From owner-proteins@net.bio.net Sun Mar 01 22:00:00 1998 Path: biosci!agate!newsfeed.kornet.nm.kr!news.maxwell.syr.edu!newsfeed.internetmci.com!192.87.106.104!surfnet.nl!highway.leidenuniv.nl!ruly46!masc From: Martijn Scheffers Newsgroups: bionet.molbio.proteins Subject: Signal sequences Date: Mon, 2 Mar 1998 11:25:46 +0100 Organization: Leiden University, The Netherlands Lines: 22 Message-ID: References: <6d25t5$2k@newsstand.cit.cornell.edu> NNTP-Posting-Host: ruly46.medfac.leidenuniv.nl Mime-Version: 1.0 Content-Type: TEXT/PLAIN; charset=US-ASCII X-Sender: masc@ruly46 In-Reply-To: <6d25t5$2k@newsstand.cit.cornell.edu> Hi there, I'am looking for information about signal sequences and their functions in protein transport aross the ER and to the plasma membrane. Thanks, Martijn. ************************************************** Martijn Scheffers RUL, fakulteit geneeskunde Anthropogenetica Wassenaarseweg 72 Postbus 9503 2300 RA Leiden tel. : 071-5276203 e-mail: M.Scheffers@ruly46.medfac.leidenuniv.nl ************************************************** From owner-proteins@net.bio.net Sun Mar 01 22:00:00 1998 Path: biosci!agate!logbridge.uoregon.edu!news.maxwell.syr.edu!nntp.news.xara.net!xara.net!server5.netnews.ja.net!mserv1.dl.ac.uk!not-for-mail From: ctzhang@tju.edu.cn (chun ting zhang) Newsgroups: bionet.molbio.proteins Subject: How to calculate the ratio of parallel to antiparallel strands Date: 2 Mar 1998 13:56:33 -0000 Lines: 10 Sender: lpddist@mserv1.dl.ac.uk Distribution: bionet Message-ID: <6dedqh$sbc@mserv1.dl.ac.uk> Apparently-To: proteins@dl.ac.uk Dear Netters: I need a program to calculate the ratio of the parallel to anti-parallel strands from a PDB file. Those who can show me the relevant Internet address by which I can download the program will be highly appreciated. Thank you very much. R. Zhang (ctzhang@tju.edu.cn) From owner-proteins@net.bio.net Sun Mar 01 22:00:00 1998 Path: biosci!agate!howland.erols.net!news-peer.gip.net!news-lond.gip.net!news.gsl.net!gip.net!nntp.news.xara.net!xara.net!server6.netnews.ja.net!server4.netnews.ja.net!server2.netnews.ja.net!lyra.csx.cam.ac.uk!hgmp.mrc.ac.uk!guardian.dcs.warwick.ac.uk!warwick!leicester!usenet From: "Dr E. Buxbaum" Newsgroups: bionet.molbio.proteins Subject: Re: How to purified a protein by its anti-body? Date: Mon, 02 Mar 1998 10:39:13 -0800 Organization: University of Leicester (PCFS User) Lines: 34 Message-ID: <34FAFCD1.79B0@le.ac.uk> References: <6p5K.530$wu2.1980305@carnaval.risq.qc.ca> NNTP-Posting-Host: pc51.msb.le.ac.uk Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 3.01 (Win16; I) XIAOMING YANG wrote: > > I had purified a human protein from two different cell lines. The way i > took was to employ two different columns: DEAE sepharose and DNA agarose > columns. But, the results show that there are other two more bands on > SDS-PAGE gel. I want to take immunopercipitation way to make a further > purification. How can I do? Protocal? Thank! The first thing I would do is to make a western blot to see whether or not the antibody binds to the other bands (this could happen for example after partial proteolysis). If they don't, you can use the antibody for further purification. Immunoprecipitation is suitable only for very small scale preparations (ng-ug amounts). You incubate the protein with the antibody, then fish out the complex with Protein A (or Protein G) agarose beads. Wash a couple of times with high salt/mild detergent, then use the pellet directly for example for SDS-PAGE. Alternatively, you can use dead Staph. aureus bacteria (Pansorbin, Calbiochem) instead of Protein A agarose (cheaper). Calbiochem has a free booklet on Pansorbin, which contains suggestions for washing buffer composition. For larger scale preps, couple your antibody to activated agarose (several chemistries are available commercially, for example from Pharmacia) and pack the gel into a small column. Load the column with your sample, wash with high salt/mild detergent and elute with a denaturing buffer (Diiodosalicylate or Glycine/HCl pH 4). In any case, have some neutralising buffer in the tubes you use to collect the fractions to minimize exposure of your protein to the elution buffer and thoroughly wash the column with PBS or similar immediately afterwards. With luck, both your protein and the antibody will withstand this procedure without permanent damage. If you find the different bands again after these procedures, then they may be part of an multi-subunit protein complex. From owner-proteins@net.bio.net Sun Mar 01 22:00:00 1998 Path: biosci!agate!logbridge.uoregon.edu!Cabal.CESspool!bofh.vszbr.cz!lyra.csx.cam.ac.uk!hgmp.mrc.ac.uk!guardian.dcs.warwick.ac.uk!warwick!leicester!usenet From: "Dr E. Buxbaum" Newsgroups: bionet.molbio.proteins Subject: Re: Ligand Binding Date: Mon, 02 Mar 1998 10:22:52 -0800 Organization: University of Leicester (PCFS User) Lines: 10 Message-ID: <34FAF8FC.3E4D@le.ac.uk> References: <6d27os$nla$1@agate.berkeley.edu> NNTP-Posting-Host: pc51.msb.le.ac.uk Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 3.01 (Win16; I) Louis Hom wrote: > > Basic question: > Is there an activation energy associated with ligand binding? Yes, there is, and it is easy to measure. Simply determine the Kd at various temperatures and plot ln(Kd) vs the reciprocal temperature (absolute temperature, i.e. in Kelvin). If certain conditions are met, then the data points will be on a straight line, from the slope of which you can calculate the activation energy. From owner-proteins@net.bio.net Sun Mar 01 22:00:00 1998 Path: biosci!agate!logbridge.uoregon.edu!newsfeed2.uk.ibm.net!ibm.net!nntp.news.xara.net!xara.net!rill.news.pipex.net!pipex!server1.netnews.ja.net!hgmp.mrc.ac.uk!guardian.dcs.warwick.ac.uk!warwick!leicester!usenet From: "Dr E. Buxbaum" Newsgroups: bionet.molbio.proteins Subject: Re: Algal Biliproteins Date: Mon, 02 Mar 1998 10:15:46 -0800 Organization: University of Leicester (PCFS User) Lines: 16 Message-ID: <34FAF752.4BC7@le.ac.uk> References: <6d079e$cqm$1@Venus.mcs.net> NNTP-Posting-Host: pc51.msb.le.ac.uk Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 3.01 (Win16; I) Deirdre Sholto-Douglas wrote: > He describes using a Calcium phosphate gel column and the addition > of stepwise phosphate buffers to elute the coloured proteins from > _Porphyra_. His methodology states that Celite was mixed to the > gel "as support" but its unclear exactly how much was used. > Admittedly, the gel is decidedly unstable as is and would probably > benefit from the addition of diatomaceous earth. > > Has anyone done something similar or know the rough proportions > of such a mixture? If by Calcium phosphate you mean Hydroxyapatite, then there are nice beaded forms of this material on the market, for example from BioRad. An introduction into HA chromatography can be found in Deutscher's "Guide to Protein Purification" (in the Methods in Enzymology series). From owner-proteins@net.bio.net Sun Mar 01 22:00:00 1998 Path: biosci!agate!logbridge.uoregon.edu!newsfeed2.uk.ibm.net!ibm.net!nntp.news.xara.net!xara.net!rill.news.pipex.net!pipex!server1.netnews.ja.net!hgmp.mrc.ac.uk!guardian.dcs.warwick.ac.uk!warwick!leicester!usenet From: "Dr E. Buxbaum" Newsgroups: bionet.molbio.proteins Subject: Re: stable protein complex during SDS-PAGE Date: Mon, 02 Mar 1998 10:09:01 -0800 Organization: University of Leicester (PCFS User) Lines: 11 Message-ID: <34FAF5BD.1542@le.ac.uk> References: <01bd4167$1c88b480$df80808f@bchm7.bch.unp.ac.za> NNTP-Posting-Host: pc51.msb.le.ac.uk Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 3.01 (Win16; I) brendon wrote: > > Thomas wrote in article > HI - I HAVE HAD THE SAME TROUBLE IN THE PAST. YOU CAN DENATURE YOUR > PROTEINS IN 6M GUANIDINE HCL, 5 mM EDTA, pH 8.1. FROM THERE, REDUCE WITH > DITHIOTHREITOL AND CARBOXMETHYLATE. GOOD LUCK Guanidinum HCl may not be the ideal reagent for use in electrophoresis. Urea, being non-charged, might be the better choice. The other little trick I found usefull is to incubate the sample in Laemli buffer for 30 min at 37 degrees, instead of boiling them. From owner-proteins@net.bio.net Sun Mar 01 22:00:00 1998 Newsgroups: bionet.molbio.methds-reagnts,bionet.molbio.proteins Path: biosci!agate!spool.mu.edu!uwm.edu!news1.chicago.iagnet.net!news2.chicago.iagnet.net!qual.net!iagnet.net!newsxfer3.itd.umich.edu!newsxfer.itd.umich.edu!gumby!newspump.wustl.edu!biko.cc.rochester.edu!news.acsu.buffalo.edu!srv1.drenet.dnd.ca!crc-news.doc.ca!nott!kwon!utnut!utinfo!nntp From: Sam Lee Subject: pRK5 Expression Vector X-Nntp-Posting-Host: pharm4358b.med.utoronto.ca Content-Type: text/plain; charset=us-ascii Message-ID: <34FB4315.94B62778@utoronto.ca> Sender: nntp@utcc.utoronto.ca (News) Content-Transfer-Encoding: 7bit Organization: University of Toronto Mime-Version: 1.0 Date: Mon, 2 Mar 1998 23:39:01 GMT X-Mailer: Mozilla 4.03 [en] (Win95; I) Lines: 22 Xref: biosci bionet.molbio.methds-reagnts:65396 bionet.molbio.proteins:12387 Hello All, Does anyone have any information on the pRK5 expression vector? Most importantly, what is its antibiotic resistance? What are the commercial sources of this vector. I have seen questions about pRK5 asked before in usenet discussion groups, but I have not seen an answer. I have also thoroughly searched the internet and can find no mention of it. Is there a reason why, apparently, no one uses this vector these days? Any help would be greatly appreciated. Sam. ------------------------------------- samuel.lee@utoronto.ca Department of Pharmacology University of Toronto Toronto, Ontario Canada From owner-proteins@net.bio.net Sun Mar 01 22:00:00 1998 Path: biosci!ACC.WUACC.EDU!zzbart From: zzbart@ACC.WUACC.EDU (barton jan) Newsgroups: bionet.molbio.proteins Subject: Re: Ligand Binding Date: 2 Mar 1998 08:10:48 -0800 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 31 Sender: daemon@net.bio.net Distribution: world Message-ID: References: <34FAF8FC.3E4D@le.ac.uk> NNTP-Posting-Host: net.bio.net On Mon, 2 Mar 1998, Dr E. Buxbaum wrote: > Louis Hom wrote: > > > > Basic question: > > Is there an activation energy associated with ligand binding? > > Yes, there is, and it is easy to measure. Simply determine the Kd at > various temperatures and plot ln(Kd) vs the reciprocal temperature > (absolute temperature, i.e. in Kelvin). If certain conditions are met, > then the data points will be on a straight line, from the slope of which > you can calculate the activation energy. > > There seems to be some confusion about kinetic and equilibrium processes. Kinetic processes yield the activation energy when the ln(rate constant) if plotted vs i/T. Equilibrium processes yield the standard enthalpy change when the equilibrium/dissociation constant is substituted for the rate constant in the ln term. Janice S. Barton, Ph.D. Professor and Chair of Chemistry Department of Chemistry Washburn University, Topeka, KS 66621 zzbart@washburn.edu 785-231-1010-x1269 From owner-proteins@net.bio.net Mon Mar 02 22:00:00 1998 Path: biosci!mpih-frankfurt.mpg.de!Junghans From: Junghans@mpih-frankfurt.mpg.de (Junghans) Newsgroups: bionet.molbio.proteins Subject: Staining of PVDF after transfer? Date: 3 Mar 1998 06:08:03 -0800 Organization: mpih-frankfurt Lines: 22 Sender: daemon@net.bio.net Distribution: world Message-ID: <34FC0E1D.D33FCCE3@mpih-frankfurt.mpg.de> Reply-To: Junghans@mpih-frankfurt.mpg.de NNTP-Posting-Host: net.bio.net Hi I have a problem with staining of PVDF-membranes after western-blot transfer. How can I stain the marker or in general my proteins before immunodetection. I tried PonceauS but this didn't work even after adding some methanol the result was not very promising, because only very strong bands were recognized. Does anybody know a more sensitive method? Thanks, Dirk Junghans -- ************************************** Dirk Junghans Max-Planck-Institut f. Hirnforschung Abt. Neurochemie Deutschordenstr. 