.uk Thu Oct 3 17:45:16 1996 Date: Mon, 30 Sep 1996 17:42:26 +0100 From: "Ross D. King" To: m.sternberg@icrf.icnet.uk Subject: DSC DSC Protein Secondary Structure Prediction I am pleased to announce the opening of a new web site for the prediction of protein secondary structure. Two prediction modes are available: 1) Given a single sequence. A multiple sequence alignmentt will be formed and DSC used to predict secondary structure. http://www.icnet.uk/bmm/dsc/dsc_form_align.html 2) Given a multiple sequence alignmnet. DSC will use this alignment to predict secondary structure. http://www.icnet.uk/bmm/dsc/dsc_read_align.html The advantages of DSC are: 1) It is very accurate. DSC has a prediction accuracy of 70.1% on a standard set of 126 proteins. This was not significantly different from PHD, a popular prediction method. For medium length sequences DSC was more accurate than PHD, and combining DSC and PHD produced a prediction method more accurate than either. 2) It is free. There is no charge for using DSC. 3) The C source code is available. This would allow you to run DSC on your own system if you had confidential sequences. ftp://ftp.icnet.uk/icrf-public/bmm/king/dsc/dsc.tar.z DSC is based on simple linear statistics. A paper on the scientific basis of DSC will appear in Protein Science: "Identification and Application of the Concepts Important for Accurate and Reliable Protein Secondary Structure Prediction" by Ross D. King & Michael J.E. Sternberg Ross D. King Biomolecular Modelling Laboratory Imperial Cancer Research Fund Lincoln's Inn Fields, P. O. Box 123, London, WC2A 3PX, U.K. Tel: +44 171 269 3565, Fax: +44 171 269 3479, rd_king@icrf.icnet.uk m.sternberg@icrf.icnet.uk (contact until 25 Oct 96) -- ------------------------------------------------------------------------ * from e-mail address: m.sternberg@icrf.icnet.uk * * * * Mike Sternberg Tel +44-71-269-3565 / 3591 * * Biomolecular Modelling Laboratory FAX +44-71-269-3479 * * Imperial Cancer Research Fund Others in group who can help : * * 44 Lincoln's Inn Fields Suhail Islam Tel +44-71-269-3380 * * London WC2A 3PX, England Paul Bates Tel +44-71-269-3348 * ************************************************************************ From owner-proteins@net.bio.net Thu Oct 03 23:00:00 1996 Newsgroups: bionet.molbio.proteins Path: biosci!agate!howland.erols.net!newsfeed.internetmci.com!news-in2.uu.net!utcsri!utgpu!utinfo!bloch.med.utoronto.ca!gardner From: gardner@bloch.med.utoronto.ca (Kevin Gardner) Subject: Re: protein-bound Zn measurement needed X-Nntp-Posting-Host: bloch.med.utoronto.ca Message-ID: Sender: nntp@utcc.utoronto.ca (News) Organization: UTCC Campus Access X-Newsreader: TIN [version 1.2 PL2] References: <52u34s$gvl@mserv1.dl.ac.uk> Distribution: bionet Date: Fri, 4 Oct 1996 04:10:42 GMT Lines: 34 Zophonias O. Jonsson (zjons@vetbio.unizh.ch) wrote: : In article <52u34s$gvl@mserv1.dl.ac.uk>, ditlev@kemi.aau.dk (Ditlev : Brodersen) wrote: : > >Does anyone konw of a lab which can identify protein-bound metals using : > >ug quantities of sample? : > > : > : > Why don't you somehow dissociate the metalion from the protein and run an : > atom absorbance spectroscopy - they can determine metals down to ppm/ppb : > which should suffice : Why would you have to dissociate the metal from the protein for atomic : absorbtion? There will probably not be very much left of the protein in : the flame or in the carbon arc anyway by the time you vaporise the metal. : Or am I missing something obvious?? As to the sensitivity I agree, if the : protein in question has reasonable affinity for the metal. I actually : tried to detect Zn in this way once, but didn't find any, probably because : the protein didn't bind Zink. At least it was an easy experiment! Correct on the point of not needing to dissociate the metal away from the protein coordinating it prior to use in flame atomic absorption spectroscopy. As to the sensitivity, machines are fairly(?) routinely available to detect uM concentrations of zinc from ca. 500uL - 1mL of sample (e.g. nanomoles of Zn required). Hats off to the mighty d10 crew, Kevin -- ************************************************************************* Kevin Gardner gardner@bloch.med.utoronto.ca University of Toronto http://abragam.med.utoronto.ca/~gardner Dept. of Medical Genetics & Microbiology phone: 416-978-0642/FAX: -6885 From owner-proteins@net.bio.net Thu Oct 03 23:00:00 1996 Path: biosci!agate!spool.mu.edu!newspump.sol.net!www.nntp.primenet.com!nntp.primenet.com!howland.erols.net!news3.cac.psu.edu!news.cse.psu.edu!news.ece.nwu.edu!newsfeed.acns.nwu.edu!news.cc.uic.edu!usenet From: Keld Sorensen Newsgroups: bionet.molbio.proteins Subject: Re: Wanted:peptide biotinylation or fluorocinylation method Date: Fri, 04 Oct 1996 09:54:21 +0000 Organization: University of Illinois, College of Medicine Lines: 5 Message-ID: <3254DECD.4319@uic.edu> References: NNTP-Posting-Host: sliprock-01.dialin.uic.edu Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 2.01 (Macintosh; I; PPC) To: "W.H. Colledge" [snip Q about peptide biotinylation and fluorescent labeling with fluorescein] Peptides can be labeled through amino groups (NH2) with NHS-Biotin and NHS-Fluorescein or if water solubility is an issue use: Sulfo-NHS-Biotin or Sulfo-NHS-Flurorescein. From owner-proteins@net.bio.net Fri Oct 04 23:00:00 1996 Path: biosci!agate!howland.erols.net!www.nntp.primenet.com!nntp.primenet.com!ddsw1!news.mcs.net!juniper.cis.uab.edu!maze.dpo.uab.edu!bmg146.cmc.uab.edu!user From: siyer@bmg.bhs.uab.edu (Arioch) Newsgroups: bionet.molbio.methds-reagnts,bionet.molbio.proteins Subject: de-salting prior to SDS-PAGE loading... Date: 5 Oct 1996 22:45:40 GMT Organization: Lord of Chaos Lines: 18 Message-ID: NNTP-Posting-Host: bmg146.cmc.uab.edu Xref: biosci bionet.molbio.methds-reagnts:49943 bionet.molbio.proteins:8922 hey all, anybody know of a good protocol to remove the salt from samples before they are loaded on to SDS-PAGEs?? i have found out that hi salt tends to distort the way the bands run on the gel (screwing up the electrical field and therefore affecting protein mobility). i found this one protocol that tells you to add 0.15% deoxycholic acid, incubate for 10 min followed by addition of TCA ---> spin for about 15 min at 3000g ---> decant all the TCA and resuspend pellet (containing precipitated protein with presumably no salt). this however has not been too successful (for me) and still causes distortion of my gel bands (much annoyance). anybody know of any modicfications to this protocol or other protocols that work?? thanks much... -- Arioch, Lord of Chaos siyer@bmg.bhs.uab.edu graduate student; dept of biochemistry and molecular genetics university of alabama at birmingham From owner-proteins@net.bio.net Fri Oct 04 23:00:00 1996 Path: biosci!agate!howland.erols.net!nntp.coast.net!netnews.worldnet.att.net!newsadm From: David Weber Newsgroups: bionet.molbio.proteins Subject: Re: Protein L Date: Sat, 05 Oct 1996 11:19:00 -0700 Organization: AT&T WorldNet Services Lines: 15 Message-ID: <3256A694.175F@worldnet.att.net> References: <52rj8r$9q4@rockall.cc.strath.ac.uk> Reply-To: david.weber@worldnet.att.net NNTP-Posting-Host: 8.bridgeton-056.mo.dial-access.att.net Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 3.0 (Win16; U) linda brooks wrote: > > Hello, > I wish to use protein L to create an immunoaffinity column > for purification of recombinant antibody variable light chains - > does anyone know of a commercial supplier of protein L? > > Thanks > Linda What's protein L? Where does it come from and what is its affinity? Thanks, David From owner-proteins@net.bio.net Fri Oct 04 23:00:00 1996 Path: biosci!agate!spool.mu.edu!uwm.edu!cs.utexas.edu!howland.erols.net!nntp.coast.net!oleane!jussieu.fr!univ-lyon1.fr!in2p3.fr!swidir.switch.ch!scsing.switch.ch!rzunews.unizh.ch!NewsWatcher!user From: zjons@vetbio.unizh.ch (Zophonias O. Jonsson) Newsgroups: bionet.molbio.proteins Subject: Re: protein-bound Zn measurement needed Date: Sat, 05 Oct 1996 16:26:09 +0200 Organization: Universitat Zurich-Irchel Lines: 28 Distribution: bionet Message-ID: References: <52u34s$gvl@mserv1.dl.ac.uk> NNTP-Posting-Host: 130.60.120.18 In article , gardner@bloch.med.utoronto.ca (Kevin Gardner) wrote: > As to the sensitivity, machines are fairly(?) routinely > available to detect uM concentrations of zinc from ca. 500uL - 1mL > of sample (e.g. nanomoles of Zn required). That's why an electrothermal atomizer would be more practical, one can use sample volumes of less than a uL up to about 10 uL. Using a laminar burner would be an awful waste of material, since a large part of it will flow directly down the drain. According to my old Skoog and West analytical chemistry textbook the sensitivity is 0.000002 ppm (ug/mL) for Zn about 20.000 times better than with flame (=> in the femtomole range). I don't think a machine like this will be hard to find either, they have been around for a long time and many analytical chemistry labs will have them. The main problem is likely to be background Zn or lack of accuracy in the measurements. Zophonias _____________________________________________________________________ Zophonias O. Jonsson Institut fur Veterinarbiochemie Tel: (41-1)-257-54-75 Universitat Zurich-Irchel Fax: (41-1)-362-05-01 Winterthurerstrasse 190 CH-8057 Zurich Switzerland _____________________________________________________________________ From owner-proteins@net.bio.net Fri Oct 04 23:00:00 1996 Path: biosci!agate!newsgate.duke.edu!solaris.cc.vt.edu!usenet From: mthorste@vt.edu (mthorste) Newsgroups: bionet.molbio.proteins Subject: ISO:Anti-oxidase antibodies Date: 5 Oct 1996 22:04:47 GMT Organization: Virginia Tech/Blacksburg Electronic Village Lines: 13 Message-ID: <536m1v$pl6@solaris.cc.vt.edu> Reply-To: Marc Thorsteinsson NNTP-Posting-Host: as2511-36.sl011.cns.vt.edu X-Newsreader: WinVN 0.92.1 Hi Protein Chemists in Netland, I am trying to characterize terminal (aa3, d, and o) oxidases in membrane preparations of nitrogen fixing cyanobacteria. Are there vendors who specialize in these? No luck with the common vendors. Source is not critical as terminal oxidase Ab have a tendency to cross-react among species. Any investigators out there working with these ?? Please help if you can and thanks in advance. Marc Thorsteinsson Dept. of Biochemistry VPI&SU mthorste@vt.edu From owner-proteins@net.bio.net Fri Oct 04 23:00:00 1996 Path: biosci!agate!howland.erols.net!vixen.cso.uiuc.edu!newsfeed.internetmci.com!news.msfc.nasa.gov!info.uah.edu!maze.dpo.uab.edu!bmg146.cmc.uab.edu!user From: siyer@bmg.bhs.uab.edu (Arioch) Newsgroups: bionet.molbio.methds-reagnts,bionet.molbio.proteins Subject: IMPACT kit from new england biolabs... Date: 5 Oct 1996 22:56:49 GMT Organization: Lord of Chaos Lines: 16 Message-ID: NNTP-Posting-Host: bmg146.cmc.uab.edu Xref: biosci bionet.molbio.methds-reagnts:49944 bionet.molbio.proteins:8923 hey all, just saw this in the september 6th issue of cell. this is a new kit from NEB that claims that you can get pure NATIVE protein (no fusions) in "one" step. basically they use proteins called inteins that are fused to YPOI. this undergoes directed cleavage when you add DTT (cleave a S-S bond somewhere right before the fused region) and you get native protein with no fusion domain to it. anybody try this and if so, what is the yield like?? how good is this system? seems like a cool idea, but as alwayz with any kit, i'm sceptical...any info on this would be apprciated...thanx much... -- Arioch, Lord of Chaos siyer@bmg.bhs.uab.edu graduate student; dept of biochemistry and molecular genetics university of alabama at birmingham From owner-proteins@net.bio.net Sat Oct 05 23:00:00 1996 Path: biosci!agate!howland.erols.net!news-peer.gsl.net!news.gsl.net!news-hk.gsl.net!news.gsl.net!newsgate.cuhk.edu.hk!newsfeeder.ust.hk!nntp.hk.super.net!uunet!news-in2.uu.net!netnews.nwnet.net!news-hub.interserv.net!news.sprynet.com!news From: Louis Geller Newsgroups: bionet.molbio.yeast,bionet.molbio.proteins Subject: How much Crypto to grow? Date: Sun, 06 Oct 1996 09:52:06 -0700 Organization: Cal State Univ. Long Beach Lines: 10 Message-ID: <3257E3B5.56B4@sprynet.com> NNTP-Posting-Host: dd63-020.compuserve.com Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 3.0 (Macintosh; I; 68K) Xref: biosci bionet.molbio.yeast:5919 bionet.molbio.proteins:8925 Please disregard the prior posting "Required Culture volume". My colleague needs to know what volume of Cryptococcus neoformans to grow, in order to partially isolate an enzyme, and run enzyme activities. The protein is not particularly abundant, and the assay is fairly sensitive. Thank you very much for your help, Louis Geller... From owner-proteins@net.bio.net Sat Oct 05 23:00:00 1996 Path: biosci!agate!newsxfer2.itd.umich.edu!www.nntp.primenet.com!nntp.primenet.com!swrinde!news-peer.gsl.net!news.gsl.net!news-stkh.gsl.net!news.gsl.net!nntp-oslo.UNINETT.no!nntp-trd.UNINETT.no!nntp.uio.no!nntp.zit.th-darmstadt.de!