46 60528 Frankfurt am Main e-mail: junghans@mpih-frankfurt.mpg.de Tel. ++49 - 69-96769-457 Fax. ++49 - 69-96769-441 From owner-proteins@net.bio.net Mon Mar 02 22:00:00 1998 Path: biosci!agate!logbridge.uoregon.edu!ais.net!newsm.ibm.net!ibm.net!newsjunkie.ans.net!newsfeeds.ans.net!news.pprd.abbott.com!unixadmin@pprd.abbott.com From: Seth Crosby Newsgroups: bionet.molbio.methds-reagnts,bionet.molbio.proteins,bionet.cellbio Subject: Tet inducible mouse Date: Tue, 03 Mar 1998 08:55:20 -0500 Organization: Abbott Labs Lines: 18 Message-ID: <34FC0BB5.7F492432@ugene1.pprd.abbott.com> Reply-To: crosbys@ugene1.pprd.abbott.com NNTP-Posting-Host: lc922990.pprd.abbott.com Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii; x-mac-type="54455854"; x-mac-creator="4D4F5353" Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 4.04 (Macintosh; I; PPC) Xref: biosci bionet.molbio.methds-reagnts:65421 bionet.molbio.proteins:12394 Hello All, We would like to purchase a mouse transgenic for reverse tet repressor (rTetR a.k.a. "pTet-on"). If your lab has any to spare, please contact me. Thanks!! -- Seth Crosby, MD Senior Research Scientist Genomics/Bioinformatics Dept. 04MD, Bldg. AP9A/3 Abbott Labs Abbott Park, IL (847) 938-6999 FAX 938-6046 From owner-proteins@net.bio.net Mon Mar 02 22:00:00 1998 Path: biosci!agate!news.Stanford.EDU!nntp.Stanford.EDU!not-for-mail From: jladasky@pmgm.Stanford.EDU (John Ladasky) Newsgroups: bionet.cellbiol,bionet.molbio.proteins Subject: Re: Gradient separations of Organelles Date: 3 Mar 1998 11:53:45 -0800 Organization: Stanford University, CA 94305, USA Lines: 22 Message-ID: <6dhn49$vv@pmgm.Stanford.EDU> References: <6dhim0$oqi$1@news.duke.edu> NNTP-Posting-Host: pmgm.stanford.edu Xref: biosci bionet.cellbiol:8980 bionet.molbio.proteins:12392 In article <6dhim0$oqi$1@news.duke.edu>, William P. Tschantz wrote: >Hi > >I am looking for a reletively easy protocal to separate different membrane >fractions by gradient centrifugation preferably using sucrose or percoll >gradients. Mostly want to separate PM, golgi, mitochondria and lysosomes. >Any references to point me in the right direction would be greatly >appreciated. Ira Mellman's group has succeeded in isolating endosomal organelles using an electrophoresis technique. I heard him speak in 1996, about the time that Immunity 1996 Mar;4(3):229-239 came out, so you might want to start with that reference. I don't know whether this would suit your needs, since you seem to want to use gradient centrifugation, but check it out. Good luck! -- Rainforest laid low. "Wake up and smell the ozone," Says man with chainsaw. - John Ladasky From owner-proteins@net.bio.net Mon Mar 02 22:00:00 1998 Path: biosci!agate!newsgate.duke.edu!galactose.mc.duke.edu.uucp!tschantz From: tschantz@galactose.mc.duke.edu (William P. Tschantz) Newsgroups: bionet.cellbiol,bionet.molbio.proteins Subject: Gradient separations of Organelles Date: 3 Mar 1998 18:37:52 GMT Organization: Duke University; Durham, N.C. Lines: 17 Distribution: world Message-ID: <6dhim0$oqi$1@news.duke.edu> NNTP-Posting-Host: galactose.mc.duke.edu Xref: biosci bionet.cellbiol:8979 bionet.molbio.proteins:12391 Hi I am looking for a reletively easy protocal to separate different membrane fractions by gradient centrifugation preferably using sucrose or percoll gradients. Mostly want to separate PM, golgi, mitochondria and lysosomes. Any references to point me in the right direction would be greatly appreciated. Thanks in advance Bill -- | William R. Tschantz, Ph.D. * | | Dept of Pharmacology and Cancer Biology * | | Duke University, Durham, NC * Got Homebrew ?? | | tschantz@galactose.mc.duke.edu * | From owner-proteins@net.bio.net Mon Mar 02 22:00:00 1998 Path: biosci!agate!howland.erols.net!newsfeed.internetmci.com!193.174.75.126!news-was.dfn.de!news-fra1.dfn.de!zeus.rbi.informatik.uni-frankfurt.de!grapool30.rz.uni-frankfurt.de!parcej From: parcej@biophys.mpg.de (Dr Dave Parcej) Newsgroups: bionet.molbio.proteins Subject: Re: Staining of PVDF after transfer? Date: Wed, 04 Mar 1998 08:46:17 +0100 Organization: MPI fuer Biophysik Lines: 18 Distribution: world Message-ID: References: <34FC0E1D.D33FCCE3@mpih-frankfurt.mpg.de> NNTP-Posting-Host: mac-kb5.biophys.mpg.de X-Newsreader: MT-NewsWatcher 2.3.5 In article , parcej@biophys.mpg.de (Dr Dave Parcej) wrote: Thats 0.01% sulphorhodamine!!!!!! > Dirk > Its odd that Ponceau S didnt work, but maybe youd like to try this: > Briefly dip the blot in a 0.01%(w/v) prepared in 30% methanol. I think (ie > you should test it with an unimportant sample first!) that you can destain > with either water or methanol. We used this for preparing samples for > sequencing. > Heres the reference: > Pappin et al (1990) Analyt Biochem. 187, 10-19 > Let me know how it works! > Dave > > > From owner-proteins@net.bio.net Mon Mar 02 22:00:00 1998 Path: biosci!agate!ihnp4.ucsd.edu!news.scripps.edu!not-for-mail From: "Geoffrey Swanson" Newsgroups: bionet.molbio.methds-reagnts,bionet.molbio.proteins Subject: Re: pRK5 Expression Vector Date: Tue, 3 Mar 1998 08:43:57 -0800 Organization: The Salk Institute Lines: 15 Message-ID: <6dhc1c$8r9$1@hermes.scripps.edu> References: <34FB4315.94B62778@utoronto.ca> NNTP-Posting-Host: cornetto.salk.edu Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Newsreader: Microsoft Outlook Express 4.71.1712.3 X-MimeOLE: Produced By Microsoft MimeOLE V4.71.1712.3 Xref: biosci bionet.molbio.methds-reagnts:65426 bionet.molbio.proteins:12395 Sam Lee wrote in message <34FB4315.94B62778@utoronto.ca>... >Hello All, > >Does anyone have any information on the pRK5 expression vector? ....> I have a more-or-less correct map for pRK5. Email your details and I will send a copy to you. The antibiotic resistance is amp. As far as I know, there are no commercial sources for this vector. There isn't much of a useful MCS, which is probably why not many people use it any more. Geoff Swanson Email: swanson@salk.edu From owner-proteins@net.bio.net Mon Mar 02 22:00:00 1998 Path: biosci!agate!newsfeed.kornet.nm.kr!nntp.kreonet.re.kr!news.maxwell.syr.edu!news.algonet.se!newsfeed.sollentuna.se!newsfeed.interlog.com!news.interlog.com!lemmings From: lemmings@interlog.com (Carol Thomas) Newsgroups: bionet.molbio.proteins Subject: Anyone doing proteomics research? Followup-To: bionet.molbio.proteins Date: Tue, 03 Mar 1998 20:33:37 GMT Organization: Interlog Internet Services Lines: 28 Message-ID: <6dhoum$688$1@news.interlog.com> Reply-To: lemmings@interlog.com NNTP-Posting-Host: lemmings.interlog.com X-Newsreader: News Xpress 2.01 ..and willing to be interviewed about it via email? Forgive me for invading your newsgroup, but I'm a science writer with a tough assignment: 1500 words on proteomics by Friday, March 6th. Since I normally do medical writing, this is slightly out of my field. I've researched proteomics and do know what it is and how it's generally done, but I really need to find a few kind souls who wouldn't mind being interviewed, since my usual sources aren't appropriate. What I was thinking of was a short list of questions -- what do you think the benefits of proteomics are, how can it help in the drug discovery process, how can it be done faster/more effectively, that kind of thing -- sent by email. If you prefer, I'll call you long-distance to talk. If you're willing to help, I'll use all your quotes accurately (I'm a perfectionist about quote accuracy!) and cite you with your full title as a source. The magazine is Laboratory Focus, based in Ontario, but non-Canadian sources are entirely welcome. I'd be very grateful for any interviewees. Email me at lemmings@interlog.com or post here -- I'll be checking the newsgroup every day. Many thanks, Carol Thomas, B.Sc. From owner-proteins@net.bio.net Mon Mar 02 22:00:00 1998 Path: biosci!agate!logbridge.uoregon.edu!news.maxwell.syr.edu!fu-berlin.de!news.tu-darmstadt.de!grapool30.rz.uni-frankfurt.de!parcej From: parcej@biophys.mpg.de (Dr Dave Parcej) Newsgroups: bionet.molbio.proteins Subject: Re: Staining of PVDF after transfer? Date: Tue, 03 Mar 1998 15:41:17 +0100 Organization: MPI fuer Biophysik Lines: 36 Distribution: world Message-ID: References: <34FC0E1D.D33FCCE3@mpih-frankfurt.mpg.de> NNTP-Posting-Host: mac-kb5.biophys.mpg.de X-Newsreader: MT-NewsWatcher 2.3.5 Dirk Its odd that Ponceau S didnt work, but maybe youd like to try this: Briefly dip the blot in a 0.01%(w/v) prepared in 30% methanol. I think (ie you should test it with an unimportant sample first!) that you can destain with either water or methanol. We used this for preparing samples for sequencing. Heres the reference: Pappin et al (1990) Analyt Biochem. 187, 10-19 Let me know how it works! Dave article <34FC0E1D.D33FCCE3@mpih-frankfurt.mpg.de>, Junghans@mpih-frankfurt.mpg.de wrote: > Hi > I have a problem with staining of PVDF-membranes after western-blot > transfer. > How can I stain the marker or in general my proteins before > immunodetection. I tried PonceauS but this didn't work even after adding > some methanol the result was not very promising, because only very > strong bands were recognized. > Does anybody know a more sensitive method? > Thanks, Dirk Junghans > -- > ************************************** > Dirk Junghans > Max-Planck-Institut f. Hirnforschung > Abt. Neurochemie > Deutschordenstr. 46 > 60528 Frankfurt am Main > > e-mail: junghans@mpih-frankfurt.mpg.de > Tel. ++49 - 69-96769-457 > Fax. ++49 - 69-96769-441 > From owner-proteins@net.bio.net Mon Mar 02 22:00:00 1998 Path: biosci!agate!logbridge.uoregon.edu!feed2.news.erols.com!erols!newsxfer.visi.net!nntp.news.xara.net!xara.net!server5.netnews.ja.net!server3.netnews.ja.net!server1.netnews.ja.net!bham!med1000.bham.ac.uk!user From: noone@cancer.bham.ac.uk (noone) Newsgroups: bionet.molbio.proteins Subject: Re: Staining of PVDF after transfer? Date: Tue, 03 Mar 1998 17:49:14 +0000 Organization: noone Lines: 24 Distribution: world Message-ID: References: <34FC0E1D.D33FCCE3@mpih-frankfurt.mpg.de> NNTP-Posting-Host: med1000.bham.ac.uk In article , parcej@biophys.mpg.de (Dr Dave Parcej) wrote: > > Hi > > I have a problem with staining of PVDF-membranes after western-blot > > transfer. > > How can I stain the marker or in general my proteins before > > immunodetection. I tried PonceauS but this didn't work even after adding > > some methanol the result was not very promising, because only very > > strong bands were recognized. > > Does anybody know a more sensitive method? > > Thanks, Dirk Junghans Hi Dirk, what's your recipe for the Ponceau? You have to add something like Sulfosalicylic acid or TCA to precipitate the protein. Anyway, the Ponceau stain is very insensitive compared to Amidoblack T etc. Peter p.weber@bham.ac.uk From owner-proteins@net.bio.net Tue Mar 03 22:00:00 1998 Path: biosci!daresbury!uninett.no!news.maxwell.syr.edu!korova.insync.net!news.io.com!news.tamu.edu!news.ttu.edu!usenet From: "Benan Dincturk" Newsgroups: bionet.molbio.proteins Subject: Re: Signal sequences Date: 4 Mar 1998 21:04:44 GMT Organization: Texas Tech Academic Computing Services Lines: 5 Message-ID: <01bd47b0$7c1af6c0$5d227681@chm407-1.chem.ttu.edu> References: <6d25t5$2k@newsstand.cit.cornell.edu> NNTP-Posting-Host: chm407-1.chem.ttu.edu X-Newsreader: Microsoft Internet News 4.70.1161 Anthony Pugsley's book called "Protein Targeting" which is published in 1989 by Academic Press is a very good reference for signal peptides. Benan knben@ttacs.ttu.edu From owner-proteins@net.bio.net Tue Mar 03 22:00:00 1998 Path: biosci!agate!howland.erols.net!news.idt.net!woodstock.news.demon.net!demon!dispose.news.demon.net!demon!nntp.news.xara.net!xara.net!server5.netnews.ja.net!server3.netnews.ja.net!server1.netnews.ja.net!hgmp.mrc.ac.uk!guardian.dcs.warwick.ac.uk!warwick!leicester!usenet From: "Dr E. Buxbaum" Newsgroups: bionet.molbio.proteins Subject: Re: Staining of PVDF after transfer? Date: Wed, 04 Mar 1998 13:47:54 -0800 Organization: University of Leicester (PCFS User) Message-ID: <34FDCC0A.EAE@le.ac.uk> References: <34FC0E1D.D33FCCE3@mpih-frankfurt.mpg.de> NNTP-Posting-Host: pc48.msb.le.ac.uk Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 3.01 (Win16; I) Lines: 15 Junghans wrote: > > Hi > I have a problem with staining of PVDF-membranes after western-blot > transfer. > How can I stain the marker or in general my proteins before > immunodetection. I would use prestained marker proteins, which are available commercialy from Sigma, BioRad and other companies. Slightly more expensive than the non-stained version, but time saving and convenient. The "rainbow markers" from BioRad are particularly convenient imho, as the different proteins have different colours and can be easily and unambigously identified, even if the gel has been left to run a little too long, so that the front is no longer visible. From owner-proteins@net.bio.net Tue Mar 03 22:00:00 1998 Path: biosci!agate!newsfeed.kornet.nm.kr!news.maxwell.syr.edu!news.mel.connect.com.au!news.labyrinth.net.au!news.alphalink.com.au!not-for-mail From: "Justin Clift" Newsgroups: bionet.molbio.proteins Subject: Anyone heard from Harry Banaharis lately? Know about the protein folding website? Date: 5 Mar 1998 03:00:53 GMT Organization: Alphalink Australia Lines: 13 Message-ID: <01bd4786$95873da0$e1b73ecb@knight> NNTP-Posting-Host: ppp18-9-3.alphalink.com.au X-Newsreader: Microsoft Internet News 4.70.1161 :-) Just as the subject says... I'm trying to find Harry John Banaharis and the protein folding site he was going to create.... Any news or updates? Regards and best wishes, + Justin Clift Digital Distribution From owner-proteins@net.bio.net Tue Mar 03 22:00:00 1998 Path: biosci!bloom-beacon.mit.edu!news.kodak.com!news-pen-14.sprintlink.net!news.sprintlink.net!Sprint!128.122.253.90!newsfeed.nyu.edu!news.idt.net!news.maxwell.syr.edu!uninett.no!Cabal.CESspool!bofh.vszbr.cz!lyra.csx.cam.ac.uk!hgmp.mrc.ac.uk!guardian.dcs.warwick.ac.uk!warwick!not-for-mail From: bssbc@csv.warwick.ac.uk (Dr D T Jones) Newsgroups: bionet.molbio.proteins Subject: UK Studentships Available Date: 4 Mar 1998 21:28:15 -0000 Organization: University of Warwick, UK Lines: 41 Message-ID: <6dkh1f$ibp@lupin.csv.warwick.ac.uk> NNTP-Posting-Host: lupin-fddi.csv.warwick.ac.uk Mime-Version: 1.0 Content-Type: text/plain; charset=US-ASCII Content-Transfer-Encoding: 7bit X-Newsreader: TIN [version 1.2 PL2] UNIVERSITY OF WARWICK DEPARTMENT OF BIOLOGICAL SCIENCES PROTEIN STRUCTURE GROUP BBSRC STUDENTSHIPS IN PROTEIN STRUCTURE & BIOINFORMATICS AVAILABLE Applications are invited for a number of BBSRC Studentships available in the Department of Biological Sciences. Studentships are avilable for projects in the following areas: Viral Crystallography - Supervisor Dr Rob Mckenna, E-mail: mckenna@globin.bio.warwick.ac.uk Protein Crystallography - Supervisor Dr Vilmos Fulop, E-mail: vilmos@globin.bio.warwick.ac.uk Protein Structure Prediction - Supervisor Dr David Jones, (Bioinformatics) E-mail: jones@globin.bio.warwick.ac.uk For general details on the Protein Structure Group, please check the following Web page: http://globin.bio.warwick.ac.uk/structure/structure.html PLEASE NOTE: Eligibility 1. Candidates must have (or expect to get) a 1st or upper second class honours degree (or equivalent). 