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.proteins Subject: Re: Protein Viewer Date: 6 Oct 1996 22:29:46 GMT Organization: University of Wuerzburg, Germany Lines: 12 Message-ID: <539bsq$a9a@winx03.informatik.uni-wuerzburg.de> References: <538hmt$nd@newsbf02.news.aol.com> NNTP-Posting-Host: wpxx02.toxi.uni-wuerzburg.de X-Newsreader: TIN [UNIX 1.3 950824BETA PL0] BLDUNCCH (blduncch@aol.com) wrote: > Can any one tell me how to obtain a 3D protein viewer to view Brookhaven > PD files? I want to be able to color the aa residues that I select. Look for Rasmol, available for Macs, Windows and Unix (X). --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 Sat Oct 05 23:00:00 1996 Path: biosci!AESOP.RUTGERS.EDU!pwang From: pwang@AESOP.RUTGERS.EDU (pinger wang) Newsgroups: bionet.molbio.proteins Subject: My protein is an RNase or contaminated with RNase? Date: 6 Oct 1996 10:20:55 -0700 Organization: Rutgers University Lines: 15 Sender: daemon@net.bio.net Distribution: world Message-ID: <3257EB9F.796A@aesop.rutgers.edu> Reply-To: pwang@AESOP.RUTGERS.EDU NNTP-Posting-Host: net.bio.net Dear Netters, I am working on a ribosome-inactivating protein which has been already demonstrated to has a specific N-glycosidase activity. Recently, I found that this protein can also degrade RNA non- specifcally. RNase activity gel analysis of this protein sample with yeast RNA as substrate showed that this protein has week RNase activity and no other RNase activity was observed in the gel. My question is that the evidence of RNase activity gel is good enough to exculde other RNase contamination? Are there any other good way to discriminate them? Thanks for help Pinger From owner-proteins@net.bio.net Sat Oct 05 23:00:00 1996 Path: biosci!agate!howland.erols.net!news-peer.gsl.net!news.gsl.net!portc01.blue.aol.com!newstf01.news.aol.com!newsbf02.news.aol.com!not-for-mail From: blduncch@aol.com (BLDUNCCH) Newsgroups: bionet.molbio.proteins Subject: Protein Viewer Date: 6 Oct 1996 11:02:53 -0400 Organization: America Online, Inc. (1-800-827-6364) Lines: 2 Sender: root@newsbf02.news.aol.com Message-ID: <538hmt$nd@newsbf02.news.aol.com> Reply-To: blduncch@aol.com (BLDUNCCH) NNTP-Posting-Host: newsbf02.mail.aol.com Can any one tell me how to obtain a 3D protein viewer to view Brookhaven PD files? I want to be able to color the aa residues that I select. From owner-proteins@net.bio.net Sun Oct 06 23:00:00 1996 Path: biosci!agate!spool.mu.edu!newspump.sol.net!www.nntp.primenet.com!nntp.primenet.com!nntp.coast.net!oleane!jussieu.fr!citi2.fr!federici.cochin.inserm.fr!user From: federici@icgm.cochin.inserm.fr (Federici) Newsgroups: bionet.molbio.proteins Subject: Re: Elution of protein from dry SDS_PAGE Date: 7 Oct 1996 08:38:40 GMT Organization: ICGM CNRS UPR415 Lines: 30 Message-ID: References: <4104vsav-3009962009010001@mac118.envir.ulaval.ca> NNTP-Posting-Host: federici.cochin.inserm.fr X-Newsreader: Yet Another NewsWatcher F2.0 In article (Dans l'article) <4104vsav-3009962009010001@mac118.envir.ulaval.ca>, 4104vsav@vm1.ulaval.ca (Savita Visal) wrote (écrivait) : > Dear Friends, > I want to sequence protein of interest. I have my protein of interest as a > single band on SDS_PAGE. I've dried this gel. I would like sequence this > protein..the problem is elution from the SDS-PAGE. I have read so many > protocols and at this stage I'm totally confused ..which protocol do I > use? MW of my protein is 55kDa. Could anyone share their experiences and > guide me to which is the best protocol I could use? > Thanks > > Savita > 4104vsav@vm1.ulaval.ca First of all, you must humidify the gel. To do this step it will be better to use the SDS-PAGE buffer. Then, to elute the protein, transfer to hydroxyapatit gel is an efficient procedure; hydroxyapatit is a calcium phosphate crystal that is used as chromatography gel (Sigma) . To realize this step, you must use SDS-PAGE procedure, but on a tube. Fill the third of it with SDS-PAGE gel, then add 1 ml of hydroxyapatit that was previously washed by SDS-PAGE buffer, and finally cut the gel containing your protein into small pieces (2 millimetres sided) and load them onto hydroxyapatit. Then, run SDS_PAGE (at the same current intensity that your initial SDS-PAGE). At the end of elution, recuperate the hydroxyapatit (take care, hydroxyapatit is fragile), load it on a small column,taking care not to dry it; and elute the protein using 2ml of a 0,5M sodium phosphate pH:6.8, 0,1 %SDS, 1mM DTT buffer. If you want to eliminate the 0.5M sodium phosphate, you can filtrate your fraction upon suitable menbrane but keep the presence of SDS and DTT. From owner-proteins@net.bio.net Sun Oct 06 23:00:00 1996 Path: biosci!agate!howland.erols.net!EU.net!usenet2.news.uk.psi.net!uknet!usenet1.news.uk.psi.net!uknet!dispatch.news.demon.net!demon!genesys.demon.co.uk!duncan From: "Dr. Duncan Clark" Newsgroups: bionet.molbio.methds-reagnts,bionet.molbio.proteins Subject: Re: IMPACT kit from new england biolabs... Date: Mon, 7 Oct 1996 11:26:14 +0100 Organization: GeneSys Ltd Lines: 29 Distribution: world Message-ID: References: NNTP-Posting-Host: genesys.demon.co.uk X-NNTP-Posting-Host: genesys.demon.co.uk MIME-Version: 1.0 X-Newsreader: Turnpike Version 3.01 beta 1 Xref: biosci bionet.molbio.methds-reagnts:49968 bionet.molbio.proteins:8930 In article , Arioch writes >hey all, > just saw this in the september 6th issue of cell. this is a new >kit from NEB that claims that you can get pure NATIVE protein (no fusions) >in "one" step. basically they use proteins called inteins that are fused >to YPOI. this undergoes directed cleavage when you add DTT (cleave a S-S >bond somewhere right before the fused region) and you get native protein >with no fusion domain to it. anybody try this and if so, what is the >yield like?? how good is this system? seems like a cool idea, but as >alwayz with any kit, i'm sceptical...any info on this would be >apprciated...thanx much... They are using a modified intein and from data I saw presented at a conference a few weeks back it is a very promising system. My only queries are: 1. Is if the affinity column is reusable? 2. How expensive is the affinity column 'cos it's OK purifying proteins on a shake flask scale but I need to work on a much larger scale. Duncan ----------------------------------------------------------------------------- My mind's made up. Don't confuse me with the facts! Duncan Clark DNAmp Ltd. http://www.dnamp.com From owner-proteins@net.bio.net Sun Oct 06 23:00:00 1996 Path: biosci!daresbury!lyra.csx.cam.ac.uk!news.ox.ac.uk!news From: "Simon M. Brocklehurst" Newsgroups: bionet.molbio.proteins Subject: NAOMI - Important messages Date: Mon, 07 Oct 1996 09:41:44 +0100 Organization: University of Oxford Lines: 65 Message-ID: <3258C248.446B9B3D@bioch.ox.ac.uk> NNTP-Posting-Host: nmrv.ocms.ox.ac.uk Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 3.0 (X11; I; SunOS 4.1.3_U1 sun4m) NAOMI - Important messages to all users - please read. _____________________________________________________________________________ **** The NAOMI anonymous FTP server is now back up again after some considerable downtime. Sorry for all the inconvenience this caused. ----------------------------------------------------------------------------- **** I've taken this opportunity to make available the Version 3.0 User Guide which is complete reworking of the old User Guide. It is much improved (fast loading, better organised etc). It is available only on the Web site (select the User Guide - Europe link) at present. NAOMI Web site URL given below. Version 3.0 of the program is not yet available (release soon), but the new User Guide is applicable to the current version. ----------------------------------------------------------------------------- **** Due to bandwidth problems, especially across the Atlantic and to the Pacific Rim, I'm looking for places that would be prepared to act as mirror sites for the NAOMI distribution (you need an anonymous ftp server to do this). If you are interested in doing this, please e-mail me at: smb@bioch.ox.ac.uk ------------------------------------------------------------------------------ ****** What is NAOMI ? ******************* NAOMI is a computer program system for studying 3-D structures of proteins. NAOMI is available free of charge for academic users. NAOMI Version 2.4c is available as of now from the NAOMI Web site at: http://www.ocms.ox.ac.uk/~smb/Software/N_details/naomi.html or via anonymous ftp ftp://nmrz.ocms.ox.ac.uk/pub/smb/naomi i.e. at nmrz.ocms.ox.ac.uk in directory pub/smb/naomi/ _____________________________________________________________________________ | | ,_ o Simon M. Brocklehurst, | / //\, Oxford Centre for Molecular Sciences, Department of Biochemistry, | \>> | University of Oxford, Oxford, UK. | \\, E-mail: smb@bioch.ox.ac.uk | WWW: http://www.ocms.ox.ac.uk/~smb/ |____________________________________________________________________________ From owner-proteins@net.bio.net Sun Oct 06 23:00:00 1996 Path: biosci!ihnp4.ucsd.edu!agate!spool.mu.edu!newspump.sol.net!news.inc.net!news.sprintlink.net!news-chi-8.sprintlink.net!moonbeam.aecom.yu.edu!usenet From: Alan Schoenfeld Newsgroups: bionet.molbio.proteins Subject: (no subject) Date: 7 Oct 1996 18:13:51 GMT Organization: AECOM Lines: 8 Message-ID: <53bh8v$9vg@moonbeam.aecom.yu.edu> NNTP-Posting-Host: darkstar.aecom.yu.edu Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 1.1N (Macintosh; I; 68K) X-URL: news:bionet.molbio.proteins I want to check if the protein that I'm studying has any consensus phosphorylation sites. Does anybody know a way to to do this (either over the web or by email)? Alan From owner-proteins@net.bio.net Sun Oct 06 23:00:00 1996 Path: biosci!WESLEYAN.EDU!tguina From: tguina@WESLEYAN.EDU (Tina Guina) Newsgroups: bionet.molbio.proteins Subject: HELP! protein purification with gelatin/BSA Date: 7 Oct 1996 08:29:52 -0700 Organization: Wesleyan university Lines: 22 Sender: daemon@net.bio.net Distribution: world Message-ID: <325923C5.561D@wesleyan.edu> Reply-To: tguina@wesleyan.edu NNTP-Posting-Host: net.bio.net Dear colleagues! I am trying to purify a small cationic and hydrophobic protein from bacterial culture supernatants. The culture medium contains insect a mammalian cell culture components including 50g/L of BSA and 0.4-0.5% GELATIN. The protein of interest is very likely to be produced in small amounts and associating with one or more medium components of higher MWt. When expressed in E. coli, this protein is soluble, but it partitions into the detergent phase after the triton X114 extraction. I need to show the presence of this protein in culture supernantants. I have a good affinity purified antiserum against the protein. My questions are: 1) what would be the best method to concentrate the protein? (is ammonium sulfate good enough and would it precipitate the gelatin as well) 2) is there a simple, for the protein harmless way to get rid of gelatin without heating the solution? Thanks a lot in advance! Tina Guina From owner-proteins@net.bio.net Sun Oct 06 23:00:00 1996 Path: biosci!bcm.tmc.edu!news.msfc.nasa.gov!news.sgi.com!www.nntp.primenet.com!nntp.primenet.com!nntp.uio.no!nntp.zit.th-darmstadt.de!fu-berlin.de!zrz.TU-Berlin.DE!news.tu-chemnitz.de!News.HTWM.De!news.uni-weimar.de!news.uni-jena.de!news.uni-leipzig.de!mlucom4.urz.uni-halle.de!dnennsti From: dnennsti@ipb.uni-halle.de (Dirk Nennstiel) Newsgroups: bionet.molbio.proteins Subject: Re: Wanted:peptide biotinylation or fluorocinylation method Date: Mon, 7 Oct 1996 20:15:12 +0200 Organization: YES Lines: 20 Message-ID: <1996100720151221429@dial-4.urz.uni-halle.de> References: NNTP-Posting-Host: dial-4.urz.uni-halle.de X-Newsreader: MacSOUP 2.2b3 W.H. Colledge wrote: > Does anyone have a simple method for labelling peptides with biotin or > fluorocine? How can the labelled peptide be purified from the reaction > products? For both topics I use the products from Pierce (the NHS coupling to primary amino groups). I purify the products by HPLC. I hope this helps. Dirk -- Dirk Nennstiel Inst. of plant biochemistry Weinberg 3 06110 Halle/Saale Germany From owner-proteins@net.bio.net Sun Oct 06 23:00:00 1996 Path: biosci!SFSU.EDU!c07craig From: c07craig@SFSU.EDU (Craig) Newsgroups: bionet.molbio.proteins Subject: I would like to subscribe Date: 7 Oct 1996 18:15:50 -0700 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 5 Sender: daemon@net.bio.net Distribution: world Message-ID: NNTP-Posting-Host: net.bio.net Hello, I would like to subscribe to the proteins newsgroup. Thanks Craig From owner-proteins@net.bio.net Sun Oct 06 23:00:00 1996 Path: biosci!botany.uq.edu.au!J.Marcus From: J.Marcus@botany.uq.edu.au ("Marcus, Dr J.") Newsgroups: bionet.molbio.proteins Subject: Re: HELP! protein purification with gelatin/BSA Date: 7 Oct 1996 15:44:12 -0700 Organization: Dept of Botany, Univ of Qld Lines: 58 Sender: daemon@net.bio.net Distribution: world Message-ID: <9FB752C06D2@botany.uq.edu.au> Reply-To: Marcus@tpp.uq.edu.au NNTP-Posting-Host: net.bio.net Hi Tina, > I am trying to purify a small cationic and hydrophobic protein from > bacterial culture supernatants. The culture medium contains insect a > mammalian cell culture components including 50g/L of BSA and 0.4-0.5% > GELATIN. Can you grow your bacteria in another medium? > The protein of interest is very likely to be produced in small amounts > and associating with one or more medium components of higher MWt. > When expressed in E. coli, this protein is soluble, but it partitions > into the detergent phase after the triton X114 extraction. Cation exchange might be something to think about. An anionic detergent would be compatible with cation-exchange step which would concentrate your protein. > > I need to show the presence of this protein in culture supernantants. I > have a good affinity purified antiserum against the protein. > My questions are: > 1) what would be the best method to concentrate the protein? > (is ammonium sulfate good enough and would it precipitate the gelatin > as well) Have a go. By using different concentrations of amm.sulfate you might get the gelatin to precipitate out leaving your protein behind. Then with higher concentrations of amm.sulfate (say 90%) you might precipitate your protein. However, even if your protein precipitates with the gelatin you may still be able to load it on a gel and do a western blot on it. > 2) is there a simple, for the protein harmless way to get rid of gelatin > without heating the solution? Maybe size exclusion. Hope that is helpful. Regards, John _________________________________________________________ John Marcus Marcus@tpp.uq.edu.au (Dr J.Marcus) Cooperative Research Centre for Tropical Plant Pathology 5th Level John Hines Building University of Queensland St. Lucia, QLD 4072 AUSTRALIA Fax: 61-7-3365-4771 Phone: 61-7-3365-4764 From owner-proteins@net.bio.net Sun Oct 06 23:00:00 1996 Path: biosci!agate!spool.mu.edu!howland.erols.net!vixen.cso.uiuc.edu!saluki-news.wham.siu.edu!mac50.biochem.siu.edu!user From: sedmondson@som.siu.edu (Stephen Edmondson) Newsgroups: bionet.molbio.proteins Subject: Re: My protein is an RNase or contaminated with RNase? Date: Mon, 07 Oct 1996 14:07:45 -0600 Organization: Southern Illinois University Lines: 19 Distribution: world Message-ID: References: <3257EB9F.796A@aesop.rutgers.edu> NNTP-Posting-Host: mac50.biochem.siu.edu Try HPLC to separate RNase activity from your protein. If your protein lacks Cys, try adding mercaptoethanol to reduce disulfides of RNase. If your protein is easily denatured, see if loss of RNase activity parallels unfolding. RNase can have a very high specific activity so a very small amount of contamination is quite possible. In article <3257EB9F.796A@aesop.rutgers.edu>, pwang@AESOP.RUTGERS.EDU wrote: > I am working on a ribosome-inactivating protein which has been > already demonstrated to has a specific N-glycosidase activity. > Recently, I found that this protein can also degrade RNA non- > specifcally. RNase activity gel analysis of this protein sample with > yeast RNA as substrate showed that this protein has week RNase > activity and no other RNase activity was observed in the gel. My > question is that the evidence of RNase activity gel is good enough > to exculde other RNase contamination? Are there any other good way > to discriminate them? Steve Edmondson sedmondson@som.siu.edu From owner-proteins@net.bio.net Sun Oct 06 23:00:00 1996 Path: biosci!daresbury!not-for-mail From: vachon@bio.grenet.fr (Gilles VACHON) Newsgroups: bionet.molbio.proteins Subject: Gel retardation: specific binding but no competition Date: 7 Oct 1996 14:03:42 +0100 Lines: 25 Sender: lpddist@mserv1.dl.ac.uk Distribution: bionet Message-ID: <53av3e$s8d@mserv1.dl.ac.uk> Original-To: proteins@dl.ac.uk Posted-Date: Mon, 7 Oct 96 15:03:34 +0100 Dear all, we are working on a transcription factor that is known to bind to a specific sequence called A and not to the mutated version calld B. When we labele A and B and perform a bandshifht, A is retarded but not B. BUT when we label A only and perform conpetition assays with cold A or cold B we se no competition with either cold A or cold B. This competition has been done by other groups but don't work in our hands. Any suggestions why this isn't working? Please forward your message to the list AND to me since I am not on this list. Thanks Dr. Gilles Vachon Laboratoire de Genetique Moleculaire des Plantes CERMO, 3eme etage Universite J. Fourier, BP 53X 38041 GRENOBLE CEDEX FRANCE Tel: (33) 76 63 56 58 fax: (33) 76 51 43 36 e-mail: vachon@bio.grenet.fr From owner-proteins@net.bio.net Sun Oct 06 23:00:00 1996 Path: biosci!agate!howland.erols.net!www.nntp.primenet.com!nntp.primenet.com!news-peer.gsl.net!news.gsl.net!news-stkh.gsl.net!news.gsl.net!nntp-oslo.UNINETT.no!nntp-trd.UNINETT.no!daresbury!not-for-mail From: ditlev@kemi.aau.dk (Ditlev Brodersen) Newsgroups: bionet.molbio.proteins Subject: Re: de-salting prior to SDS-PAGE loading... Date: 7 Oct 1996 13:28:18 +0100 Lines: 41 Sender: lpddist@mserv1.dl.ac.uk Distribution: bionet Message-ID: <53at12$ptt@mserv1.dl.ac.uk> X-Sender: ditlev@beauty.kemi.aau.dk Original-To: proteins@dl.ac.uk >hey all, > anybody know of a good protocol to remove the salt from samples >before they are loaded on to SDS-PAGEs?? i have found out that hi salt >tends to distort the way the bands run on the gel (screwing up the >electrical field and therefore affecting protein mobility). i found this >one protocol that tells you to add 0.15% deoxycholic acid, incubate for 10 >min followed by addition of TCA ---> spin for about 15 min at 3000g ---> >decant all the TCA and resuspend pellet (containing precipitated protein >with presumably no salt). this however has not been too successful (for >me) and still causes distortion of my gel bands (much annoyance). anybody >know of any modicfications to this protocol or other protocols that >work?? thanks much... > >-- >Arioch, Lord of Chaos >siyer@bmg.bhs.uab.edu >graduate student; dept of biochemistry and molecular genetics >university of alabama at birmingham Dear Arioch, Maybe the high ionic strength of TCA affects your SDS-PAGE run...Have you tried precipitation by acetone instead? Add 33% acetone (1 acetone: 2 sample) and precipitate app. 10' at 0'C Spin 5' at 13 krpm. Resuspend pellet in whatever loading-buffer you use. -- Ditlev ------------------------------------------------------------------- Ditlev Brodersen Phone: +45 8942 3871 Dept. of Struct. and Mol. Biology Fax: +45 8619 6199 c/o Dept. of Chemistry Email: ditlev@kemi.aau.dk Langelandsgade 140 ditlev@beauty.kemi.aau.dk DK-8000 Aarhus C Denmark WWW:http://kaktus.kemi.aau.dk/~ditlev From owner-proteins@net.bio.net Sun Oct 06 23:00:00 1996 Path: biosci!daresbury!not-for-mail From: l.kempster@ic.ac.uk (L.Kempster) Newsgroups: bionet.molbio.proteins Subject: Ub-conjugation Date: 7 Oct 1996 14:53:35 +0100 Lines: 14 Sender: lpddist@mserv1.dl.ac.uk Distribution: bionet Message-ID: <53b20v$2en@mserv1.dl.ac.uk> X-Sender: lkhl@tolstoy.nhli.ic.ac.uk Original-To: proteins@dl.ac.uk Hi. Can anyone give me information on how mammalian ubiquitin molecules conjugate with misfolded proteins retained in the ER membrane? Is there a mammalian ubc6p homologue? Thanks Lee Kempster Imperial College. From owner-proteins@net.bio.net Mon Oct 07 23:00:00 1996 Path: biosci!agate!spool.mu.edu!howland.erols.net!news.sprintlink.net!news-peer.sprintlink.net!uunet!in1.uu.net!fu-berlin.de!news.belwue.de!news.uni-freiburg.de!sun2.ruf.uni-freiburg.de!eschiltz From: eschiltz@sun2.ruf.uni-freiburg.de (Emile Schiltz) Newsgroups: bionet.molbio.proteins Subject: Re: enzyme catalytic cycle movies? Date: 8 Oct 1996 09:37:25 GMT Organization: Rechenzentrum der Universitaet Freiburg, Germany Lines: 25 Message-ID: <53d7cl$340@n.ruf.uni-freiburg.de> References: <52f1hs$mpu@r02n01.cac.psu.edu> NNTP-Posting-Host: sun2.ruf.uni-freiburg.de X-Newsreader: TIN [version 1.2 PL2] Ira Ropson (iropson@bcmic.hmc.psu.edu) wrote: : Hi folks, : Does anyone know of any web sites where a "movie" of the catalytic : action of an enzyme can be found? I have in mind a relativly simple : active site picture showing just those residues involved in catalysis : and the binding of substrate, and the substrate structure for the first : frame, the change in conformation and structure going to the transition : state, and then changing conformation and structure to the product. : Such movies would be real useful for the course I'm teaching. : Thanks, : Ira Ropson : iropson@bcmic.hmc.psu.edu : If I had time for a fancy Sig file I problably wouldn't have a job. Hi Ira, try http://bio5.chemie.uni-freiburg.de, then the ak-movie it shows an enzyme in motion, based on x-ray structures. Regards Mil Schiltz -- Emile Schiltz or Institute of Organic Chemistry and Biochemistry Albertstrasse 21, D-79104 Freiburg, Germany phone: +49 761 203 6090 From owner-proteins@net.bio.net Mon Oct 07 23:00:00 1996 Path: biosci!agate!howland.erols.net!news.sgi.com!enews.sgi.com!decwrl!tribune.usask.ca!rover.ucs.ualberta.ca!news From: Simon Rabinovitch Newsgroups: bionet.molbio.proteins Subject: keep your shit off my newsgroup Date: Tue, 08 Oct 1996 02:09:12 -0700 Organization: University of Alberta Lines: 1 Message-ID: <325A1A38.36DF@gpu.srv.ualberta.ca> References: NNTP-Posting-Host: async8-6.remote.ualberta.ca Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 3.0 (Win95; I) To: KIM NORDSTR™M you fuck From owner-proteins@net.bio.net Mon Oct 07 23:00:00 1996 Path: biosci!agate!howland.erols.net!news.sprintlink.net!news-peer.sprintlink.net!uunet!news-in2.uu.net!boulder!rikki.cc.colorado.edu!MDOUGHERTY From: mdougherty@cc.colorado.edu Newsgroups: bionet.molbio.proteins Subject: Looking for Bob Pedersen Date: 8 Oct 1996 14:53:18 GMT Organization: The Colorado College, Colorado Springs, CO Lines: 8 Message-ID: <53dpsu$1ck@lace.colorado.edu> Reply-To: mdougherty@cc.colorado.edu NNTP-Posting-Host: rikki.cc.colorado.edu I am looking for a molecular biologist named Bob Pedersen; we used to work together at the Univ. of Massachusetts. Three years ago Bob was working for a biotech firm in the Boston area, and I would guess that he still is in the Boston area. If anyone has any information on where Bob currently is working, I certainly would appreciate knowing. Thanks. From owner-proteins@net.bio.net Mon Oct 07 23:00:00 1996 Path: biosci!daresbury!not-for-mail From: l.kempster@ic.ac.uk (L.Kempster) Newsgroups: bionet.molbio.proteins Subject: UDP-glucose:glycoprotein glucosyltransferase Date: 8 Oct 1996 16:34:19 +0100 Lines: 11 Sender: lpddist@mserv1.dl.ac.uk Distribution: bionet Message-ID: <53ds9r$ctg@mserv1.dl.ac.uk> X-Sender: lkhl@tolstoy.nhli.ic.ac.uk Original-To: proteins@dl.ac.uk Hi Does anyone know