2. The candidate must be a UK Citizen who has been ordinarily resident in Great Britain for at least 3 years. For general information about the Department of Biological Sciences, please browse the following address: http://www.oikos.warwick.ac.uk From owner-proteins@net.bio.net Tue Mar 03 22:00:00 1998 Path: biosci!agate!newsfeed.kornet.nm.kr!news.maxwell.syr.edu!newsfeed.nacamar.de!fu-berlin.de!news.uni-stuttgart.de!news.ruhr-uni-bochum.de!not-for-mail From: Markus Piotrowski Newsgroups: bionet.molbio.proteins Subject: Re: Staining of PVDF after transfer? Date: Wed, 04 Mar 1998 18:19:59 +0100 Organization: Ruhr-Universitaet Bochum Lines: 14 Message-ID: <34FD8D3F.49CAE222@ruhr-uni-bochum.de> References: <34FC0E1D.D33FCCE3@mpih-frankfurt.mpg.de> NNTP-Posting-Host: dialppp-5-37.rz.ruhr-uni-bochum.de Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 4.03 [en] (Win95; I) To: Junghans@mpih-frankfurt.mpg.de Xcanpos: shelf.01/199803182101!0023565321 Hi Dirk, proteins on PVDF can be stained with Coomassie-blue (Ponceau S will not work)! Simply use your gel stain- and destain-solutions. Unfortunately, I don't know how good it can be destained and if remaining blue color will interfere with your following detection (for example in Westerns). If you want to make Westerns and you use an ECL-system, remaining blue color will be no problem for you. Alternatively, use amido-black. Give it a try, hope this will help you Markus From owner-proteins@net.bio.net Wed Mar 04 22:00:00 1998 Path: biosci!newshost.lanl.gov!news.ttu.edu!usenet From: "Benan Dincturk" Newsgroups: bionet.molbio.proteins Subject: ideal yeast expression system for (plant) protein expression Date: 5 Mar 1998 17:47:41 GMT Organization: Texas Tech Academic Computing Services Lines: 11 Message-ID: <01bd485e$29aef720$5d227681@chm407-1.chem.ttu.edu> NNTP-Posting-Host: chm407-1.chem.ttu.edu X-Newsreader: Microsoft Internet News 4.70.1161 Hello netters E. coli expression does not seem to work for my huge protein. So while I am playing with E. coli vectors, I plan to start with yeast expression. Who has experience with yeast? Can you suggest some vectors with strong promoters? What are the important points in yeast transformation? I know that is the thoughest part usually. I am a beginner so every suggestion will be appreciated. Thank you all. Benan From owner-proteins@net.bio.net Wed Mar 04 22:00:00 1998 Path: biosci!agate!howland.erols.net!news.net.uni-c.dk!news.uni-c.dk!not-for-mail From: "Claus S. Nielsen" Newsgroups: bionet.molbio.proteins Subject: HPLC Hitachi help me Date: Thu, 05 Mar 1998 20:08:49 +0100 Organization: Schafer-N Lines: 12 Message-ID: <34FEF841.74CA@SYMBION.DK> Reply-To: SCHAFER-N@SYMBION.DK NNTP-Posting-Host: schafer-n.symbion.dk Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 3.0Gold (Win95; I) We got an old HPLC from Hitachi, but the manual is lost. Does anyone have operating instructions for the 655-61 Processor A controller? Claus Nielsen Schafer-N Symbion Science Park 2100 Copenhagen O Denmark Fax + 45 39273801 Tel + 45 30273800 shafer-n@symbion.dk From owner-proteins@net.bio.net Wed Mar 04 22:00:00 1998 Path: biosci!agate!newsfeed.kornet.nm.kr!news.maxwell.syr.edu!fu-berlin.de!news.tu-darmstadt.de!News.Uni-Marburg.DE!not-for-mail From: Marco Bocola Newsgroups: bionet.molbio.proteins Subject: Re: Ligand Binding Date: 5 Mar 1998 17:36:00 GMT Organization: HRZ Uni Marburg Lines: 24 Message-ID: <34FEE20C.6B4EE985@mailer.uni-marburg.de> References: <6d27os$nla$1@agate.berkeley.edu> NNTP-Posting-Host: pc1661.pharmazie.uni-marburg.de Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 4.03 [en] (WinNT; I) To: Louis Hom Louis Hom wrote: > Basic question: > Is there an activation energy associated with ligand binding? > There has to be, right? Since you have to displace solvent? > This came up after I was talking to an undergrad about basic > enzyme behavior and the effects on the activation energy of the catalyzed > reaction and I got to thinking about the reaction that catalyzes the > reaction. > meta-reactions. > -- > _______________________________________________________________________________ > Lou Hom >K '93 > lhom@nature.berkeley.edu > http://www.ocf.berkeley.edu/~lhom Don't forget about conformational changes of proteins (receptors,channels etc.) during the binding process (induced fit !!!) which are believed to play the central role in binding thermodynamics besides the solvation-desolvation process. From owner-proteins@net.bio.net Wed Mar 04 22:00:00 1998 Path: biosci!agate!newsfeed.kornet.nm.kr!nntp.kreonet.re.kr!news.maxwell.syr.edu!fu-berlin.de!news.tu-darmstadt.de!News.Uni-Marburg.DE!not-for-mail From: Marco Bocola Newsgroups: bionet.molbio.proteins Subject: Re: Ligand Binding Date: 5 Mar 1998 17:35:40 GMT Organization: HRZ Uni Marburg Lines: 19 Message-ID: <34FEE1F7.F648B92@mailer.uni-marburg.de> References: <6d27os$nla$1@agate.berkeley.edu> <34FAF8FC.3E4D@le.ac.uk> NNTP-Posting-Host: pc1661.pharmazie.uni-marburg.de Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 4.03 [en] (WinNT; I) Dr E. Buxbaum wrote: > Louis Hom wrote: > > > > Basic question: > > Is there an activation energy associated with ligand binding? > > Yes, there is, and it is easy to measure. Simply determine the Kd at > various temperatures and plot ln(Kd) vs the reciprocal temperature > (absolute temperature, i.e. in Kelvin). If certain conditions are met, > then the data points will be on a straight line, from the slope of which > you can calculate the activation energy. under the assumption that the heat capacity of the system, the local environment and solvation and furthermore the conformation and folding of the protein is temperature independent...... From owner-proteins@net.bio.net Wed Mar 04 22:00:00 1998 Path: biosci!agate!logbridge.uoregon.edu!news-feed.inet.tele.dk!bofh.vszbr.cz!news.eecs.umich.edu!news.niehs.nih.gov!not-for-mail From: Mara Casar Newsgroups: bionet.molbio.proteins Subject: Re: Staining of PVDF after transfer? Date: Thu, 05 Mar 1998 11:56:07 -0500 Organization: NIEHS Lines: 32 Message-ID: <34FED927.35B4@niehs.nih.gov> References: <34FC0E1D.D33FCCE3@mpih-frankfurt.mpg.de> NNTP-Posting-Host: ecarballo-jane.niehs.nih.gov Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 3.02 (Macintosh; I; PPC) You can also stain a blot AFTER doing your Western (ECL, HRP, etc.) detection. Use Coomassie G-250, not R-250 - R is the one you use for gels. Make up the stain and destains the same way as you would for gels, but then stain your blot for just a few minutes. It's not reversible, though, so it's best to do it last. -Mara ------ Junghans wrote: > > Hi > I have a problem with staining of PVDF-membranes after western-blot > transfer. > How can I stain the marker or in general my proteins before > immunodetection. I tried PonceauS but this didn't work even after adding > some methanol the result was not very promising, because only very > strong bands were recognized. > Does anybody know a more sensitive method? > Thanks, Dirk Junghans > -- > ************************************** > Dirk Junghans > Max-Planck-Institut f. Hirnforschung > Abt. Neurochemie > Deutschordenstr. 46 > 60528 Frankfurt am Main > > e-mail: junghans@mpih-frankfurt.mpg.de > Tel. ++49 - 69-96769-457 > Fax. ++49 - 69-96769-441 > From owner-proteins@net.bio.net Wed Mar 04 22:00:00 1998 Path: biosci!newshost.lanl.gov!news.ttu.edu!usenet From: "Benan Dincturk" Newsgroups: bionet.molbio.proteins Subject: Best expression system for (plant) protein expression in yeast Date: 5 Mar 1998 17:41:58 GMT Organization: Texas Tech Academic Computing Services Lines: 11 Message-ID: <01bd485d$5d6e25a0$5d227681@chm407-1.chem.ttu.edu> NNTP-Posting-Host: chm407-1.chem.ttu.edu X-Newsreader: Microsoft Internet News 4.70.1161 Hello netters E. coli expression does not seem to work for my huge protein. So while I am playing with E. coli vectors, I plan to start with yeast expression. Who has experience with yeast? Can you suggest some vectors with strong promoters? What are the important points in yeast transformation? I know that is the thoughest part usually. I am a beginner so every suggestion will be appreciated. Thank you all. Benan From owner-proteins@net.bio.net Thu Mar 05 22:00:00 1998 Path: biosci!agate!logbridge.uoregon.edu!news-peer.gip.net!news-raspail.gip.net!news.gsl.net!gip.net!newscore.univie.ac.at!news-ge.switch.ch!news.unige.ch!news From: "Andreas Mühlemann" Newsgroups: bionet.molbio.proteins Subject: How can I isolate targets of proteins (cytosolic) ? Date: Fri, 6 Mar 1998 13:40:47 +0100 Organization: University of Geneva Lines: 9 Message-ID: <6doqrc$mdo@uni2f.unige.ch> NNTP-Posting-Host: 129.194.52.82 X-Newsreader: Microsoft Outlook Express 4.72.2106.4 X-MimeOLE: Produced By Microsoft MimeOLE V4.72.2106.4 Hello ! I'm working on a neuronal cytosolic protein and I'd like to isolate its unknown target in vivo. Any ideas on the approach I could choose ? Tanx. Andreas. From owner-proteins@net.bio.net Thu Mar 05 22:00:00 1998 Path: biosci!agate!logbridge.uoregon.edu!news.maxwell.syr.edu!newsxfer3.itd.umich.edu!rill.news.pipex.net!pipex!server1.netnews.ja.net!bham!med1018.bham.ac.uk!user From: noone@cancer.bham.ac.uk (noone) Newsgroups: bionet.molbio.proteins Subject: Re: How can I isolate targets of proteins (cytosolic) ? Date: Fri, 06 Mar 1998 14:55:10 +0000 Organization: noone Lines: 22 Message-ID: References: <6doqrc$mdo@uni2f.unige.ch> NNTP-Posting-Host: med1018.bham.ac.uk In article <6doqrc$mdo@uni2f.unige.ch>, "Andreas Mühlemann" wrote: > Hello ! > I'm working on a neuronal cytosolic protein and I'd like to > isolate its unknown target in vivo. > Any ideas on the approach I could choose ? > > Tanx. > Andreas. Hi Andreas, I think some of the usual approaches are Immunoprecipitation Yeast two hybrid Expression of tagged protein and pull-out assay BiaCore Peter From owner-proteins@net.bio.net Thu Mar 05 22:00:00 1998 Path: biosci!agate!logbridge.uoregon.edu!howland.erols.net!fastnet!ptdnetP!newsgate.ptd.net!rill.news.pipex.net!pipex!server1.netnews.ja.net!bham!med1018.bham.ac.uk!user From: noone@cancer.bham.ac.uk (noone) Newsgroups: bionet.molbio.proteins Subject: subcellular fractionation Date: Fri, 06 Mar 1998 14:57:47 +0000 Organization: noone Lines: 9 Message-ID: NNTP-Posting-Host: med1018.bham.ac.uk Hi, I'm looking for a reliable method for subcellular fractionation (samples for Western) of adherent cells (like human kidney cells, 293's etc.) Any suggestions would be appreciated, Peter p.weber@bham.ac.uk From owner-proteins@net.bio.net Thu Mar 05 22:00:00 1998 Path: biosci!agate!newsgate.duke.edu!interpath.net!nntp.news.xara.net!xara.net!server5.netnews.ja.net!server3.netnews.ja.net!server1.netnews.ja.net!news.nott.ac.uk!pmbfjd0.nottingham.ac.uk!user From: mbzrl@mbn1.biochem.nottingham.ac.uk (Rob) Newsgroups: bionet.molbio.proteins Subject: Disappearing His-tagged proteins Followup-To: bionet.molbio.proteins Date: 6 Mar 1998 11:50:18 GMT Organization: University of Nottingham Lines: 8 Message-ID: NNTP-Posting-Host: pmbfjd0.nottingham.ac.uk Hi Has anyone experienced his-tagged proteins apparently 'disappearing' on FPLC gel filtration columns (eg Superose 6 or 12 columns). I assume they're coming out of solution, but it's happened now for the last 2 his-tagged proteins I've tried to purify but doesn't happen for the