whether a specific inhibitor of UDP-glucose:glycoprotein glucosyltransferase exists? Thanks Lee Kempster Imperial College. From owner-proteins@net.bio.net Mon Oct 07 23:00:00 1996 Path: biosci!daresbury!nntp-trd.UNINETT.no!sn.no!nntp.uio.no!www.nntp.primenet.com!nntp.primenet.com!feed1.news.erols.com!howland.erols.net!news.mathworks.com!newsfeed.internetmci.com!news.msfc.nasa.gov!elroy.jpl.nasa.gov!lll-winken.llnl.gov!fnnews.fnal.gov!nntp-server.caltech.edu!seqaxp.bio.caltech.edu!MATHOG From: mathog@seqaxp.bio.caltech.edu Newsgroups: bionet.molbio.proteins,comp.sys.sgi.hardware,bionet.software Subject: Evans & Sutherland dials on SGI??? Date: 8 Oct 1996 22:53:31 GMT Organization: Biology Division, Caltech, Pasadena CA 91125 Lines: 17 Message-ID: <53em1b$a70@gap.cco.caltech.edu> Reply-To: mathog@seqaxp.bio.caltech.edu NNTP-Posting-Host: seqaxp.bio.caltech.edu Xref: biosci bionet.molbio.proteins:8948 comp.sys.sgi.hardware:22444 bionet.software:16758 We have several defunct Evans & Sutherland graphics workstations about (PS300's, I think). Each has a set of 8 dials on it that look quite a bit like an older version of the dial boxes on our SGIs. We have one SGI that is currently sans dials and it would be great if we could scavenge one of these old dial sets for it, rather than having to shell out $2000 for a new set (with the buttons, which we basically don't use). So.... Anybody know if the old E&S dials can be used on an SGI? Obviously the connector is different, but are the electronics compatible? Thanks, David Mathog mathog@seqaxp.bio.caltech.edu Manager, sequence analysis facility, biology division, Caltech From owner-proteins@net.bio.net Mon Oct 07 23:00:00 1996 Path: biosci!bcm.tmc.edu!news.msfc.nasa.gov!news.sgi.com!howland.erols.net!cs.utexas.edu!news.tamu.edu!news From: Dawn Capp Newsgroups: bionet.molbio.proteins Subject: Free Web service to research authors Date: Tue, 08 Oct 1996 09:27:44 -0500 Organization: Texas A&M University Lines: 6 Message-ID: <325A64E0.297B@tam2000.tamu.edu> NNTP-Posting-Host: ppp1b-12.rns.tamu.edu Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 3.0 (Win95; I) Capp Scientific offers a free WEB service to research authors who have difficulty getting their work published in scientific journals. Go To http://members.aol.com/CappSci for more information. From owner-proteins@net.bio.net Mon Oct 07 23:00:00 1996 Path: biosci!bcm.tmc.edu!news.msfc.nasa.gov!news.sgi.com!nntp-hub2.barrnet.net!cpk-news-hub1.bbnplanet.com!www.nntp.primenet.com!nntp.primenet.com!arclight.uoregon.edu!news.uoregon.edu!flynn-lab.uoregon.edu!user From: tedm@darkwing.uoregon.edu (Ted Michelini) Newsgroups: bionet.molbio.methds-reagnts,bionet.molbio.proteins Subject: Re: IMPACT kit from new england biolabs... Date: 9 Oct 1996 00:16:49 GMT Organization: University of Oregon Lines: 45 Distribution: world Message-ID: References: NNTP-Posting-Host: flynn-lab.uoregon.edu Xref: biosci bionet.molbio.methds-reagnts:50065 bionet.molbio.proteins:8949 In article , "Dr. Duncan Clark" wrote: > In article , Arioch > writes > >hey all, > > just saw this in the september 6th issue of cell. this is a new > >kit from NEB that claims that you can get pure NATIVE protein (no fusions) > >in "one" step. basically they use proteins called inteins that are fused > >to YPOI. this undergoes directed cleavage when you add DTT (cleave a S-S > >bond somewhere right before the fused region) and you get native protein > >with no fusion domain to it. anybody try this and if so, what is the > >yield like?? how good is this system? seems like a cool idea, but as > >alwayz with any kit, i'm sceptical...any info on this would be > >apprciated...thanx much... > > They are using a modified intein and from data I saw presented at a > conference a few weeks back it is a very promising system. My only > queries are: > 1. Is if the affinity column is reusable? > 2. How expensive is the affinity column 'cos it's OK purifying > proteins on a shake flask scale but I need to work on a much larger > scale. > > Duncan Duncan, I too think this is neato stuff, the acrobat file from NEB's web site contains the full protocol and they say the chitin beads are reusable 5 times. Since we use Ni columns roughly 10X what we're supposed too, maybe the same math holds (?). The chitin should be cheap, although I haven't found a alt. supplier yet, perhaps one could use roughly ground cockroaches or crabs...The manual does state the system doesn't work well with eukaryotic proteins and that certain flanking AA's at the amino terminus give premature cleavage. I'm going to give it a try and will post my results soon. The promise of one step purification/ C terminal tag cleavage with one or no added amino acids is exciting, I'm going to give it a whirl. regards, Ted Michelini Institute of Molecular Biology University of Oregon tedm@darkwing.uoregon.edu From owner-proteins@net.bio.net Mon Oct 07 23:00:00 1996 Path: biosci!daresbury!nntp-trd.UNINETT.no!sn.no!nntp.uio.no!www.nntp.primenet.com!nntp.primenet.com!mr.net!news.mr.net!cruise.biol.stthomas.edu!jlcruise From: JLCruise Newsgroups: bionet.molbio.proteins Subject: membrane preps from cultured mammalian cells Date: 8 Oct 1996 22:25:08 GMT Organization: University of St. Thomas Lines: 30 Distribution: world Message-ID: <53ekc4$81s@news.mr.net> NNTP-Posting-Host: cruise.biol.stthomas.edu Mime-Version: 1.0 Content-Type: text/plain; charset=ISO-8859-1 Content-Transfer-Encoding: 8bit X-Newsreader: Nuntius 2.0.4_PPC X-XXMessage-ID: X-XXDate: Tue, 8 Oct 1996 22:29:05 GMT I am looking for suggested methods of making rapid membrane preps from various mammalian cells in culture. The uses I have in mind for these preps are 1) receptor-binding studies and 2) western blotting of membrane-associated proteins. Most of the lines I am using are adherent. While I have experience with standard protocols (modified Neville method, Prpic membrane prep, crude microsomal preps) that work well for me when I start with tissues, my limited experience with cell cultures is of low yield and probably cleaner preps than I may need. Searching the literature on adrenergic receptor studies (which is what I'm doing the binding assays on) has produced the full spectrum from hypotonic lysates without ANY fractionation to very pure plasma membrane fractions produced by long protocols. Can anyone recommend a rapid, fairly simple membrane prep with decent yield from cultures that they know to produce membranes suitable for adrenergic (or other) binding studies? For the westerns, my need is merely for a prep with good yield, that is clearly membrane-enriched, relative to total cell lysate. Any tips will be appreciated. Please email me directly or post here. Thanks. jlcruise@stthomas.edu Jennifer L. Cruise Dept. Biology Univ. St. Thomas St. Paul, MN From owner-proteins@net.bio.net Mon Oct 07 23:00:00 1996 Path: biosci!agate!spool.mu.edu!newspump.sol.net!www.nntp.primenet.com!nntp.primenet.com!howland.erols.net!EU.net!usenet2.news.uk.psi.net!uknet!usenet1.news.uk.psi.net!uknet!uknet!warwick!news.nott.ac.uk!NewsWatcher!user From: kevin.bailey@nottingham.ac.uk () Newsgroups: bionet.molbio.proteins Subject: Re: signal peptides Followup-To: bionet.molbio.proteins Date: Tue, 08 Oct 1996 16:38:29 +0100 Organization: Cripps Computing Centre, The University of Nottingham Lines: 32 Message-ID: References: <01bbaf81$cb906320$ced6ebc2@guidok.telebyte.nl> NNTP-Posting-Host: wpbjwk1.biochem.nottingham.ac.uk In article <01bbaf81$cb906320$ced6ebc2@guidok.telebyte.nl>, "Calimero" wrote: > Hi, > > I used a computer program to find signal peptides in a putative protein and > it gave the following results: > > Maximum score 3.5 at residue 46 > > Sequence: ACLFRAVADQVYG-DQDMHEVVRKHCMDYLMKNADYFSNYVTEDFTTYINRKRKNNCHGNHIEM > | (signal) | (mature peptide) > 33 46 > > Score 3.5 at residue 166 > > Sequence: VGLGLPSFKPGFA-EQSLMKNAIKTSEESWIEQQMLEDKKRATDWEATNESIEEQVARESYLQW > | (signal) | (mature peptide) > 153 166 > > Does anybody know what these signals mean. > > > Guido > > guidok@telebyte.nl Not sure if this is the answer you are after, but the signal part is that which is transcribed at some point but which is removed before the mature peptide is released. Hence the amino acid noted as the start of the mature peptide/protein should be the one which would be residue 1 if the protein were recovered naturally in it's mature "finished" form. From owner-proteins@net.bio.net Mon Oct 07 23:00:00 1996 Path: biosci!bcm.tmc.edu!cs.utexas.edu!newshost.convex.com!newsgate.duke.edu!news.duq.edu!usenet From: "Frank R. Gorga" Newsgroups: bionet.molbio.proteins Subject: Re: membrane preps from cultured mammalian cells Date: Tue, 08 Oct 1996 20:21:24 -0700 Organization: Duquesne University Lines: 24 Message-ID: <325B1A34.4DDF@next.duq.edu> References: <53ekc4$81s@news.mr.net> 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 3.0Gold (Win16; I) JLCruise wrote: > > I am looking for suggested methods of making rapid membrane preps from > various mammalian cells in culture. For fibroblasts, try the method of Thom, et al. (1977) Biochem. J. 168:187. This method has worked well for me using a variety of mose and rat fibroblast lines (3T3, F111, and others). I do nnot know if it will work with other types of cells, but suspect that with small modifications it could be used on many adherant cell lines. Hope this helps, --- Frank Gorga *********************************** Frank R. Gorga, Ph.D. Dept. of Chemistry & Biochemistry Duquesne University Pittsburgh, PA 15282 412-396-5858 / 412-396-5683 (fax) gorga@next.duq.edu From owner-proteins@net.bio.net Tue Oct 08 23:00:00 1996 Path: biosci!faseb.org!cpk-news-feed1.bbnplanet.com!cpk-news-hub1.bbnplanet.com!www.nntp.primenet.com!nntp.primenet.com!howland.erols.net!EU.net!usenet2.news.uk.psi.net!uknet!usenet1.news.uk.psi.net!uknet!uknet!yama.mcc.ac.uk!bignews.shef.ac.uk!usenet From: Antonio Pregueiro Newsgroups: bionet.molbio.proteins Subject: regulation of E.coli's ndh gene Date: Wed, 09 Oct 1996 19:12:51 +0100 Organization: Molecular Biology & Biotechnology, University of Sheffield , UK Lines: 5 Message-ID: <325BEB23.772E@shef.ac.uk> NNTP-Posting-Host: pc113051.shef.ac.uk Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 3.0 (Win16; I) I would just like to ask for information about the regulatory mechanisms of E.coli's NADH dehydrogenase gene. If anyone is working in that are or simply with the FNR protein please share any info you possess. Thank you! From owner-proteins@net.bio.net Tue Oct 08 23:00:00 1996 Path: biosci!agate!spool.mu.edu!howland.erols.net!EU.net!uunet!news-in2.uu.net!nntp.inet.fi!news.funet.fi!news.utu.fi!newsmaster From: heihei@utu.fi (Heikki Heimo) Newsgroups: bionet.molbio.proteins,bionet.molbio.methds-reagnts Subject: Mimetic Blue AP A6XL Date: Wed, 09 Oct 1996 18:04:55 GMT Organization: Univ. of Turku, Finland Lines: 16 Message-ID: <53fm0q$i7b@castor.utu.fi> Reply-To: heihei@utu.fi NNTP-Posting-Host: nasu.bk.utu.fi X-Newsreader: Forte Free Agent 1.0.82 Xref: biosci bionet.molbio.proteins:8951 bionet.molbio.methds-reagnts:50082 Does anybody know the address/E-mail/fax details for a company called ACL, situated in Isle of Man, UK. It is claimed to be manufacturer of an affinity matrix called Mimetic Blue AP A6XL (Pozidis & Bouriotis, Bioseparation 5, 89-93, 1995). Or, alternatively, any other provider of that matrix or any other matrix intended for affinity purification of alkaline phospatases? Any clues welcome! Heikki Heimo, M.Sc. heihei@utu.fi Department of Biochemistry University ofTurku Turku, Finland Tel. +358 21 633 6847 Fax. +358 21 633 6860 From owner-proteins@net.bio.net Tue Oct 08 23:00:00 1996 Path: biosci!daresbury!nntp-trd.UNINETT.no!sn.no!www.nntp.primenet.com!nntp.primenet.com!howland.erols.net!surfnet.nl!swidir.switch.ch!in2p3.fr!univ-lyon1.fr!pasteur.fr!60w1-3-105.dyn.pasteur.fr!dick From: Dick Dickson Newsgroups: bionet.molbio.proteins Subject: Re: eucaryotic cell membranes? Date: 9 Oct 1996 18:07:36 GMT Organization: state university Lines: 1 Distribution: world Message-ID: <53gpl8$hn9$1@montespan.pasteur.fr> References: <53gnpe$75v$2@montespan.pasteur.fr> NNTP-Posting-Host: 60w1-3-105.dyn.pasteur.fr Mime-Version: 1.0 Content-Type: text/plain; charset=ISO-8859-1 Content-Transfer-Encoding: 8bit X-Newsreader: Nuntius 2.0.4_PPC X-XXMessage-ID: X-XXDate: Wed, 9 Oct 1996 20:06:14 GMT ignore From owner-proteins@net.bio.net Tue Oct 08 23:00:00 1996 Newsgroups: bionet.molbio.methds-reagnts,bionet.molbio.proteins Path: biosci!bcm.tmc.edu!news.msfc.nasa.gov!news.sgi.com!nntp-hub2.barrnet.net!news.PBI.net!samba.rahul.net!rahul.net!a2i!bug.rahul.net!rahul.net!a2i!ns2.mainstreet.net!viper.inow.com!news.he.net!news2.qtm.net!news1!ind-017-238-158 From: vanfrank@iquest.net (Richard Van Frank) Subject: Re: de-salting prior to SDS-PAGE loading... X-Nntp-Posting-Host: ind-017-238-158.iquest.net Message-ID: <53es7m$87o_001@ind-017-238-158.iquest.net> Sender: news@iquest.net (News Admin) Organization: IQuest Network Services X-Newsreader: News Xpress Version 1.0 Beta #4 References: Date: Wed, 9 Oct 1996 01:56:55 GMT Lines: 17 Xref: biosci bionet.molbio.methds-reagnts:50128 bionet.molbio.proteins:8955 In article , siyer@bmg.bhs.uab.edu (Arioch) wrote: >hey all, > anybody know of a good protocol to remove the salt from samples >before they are loaded on to SDS-PAGEs?? i have found out that hi salt >tends to distort the way the bands run on the gel (screwing up the >electrical field and therefore affecting protein mobility). i found this >one protocol that tells you to add 0.15% deoxycholic acid, incubate for 10 >min followed by addition of TCA ---> spin for about 15 min at 3000g ---> >decant all the TCA and resuspend pellet (containing precipitated protein >with presumably no salt). this however has not been too successful (for >me) and still causes distortion of my gel bands (much annoyance). anybody >know of any modicfications to this protocol or other protocols that >work?? thanks much... >You might try microfiltration or dialysis on a micro scale. RMVF From owner-proteins@net.bio.net Tue Oct 08 23:00:00 1996 Path: biosci!daresbury!not-for-mail From: Cormac Shaw Newsgroups: bionet.molbio.proteins Subject: Re: How much azide to use? Date: 9 Oct 1996 18:07:40 +0100 Organization: University College Dublin Lines: 23 Sender: lpddist@mserv1.dl.ac.uk Distribution: bionet Message-ID: <53gm4s$iga@mserv1.dl.ac.uk> Original-To: parker@topaz.microbio.uab.edu, PROTEINS@DL.AC.UK On Wed, 09 Oct 1996 10:03:31 -0500 Matthew Parker wrote: >I would like to add azide to my protein preps, as we get a lot of >breakdown when it is stored at 4 degrees. I was thinking of adding 1 or >2 mM; is this enough to keep bacteria down? (I'd like to use as little >as possible.) In our labs we conventionally use sodium azide at 0.02% (w/v) which is a whisker over 3mM. As far as I know, that's a widespread proctice. However, I don't know if its the lowest limit for prevention of contamination. ____________________________________________________ Cormac Shaw, B.Sc., Dept. of Industrial Microbiology, University College Dublin, Belfield, Dublin 4, Ireland. e-mail: cshaw@acadamh.ucd.ie, phone: +353 1 706 1796 fax: +353 1 706 1183. From owner-proteins@net.bio.net Tue Oct 08 23:00:00 1996 Path: biosci!FREENET.TORONTO.ON.CA!bz182 From: bz182@FREENET.TORONTO.ON.CA (Wei Hu) Newsgroups: bionet.molbio.proteins Subject: unsubscribe Date: 9 Oct 1996 02:58:10 -0700 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 2 Sender: daemon@net.bio.net Distribution: world Message-ID: NNTP-Posting-Host: net.bio.net unsubscribe From owner-proteins@net.bio.net Tue Oct 08 23:00:00 1996 Path: biosci!bcm.tmc.edu!cs.utexas.edu!howland.erols.net!feed1.news.erols.com!arclight.uoregon.edu!netnews.worldnet.att.net!newsadm From: Jeffrey J Ayres Newsgroups: bionet.molbio.proteins Subject: Re: HELP! protein purification with gelatin/BSA Date: 9 Oct 1996 21:34:30 GMT Organization: AT&T WorldNet Services Lines: 13 Message-ID: <53h5p6$1n4@mtinsc01-mgt.ops.worldnet.att.net> References: <325923C5.561D@wesleyan.edu> NNTP-Posting-Host: 173.san-francisco-005.ca.dial-access.att.net Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 1.22ATT (Windows; U; 16bit) To: tguina@wesleyan.edu Tina, Since the compound maybe in the triton supernantant you may want to change the detergent. A non polar detergent is the Emulphogen BC series. Since the compound was cationic and hydrophobic you may also want to try a cationic detergent such as Cetylpyridinium Cl, Cetylpyridinium is available from Sigma (www.sigma.com). Centrifuge the solution retaining the supernatant. Follow up with size exclusion chromatography. Another cationic ,or CTAB, CTab is a strong denaturant. A text "Protein Purification Applications" Harris E.L.V, and Angal, S.(Oxford University Press, Oxford) 1990 pp 66-9,76-7 may help with the choice of detergent. Centrifuging the gelatin and decanting off the gelatin, is a way of removing the gelatin from the bacterial culture. From owner-proteins@net.bio.net Tue Oct 08 23:00:00 1996 Path: biosci!agate!spool.mu.edu!howland.erols.net!news.sgi.com!sdd.hp.com!night.primate.wisc.edu!newsspool.doit.wisc.edu!news.doit.wisc.edu!alpha.oncology.wisc.edu!user From: mcmahan@oncology.wisc.edu (Scott McMahan) Newsgroups: bionet.molbio.proteins Subject: (Stept)avidin detection of non-biotinylated proteins Date: Thu, 10 Oct 1996 00:27:00 -0500 Organization: University of Wisconsin-Madison Lines: 10 Message-ID: NNTP-Posting-Host: alpha.oncology.wisc.edu X-Newsreader: Yet Another NewsWatcher 2.1.8 For some experiments I'm doing, I'm trying to detect biotinylated proteins on a western blot. But the (strept)avidins I've tried all also detect the BSA (or lysozyme) I have in my reactions as a non-specific protein. This interaction only occurs with denatured proteins. If I dot-blot the native proteins, I see no signal. Has anyone ever had a similar problem, and if so, what worked to prevent it? -- Scott McMahan mcmahan@oncology.wisc.edu From owner-proteins@net.bio.net Tue Oct 08 23:00:00 1996 Path: biosci!faseb.org!cpk-news-feed1.bbnplanet.com!cpk-news-feed2.bbnplanet.com!cpk-news-hub1.bbnplanet.com!www.nntp.primenet.com!nntp.primenet.com!howland.erols.net!surfnet.nl!swidir.switch.ch!in2p3.fr!univ-lyon1.fr!pasteur.fr!60w1-1-80.dyn.pasteur.fr!fverrier From: Florence Verrier Newsgroups: bionet.molbio.proteins Subject: eucaryotic cell membranes? Date: 9 Oct 1996 17:35:42 GMT Organization: Institut Pasteur Lines: 14 Distribution: world Message-ID: <53gnpe$75v$2@montespan.pasteur.fr> NNTP-Posting-Host: 60w1-1-80.dyn.pasteur.fr Mime-Version: 1.0 Content-Type: text/plain; charset=ISO-8859-1 Content-Transfer-Encoding: 8bit X-Newsreader: Nuntius 2.0.4_PPC X-XXMessage-ID: X-XXDate: Wed, 9 Oct 1996 18:35:55 GMT Dear collueges, I am trying to isolate eucaryotic cell membranes (e.g. BHK cells) before extraction of membrane proteins. For that reason I would like to get a convinient protocoll to isolate the whole membranes without other intra-cellular proteins. I tried to find a good protocoll, but so far I was not satisfied. So I wonder if there might be a colluege out there who has done anything similar and would share his/her protocoll with me, or would give me some references. Thank you very much in advance for your help. Could you please send any suggestions directly to my e-mailadress: fverrier@pasteur.fr Best regards, FV From owner-proteins@net.bio.net Tue Oct 08 23:00:00 1996 Path: biosci!faseb.org!cpk-news-feed1.bbnplanet.com!cpk-news-hub1.bbnplanet.com!newsfeed.internetmci.com!feed1.news.erols.com!hunter.premier.net!netnews.worldnet.att.net!newsadm From: Jeffrey J Ayres Newsgroups: bionet.molbio.proteins Subject: Re: HELP! protein purification with gelatin/BSA Date: 9 Oct 1996 21:38:02 GMT Organization: AT&T WorldNet Services Lines: 13 Message-ID: <53h5vq$1n4@mtinsc01-mgt.ops.worldnet.att.net> References: <325923C5.561D@wesleyan.edu> NNTP-Posting-Host: 173.san-francisco-005.ca.dial-access.att.net Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 1.22ATT (Windows; U; 16bit) To: tguina@wesleyan.edu Tina, Since the compound maybe in the triton supernantant you may want to change the detergent. A non polar detergent is the Emulphogen BC series. Since the compound was cationic and hydrophobic you may also want to try a cationic detergent such as Cetylpyridinium Cl, Cetylpyridinium is available from Sigma (www.sigma.com). Centrifuge the solution retaining the supernatant. Follow up with size exclusion chromatography. Another cationic ,or CTAB, CTab is a strong denaturant. A text "Protein Purification Applications" Harris E.L.V, and Angal, S.(Oxford University Press, Oxford) 1990 pp 66-9,76-7 may help with the choice of detergent. Centrifuging the gelatin and decanting off the gelatin, is a way of removing the gelatin from the bacterial culture. From owner-proteins@net.bio.net Tue Oct 08 23:00:00 1996 Path: biosci!bcm.tmc.edu!cs.utexas.edu!howland.erols.net!news-peer.gsl.net!news.gsl.net!hunter.premier.net!netnews.worldnet.att.net!newsadm From: Jeffrey J Ayres Newsgroups: bionet.molbio.proteins Subject: Re: HELP! protein purification with gelatin/BSA Date: 9 Oct 1996 22:47:11 GMT Organization: AT&T WorldNet Services Lines: 21 Message-ID: <53ha1f$7qa@mtinsc01-mgt.ops.worldnet.att.net> References: <325923C5.561D@wesleyan.edu> NNTP-Posting-Host: 173.san-francisco-005.ca.dial-access.att.net Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 1.22ATT (Windows; U; 16bit) Tina, Since the compound maybe binding in the triton supernatant you may want to change the detergent. Since the compound was cationic you may want to try a cationic detergent such as cetylpyridinium Cl, cetylpyridinium is available fro Sigma(www.sigma.com). Centrifuge the solution retaining the supernatant. Follow up with size exclusion chromatography. Another cationic detergent is MTAB, nTABs are strong denaturants, they are also available from Sigma. A text "Protein Purification Applications" Harris, E.L.V. and Angal, S. (Oxford University Press, Oxford) 1990 pp66-9, 76-7 may help with the choice of detergents. Using the cell culture sample inoculate a cell growth media. A growth media such as L-broth was successful in producing large amounts of e.coli cells in previous experiments (this may also help if the amount of protein is small). L-broth contains 10g tryptone, 5g yeast extract, 5g NaCl. Centrifuging the gelatin-bacterial culture and decanting off the gelatin is the easiest way of removing the media. Further details pertaining to using the growth media are necessary such as temperature, incubation period and volume innoculate write to JeffAyres@worldnet.att.net From owner-proteins@net.bio.net Wed Oct 09 23:00:00 1996 Path: biosci!agate!howland.erols.net!news3.cac.psu.edu!news.cse.psu.edu!news.ece.nwu.edu!newsfeed.acns.nwu.edu!news.cc.uic.edu!usenet From: Keld Sorensen Newsgroups: bionet.molbio.proteins Subject: Re: (Stept)avidin detection of non-biotinylated proteins Date: Thu, 10 Oct 1996 09:29:02 +0000 Organization: University of Illinois, College of Medicine Lines: 22 Message-ID: <325CC1DE.7313@uic.edu> References: NNTP-Posting-Host: sliprock-01.dialin.uic.edu Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 2.01 (Macintosh; I; PPC) To: Scott McMahan [snip question on non specific interaction between blotted proteins and avidin/streptavidin] Non specific interactions are not that uncommon - ways to solve: Use high dilutions of the Avidin (requires a very sensitive substrate) Use deglycosylated avidin (has almost neutral pI) Use additives to the diluent for the avidin such as - high salt (eg 100 mM phos plus 150 mM NaCl or even more salt) - Non-ionic detergents, such as Tween 20 at 0.05% or 0.1% Also - double check that the problem is indeed where you think it is by simply doing the blot and leave out key elements, such as the avidin etc. Keld. From owner-proteins@net.bio.net Wed Oct 09 23:00:00 1996 Path: biosci!ihnp4.ucsd.edu!munnari.OZ.AU!news.ecn.uoknor.edu!news.cis.okstate.edu!usenet From: Ron Tate Newsgroups: bionet.molbio.proteins Subject: Re: Unblocking acetylated amino termini of proteins Date: Thu, 10 Oct 1996 13:04:38 -0500 Organization: Oklahoma State University, Biochemistry and Molecular Biology Lines: 24 Message-ID: <325D3AB6.EAE@bmb-fs1.biochem.okstate.edu> References: <199610101331.GAA12051@net.bio.net> NNTP-Posting-Host: firefly3.biochem.okstate.edu Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 3.0 (Win95; I) Paul Sehnke wrote: > > I am interested in a