GST-tagged equivalents.... Very strange! Thanks in advance Rob From owner-proteins@net.bio.net Thu Mar 05 22:00:00 1998 Path: biosci!agate!howland.erols.net!news-peer.sprintlink.net!news.sprintlink.net!Sprint!newsfeed.nacamar.de!news-kar1.dfn.de!news-fra1.dfn.de!news-ber1.dfn.de!news-ham1.dfn.de!news.mu-luebeck.de!not-for-mail From: "Lars Komorowski" Newsgroups: bionet.molbio.methds-reagnts,bionet.molbio.proteins,bionet.software Subject: Proposal for a new bionet newsgroup Date: Fri, 6 Mar 1998 09:13:33 +0100 Organization: Med. Universitaet zu Luebeck Lines: 7 Message-ID: <6dobf1$4cj$1@gwsun.medinf.mu-luebeck.de> NNTP-Posting-Host: 141.83.167.168 X-Newsreader: Microsoft Outlook Express 4.71.1712.3 X-MimeOLE: Produced By Microsoft MimeOLE V4.71.1712.3 Xref: biosci bionet.molbio.methds-reagnts:65549 bionet.molbio.proteins:12408 bionet.software:20661 I propose to set up a new newsgroup for used lab euipment. Who is interested ? Lars Komorowski Ins. f. Biochemie Med. Univ. Luebeck, Germany From owner-proteins@net.bio.net Sat Mar 07 22:00:00 1998 Path: biosci!rutgers!uwm.edu!news1.chicago.iagnet.net!qual.net!iagnet.net!btnet-peer!btnet!newsfeed.metronet.de!news.metronet.de!not-for-mail From: "J. Jockers" Newsgroups: bionet.microbiology,bionet.molbio.methds-reagnts,bionet.molbio.proteins Subject: Problems with beta-Galactosidase and X-Gal Followup-To: bionet.microbiology Date: Mon, 9 Mar 1998 01:27:25 +0100 Organization: Metronet Lines: 39 Message-ID: <6dvd15$74m$1@news.metronet.de> NNTP-Posting-Host: kaiserslautern1.pop.metronet.de X-Newsreader: Microsoft Outlook Express 4.71.1712.3 X-MimeOLE: Produced By Microsoft MimeOLE V4.71.1712.3 Xref: biosci bionet.microbiology:12663 bionet.molbio.methds-reagnts:65596 bionet.molbio.proteins:12413 Hello! I am searching for a method to examine funghi for extracellular beta-galactosidase activity. Right now I am using X-Gal (5-bromo-4-chloro-3-indonyl-beta-D-galactopyanoside) as substrate for this enzyme. I add a (low concentrated) X-Gal solution to the medium of the funghi. 24 hours later I check the medium for blue coloring. I think that I see blue enzyme-pigment complexes in the medium, because of the unsolubility of the blue complex. Well, at least it DOES work, but the results are unsufficient reproducible, quantitative and reliable. I know that the use of X-Gal in this case is rather unusual, but I saw no other way to carry out this task ia a easy and simple way. - I know that X-Gal is actual unsoluble in water. But what about the blue complex after X-Gal has been spit by the enzyme? - Does anybody know a "smart" method to do this thing another way or improve the reliability of this test? - Does anybody know something about the stability of aquatic X-Gal-solutions? (In my opinion they degrade quite rapidly even at storage temperatures of 2-8 degrees celsius) I would be very happy for any tips and hints that could help me to solve this problems! thank you very much! Jochen, Germany (please forgive me my very awkward English!) <- is this correct, after all? :-) From owner-proteins@net.bio.net Sun Mar 08 22:00:00 1998 Path: biosci!agate!howland.erols.net!vixen.cso.uiuc.edu!Lightyear From: Lightyear@uiuc.edu.nospam (Matt) Newsgroups: bionet.molbio.proteins Subject: Re: Best expression system for (plant) protein expression in yeast Date: Mon, 09 Mar 1998 13:19:51 -0600 Organization: US Robotics and Mechanical Men Lines: 31 Message-ID: References: <01bd485d$5d6e25a0$5d227681@chm407-1.chem.ttu.edu> NNTP-Posting-Host: bushlab1.life.uiuc.edu X-Newsreader: MT-NewsWatcher 2.3.5 X-Face: $VCBL>c'm[sO:~:%UH#A--p2Mjs163<#Yal[|)8@\?Yj"&7`l(h:-VDzvSu3! 2AU,zEj|^Fu`Igw%qgr'0e)V:pp@$OG&,gIYy>Z"$VO#CX0"&:4vH|sekTYL[r"tT} Lambda-YES is a great single-copy plasmid with a Gal-Inducible Promoter and a URA marker.. we use it all the time here. As far as transformation is concerned, electroporation a la E. coli is fairly effective.. we have a protocol or two if you're interested. Matt Vaughn In article <01bd485d$5d6e25a0$5d227681@chm407-1.chem.ttu.edu>, "Benan Dincturk" wrote: > Hello netters > > E. coli expression does not seem to work for my huge protein. So while I am > playing with E. coli vectors, I plan to start with yeast expression. Who > has experience with yeast? Can you suggest some vectors with strong > promoters? What are the important points in yeast transformation? I know > that is the thoughest part usually. I am a beginner so every suggestion > will be appreciated. Thank you all. > > Benan In theory, there is no difference between theory and practice; in practice, however, there is no similarity Matthew Vaughn University of Illinois Department of Biology 190 ERML 1201 W. Gregory Drive Urbana, IL 61801 PS: You know the part about removing the nospam from the end of my email don't you? I hate to do it but I have gotten some TRULY XXX offensive spams lately and can't tolerate them anymore. From owner-proteins@net.bio.net Sun Mar 08 22:00:00 1998 Path: biosci!agate!newsgate.duke.edu!nntp-out.monmouth.com!newspeer.monmouth.com!ais.net!uunet!in4.uu.net!hearst.acc.Virginia.EDU!murdoch.acc.Virginia.EDU!not-for-mail From: John S Walker Newsgroups: bionet.molbio.proteins Subject: "Halos" in Western Blots; What are they? Date: Mon, 09 Mar 1998 16:57:01 -0500 Organization: University of Virginia Lines: 23 Message-ID: <350465AD.5DED9FB5@virginia.edu> NNTP-Posting-Host: mars.med.virginia.edu Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 3.01GoldC-Caldera (X11; I; Linux 2.0.30 i486) G'day, I'm a newbie to doing Western blots. At present I'm trying to optimise a technique for quantifying the percentage phosphorylation of a protein. The protocol i'm following consists of an isoelectric focusing gel that is then transferred to a nitrocellulose membrane. The membrane is then exposed to primary and secondary antibodies. The detection system uses enhanced chemiluminescence with a horseradish peroxidase linked secondary. The light is detected using x-ray film. At present we are having problems with linearity of response and a steep response curve (i.e. we only go from detection limit to saturation over a five-fold range of protein concentrations) In addition, and this brings me to my question, we have halo's around the detection bands, i.e. a light band just above the more heavily stained 'normal' location of the band. I've read some accounts that say that this is due to protein overloading and some that say it's due to excessive antibody. Can anybody give me a clear answer as to what causes these halo's and why? I don't think that this is a 'real' band. John Walker Dept of Physiology University of Virginia From owner-proteins@net.bio.net Sun Mar 08 22:00:00 1998 Path: biosci!NIC.BMI.AC.CN!zhangzg From: zhangzg@NIC.BMI.AC.CN ("Fei WANG") Newsgroups: bionet.molbio.proteins Subject: help to concentration Date: 9 Mar 1998 00:32:51 -0800 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 4 Sender: daemon@net.bio.net Distribution: world Message-ID: <01bd4b35$7f0d7460$ae247aca@pupo> NNTP-Posting-Host: net.bio.net hi, every bodies. i want to concentrate a protein for medicine use,for injection. i want a method not increase the pyrogen level or DNA content in final product and at the same time to get the protein product to 2 mg/ml. From owner-proteins@net.bio.net Sun Mar 08 22:00:00 1998 Path: biosci!agate!news.ucsc.edu!tjandra From: tjandra@darwin.ucsc.edu (hT) Newsgroups: bionet.molbio.proteins,bionet.molbio.methds-reagnts Subject: pCAL or CaM-binding Peptide fusion proteins Date: Mon, 09 Mar 1998 15:01:03 -0700 Organization: biology Lines: 5 Message-ID: NNTP-Posting-Host: kellogg-lab1.ucsc.edu Mime-Version: 1.0 Content-Type: text/plain; charset=ISO-8859-1 Content-Transfer-Encoding: 8bit X-Newsreader: Yet Another NewsWatcher 2.4.0 Xref: biosci bionet.molbio.proteins:12418 bionet.molbio.methds-reagnts:65620 Hi Everyone: I am soliciting comments from those who have used the pCAL-n/c plasmid that is available from Stragene, San Diego. i am wondering if there is any good and bad thing* I should know of before I order the plasmid. Please cc your comments to tjandra@darwin.ucsc.edu and i will summarize any 'interesting' point in a later posting. Many thanks. Hendri From owner-proteins@net.bio.net Sun Mar 08 22:00:00 1998 Path: biosci!CC.USU.EDU!sl86b From: sl86b@CC.USU.EDU (Ben) Newsgroups: bionet.molbio.proteins Subject: Gigapite Column Date: 9 Mar 1998 13:47:14 -0800 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 14 Sender: daemon@net.bio.net Distribution: world Message-ID: NNTP-Posting-Host: net.bio.net Is there anyone out there who knows what a Gigapite column is, and if so who manufactures it? Came across a paper by a Japanese group which used the column to purify a lipase. Thanks, Benjamin Dias SL86B@cc.usu.edu Dept. of Nutrition and Food Science Utah State University, Logan, UT 84322 (801) 797 2123 From owner-proteins@net.bio.net Sun Mar 08 22:00:00 1998 Path: biosci!newshost.lanl.gov!news.ttu.edu!usenet From: "Benan Dincturk" Newsgroups: bionet.molbio.proteins Subject: Re: Best expression system for (plant) protein expression in yeast Date: 9 Mar 1998 23:45:23 GMT Organization: Texas Tech Academic Computing Services Lines: 13 Message-ID: <01bd4bb4$b008b8c0$5d227681@chm407-1.chem.ttu.edu> References: <01bd485d$5d6e25a0$5d227681@chm407-1.chem.ttu.edu> NNTP-Posting-Host: chm407-1.chem.ttu.edu X-Newsreader: Microsoft Internet News 4.70.1161 Matt wrote in article ... > Lambda-YES is a great single-copy plasmid with a Gal-Inducible Promoter > and a URA marker.. we use it all the time here. As far as transformation > is concerned, electroporation a la E. coli is fairly effective.. we have > a protocol or two if you're interested. > > Matt Vaughn > > Yes, I am interested. Can you possibly send me the vector? From owner-proteins@net.bio.net Mon Mar 09 22:00:00 1998 Path: biosci!agate!howland.erols.net!news-peer.gip.net!news-lond.gip.net!news.gsl.net!gip.net!peer.news.u-net.net!u-net!news.u-net.com!not-for-mail From: arnoud@vanvliet.u-net.com (Arnoud van Vliet) Newsgroups: bionet.molbio.proteins Subject: Re: His-tagged protein and SDS-PAGE migration Date: Tue, 10 Mar 1998 16:43:37 GMT Organization: (Posted Via) U-NET Ltd. Lines: 25 Message-ID: <35056d1f.799515@news.u-net.com> References: <350540de.110595@news.columbia.edu> Reply-To: arnoud@vanvliet.u-net.com NNTP-Posting-Host: p112.nas2.is3.u-net.net X-Newsreader: Forte Free Agent 1.1/32.230 Hi, I don't know the system, but if it is a 6-His system, then it can run a bit higher on SDS-PAGE. My 10 and 14 kDa proteins (low histidine content) ran at 16 and 20 kDa, respectively. However, there are exceptions to the rule, as a third protein I expressed (high histidine content) ran at the expected MW. So summarizing: yes, up to 6 kDa higher (consult the QiaExpress manual from Qiagen). Hope this helps Arnoud On Tue, 10 Mar 1998 13:33:45 GMT, ct133@columbia.edu wrote: >Hi, > >Does anybody know whether his-tagged protein expressed from pRSET >vector (Invitrogen) can migrate aberrantly on SDS-PAGE? > >Thanks, > >C. Tunyaplin From owner-proteins@net.bio.net Mon Mar 09 22:00:00 1998 Path: biosci!agate!howland.erols.net!news-peer.gip.net!news.gsl.net!gip.net!news.new-york.net!news.columbia.edu!news-not-for-mail From: ct133@columbia.edu Newsgroups: bionet.molbio.proteins Subject: His-tagged protein and SDS-PAGE migration Date: Tue, 10 Mar 1998 13:33:45 GMT Organization: Columbia University Lines: 8 Message-ID: <350540de.110595@news.columbia.edu> NNTP-Posting-Host: tw116cct.cpmc.columbia.edu Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Newsreader: Forte Agent 1.5/32.478 Hi, Does anybody know whether his-tagged protein expressed from pRSET vector (Invitrogen) can migrate aberrantly on SDS-PAGE? Thanks, C. Tunyaplin From owner-proteins@net.bio.net Mon Mar 09 22:00:00 1998 Path: biosci!agate!logbridge.uoregon.edu!news-peer.gip.net!news.gsl.net!gip.net!news-peer.sprintlink.net!news.sprintlink.net!Sprint!newsfeed.nacamar.de!fu-berlin.de!news.tu-darmstadt.de!News.Uni-Marburg.DE!not-for-mail From: Marco Bocola Newsgroups: bionet.molbio.proteins Subject: E280 Protein quantitation on microplates ?! Date: 10 Mar 1998 19:22:11 GMT Organization: HRZ Uni Marburg Lines: 25 Message-ID: <35059283.FB7F6CF3@mailer.uni-marburg.de> NNTP-Posting-Host: pc1661.pharmazie.uni-marburg.de Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 4.03 [en] (WinNT; I) We build up a new biochemical workgroup and looking foorward to by an UV/VIS-Microplate Reader. Has anyone experience with UV-Microplates and especially Protein determination ? (Unfortunately we have no "normal" UV/VIS Photometer) Is it possible to measure quantitative absorption with good accuracy for small volumes? Thank you for telling me your experiences or suggestions, Marco Bocola -- Marco Bocola Inst. f. pharmaz. Chemistry AG