procedure to remove acetylation from N-terminal methionine > s of plant proteins, so they can be sequenced. Anyone know of references > for such a procedure or info on the procedure itself? I plan to blot the prote > ins after SDS -PAGE onto PVDF. Any help would be most appreciated. > Thank you for your time > Paul Sehnke (PCS@nervm.nerdc.ufl.edu) I don't know about procedures for removing acetyls (but then I haven't looked for any), but I might ask one question. Do you know for sure they are blocked naturally? Sometimes proteins being sequenced which turn up blocked were blocked during preparation, probably during electrophoresis. Precautions can be taken to prevent this. -- >>>>>>---------------------------> Ron Tate Lab of Franklin Leach Dept. of Biochem. & Molecular Biology Oklahoma State University rtate@bmb-fs1.biochem.okstate.edu (405) 744-9326 --------------------------------------------- From owner-proteins@net.bio.net Wed Oct 09 23:00:00 1996 Path: biosci!REX.RE.UOKHSC.EDU!v161 From: v161@REX.RE.UOKHSC.EDU ("v161@rex.uokhsc.edu") Newsgroups: bionet.molbio.proteins Subject: Immunize rabbits with Protein in SDS PAGE gel?? Date: 10 Oct 1996 08:01:52 -0700 Organization: uokhsc Lines: 9 Sender: daemon@net.bio.net Distribution: world Message-ID: <325CCB35.7B2C@rex.uokhsc.edu> Reply-To: v161@REX.RE.uokhsc.edu NNTP-Posting-Host: net.bio.net Hi, I am looking for information regarding immunization of rabbits. I have read that you can immunize them with protein in SDS PAGE gel ( lyophilized or crushed gel slice, and resuspended in water). Does any one have any experience in this matter? Will the SDS be toxic to the Rabbits? Thank you very much, SPaul. From owner-proteins@net.bio.net Wed Oct 09 23:00:00 1996 Path: biosci!NERVM.NERDC.UFL.EDU!PCS From: PCS@NERVM.NERDC.UFL.EDU (Paul Sehnke) Newsgroups: bionet.molbio.proteins Subject: Unblocking acetylated amino termini of proteins Date: 10 Oct 1996 06:36:48 -0700 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 6 Sender: daemon@net.bio.net Distribution: world Message-ID: <199610101331.GAA12051@net.bio.net> NNTP-Posting-Host: net.bio.net I am interested in a procedure to remove acetylation from N-terminal methionine s of plant proteins, so they can be sequenced. Anyone know of references for such a procedure or info on the procedure itself? I plan to blot the prote ins after SDS -PAGE onto PVDF. Any help would be most appreciated. Thank you for your time Paul Sehnke (PCS@nervm.nerdc.ufl.edu) From owner-proteins@net.bio.net Wed Oct 09 23:00:00 1996 Path: biosci!agate!howland.erols.net!surfnet.nl!ruu.nl!cble46.chem.ruu.nl!user From: f.s.wouters@chem.ruu.nl (fred wouters) Newsgroups: bionet.molbio.proteins Subject: CoA derivatization Date: Thu, 10 Oct 1996 11:52:54 +0200 Organization: ruu Lines: 9 Message-ID: NNTP-Posting-Host: cble46.chem.ruu.nl X-Newsreader: Yet Another NewsWatcher 2.1.8 Hi, does anybody have experience with the CoA derivatization of fatty acids in vitro? Is it just as simple as buying acyl CoA sythetase, add ATP and CoA and incubate? Please give me some hints on how to do this. Thanks in advance, Fred From owner-proteins@net.bio.net Wed Oct 09 23:00:00 1996 Path: biosci!faseb.org!cpk-news-feed1.bbnplanet.com!cpk-news-hub1.bbnplanet.com!cam-news-hub1.bbnplanet.com!howland.erols.net!feed1.news.erols.com!arclight.uoregon.edu!netnews.worldnet.att.net!newsadm From: Jeffrey J Ayres Newsgroups: bionet.molbio.proteins Subject: Wrong messages posted to HELP! protein purification with... Date: 10 Oct 1996 18:14:43 GMT Organization: AT&T WorldNet Services Lines: 6 Message-ID: <53jeej$piq@mtinsc01-mgt.ops.worldnet.att.net> NNTP-Posting-Host: 76.san-francisco-006.ca.dial-access.att.net Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 1.22ATT (Windows; U; 16bit) Oops! My unfamiliarity with posting messages to the newsgroup resulted in posting duplicate messages, also note the first two messages posted are not the correct procedures required, the third message is a possible scheme for purifying the protein and a simple technique for separating media from bacteria. From owner-proteins@net.bio.net Wed Oct 09 23:00:00 1996 Path: biosci!dnax.org!dave_campbell From: dave_campbell@dnax.org ("Dave Campbell") Newsgroups: bionet.molbio.proteins Subject: Thioglycolic Acid Date: 10 Oct 1996 10:45:46 -0700 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 4 Sender: daemon@net.bio.net Distribution: world Message-ID: NNTP-Posting-Host: net.bio.net Does anybody know the concentration of Thioglycolate to use in the running buffer of an SDS-PAGE gel to prevent blockage of free N-termini by acrylamide free radicals? From owner-proteins@net.bio.net Wed Oct 09 23:00:00 1996 Path: biosci!NMSU.EDU!hroychow From: hroychow@NMSU.EDU (Hiranya Roychowdhury) Newsgroups: bionet.molbio.proteins Subject: Re: Immunize rabbits with Protein in SDS PAGE gel?? Date: 10 Oct 1996 09:07:48 -0700 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 32 Sender: daemon@net.bio.net Distribution: world Message-ID: <199610101607.KAA06673@NMSU.Edu> NNTP-Posting-Host: net.bio.net At 08:01 AM 10/10/96 -0700, v161@rex.uokhsc.edu wrote: >Hi, >I am looking for information regarding immunization of >rabbits. I have read that you can immunize them with >protein in SDS PAGE gel ( lyophilized or crushed gel >slice, and resuspended in water). Does any one have any >experience in this matter? Will the SDS be toxic to the >Rabbits? >Thank you very much, >SPaul. > > It is interesting to note that you don't consider acrylamide toxic! Although I have heard that one can immunize with the acrylamide present, I would not inject the bunny with acrylamide (call me sensisitive). Acrylamide, even polymerized, is far more toxic than the little SDS that remains after elution. Proteins (polypeptides) are easily eluted from SDS-PAG slices either by electroelution or by passive diffusion. SDS containing sample does not in any way reduce the antigenicity of the polypeptide. If anything, the reduced and denatured state offers better antigen for raising polyclonals. Dr. Hiranya Sankar Roychowdhury Plant Genetic Engineering Lab. New Mexico State University Las Cruces, NM 88003 Ph. (505) 646-5785 hroychow@nmsu.edu From owner-proteins@net.bio.net Wed Oct 09 23:00:00 1996 Path: biosci!biosci!not-for-mail From: Larry Hunter Newsgroups: bionet.info-theory,bionet.molbio.bio-matrix,bionet.molbio.evolution,bionet.molbio.proteins,bionet.biophysics,bionet.general,bionet.molec-model,bionet.women-in-bio,bionet.xtallography Subject: Pacific Symposium on Biocomputing, Announcement and Call for Abstracts Date: 10 Oct 1996 16:05:44 -0700 Organization: National Library of Medicine Lines: 25 Sender: daemon@net.bio.net Distribution: world Message-ID: NNTP-Posting-Host: net.bio.net Xref: biosci bionet.info-theory:4320 bionet.molbio.bio-matrix:772 bionet.molbio.evolution:5142 bionet.molbio.proteins:8973 bionet.biophysics:2360 bionet.general:23557 bionet.molec-model:1180 bionet.women-in-bio:5564 bionet.xtallography:2939 Pacific Symposium on Biocomputing Monday, January 6 through Thursday, January 9, 1997 Ritz Carlton Kapalua, Maui, Hawaii PSB '97 is an international, multidisciplinary conference for the presentation and discussion of current research in the theory and application of computational methods in problems of biological significance. Abstracts will be accepted until November 1, 1996. Please consult our web site for further information: http://www.cgl.ucsf.edu/psb -- Lawrence Hunter, PhD. National Library of Medicine phone: +1 (301) 496-9303 Bldg. 38A, 9th fl, MS-54 fax: +1 (301) 496-0673 Bethesda. MD 20894 USA email: hunter@nlm.nih.gov From owner-proteins@net.bio.net Wed Oct 09 23:00:00 1996 Path: biosci!agate!spool.mu.edu!newspump.sol.net!howland.erols.net!feed1.news.erols.com!news.bconnex.net!clicnet!news.clic.net!rcogate.rco.qc.ca!altitude!usenet From: "Achim Recktenwald, PhD" Newsgroups: bionet.molbio.proteins Subject: Re: How much azide to use? Date: Thu, 10 Oct 1996 13:32:08 -0500 Organization: IBEX Technologies, Inc., Biochemistry, 5485 Pare, Montreal, PQ, H4P 1P7, Canada Lines: 21 Message-ID: <325D4126.68A5@ibex.ca> References: <53gm4s$iga@mserv1.dl.ac.uk> NNTP-Posting-Host: ibex.hip.cam.org Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 3.0 (Macintosh; I; PPC) Cormac Shaw wrote: > > On Wed, 09 Oct 1996 10:03:31 -0500 > Matthew Parker wrote: > > >I would like to add azide to my protein preps, as we get a lot of > >breakdown when it is stored at 4 degrees. I was thinking of adding 1 or > >2 mM; is this enough to keep bacteria down? (I'd like to use as little > >as possible.) > > In our labs we conventionally use sodium azide at 0.02% (w/v) which > is a whisker over 3mM. As far as I know, that's a widespread > proctice. However, I don't know if its the lowest limit for > prevention of contamination. > Be careful and test it first with your protein/enzyme; not all enzymes like it. Achim From owner-proteins@net.bio.net Wed Oct 09 23:00:00 1996 Path: biosci!ihnp4.ucsd.edu!news1.ucsd.edu!usenet From: risaacson@ucsd.edu (Roger Isaacson) Newsgroups: bionet.molbio.proteins,comp.sys.sgi.hardware,bionet.software Subject: Re: Evans & Sutherland dials on SGI??? Date: Fri, 11 Oct 1996 00:46:35 GMT Organization: UCSD Physics Lines: 28 Message-ID: <325d97d0.436300770@news.ucsd.edu> References: <53em1b$a70@gap.cco.caltech.edu> NNTP-Posting-Host: risaacso.extern.ucsd.edu X-Newsreader: Forte Agent .99e/32.227 Xref: biosci bionet.molbio.proteins:8975 comp.sys.sgi.hardware:22561 bionet.software:16774 On 8 Oct 1996 22:53:31 GMT, mathog@seqaxp.bio.caltech.edu wrote: ->We have several defunct Evans & Sutherland graphics workstations about ->(PS300's, I think). Each has a set of 8 dials on it that look quite a bit ->like an older version of the dial boxes on our SGIs. We have one SGI that ->is currently sans dials and it would be great if we could scavenge one of ->these old dial sets for it, rather than having to shell out $2000 for a ->new set (with the buttons, which we basically don't use). .... ->Anybody know if the old E&S dials can be used on an SGI? .......->David Mathog ->mathog@seqaxp.bio.caltech.edu ->Manager, sequence analysis facility, biology division, Caltech We also have a set of dials from a defunct E&S and an SGI without dials and would like to adapt them if possible. In addition we have a Tektronix liquid crystal shutter stereo adapter from the E&S system we would like to adapt to use instead of crystal eyes. Has anyone tried this on and SGI? Thanks, Roger Isaacson risaacson@ucsd.edu UCSD Physics Dept PH 619-534-2505 9500 GILMAN DRIVE FAX 619-822-0007 LA JOLLA CA 92093-0319 From owner-proteins@net.bio.net Wed Oct 09 23:00:00 1996 Path: biosci!faseb.org!cpk-news-feed1.bbnplanet.com!cpk-news-feed2.bbnplanet.com!cam-news-hub1.bbnplanet.com!howland.erols.net!news3.cac.psu.edu!news.cse.psu.edu!news.cc.swarthmore.edu!netnews.upenn.edu!mail.med.upenn.edu!wellsg From: wellsg@mail.med.upenn.edu (Gregg B Wells) Newsgroups: bionet.molbio.proteins Subject: myelomas for heterologous protein expression? Date: 10 Oct 1996 18:50:43 GMT Organization: University of Pennsylvania Lines: 16 Message-ID: <53jgi3$7in@netnews.upenn.edu> NNTP-Posting-Host: mail.med.upenn.edu X-Newsreader: TIN [version 1.2 PL2-upenn1.1] Has anyone done much with the production of non-immunoglobulin proteins from myeloma cell lines? Are protein expression vectors available with immunoglobulin signal sequences or immunoglobulin promoters? I think that myeloma cell lines would be excellent producers for heterologous protein expression, but I could find very little literature other than for immunoglobulin-type proteins. Thanks for insights on this topic. Gregg Wells Department of Pathology University of Pennsylvania Philadelphia, Pennsylvania USA email: wellsg@mail.med.upenn.edu From owner-proteins@net.bio.net Thu Oct 10 23:00:00 1996 Path: biosci!faseb.org!cpk-news-feed1.bbnplanet.com!cpk-news-hub1.bbnplanet.com!www.nntp.primenet.com!nntp.primenet.com!arclight.uoregon.edu!usenet.eel.ufl.edu!usenet.nerdc.ufl.edu!usenet.ufl.edu!mitzi.rsmas.miami.edu!miasun.med.miami.edu!newssun!tguettou From: Toumy Guettouche Newsgroups: bionet.molbio.proteins Subject: Re: Student needs help. Date: Fri, 11 Oct 1996 15:18:57 -0400 Organization: University