Prof. Klebe Tel: +49-6421-28-5071 Marbacher Weg 6 Fax: -8994 D-35032 Marburg NEW !!! e-mail:bocola@mailer.uni-marburg.de NEW !!! :-( There are no problems, just solutions !? :-) From owner-proteins@net.bio.net Mon Mar 09 22:00:00 1998 Path: biosci!STUDENT1.MAHIDOL.AC.TH!g3937506 From: g3937506@STUDENT1.MAHIDOL.AC.TH (Jongrak Kittiworakarn) Newsgroups: bionet.molbio.proteins Subject: Re:Gigapite Column Date: 10 Mar 1998 19:13:54 -0800 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 23 Sender: daemon@net.bio.net Distribution: world Message-ID: <3505FFB0.C6F2F34D@student.mahidol.ac.th> NNTP-Posting-Host: net.bio.net Hi Benjamin: Gigapite is hydroxy apatite, Ca18 (PO4)6 (OH)2. You may contact TOAGOSEI CHEMICAL INDUSTRY CO., LTD> 14-1 Nishi Shinbashi, 1-chome, Minoto-ku, Tokyo 105 JAPAN Tel: 03-597-7739 Fax: 03-597-7217 Hope this help, Jongrak -- -----------------------------------Mr. Jongrak Kittiworakarn g3937506@student.mahidol.ac.th Inst. of Science and Technology for Research and Development Mahidol University (Salaya Campus) Salaya, Nokornprathom, THAILAND. Tel. 001-66-2-441-9003-7 ext.1277,1278,1279 Fax. 001-66-2-441-9906, 001-66-2-441-1013 ----------------------------------------------------------------- From owner-proteins@net.bio.net Mon Mar 09 22:00:00 1998 Path: biosci!agate!logbridge.uoregon.edu!news-peer.gip.net!news.gsl.net!gip.net!news.new-york.net!news.columbia.edu!news-not-for-mail From: ct133@columbia.edu Newsgroups: bionet.molbio.proteins Subject: Re: His-tagged protein and SDS-PAGE migration Date: Wed, 11 Mar 1998 01:18:06 GMT Organization: Columbia University Lines: 14 Message-ID: <3505e5df.560449@news.columbia.edu> References: <350540de.110595@news.columbia.edu> <35056d1f.799515@news.u-net.com> NNTP-Posting-Host: tw116cct.cpmc.columbia.edu Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Newsreader: Forte Agent 1.5/32.478 >I don't know the system, but if it is a 6-His system, then it can run >a bit higher on SDS-PAGE. My 10 and 14 kDa proteins (low histidine >content) ran at 16 and 20 kDa, respectively. However, there are >exceptions to the rule, as a third protein I expressed (high histidine >content) ran at the expected MW. > >So summarizing: yes, up to 6 kDa higher (consult the QiaExpress manual >from Qiagen). The tag should be about 6 kD. My protein is 64 kD so I'm expecting a 70 kD protein. But what I saw on the gel runs at ~90 kD. This is a bit too much to be explained by the tag, isn't it? C. Tunyaplin From owner-proteins@net.bio.net Tue Mar 10 22:00:00 1998 Path: biosci!TC3NET.COM!cola From: cola@TC3NET.COM (cola) Newsgroups: bionet.molbio.proteins Subject: growth hormone Date: 11 Mar 1998 15:13:36 -0800 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 4 Sender: daemon@net.bio.net Distribution: world Message-ID: <35071AB3.C910BBBB@tc3net.com> NNTP-Posting-Host: net.bio.net I have isolate a putative growth hormone and would like to test my sample for activity. Does anyone know of a cell line or other test that would confirm the sample as being growth hormone? Thanks Cola From owner-proteins@net.bio.net Tue Mar 10 22:00:00 1998 Path: biosci!axisgenetics.co.uk!KH From: KH@axisgenetics.co.uk (Koen Hellendoorn) Newsgroups: bionet.molbio.proteins Subject: C-terminal protein sequencing Date: 11 Mar 1998 03:10:35 -0800 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 8 Sender: daemon@net.bio.net Distribution: world Message-ID: NNTP-Posting-Host: net.bio.net Hello, Can anyone tell me if there is any institute or company (preferably in the UK/ other Europe) that can do C-terminal sequencing of proteins? Thanks, Koen From owner-proteins@net.bio.net Tue Mar 10 22:00:00 1998 Newsgroups: bionet.molbio.methds-reagnts,bionet.molbio.proteins,bionet.software Path: biosci!agate!spool.mu.edu!uwm.edu!chicago-news-feed2.bbnplanet.com!cpk-news-hub1.bbnplanet.com!news.bbnplanet.com!news-peer.gip.net!news-lond.gip.net!news.gsl.net!gip.net!peer.news.u-net.net!u-net!yama.mcc.ac.uk!liv!news From: GcR Subject: Re: Proposal for a new bionet newsgroup Content-Type: text/plain; charset=us-ascii Message-ID: <35066A66.2B70@liv.ac.uk> Sender: news@liverpool.ac.uk (News System) NNTP-Posting-Host: ash-35.liv.ac.uk Content-Transfer-Encoding: 7bit Organization: The University of Liverpool References: <6dobf1$4cj$1@gwsun.medinf.mu-luebeck.de> Mime-Version: 1.0 Date: Wed, 11 Mar 1998 10:41:42 GMT X-Mailer: Mozilla 3.02 (X11; I; SunOS 5.5.1 sun4c) Lines: 30 Xref: biosci bionet.molbio.methds-reagnts:65689 bionet.molbio.proteins:12426 bionet.software:20692 Lars Komorowski wrote: > > I propose to set up a new newsgroup for used lab euipment. Who is interested > ? > Lars Komorowski > Ins. f. Biochemie > Med. Univ. Luebeck, Germany As I can recall, a few months ago somebody came up with a similar idea, he made a webpage specially for the (re)sale of used lab equipment. You can find it at http://www.geocities.com/ResearchTriangle/Lab/5656/ Sincerely, GcR ---------------------------------------------\/----- Gert Roosen /\ PhD Student /--\ Department of Haematology (====\ Royal Liverpool University Hospital \----) Duncan Building 2nd floor \--/ Prescot Street L69 3BX Liverpool \/ + 44 (0)151 706 4326 (phone) /\ + 44 (0)151 706 4311 (fax) /--\ g.roosen@liv.ac.uk (email) /====) -------------------------------------------(----/--- From owner-proteins@net.bio.net Tue Mar 10 22:00:00 1998 Path: biosci!agate!logbridge.uoregon.edu!news.maxwell.syr.edu!newsfeed.internetmci.com!152.163.199.19!portc03.blue.aol.com!audrey01.news.aol.com!not-for-mail From: alpies@aol.com (AlPiEs) Newsgroups: bionet.molbio.proteins Subject: is it the same protein? Date: 11 Mar 1998 13:16:12 GMT Lines: 9 Message-ID: <19980311131600.IAA08911@ladder03.news.aol.com> NNTP-Posting-Host: ladder03.news.aol.com X-Admin: news@aol.com Organization: AOL http://www.aol.com We are studying a human protein that we suspect has a tendency to aggregate in vivo. After size exclusion chromatography and Western blot analysis of the fractions with polyclonal antibodies,there are two strong signals: one in the first fraction, and one in later fraction. Presumably the first fraction contains an aggregated form of the protein. My question is: how do we show that the protein present in both fractions is the same? We do not want to rely on the antibodies only, and we cannot do microsequencing, the amounts are too low. Limited proteolysis is not agood idea either if it is the same protein in different conformations. Any suggestions, please? From owner-proteins@net.bio.net Tue Mar 10 22:00:00 1998 Path: biosci!ruf.uni-freiburg.de!frnicole From: frnicole@ruf.uni-freiburg.de (Nicole Frankenberg) Newsgroups: bionet.molbio.proteins Subject: Svedberg units Date: 11 Mar 1998 03:39:03 -0800 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 4 Sender: daemon@net.bio.net Distribution: world Message-ID: <350685CF.87B75362@sun2.ruf.uni-freiburg.de> NNTP-Posting-Host: net.bio.net Does anybbody know where to get svedberg units for standard proteins. any idea of a book or article is appreciated. Thanks for your help.Nicole From owner-proteins@net.bio.net Tue Mar 10 22:00:00 1998 Path: biosci!agate!newsgate.duke.edu!nntp-out.monmouth.com!newspeer.monmouth.com!news.maxwell.syr.edu!nntp.news.xara.net!xara.net!server5.netnews.ja.net!server3.netnews.ja.net!news.icnet!NewsWatcher!user From: I.McFarlane@icrf.icnet.uk (Ian McFarlane) Newsgroups: bionet.molbio.proteins Subject: Re: is it the same protein? Date: Wed, 11 Mar 1998 17:58:17 +0000 Organization: Imperial Cancer Research Fund Lines: 21 Message-ID: References: <19980311131600.IAA08911@ladder03.news.aol.com> NNTP-Posting-Host: 143.65.52.61 If you run SDS-PAGE with mercaptoethanol you should only get one band on your Western. If this doesn't work, some protein complexes will resist the treatment urea can be added to the sample buffer and gel. Ian Mc In article <19980311131600.IAA08911@ladder03.news.aol.com>, alpies@aol.com (AlPiEs) wrote: > We are studying a human protein that we suspect has a tendency to aggregate in > vivo. After size exclusion chromatography and Western blot analysis of the > fractions with polyclonal antibodies,there are two strong signals: one in the > first fraction, and one in later fraction. Presumably the first fraction > contains an aggregated form of the protein. My question is: how do we show > that the protein present in both fractions is the same? We do not want to rely > on the antibodies only, and we cannot do microsequencing, the amounts are too > low. Limited proteolysis is not agood idea either if it is the same protein in > different conformations. Any suggestions, please? From owner-proteins@net.bio.net Tue Mar 10 22:00:00 1998 Path: biosci!agate!howland.erols.net!newshub.northeast.verio.net!nntp.upenn.edu!msunews!not-for-mail From: Jonathan Walton Newsgroups: bionet.molbio.proteins Subject: carbon content of proteins? Date: Wed, 11 Mar 1998 11:34:29 -0500 Organization: Michigan State University Lines: 10 Message-ID: <3506BD15.5F0F@pilot.msu.edu> NNTP-Posting-Host: 35.8.197.111 Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 3.0 (Win95; I) X-MD5: 859d1742a9f46ed064e94ba21b450691 Can anyone tell me a generally accepted average value for the % (by weight) of carbon in proteins? The value of 15-16% for N is generally used, but how about carbon? And (related question that would give the answer) is there is a program that will calculate the elemental composition of a protein? Thanks, Jon Walton Michigan State Univ. walton@pilot.msu.edu From owner-proteins@net.bio.net Tue Mar 10 22:00:00 1998 Path: biosci!agate!newsfeed.wli.net!newsfeed.ecrc.net!newsfeed.ecrc.net!news.duesseldorf.ecrc.net!RRZ.Uni-Koeln.DE!usenet From: "Prof. Dr. Dietmar Schomburg" Newsgroups: bionet.molbio.proteins Subject: Conference in Bioinformatics Date: Wed, 11 Mar 1998 10:33:43 +0100 Organization: Universitaet zu Koeln Lines: 198 Message-ID: <35065A77.9E9F9E07@uni-koeln.de> NNTP-Posting-Host: 134.95.151.4 Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 4.03 [de] (Win95; I) GCB 98, Conference on Bioinformatics FIRST ANNOUNCEMENT AND CALL FOR PAPERS COMPUTER SCIENCE AND BIOLOGY German Conference on BIOINFORMATICS GCB '98 Cologne University October 7-10, 1998 University of Cologne, Institute of Biochemistry German Society for Molecular Biology and Biochemistry (GBM) German Society for Computer Science (GI) German Society of Chemistry (GDCh) German Society for Chemical Apparatus, Chemical Engineering and Biotechnology (DECHEMA) German Society for Medical Informatics, Biometry and Epidemiology (GMDS) The conference focuses on the following topics: * Comprehensive Genome Analysis * Comparative Genome Analysis * Protein Structure and Function * Drug and Protein Design * Simulation of biological Molecules * Gene and Target Discovery * Sequence Data Analysis * Sequence/Structure/Function Relations * Evolution of Genomes * Intelligent Biological Databases * Software for Genome Analysis ORGANIZING COMMITTEE: D. Schomburg (U Koeln) Chair + U. Lessel (U Koeln) SCIENTIFIC COMMITTEE: T. Lengauer (GMD St. Augustin), W. Mewes (MIPS, Martinsried), D. Schomburg (Univ. Koeln), M. Vingron (DKFZ Heidelberg), E. Wingender (GBF, Braunschweig) The German Conference on Bioinformatics (GCB-98) will present the state of the art in Molecular Bioinformatics. It is an international conference within the tradition of the German Workshop 'Computers in Bioscience' created more than 10 years ago. GCB'98 focuses on the bioinformatics support research in the modern molecular biosciences. The conference will present the current research in the theory and application of computational methods to genomic, sequence, and structural information. The conference language is English. Participation is invited from the scientific community active in the development of concepts and applications to the evaluation of genomic information, analysis and prediction of protein structure, and design of drugs and proteins. Submissions in the form of full papers or extended abstracts (ca. three pages) for lectures, or abstracts (1-2 pages) for posters and demos should reflect recent work to be presented to an audience of interested scientists and industrial researchers. Selected papers and presentations are rigorously peer reviewed and are published in a special issue of CABIOS. The scientific committee will evaluate submissions and recommend papers for publication in CABIOS. These manuscripts are subject to the ordinary CABIOS review procedure (3 referees). A special section for the demonstration of software in form of a plenary session will adjourn the conference. Full Papers should be 12 pages, single-spaced and set in 12 point type, including title, abstract, figures, tables, and bibliography. Submissions to be considered for CABIOS publication must follow authors instructions of that