of Miami, School of Medicine Lines: 34 Message-ID: References: <325E0569.6B9C@world-net.net> NNTP-Posting-Host: newssun.med.miami.edu Mime-Version: 1.0 Content-Type: TEXT/PLAIN; charset=US-ASCII In-Reply-To: <325E0569.6B9C@world-net.net> On Fri, 11 Oct 1996, Filippo Bicocco wrote: > I'm a Bio-chem. undergrad. student @ St. Mary's Uni. @ San Antonio, TX. > USA. And I turn to the news group in hopes to recieve some help for my > seminar that is due the 11th of November. The topic of the seminar is: > > Antibody production by B lymphocytes. The distribution and functions of > isotopes. > > If you have any info. that might/could/is helpful, I'll greatly appre. > your help. If you know of a "site", literature, or if you wish to > share your knowledge, please email me @: > > flip@world-net.net > > Thank you very much... > > Filippo B. > > Hi, try bionet.immunology, Textbooks: Janeway/Travers:Immunobiology,Garland Publ.; Abbas/Lichtman/Pober, Cellular and Molecular Immunology, Saunders; or best of all, a library. BTW you probably mean the function of isotypes (IgG,M,A.D,E), not isotopes (radioactive substances)?! Hope that helps Toumy From owner-proteins@net.bio.net Thu Oct 10 23:00:00 1996 Path: biosci!ihnp4.ucsd.edu!swrinde!nntp.primenet.com!feed1.news.erols.com!uunet!news-in2.uu.net!netnews.nwnet.net!news-hub.interserv.net!news.sprynet.com!news From: Louis Geller Newsgroups: bionet.molbio.proteins Subject: Re: Student needs help. Date: Fri, 11 Oct 1996 23:35:03 -0700 Organization: Cal State Univ. Long Beach Lines: 8 Message-ID: <325F3C17.437D@sprynet.com> References: <325E0569.6B9C@world-net.net> NNTP-Posting-Host: dd16-051.compuserve.com Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 3.0 (Macintosh; I; 68K) To: Toumy Guettouche Filippo, Try the Antibody Resource Page. Here is the URL: http://www-chem.ucsd.edu/Faculty/goodman/antibody.html/abpage.html Try a search of Altavista, or Yahoo. These are good search engines. I hope this helps. Louis Geller... From owner-proteins@net.bio.net Thu Oct 10 23:00:00 1996 Path: biosci!agate!howland.erols.net!www.nntp.primenet.com!nntp.primenet.com!news.fibr.net!news.world-net.net!usenet From: Filippo Bicocco Newsgroups: bionet.molbio.proteins Subject: Student needs help (correction) Date: Fri, 11 Oct 1996 23:15:53 -0500 Organization: World Net, Inc. - San Antonio, TX Lines: 16 Message-ID: <325F1B79.78A0@world-net.net> Reply-To: flip@world-net.net NNTP-Posting-Host: dialip192.world-net.net Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 3.0Gold (Win95; I) Errr.....sorry about my mistake. In my previews message I wrote: "isotopes" and it should be: "isotypes" Sorry about that....but writing a note at 3:30 in the morning isn't easy for me.... Thanks once more... Filippo. From owner-proteins@net.bio.net Thu Oct 10 23:00:00 1996 Path: biosci!rutgers!csn!nntp-xfer-1.csn.net!carbon!night.primate.wisc.edu!newsspool.doit.wisc.edu!news.doit.wisc.edu!alpha.oncology.wisc.edu!user From: mcmahan@oncology.wisc.edu (Scott McMahan) Newsgroups: bionet.molbio.proteins Subject: Re: (Stept)avidin detection of non-biotinylated proteins Date: Fri, 11 Oct 1996 16:41:35 -0500 Organization: University of Wisconsin-Madison Lines: 43 Message-ID: References: <325CC1DE.7313@uic.edu> NNTP-Posting-Host: alpha.oncology.wisc.edu X-Newsreader: Yet Another NewsWatcher 2.1.8 In article <325CC1DE.7313@uic.edu>, Keld Sorensen wrote: > Non specific interactions are not that uncommon - > ways to solve: > > Use high dilutions of the Avidin (requires a very > sensitive substrate) I'm using a 1:10,000, is that high enough? > > Use deglycosylated avidin (has almost neutral pI) Tried that > > Use additives to the diluent for the avidin such as > - high salt (eg 100 mM phos plus 150 mM > NaCl or even more salt) I've titrated the salt from 100 to 500 mM with no noticeble effect > - Non-ionic detergents, such as Tween 20 at > 0.05% or 0.1% > I've tried both these levels. > Also - double check that the problem is indeed > where you think it is by simply doing the blot > and leave out key elements, such as the avidin etc. I tried this to determine that it is the biotin binding protein causing the problem. I've also tried change the conjugate (both Alk Phos and HRP), so it does seem to be the avidin/streptavidin. Thank you for the suggestions, but as you can see, I've tried most already. Great minds think alike :{). -- Scott McMahan mcmahan@oncology.wisc.edu From owner-proteins@net.bio.net Thu Oct 10 23:00:00 1996 Path: biosci!rutgers!iag.net!newspump.sol.net!howland.erols.net!vixen.cso.uiuc.edu!newsfeed.internetmci.com!news.msfc.nasa.gov!info.uah.edu!maze.dpo.uab.edu!usenet From: Sean Moore Newsgroups: bionet.molbio.proteins Subject: Re: How much azide to use? Date: Fri, 11 Oct 1996 15:26:28 -0500 Organization: UAB Microbiology Lines: 6 Message-ID: <325EAD74.12DB@uab.edu> References: <53gm4s$iga@mserv1.dl.ac.uk> <325D4126.68A5@ibex.ca> NNTP-Posting-Host: fenway.microbio.uab.edu Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 3.0 (Win95; I) Achim Recktenwald, PhD wrote: I think a little ethanol (like 40-50%) should do the trick. Achim From owner-proteins@net.bio.net Thu Oct 10 23:00:00 1996 Path: biosci!bcm.tmc.edu!cs.utexas.edu!howland.erols.net!www.nntp.primenet.com!nntp.primenet.com!feed1.news.erols.com!uunet!news-in2.uu.net!brighton.openmarket.com!decwrl!tribune.usask.ca!rover.ucs.ualberta.ca!gallin2.biology.ualberta.ca!wgallin From: wgallin@gpu.srv.ualberta.ca (Warren Gallin) Newsgroups: bionet.molbio.proteins Subject: Re: Immunize rabbits with Protein in SDS PAGE gel?? Date: Fri, 11 Oct 96 14:48:36 GMT Organization: University of Alberta Lines: 36 Distribution: world Message-ID: References: <199610101607.KAA06673@NMSU.Edu> NNTP-Posting-Host: gallin2.biology.ualberta.ca X-Newsreader: VersaTerm Link v1.1.4 In Article <199610101607.KAA06673@NMSU.Edu>, hroychow@NMSU.EDU (Hiranya Roychowdhury) wrote: >At 08:01 AM 10/10/96 -0700, v161@rex.uokhsc.edu wrote: >>Hi, >>I am looking for information regarding immunization of >>rabbits. I have read that you can immunize them with >>protein in SDS PAGE gel ( lyophilized or crushed gel >>slice, and resuspended in water). Does any one have any >>experience in this matter? Will the SDS be toxic to the >>Rabbits? >It is interesting to note that you don't consider acrylamide toxic! Although >I have heard that one can immunize with the acrylamide present, I would not >inject the bunny with acrylamide (call me sensisitive). Acrylamide, even >polymerized, is far more toxic than the little SDS that remains after >elution. Proteins (polypeptides) are easily eluted from SDS-PAG slices >either by electroelution or by passive diffusion. SDS containing sample does >not in any way reduce the antigenicity of the polypeptide. If anything, the >reduced and denatured state offers better antigen for raising polyclonals. Polyacrylamide is not toxic. However, it leaves a deposit at the injection site, which can often cause development of a painful granuloma. The easiest way to inject gel purified protein is to transfer to nitrocellulose by electroblotting, pulverize the little piece of NC with the protein band and inject in PBS with an appropriate adjuvant. The advantage of this over soluble protein is that the NC-bound protein provides you with a longer exposure to the antigen (depot effect). However, it works both ways, so it's more a matter of taste than any strong advantage of one approach over the others. Warren Gallin Department of Biological Sciences University of Alberta Edmonton, Alberta T6G 2E9 Canada wgallin@gpu.srv.ualberta.ca From owner-proteins@net.bio.net Thu Oct 10 23:00:00 1996 Path: biosci!daresbury!nntp-trd.UNINETT.no!sn.no!www.nntp.primenet.com!nntp.primenet.com!howland.erols.net!vixen.cso.uiuc.edu!newsfeed.internetmci.com!news.fibr.net!news.world-net.net!usenet From: Filippo Bicocco Newsgroups: bionet.molbio.proteins Subject: Student needs help. Date: Fri, 11 Oct 1996 03:29:29 -0500 Organization: World Net, Inc. - San Antonio, TX Lines: 16 Message-ID: <325E0569.6B9C@world-net.net> Reply-To: flip@world-net.net NNTP-Posting-Host: dialip123.world-net.net Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 3.0Gold (Win95; I) I'm a Bio-chem. undergrad. student @ St. Mary's Uni. @ San Antonio, TX. USA. And I turn to the news group in hopes to recieve some help for my seminar that is due the 11th of November. The topic of the seminar is: Antibody production by B lymphocytes. The distribution and functions of isotopes. If you have any info. that might/could/is helpful, I'll greatly appre. your help. If you know of a "site", literature, or if you wish to share your knowledge, please email me @: flip@world-net.net Thank you very much... Filippo B. From owner-proteins@net.bio.net Thu Oct 10 23:00:00 1996 Path: biosci!faseb.org!cpk-news-feed1.bbnplanet.com!cpk-news-hub1.bbnplanet.com!www.nntp.primenet.com!nntp.primenet.com!howland.erols.net!news-peer.gsl.net!news.gsl.net!portc01.blue.aol.com!newsstand.cit.cornell.edu!news.acsu.buffalo.edu!news.drenet.dnd.ca!crc-news.doc.ca!nott!cunews!freenet-news.carleton.ca!FreeNet.Carleton.CA!dg628 From: dg628@FreeNet.Carleton.CA (Angela C. Murphy) Newsgroups: bionet.molbio.proteins Subject: Re: unblocking acetylated N-termini Date: 11 Oct 1996 17:45:24 GMT Organization: The National Capital FreeNet Lines: 11 Sender: dg628@freenet3.carleton.ca (Angela C. Murphy) Message-ID: <53m13k$7h3@freenet-news.carleton.ca> Reply-To: dg628@FreeNet.Carleton.CA (Angela C. Murphy) NNTP-Posting-Host: freenet3.carleton.ca I believe there is an enzymatic method for removing acetyl groups from the N-termini of proteins, but I don't have any references. You can try sending a message to abrf@aecom.yu.edu or looking at the ABRF web page, at the sequencing sections. Hope this helps. Angela C. Murphy -- ************************************************** Angela C. Murphy Bowie, MD USA angelam@capaccess.org dg628@freenet.carleton.ca ************************************************** From owner-proteins@net.bio.net Thu Oct 10 23:00:00 1996 Path: biosci!agate!spool.mu.edu!howland.erols.net!EU.net!usenet2.news.uk.psi.net!uknet!usenet1.news.uk.psi.net!uknet!uknet!bhamcs!bham!usenet From: altabios@bham.ac.uk (John E. Fox) Newsgroups: bionet.molbio.proteins Subject: Re: Unblocking acetylated amino termini of proteins Date: 11 Oct 1996 12:16:00 GMT Organization: Alta Bioscience Lines: 20 Message-ID: <53ldq0$9vg@sun4.bham.ac.uk> References: <199610101331.GAA12051@net.bio.net> NNTP-Posting-Host: bcs118.bham.ac.uk X-Newsreader: WinVN 0.90.6 In article <199610101331.GAA12051@net.bio.net>, PCS@NERVM.NERDC.UFL.EDU (Paul Sehnke) says: > >I am interested in a procedure to remove acetylation from N-terminal methionine >s of plant proteins, so they can be sequenced. Anyone know of references Not heard of any way to do this. If anyone can think of a nice clean method it could be worth a lot of money. I think you are stuck with trying to get an internal sequence after digestion. I have heard that blocking can occur during purification but I am not convinced about this. If it were the case every protein would be the same. 