journal. Poster and software demonstration abstracts should be max. 2 pages. Postal and electronic mailing addresses, telephone, and fax numbers must be included. Processing of submissions is facilitated by sending postscript files via e-mail to D.Schomburg@uni-koeln.de The conference takes place in a beautiful historical place, Kardinal-Schulte-Haus. The castle-like conference centre is surrounded by a park and located at the outskirts of a hilly area near Cologne and overlooking the city and its environment. Access from the Cologne main station or international airports Koeln/Bonn or Duesseldorf is convenient (ca. 30 min. from Cologne central station). Accommodation is provided in the conference centre or, after the rooms in the Kardinal-Schulte-Haus are booked, in a nearby hotel. The number of participants is limited to 200. Please make your reservation as soon as possible. Speakers and presenters of accepted posters will have preference, all other places will be given in order of the reception of the registration. For more, up-to-date information see our WWW Page: http://www.uni-koeln.de/math-nat-fak/biochemie/GCB98.htm DEADLINES: for Abstracts or extended Abstracts: June 30. for Registration August 31. REGISTRATION FORM: Name/Title: .............................................................................................................. First Name: .............................................................................................................. Organisation: .............................................................................................................. Organisation: .............................................................................................................. Street: .............................................................................................................. City: .............................................................................................................. Phone: .............................................................................................................. FAX: .............................................................................................................. E-mail: .............................................................................................................. Signature, if faxed Please delete as appropriate: I would like to submit an abstract for consideration as a Lecture/Poster Please reserve accomodation for the nights of October 7/8/9/10 (delete as appropriate) in single/double room (preferred partner?) Full Registration: DM 250 Students: DM 200 I enclose a cheque/transfer the money before September 15th RETURN TO: GCB 98 Secretariate Universitaet zu Koeln Institut fuer Biochemie Zuelpicher Strasse 47 50674 Koeln FAX +49 221/470-5092 E-Mail: D.Schomburg@uni-koeln.de Full accomodation includes rooms and meals for accomodation in the conference centre. We will handle your reservation. Please do not make advance payments for the accomodation. Single Room: DM 276 for 3 days Double Room: DM 228 for 3 days per person For registration please send a cheque payable to: GCB 98 Bioinformatics For more information contact: Ms. Sullivan Tel.: +49 221/470 6440 FAX : +49 221/470 5092 e-mail: Sullivan@uni-koeln.de ACCESS TO COLOGNE Cologne is served by a number of international airports including Cologne/Bonn, Duesseldorf, Frankfurt and even Amsterdam and Brussels. From Cologne/Bonn Airport, busses run every 15- 20 minutes to the city centre. From Duesseldorf there is a regular train service to the Main Station (Hauptbahnhof) in the centre of Cologne. From Frankfurt there is a train via Bonn to the main station in Cologne. Journey time is 2 hours). Good rail links also exist between Amsterdam and Brussels and Cologne. The central Euopean location of Cologne means that it can be easily reached by the major highways from the Netherlands, Belgium, France, Switzerland, Italy, Austria and Eastern Europe. The main highways are A1, 3, 4, 54, 57, and 61. Built on the banks of the Rhine River, Cologne is a beautiful historic city in the heart of Europe. Cologne has an immense cultural heritage, including many relics of the Roman city, both in museums and at their original sites. With the Dom Cathedral, twelve Romanesque churches, the medieval treasures of "holy Cologne", galleries of modern and avant-garde art, Cologne offers one of Europe's greatest art collections. The Opera, Playhouse, Cologne Philharmonia and more than 100 smaller theatres and cultural institutions ensure Cologne has a cultural present besides its past. From owner-proteins@net.bio.net Tue Mar 10 22:00:00 1998 Path: biosci!bloom-beacon.mit.edu!news.starnet.net!sdd.hp.com!vixen.cso.uiuc.edu!howland.erols.net!news.maxwell.syr.edu!Cabal.CESspool!bofh.vszbr.cz!lyra.csx.cam.ac.uk!hgmp.mrc.ac.uk!guardian.dcs.warwick.ac.uk!warwick!leicester!usenet From: "Dr E. Buxbaum" Newsgroups: bionet.molbio.proteins Subject: Re: is it the same protein? Date: Wed, 11 Mar 1998 14:04:05 -0800 Organization: University of Leicester (PCFS User) Lines: 15 Message-ID: <35070A55.6DAA@le.ac.uk> References: <19980311131600.IAA08911@ladder03.news.aol.com> NNTP-Posting-Host: pc47.msb.le.ac.uk Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 3.01 (Win16; I) AlPiEs wrote: > > We are studying a human protein that we suspect has a tendency to aggregate in > vivo. After size exclusion chromatography and Western blot analysis of the > fractions with polyclonal antibodies,there are two strong signals: one in the > first fraction, and one in later fraction. Presumably the first fraction > contains an aggregated form of the protein. My question is: how do we show > that the protein present in both fractions is the same? We do not want to rely > on the antibodies only, and we cannot do microsequencing, the amounts are too > low. Limited proteolysis is not agood idea either if it is the same protein in > different conformations. Any suggestions, please? 2D electrophoresis. The probability that two different proteins have the same subunit MW, the same pI and react with the same antibody should be very low indeed. From owner-proteins@net.bio.net Tue Mar 10 22:00:00 1998 Path: biosci!agate!logbridge.uoregon.edu!news.maxwell.syr.edu!Cabal.CESspool!bofh.vszbr.cz!lyra.csx.cam.ac.uk!hgmp.mrc.ac.uk!guardian.dcs.warwick.ac.uk!warwick!leicester!usenet From: "Dr E. Buxbaum" Newsgroups: bionet.molbio.proteins Subject: Re: E280 Protein quantitation on microplates ?! Date: Wed, 11 Mar 1998 13:57:05 -0800 Organization: University of Leicester (PCFS User) Lines: 28 Message-ID: <350708B1.F51@le.ac.uk> References: <35059283.FB7F6CF3@mailer.uni-marburg.de> NNTP-Posting-Host: pc47.msb.le.ac.uk Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 3.01 (Win16; I) Marco Bocola wrote: > > We build up a new biochemical workgroup and looking foorward to by an > UV/VIS-Microplate Reader. > Has anyone experience with UV-Microplates and especially Protein > determination ? > (Unfortunately we have no "normal" UV/VIS Photometer) > Is it possible to measure quantitative absorption with good accuracy for > small volumes? The first problem here is not the reader, but the plate, because normal 96 well plates are made of polystyrene, which has considerable (read: almost total) absorbance at 280 nm. Even if you could get plates made of polyacryl, the absorbance probably will be some 50 or 60%. Additionally, I do not know of a 96 well photometer which were able to produce light of such short a wavelength (you need a mercury or xenon arc bulb, not the normal tungsten filament lamps). You should be able to get that information from the specs of the instrument you want to buy, however. To measure A280 for small samples, we use a submicro quarz cuvette together with our Beckman DU-65 spectrophotometer. The minimal volume with this setup is 50 ul, the cuvette can acomodate up to 100 ul, the pathlength is 1 cm as for normal cuvettes. We use a "crystall" tip connected to a vacuum line to empty the cuvette between readings, the cuvette remains in the instrument. The speed is of course lower than with 96 well plates, but you can still read 3-4 samples per min if you are experienced. This system works very well for Warburg protein assays. From owner-proteins@net.bio.net Tue Mar 10 22:00:00 1998 Path: biosci!agate!logbridge.uoregon.edu!news.maxwell.syr.edu!newscore.univie.ac.at!Cabal.CESspool!bofh.vszbr.cz!lyra.csx.cam.ac.uk!hgmp.mrc.ac.uk!guardian.dcs.warwick.ac.uk!warwick!leicester!usenet From: "Dr E. Buxbaum" Newsgroups: bionet.molbio.proteins Subject: Re: Svedberg units Date: Wed, 11 Mar 1998 14:01:09 -0800 Organization: University of Leicester (PCFS User) Lines: 10 Message-ID: <350709A5.1F76@le.ac.uk> References: <350685CF.87B75362@sun2.ruf.uni-freiburg.de> NNTP-Posting-Host: pc47.msb.le.ac.uk Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 3.01 (Win16; I) Nicole Frankenberg wrote: > > Does anybbody know where to get svedberg units for standard proteins. > any idea of a book or article is appreciated. Analytical ultracentrifugation is a first principle technique, you do not need standard proteins for calibration. However, you should get examples from most books on ultracentrifugation. There is one in the "Practical Approach" series, and Svedbergs "The Ultracentrifuges" is still worth reading, even after 50 years. From owner-proteins@net.bio.net Tue Mar 10 22:00:00 1998 Path: biosci!agate!newsfeed.wli.net!news-xfer.netaxs.com!howland.erols.net!psinntp!ix.netcom.com!news From: Mark Atlas Newsgroups: bionet.molbio.proteins Subject: Online Auctions for used lab equipment Going, Going...Sold!!! Date: Wed, 11 Mar 1998 15:08:43 GMT Organization: Netcom Lines: 26 Message-ID: <6e69cq$2ca@dfw-ixnews11.ix.netcom.com> NNTP-Posting-Host: sjx-ca72-24.ix.netcom.com X-NETCOM-Date: Wed Mar 11 9:08:10 AM CST 1998 X-Newsreader: NETCOMplete/4.0 Online Auctions for used lab equipment Going, Going...Sold!!! Purchasing used equipment has never been easier or more safe. With Going, Going,...Sold!!! You Set the Price You evaluate the equipment in your lab before taking ownership You utilize a predeisigned escrow to insure your money You find great prices You get knowledgeable unbiased assistance to guide you with your purchase Items on the block include Beckman 6300 Waters HPLC systems ( 30%-40% of new price for a new model Detector, Unbelievable) Waters GPC system Varian 3400 GC Centrifuges Many new items coming up weekly. Stop buy and register for free. http://www.going-going-sold.com From owner-proteins@net.bio.net Tue Mar 10 22:00:00 1998 Path: biosci!agate!spool.mu.edu!uwm.edu!chicago-news-feed2.bbnplanet.com!su-news-hub1.bbnplanet.com!news.bbnplanet.com!logbridge.uoregon.edu!news.maxwell.syr.edu!news-ge.switch.ch!surfnet.nl!rl0001.unimaas.nl!not-for-mail From: "Henri M.H. Spronk" Newsgroups: bionet.molbio.proteins Subject: Expression problems Date: 11 Mar 1998 14:15:28 GMT Organization: RL Lines: 20 Message-ID: <01bd4cf8$2a1ebda0$2d187889@henri> NNTP-Posting-Host: bioch0045uns50.unimaas.nl X-Newsreader: Microsoft Internet News 4.70.1161 We are trying to express a eukaryotic cDNA in a prokaryotic expression system. The E. coli strain is M15 [pREP4] from QIAgen. The pQE40 expression vector consist of 6His-mouseDHFR and our cDNA (220 bp). Between the mouseDHFR and the cDNA we placed a factor Xa-digestion site. In the beginning we had poor expression. At one moment, however, we obtained very good expression. We always used the same glycerolstock which is defreezed before used. So repeated freezing and defreezing of the bacteria seems to result in better expression. Does anybody understand this? After changing the Xa-site (4 aa) into AspPro (2 aa) expression is again very low. We tried already lower expression temperature and different incubation times. Repeated freezing and defreezing didn't result in a better expression. Can someone help us? Thanks in advance, Henri M.H. Spronk Department of Biochemistry University Maastricht henri.spronk@bioch.unimaas.nl From owner-proteins@net.bio.net Tue Mar 10 22:00:00 1998 Path: biosci!uthct.edu!shaun From: shaun@uthct.edu (Shaun D. Black) Newsgroups: bionet.molbio.proteins Subject: Re: Svedberg units Date: 11 Mar 1998 07:16:51 -0800 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 24 Sender: daemon@net.bio.net Distribution: world Message-ID: <9803111515.AA24912@athena.uthct.edu> NNTP-Posting-Host: net.bio.net Greetings All, Nicole Frankenberg wrote... >Does anybbody know where to get svedberg units for standard proteins. >any idea of a book or article is appreciated. >Thanks for your help.Nicole Check the CRC Handbook of Biochemistry (tables of thousands of protein S20,w and D20,w values); the tables I have are from the 2nd Edition (1970) in Section C. You might also look at current biochem textbooks, as many of them list some values for well-known proteins. I hope this helps you. Best regards, Shaun Black =-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-= = Shaun D. Black, PhD | Internet e-mail address: shaun@uthct.edu = = Dept. of Biochemistry | University of Texas Health Center at Tyler = = Tyler, TX 75710 | World Wide Web: http://www.uthct.edu = =-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-=-= From owner-proteins@net.bio.net Wed Mar 11 22:00:00 1998 Path: biosci!agate!howland.erols.net!rill.news.pipex.net!pipex!server1.netnews.ja.net!news.nott.ac.uk!NewsWatcher!user From: kevin.bailey@nottingham.ac.uk () Newsgroups: bionet.molbio.proteins Subject: Re: C-terminal protein sequencing Followup-To: bionet.molbio.proteins Date: Thu, 12 Mar 1998 13:17:02 +0100 Organization: Cripps Computing Centre, The University of Nottingham Lines: 17 Distribution: world Message-ID: References: NNTP-Posting-Host: wpbjwk1.biochem.nottingham.ac.uk In article , KH@axisgenetics.co.uk (Koen Hellendoorn) wrote: > Hello, > > Can anyone tell me if there is any institute or company (preferably in > the UK/ other Europe) that can do C-terminal sequencing of proteins? > > Thanks, > > Koen At one time Hewlett Packard were acting as intermediaries and taking C-term samples to demonstrate their sequencer, not sure if this is still the case though. Try contacting HP UK. At one time there was a service at the University of Michigan, again not clear if this still runs. From owner-proteins@net.bio.net Wed Mar 11 22:00:00 1998 Path: biosci!agate!howland.erols.net!rill.news.pipex.net!pipex!server1.netnews.ja.net!news.nott.ac.uk!NewsWatcher!user From: kevin.bailey@nottingham.ac.uk () Newsgroups: bionet.molbio.proteins Subject: Re: Online Auctions for used lab equipment Going, Going...Sold!!! Followup-To: bionet.molbio.proteins Date: Thu, 12 Mar 1998 13:23:55 +0100 Organization: Cripps Computing Centre, The University of Nottingham Lines: 10 Message-ID: References: <6e69cq$2ca@dfw-ixnews11.ix.netcom.com> NNTP-Posting-Host: wpbjwk1.biochem.nottingham.ac.uk In article <6e69cq$2ca@dfw-ixnews11.ix.netcom.com>, Mark Atlas wrote: > > Many new items coming up weekly. Stop buy and register for free. > > http://www.going-going-sold.com Note for UK/European labs: last time I checked this site it was *only* available to US sites. From owner-proteins@net.bio.net Wed Mar 11 22:00:00 1998 Path: biosci!agate!newsfeed.wli.net!feed2.news.erols.com!erols!news-peer.sprintlink.net!news.sprintlink.net!Sprint!newsfeed.nacamar.de!fu-berlin.de!news-ber1.dfn.de!news-ham1.dfn.de!news-han1.dfn.de!news.tu-bs.de!rzlimes.gbf-braunschweig.de!not-for-mail From: kaerst@gbf.de (Uwe Kaerst) Newsgroups: bionet.molbio.proteins Subject: Re: E280 Protein quantitation on microplates ?! Date: 12 Mar 1998 12:29:34 GMT Organization: GBF Lines: 54 Message-ID: <6e8kfe$1tv$1@rzlimes.gbf-braunschweig.de> References: <35059283.FB7F6CF3@mailer.uni-marburg.de> NNTP-Posting-Host: etnuka.gbf-braunschweig.de Mime-Version: 1.0 Content-Type: Text/Plain; charset=US-ASCII X-Newsreader: WinVN 0.99.8 (x86 32bit) In article <35059283.FB7F6CF3@mailer.uni-marburg.de>, bocola@mailer.uni-marburg.de says... > >We build up a new biochemical workgroup and looking foorward to by an >UV/VIS-Microplate Reader. >Has anyone experience with UV-Microplates and especially Protein >determination ? >(Unfortunately we have no "normal" UV/VIS Photometer) >Is it possible to measure quantitative absorption with good accuracy for >small volumes? > >Thank you for telling me your experiences or suggestions, >Marco Bocola > >-- >Marco Bocola Inst. f. pharmaz. >Chemistry > AG >Prof. Klebe >Tel: +49-6421-28-5071 Marbacher Weg 6 >Fax: -8994 D-35032 >Marburg > >NEW !!! e-mail:bocola@mailer.uni-marburg.de NEW !!! > >:-( There are no problems, just solutions !? :-) > > Molecular Dynamics offers a SpectraMax 250 Reader that goes down to 250 nm. Plates are also available, and we use this routinely and find it very reliable. Hope this helps (and NO AFFILIATION, of cause!) Uwe -- Uwe Kaerst Dept. Enzymology GBF - Gesellschaft fuer Biotechnologische Forschung Mascheroder Weg 1 D-38124 Braunscheig, Germany Tel: +(49) 531 6181 318 Fax: +(49) 531 2612313 EMAIL: kaerst@gbf-braunschweig.de -- Disclaimer -- Standard > Keyword : Opinions, my own, nobody else's, no affiliation, whatsoever ... ------------------------------------------------------------------------------ ------------------------------------- "It is a mistake to allow any mechanical object to realize that you are in a hurry !" From owner-proteins@net.bio.net Wed Mar 11 22:00:00 1998 Path: biosci!agate!howland.erols.net!europa.clark.net!164.67.42.145!awabi.library.ucla.edu!128.120.8.185!mark.ucdavis.edu.MISMATCH!news.ucdavis.edu!not-for-mail From: Jim Kami Newsgroups: bionet.molbio.methds-reagnts,bionet.molbio.proteins,bionet.software Subject: Re: Proposal for a new bionet newsgroup Date: Thu, 12 Mar 1998 08:38:56 -0800 Organization: University of California, Davis Lines: 37 Message-ID: <35080FA0.2886@ucdavis.edu> References: <6dobf1$4cj$1@gwsun.medinf.mu-luebeck.de> <35066A66.2B70@liv.ac.uk> NNTP-Posting-Host: reqd-091.ucdavis.edu Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 3.04Gold (Win95; I) Xref: biosci bionet.molbio.methds-reagnts:65739 bionet.molbio.proteins:12443 bionet.software:20708 Try sci.chem.labware for used equipment. Jim Kami Blue Rose Biotech GcR wrote: > > Lars Komorowski wrote: > > > > I propose to set up a new newsgroup for used lab euipment. Who is interested > > ? > > Lars Komorowski > > Ins. f. Biochemie > > Med. Univ. Luebeck, Germany > > As I can recall, a few months ago somebody came up with a similar idea, > he made a webpage specially for the (re)sale of used lab equipment. You > can find it at http://www.geocities.com/ResearchTriangle/Lab/5656/ > > Sincerely, > > GcR > > > ---------------------------------------------\/----- > Gert Roosen /\ > PhD Student /--\ > Department of Haematology (====\ > Royal Liverpool University Hospital \----) > Duncan Building 2nd floor \--/ > Prescot Street L69 3BX Liverpool \/ > + 44 (0)151 706 4326 (phone) /\ > + 44 (0)151 706 4311 (fax) /--\ > g.roosen@liv.ac.uk (email) /====) > -------------------------------------------(----/--- From owner-proteins@net.bio.net Wed Mar 11 22:00:00 1998 Path: biosci!bloom-beacon.mit.edu!hecate.umd.edu!not-for-mail From: Hongyu Zhang Newsgroups: bionet.molbio.proteins Subject: Active site residue Date: Thu, 12 Mar 1998 14:50:34 -0500 Organization: University of Maryland, College Park Lines: 18 Message-ID: <35083C8A.41C6@carb.nist.gov> NNTP-Posting-Host: tabasco.carb.nist.gov Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 3.01SGoldC-SGI (X11; I; IRIX64 6.4 IP30) Dear Molecular Biologists, I have a bit of knowledge that it's only possible for some specific amino acids to form the active sites of proteins. Can anyone enumerate all the possible resiude types? Thanks ! Hongyu -- ------------------------------------------------------------------ Hongyu Zhang, Ph.D. | Tel: (301) 738-6117 (w) ^/..\^ CARB, University of Maryland | (301) 987-0179 (h) -m( 00 )m- 9600 Gudelsky Drive | Fax: (301) 738-6255 Rockville, Maryland 20850 | Email: hyzhang@carb.nist.gov URL: http://indigo5.carb.nist.gov/~hyzhang ------------------------------------------------------------------ From owner-proteins@net.bio.net Wed Mar 11 22:00:00 1998 Path: biosci!CLEMSON.EDU!walkerc From: walkerc@CLEMSON.EDU ("Caroline J. Walker") Newsgroups: bionet.molbio.proteins Subject: FPLC Date: 12 Mar 1998 06:30:48 -0800 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 11 Sender: daemon@net.bio.net Distribution: world Message-ID: NNTP-Posting-Host: net.bio.net We are buying an FPLC and are comparing the Biorad Biologic system with the Pharmacia FPLC system. There are some small differences on the specs and the prices are about the same so we could use either. What I would like to hear is if anyone has feedback on how easy either of these systems is to use. Is the Pharmacia software (OS/2) OK, or is the Windows 95 with the Biorad better? How are the leaks? Design faults?? Reliability? Service? Would be happy to hear any comments. Thanks, Caroline From owner-proteins@net.bio.net Wed Mar 11 22:00:00 1998 Path: biosci!CLEMSON.EDU!walkerc From: walkerc@CLEMSON.EDU ("Caroline J. Walker") Newsgroups: bionet.molbio.proteins Subject: Re: is it the same protein Date: 12 Mar 1998 10:15:08 -0800 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 12 Sender: daemon@net.bio.net Distribution: world Message-ID: NNTP-Posting-Host: net.bio.net Can you take your high mw fraction off the gel filtration column, concentrate it and run it through again - see if you generate lower mw monomers from it i.e. you may be able to promote some futher dissociation of the aggregated form. I got a similar kind of result with a protein, and found that the Mg concentration was critical in determining the level of aggregation (in its absence it ran as monomers). The other possibility you may consider is that in crude preps it is possible that your protein may be associating with other proteins rather than just itself e.g. if its a part of a metabolon or other large complex (even if this association is loose). From owner-proteins@net.bio.net Thu Mar 12 22:00:00 1998 Path: biosci!agate!newsfeed.wli.net!news-out.internetmci.com!newsfeed.internetmci.com!193.174.75.126!news-was.dfn.de!news-kar1.dfn.de!hades.rz.uni-sb.de!ukmhg03.med-hg.uni-sb.de!user From: hgtsei@med-rz.uni-sb.de (Thomas Seib) Newsgroups: bionet.molbio.proteins Subject: Re: FPLC Date: 13 Mar 1998 15:04:16 GMT Organization: University of Saarland, Computing Center, Germany. Lines: 36 Distribution: world Message-ID: References: NNTP-Posting-Host: ukmhg03.med-hg.uni-sb.de In article , walkerc@CLEMSON.EDU ("Caroline J. Walker") wrote: > We are buying an FPLC and are comparing the Biorad Biologic system with the > Pharmacia FPLC system. There are some small differences on the specs and > the prices are about the same so we could use either. What I would like to > hear is if anyone has feedback on how easy either of these systems is to > use. Is the Pharmacia software (OS/2) OK, or is the Windows 95 with the > Biorad better? How are the leaks? Design faults?? Reliability? Service? > Would be happy to hear any comments. > Thanks, > Caroline We had to make the same decision and finally bought the Pharmacia FPLC system. The main reason was in my hands, that the BioRad pump was very sensible to air bubbles trapped, just like a HPLC system, resulting in strong pulsation of the UV monitor baseline. Hence, you have to degas buffer solutions very thoroughly before use. This may be feasible, but sometimes it is of advantage to apply a large-volume sample via the pump, and then it is very problematic to degas because of possible denaturation of the proteins. I also found it very cumbersome to remove air bubbles from the pump once they were trapped. With the Pharmacia system, air bubbles are no problem at all. I never degas any solution. It works very reliable. The only disadvantage is the computer software running under OS/2, which is a dying platform and of no use for other purposes. However, Windows3.1 is also installed and most programs run without problems. I hope this helps. Matthias -- Thomas Seib Kantstr. 