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 Thu Oct 10 23:00:00 1996 Path: biosci!bloom-beacon.mit.edu!howland.erols.net!news-peer.gsl.net!news.gsl.net!news.sgi.com!esiee.fr!jussieu.fr!univ-lyon1.fr!in2p3.fr!swidir.switch.ch!scsing.switch.ch!ubaclu.unibas.ch!ubaclu.unibas.ch!nntp Newsgroups: bionet.molbio.proteins Subject: PhD student position in Berlin (GERMANY) Message-ID: <1996Oct11.084445.46981@yogi.urz.unibas.ch> From: birgit@pepper.ifi.unibas.ch (Birgit Westermann) Date: 11 Oct 96 08:44:45 MET Nntp-Posting-Host: pepper.ifi.unibas.ch X-Newsreader: Alexandra.app (Version 0.82) Lines: 27 Max Delbrueck Centre for Molecular Medicine The research group "Vesicle transport" at the Max Delbrueck Centre for Molecular Medicine in Berlin (Germany) offers a doctorand position for two years (BAT IIa/2) within the project "Regulation of intracellular protein transport by PKC isoforms". The applicant should have completed his (her) studies in biochemistry, molecular biology or biology. Experience in molecular biology and cell culture techniques would be advantagous. This position has no teaching requirements. If you are interested to work in this research project, please send your c.v. and diploma certificate to Max.Delbrueck-Centrum Dr. Peter Westermann Robert-Roessle-Str. 10 13125 Berlin Germany (e-mail: westerm@mdc-berlin.de) For further information, please contact me (informally, via e-mail) or using the above mentioned address. From owner-proteins@net.bio.net Fri Oct 11 23:00:00 1996 Path: biosci!daresbury!lyra.csx.cam.ac.uk!moose.bioc.cam.ac.uk!user From: nef1002@cus.cam.ac.uk (Nick Fisher) Newsgroups: bionet.molbio.proteins Subject: Re: Thioglycolic Acid Date: Sat, 12 Oct 1996 16:03:01 +0100 Organization: Cambridge University Lines: 14 Distribution: world Message-ID: References: NNTP-Posting-Host: moose.bioc.cam.ac.uk In article , dave_campbell@dnax.org ("Dave Campbell") wrote: > Does anybody know the concentration of Thioglycolate to use in the running > buffer of an SDS-PAGE gel to prevent blockage of free N-termini by acrylamide > free radicals? I usually include 2 mM mercaptoacetic acid to prevent N-blockage. Nick Fisher Biochemistry Dept. Cambridge -- From owner-proteins@net.bio.net Fri Oct 11 23:00:00 1996 Path: biosci!agate!ihnp4.ucsd.edu!news1.ucsd.edu!usenet From: Kevin Shreder Newsgroups: bionet.molbio.proteins Subject: Updated WWWsite: The Antibody Resource Pag Date: Sat, 12 Oct 1996 12:10:51 -0700 Organization: University of California at San Diego Lines: 31 Message-ID: <325FED3B.1507@ucsd.edu> NNTP-Posting-Host: chedccfx.ucsd.edu Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 2.0 (Macintosh; I; PPC) Updated WWWsite: The Antibody Resource Page The Antibody Resource Page, the first website devoted to bringing together the vast number of resources about antibodies on the net, has recently been updated. The page is designed to be browsed by the novice or expert and is divided into several sections: EDUCATIONAL RESOURCES ONLINE JOURNALS THE STUDY OF ANTIBODY RECOGNITION RESOURCES TO FIND ANTIBODIES SEARCHABLE DATABANKS AND DATABASES ANNOUNCEMENTS MISCELLANEOUS RESOURCES The fourth section is particularly large and contains information on ways to find antibodies (including how to obtain Linscott's Directory and the Manufacturers' Specifications and Reference Synopsis (MSRS) catalog), a large list of online companies that sell antibodies or antibody related products (over 60 links!), and a list of companies that sell antibodies that are not online. I am always looking for new information and new links, so if you know of something, please contact me. Or just contact me to let me know what you think of the page! The URL for the Antibody Resource Page is: http://www-chem.ucsd.edu/Faculty/goodman/antibody.html/abpage.html Kevin Shreder, Ph.D. University of California at San Diego kshreder@ucsd.edu From owner-proteins@net.bio.net Fri Oct 11 23:00:00 1996 Path: biosci!UCONNVM.UCONN.EDU!RHODES From: RHODES@UCONNVM.UCONN.EDU Newsgroups: bionet.molbio.proteins Subject: inverse lysine Date: 12 Oct 1996 09:59:56 -0700 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 20 Sender: daemon@net.bio.net Distribution: world Message-ID: <961012.125815.EDT.RHODES@UConnVM.UConn.Edu> NNTP-Posting-Host: net.bio.net I'm looking for either (1) a simple procedure (for the organic-challenged) to convert a lysine NH3+ to a COO- (or some other anion). The process could add a few extra CH2's, but I'd like to avoid the more polar stuff and definitely avoid leaving the cations around. OR (2) a synthetic polymer with a comparable basic structure (i.e. a backbone of _some_ kind with anions at the end of flexible tethers.) A colleague suggested polystyrene sulfinate, but I'm a little concerved about the aromatics. Thanks! | O==O | | DAVID G. RHODES O==O PHONE 860-486-5413 | | SCHOOL OF PHARMACY; U-92 O==O FAX 860-486-4998 | | UNIVERSITY OF CONNECTICUT O==O | | STORRS, CT 06269-2092 O==O RHODES@UCONNVM.UCONN.EDU | | O==O | From owner-proteins@net.bio.net Fri Oct 11 23:00:00 1996 Path: biosci!agate!newsgate.cuhk.edu.hk!newsfeeder.ust.hk!news.ust.hk!not-for-mail From: salty@uxmail.ust.hk (@Salty Bean@) Newsgroups: bionet.molbio.proteins Subject: A question Date: 12 Oct 1996 18:50:31 GMT Organization: Hong Kong University of Science and Technology Lines: 19 Message-ID: <53op9n$fq6@ustsu10.ust.hk> NNTP-Posting-Host: ustsu4.ust.hk X-Newsreader: TIN [UNIX 1.3 950824BETA PL0] To all biological science experts: I have a question on protein/strutrue relationships and it is as follow: "For a long time it was imagined that overall protein folding plays an essential role in production of the correct secondary and tertiary structures requires for protein functio. More recently, it has become apparent that many proteins have a highly flexible structure thus allowing individual domains( or regions) of the protein to function independently. Discuss these contrasting aspects of protein structure with reference to structure/function relationships that apply to the tumor suppressor protein and the yeast transcriptional activator protein Gal4." I really don't know how to get started. So could you all kindly give me some hints on answering such a question. Thanks in advance. From owner-proteins@net.bio.net Fri Oct 11 23:00:00 1996 Path: biosci!UCONNVM.UCONN.EDU!RHODES From: RHODES@UCONNVM.UCONN.EDU Newsgroups: bionet.molbio.proteins Subject: inverse lysine (fwd) Date: 12 Oct 1996 12:57:19 -0700 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 17 Sender: daemon@net.bio.net Distribution: world Message-ID: <961012.155533.EDT.RHODES@UConnVM.UConn.Edu> NNTP-Posting-Host: net.bio.net OOPS ! ! ! ---------8<-----------------(snip)---------------------------- > sulfinate, but I'm a little concerved about the aromatics. They're honest typos, but _BOY_ do they look bad... Even though the i/o and n/v pairs are nearby key pairs, it looks like the poster (a) can't even spell the group he's looking for and (b) is apparently conservative - - - obviously I meant sulfunate and coacervation }:) ((Thanks in advance for any assistance...)) | O==O | | DAVID G. RHODES O==O PHONE 860-486-5413 | | SCHOOL OF PHARMACY; U-92 O==O FAX 860-486-4998 | | UNIVERSITY OF CONNECTICUT O==O | | STORRS, CT 06269-2092 O==O RHODES@UCONNVM.UCONN.EDU | | O==O | From owner-proteins@net.bio.net Fri Oct 11 23:00:00 1996 Path: biosci!ODYSSEE.NET!dellaire From: dellaire@ODYSSEE.NET (Graham Dellaire) Newsgroups: bionet.molbio.proteins Subject: Genome Structure/Function Call for Discussion on New Group Date: 12 Oct 1996 21:03:00 -0700 Organization: McGill Div. of Experimental Medicine Lines: 124 Sender: daemon@net.bio.net Distribution: world Message-ID: <326111DB.392C@odyssee.net> Reply-To: dellaire@odyssee.net NNTP-Posting-Host: net.bio.net Dear Bionet readers, I am posting this note to ask for comments and suggestions for a new news group on genome/chromatin structure and function: tentatively called bionet.genome.structure The importance of chromatin/genomes structure for the processes of replication, transcription and recombination is becoming more and more apparent. Chromatin context can affects the expression and replication timing of a gene domain. Such interelationship between gene function and changes in chromatin structure have been demonstrated through an evolution of techniques from Dnase I sensitivity mapping to fluorescent in situ hybridization (FISH. I believe with such far reaching affects on many areas of molecular biology, it is time we devoted a news group to the study of genome structure and function. The following is a list of "possible" topics. If you feel we should include others or have suggestions as to the format of the news group please reply via e-mail to: dellaire@odyssee.net In addition to myself three tentative discussion leaders have already been contacted and wish to encourage the formation of such a group. They are: Dr. Eric Milot (Erasmus, Neatherlands) Dr. Ronald Hancock (U of Laval, Quebec, Canada) Dr. Peter Cook (Oxford, England) Cheers, Graham Dellaire ________________________ Here is the list of topics so far. 1. Genome/chromatin accessibility and recombination -recombination hotspots (mieotic and mitotic) -fragile sites -imprinting and recombination rates -ectopic gene targeting and chromatin structure 3. Effect of DNA topology/structure(Triple strand, Z-DNA, cruciform, bent etc) on biological processes such as: -replication -transcription -recombination 4. Histones and Nucleosomes and chromatin structure/function -H1 repression of transcription -Post translational modification of histones acetylation (H4, H3), phosphorylation (H1, H3) and ubiquitination (H2A, H2B) -Histone variants (ex. H2A.Z in mammals, H5 of chicken) 5. Models of genome structure (Loop Domain Model, Channel Model, MegaBase Giant Loop Model, etc.) 6. Evolution of the Genome -isochores and base-content (GC vs. AT) -formation of gene clusters and syntenic mapping -repetitive elements (satellites, telomeric and centromeric (alpha) repeats, lines and sines) 7. Biologically important mutants and knockouts that affect genome/chromatin structure -ex. SNF/SWI, TOPO mutants in yeast -RAD 51,52,54 knockout mice -AT, BLM, FA mouse models 8. Techniques for genome/chromatin analysis -Fluorescent Insitu Analysis -psoralen, polyamine crosslinking -In vivo nucleosome foot printing -Dnase I/Micrococcal Nuclease sensitivity -VM26 Topoisomerase II site mapping 9. Chromatin/DNA binding proteins and their effects on chromatin structure and/or gene expression -Polycomb proteins -Rap1 (telomere silencing) -alpha2-MCM1 (repression of MAT locus) -CENP A/B/C (centromere structure/function) -XCAP-C/E, SMC1/2 (chromatin Condensation) -remodeling of chromatin by SWI/SNF proteins 10. Matrix attatchent regions (MAR's), domain boundaries and locus control regions (LCR's) and their relationship to gene structure and function. -definition of transcription/replication domains -model systems ex. betaglobin (LCR) SCS/SCS' of the Drosophila Heat Shock Locus (HS87a7) 11. Phenomenon of Position Effect and Transvection -in drosophila (HP1, polycomb, heterochromatization) -in mammalian systems (silencing or variegated expression of transgenes) 12. Epigenetic effects on gene function -imprinting -methylation -maintenance of early/late replication 13. Dosage compensation mechanisms and X chromosome inactivation -MSL proteins of Drosophila -XIC (Xist RNA) in mammals -CpG methylation 14. Chromatin structure and DNA replication -ORC1 protein of yeast 15. DNA repair and chromatin structure -TFIIH (transcription coupled repair) -p53 -BLM and AT genes -poly-ADP-polymerase (PARP) From owner-proteins@net.bio.net Sat Oct 12 23:00:00 1996 Path: biosci!agate!howland.erols.net!newspump.sol.net!news.mindspring.com!mindspring!uunet!in1.uu.net!fu-berlin.de!zrz.TU-Berlin.DE!news.dfn.de!news.ruhr-uni-bochum.de!news.rwth-aachen.de!news.mu-luebeck.de!photon.ON-Luebeck.DE!msms From: Manfred Schuermann Newsgroups: bionet.molbio.proteins Subject: Orosomucoid genotyping Date: 13 Oct 1996 05:23:57 GMT Organization: Offenes Netz Luebeck e.V. Lines: 21 Message-ID: <53pudd$n9t@photon.ON-Luebeck.DE> NNTP-Posting-Host: on.on-luebeck.de Mime-Version: 1.0 Content-Type: text/plain; charset=iso-8859-1 Content-Transfer-Encoding: 8bit X-Newsreader: TIN [UNIX 1.3 unoff BETA release 961005] Orosomucoid (ORM) is a highly polymorphic protein. Genotyping is usually done by isoelectric focusing. I would like to know, if there is a *DNA / PCR based* method to demonstrate ORM alleles. (The Genome Data Base -GDB- does not list ORM variants) Thank you very much for helpful hints! Manfred Schuermann, M.D. Institute of Human Genetics University of Luebeck Medical School, Germany e-mail msms@on-luebeck.de phone +49 451 500 2620 fax +49 451 500 4187 From owner-proteins@net.bio.net Sat Oct 12 23:00:00 1996 Path: biosci!internet!biosci!not-for-mail From: biohelp (BIOSCI Administrator) Newsgroups: bionet.molbio.proteins Subject: BIOSCI/bionet miniFAQ & Fundraiser Date: 13 Oct 1996 02:00:41 -0700 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 239 Sender: daemon@net.bio.net Distribution: world Message-ID: <199610130900.CAA24982@net.bio.net> NNTP-Posting-Host: net.bio.net (LAST REVISION: 30-JUL-95) This BIOSCI "miniFAQ" is designed to answer the questions that come up the *most frequently*. The main BIOSCI FAQ (Frequently Asked Questions) is