12 66292 Riegelsberg From owner-proteins@net.bio.net Thu Mar 12 22:00:00 1998 Path: biosci!agate!arclight.uoregon.edu!news.cc.ukans.edu!not-for-mail From: PGegen@RNAworld.edu (Dr. Peter Gegenheimer) Newsgroups: bionet.molbio.proteins Subject: Re: FPLC Date: 14 Mar 1998 03:35:18 GMT Organization: Univ. Kansas (Biochemistry) Lines: 60 Message-ID: <7opiGDf98QgB-pn2-O54PednwKmWS@rnaworld.bio.ukans.edu> References: In article , walkerc@CLEMSON.EDU > ("Caroline J. Walker") wrote: > > With the Pharmacia system, air bubbles are no problem at all. I never > degas any solution. It works very reliable. > The only disadvantage is the computer software running under OS/2, > which is a dying platform and of no use for other purposes. However, > Windows3.1 is also installed and most programs run without problems. Oops! OS/2 is definitely NOT a dying platform. First, It is a non-Windows platform in a Windows world. There are as many OS/2 users as Mac users, but the OS/2 users are all in the large corporations which IBM somehow prefers to deal with. OS/2 is also the second-best-selling network OS for the Intel processor (behind Windows NT). IBM aggressively markets & develops OS/2, just not to individual users. OS/2 uses a dynamic desktop like the Mac, but with real object-orientation (unlike Mac or Windows). Second, OS/2 is a _complete_ operating environment; you can do virtually anything with it more easily than with Win95. (Its only competitor is WinNT.) For example, I use a native OS/2 word processor, spreadsheet (better than any Win?? app), graphics apps, network & internet apps. I also use a few Win3.1 and DOS apps for which there are no substitutes (not for the Mac, either). My web site is maintained & run on the OS/2 PC we use in our lab. Also, don't forget that the LICOR DNA sequencer uses OS/2 to collect and interpret sequence data from multiple gels simultaneously. Third, you are right in that IBM does not plan to support Win95 or NT programs under OS/2. However, some of the best Windows NT software was developed on OS/2 and ported to NT. Finally, OS/2 is one of the few OS's with built-in support for Java programs (i.e. it runs Java without a browser), and will support--for example--the Java Molecular Biology Workbench. Basically, the OS should be the least of your worries o------------------------------------------------------------------- --o | Dr. Peter Gegenheimer | Vox: 785-864-3939 FAX: 785-864-5321 | | Departments of Biochemistry | PGegen@UKans.edu | | and of Botany | http://RNAworld.Bio.UKans.edu/ | | | | | University of Kansas | | | 2045 Haworth Hall | "The sleep of reason produces | | Lawrence KS 66045-2106 | monsters." Goya | o_____________________________|_____________________________________ __o From owner-proteins@net.bio.net Thu Mar 12 22:00:00 1998 Path: biosci!agate!newsfeed.kornet.nm.kr!news.maxwell.syr.edu!howland.erols.net!feed1.news.erols.com!winter.news.erols.com!not-for-mail From: Joan Evans Newsgroups: bionet.molbio.proteins Subject: Restriction enzymes study Date: Fri, 13 Mar 1998 09:19:21 +0000 Organization: Erol's Internet Services Lines: 26 Message-ID: <3508FA19.19E4@scienceboard.net> Reply-To: j.evans@scienceboard.net NNTP-Posting-Host: 207-172-57-115.s115.tnt2.ann.erols.com Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Trace: winter.news.erols.com 889798519 17402 207.172.57.115 (13 Mar 1998 14:15:19 GMT) X-Complaints-To: abuse@erols.com X-Mailer: Mozilla 3.02Gold (Macintosh; I; PPC) The use of restriction enzymes is an essential component of molecular biology research. While there appears to be little variation in quality from one vendor to another, the price of restriction enzymes varies considerably. The Science Advisory Board (http://www.scienceboard.net) will soon begin a series of studies on restriction enzymes. The goal of these studies is to better understand the preferences, expectations and unmet needs of molecular biologists, especially as they relate to reliability, selection, and packaging. The Science Advisory Board is a worldwide panel of more than 3,000 scientific experts that convenes electronically to participate in online surveys and focus groups to voice their opinions on a wide variety of topics related to new research products and emerging technologies. In order to participate in these and future studies, it is first necessary to complete the registration form which can be found at http://www.scienceboard.net. Registrants with appropriate backgrounds will be notified shortly thereafter. All responses are held in strict confidence, and participants will be compensated for their time. Joan Evans Membership Secretary The Science Advisory Board http://www.scienceboard.net From owner-proteins@net.bio.net Thu Mar 12 22:00:00 1998 Path: biosci!internet!biosci!not-for-mail From: biohelp (BIOSCI Administrator) Newsgroups: bionet.molbio.proteins Subject: BIOSCI/bionet miniFAQ & Fundraiser Date: 13 Mar 1998 02:00:08 -0800 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 233 Sender: daemon@net.bio.net Distribution: world Message-ID: <199803131000.CAA21027@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 accessible on the World Wide Web at URL http://www.bio.net/. If you can not find an answer to your question in this or other documentation, the BIOSCI technical support staff answers e-mail queries sent to biosci-help@net.bio.net We can only answer questions about the use of the newsgroups and mailing lists. We unfortunately do not have the staff to do Internet information searches or answer scientific questions. Please post those to the appropriate BIOSCI/bionet newsgroups. Contents: -------- 0) BIOSCI NEEDS YOUR SUPPORT!! 1) Using the WWW to access the BIOSCI/bionet newsgroups. 2) What to do about "spams," i.e., junk mail, ads, etc. 3) Examples of subscribing and unsubscribing to the mailing lists. 4) The BIOSCI user address and research interest directory. 0) BIOSCI NEEDS YOUR SUPPORT!! ------------------------------ BIOSCI's government funding has been expended, and we are now operating solely from advertising revenue that we have raised from our Web site at http://www.bio.net/. We need just a few minutes of your time to help us serve you. You can do two important things which will take very little time for you individually and will immensely help us continue to help you. First, please use our WWW system at http://www.bio.net/ to access the archives. You can post or reply to messages via your Web browser as described in item #1 below. Your usage helps attract sponsors. If you contact any of our sponsors, please be sure to thank them for supporting BIOSCI. It is critical for them to get this feedback if they are to continue their sponsorship for the long term. Second, if you work for a company or organization that provides products or services of interest to the biology community, please pass this message on to your marketing or marketing communications department or other appropriate group. Please ask them to help support BIOSCI by sponsoring our Web site and explain the uses and benefits of the system to the biology community. If they are interested, they can then contact us for further information at our tech support address, biosci-help@net.bio.net. 1) Using the WWW to access the BIOSCI/bionet newsgroups. -------------------------------------------------------- As of 10 December 1995, all BIOSCI/bionet full newsgroups are accessible through the World Wide Web (WWW) at URL http://www.bio.net. One can read and reply publicly or privately to both recent postings and archived messages through one's Web browser if it is configured properly to send e-mail. Each newsgroup is equipped with its own WAIS index. The main BIOSCI home page also has access to the BIO-JOURNALS Table of Contents database WAIS index and the BIOSCI user address database described in another item further below. 2) What to do about "spams," i.e., junk mail, ads, etc. ------------------------------------------------------- BIOSCI is a set of parallel USENET newsgroups (the "bionet" groups), mailing lists, and a hypermail archive at URL http://www.bio.net/. The same postings are distributed on all media (except for a small number of mailing-list-only groups at net.bio.net). Unfortunately it is becoming a despicable practice on the Internet (by a few people out to make a fast buck) to do automated mass postings to thousands of newsgroups and mailing lists. These attempts to grab free advertising are refered to as "spams" in the usual, somewhat boneheaded, net terminology. USENET is more susceptible to this practice, and many spams originate on the USENET groups and then are passed on to the mailing lists. However, spammers also get lists of mailing addresses and hit these too, so neither medium is immune. What should you do personally if you get junk mail? --------------------------------------------------- Just delete it and move on without reading it further. Filing a protest is becoming increasingly useless because spammers are often disguising the addresses where the messages are sent from. Unless you really understand Internet mail systems, your attempt at protest by sending replies to the message will often end up being sent to the address of an innocent person that the spammer is victimizing. What can BIOSCI/bionet do to protect its newsgroups? ---------------------------------------------------- The only solution currently available is to moderate the newsgroup. If this newsgroup is already moderated, then you are in good shape. Moderation protects the USENET distribution from about 95% of the spams that are being sent to date and protects the mailing lists completely. Moderation means, however, that someone has to take the time to review each message before it goes out. We have set up software here that simply allows the moderator to forward to an address at net.bio.net messages that (s)he wishes to have distributed. This takes no more time than that needed to read the message and pass it on, say about 1 min. per message. Most newsgroups currently have a discussion leader who is responsible for their newsgroup. The discussions leaders and their e-mail addresses are listed in the BIOSCI Information Sheet which is available on the Web at http://www.bio.net/. If a newsgroup is being hit with too many junk postings, please contact the discussion leader for that group and see if there is interest in moderating the group. Please do not assume that by simply posting a complaint to the newsgroup itself, anyone on the BIOSCI staff will act on your complaint. With close to 100 newsgroups to run, the BIOSCI staff has to rely on the discussion leaders of each newsgroup to report problems directly to us at biosci-help@net.bio.net. We will moderate any of our newsgroups if the discussion leader tells us that the readership of the group wishes to do so and if a moderator is willing to do the work. For most BIOSCI/bionet groups, this entails only a few minutes of work each day. Moderating a newsgroup will resolve probably 95% of the junk postings on the USENET distribution. Unfortunately there are easy ways for determined spammers to override the moderation mechanism on USENET, but we can protect our e-mail subscribers from unwanted postings if the newsgroup is moderated. You can also access our newsgroups over the WWW at URL http://www.bio.net. While this Web interface will not stop spammers from trying to post to the groups, this will give you yet another way, besides using USENET news, to keep the junk out of your personal mail files. For those of you with local USENET news systems, the Web interface will also give you faster access to new newsgroups and recent postings. 3) Examples of subscribing and unsubscribing to the mailing lists. ------------------------------------------------------------------ PLEASE NOTE: The BIOSCI management does NOT act on subscription/unsubscription requests that are posted improperly to the newsgroups and mailing lists. People who do this only bother everyone on the lists to no avail. Please be sure to follow the proper procedures below. Gory details are in the BIOSCI Information sheets on the Web at http://www.bio.net. Below we give an example utilizing the METHODS-AND-REAGENTS list at both of our two BIOSCI sites: Users in the Americas and Pacific Rim countries who use the BIOSCI ------------------------------------------------------------------ node at computer net.bio.net: ---------------------------- A) Determine the "listname" which is the <=8 character mail address ^^^^^^^^^^^^^ for the group. These can be found in the BIOSCI Info. Sheet. For the METHODS-AND-REAGENTS group the mailing address is methods@net.bio.net. The listname is the portion of the address to the left of the @ sign, i.e., "methods". The listname is used with the "subscribe" and "unsubscribe" commands illustrated below. B) Mail all commands in the body of a mail message addressed to biosci-server@net.bio.net. Do NOT send commands to the newsgroup posting addresses! Leave the Subject: line blank, any text on it will be ignored. C) In the body of your message put one or more of the following commands with an "end" command on the last line, e.g., subscribe methods unsubscribe methods end Do NOT put your e-mail address or other text on these lines. The server only allows you to cancel your subscription if the address on your mail header matches the address on our mailing list. Please ask for help at biosci-help@net.bio.net if your address has changed, e.g., if you know you are on the list but the server tells you that you are not a member. Users in Europe, Africa, and Central Asia who use the BIOSCI node at -------------------------------------------------------------------- computer daresbury.ac.uk (also known as dl.ac.uk): ------------------------------------------------- To subscribe and unsubscribe to/from the BIOSCI lists, you need to specify the full USENET newsgroup name with "bionet-news." prepended. The USENET newsgroup names are listed in the BIOSCI Information sheet on the Web at http://www.bio.net/. For the METHODS-AND-REAGENTS list the USENET newsgroup name is bionet.molbio.methds-reagnts, thus the appropriate commands are sub bionet-news.bionet.molbio.methds-reagnts unsub bionet-news.bionet.molbio.methds-reagnts These commands are included in a message addressed to mxt@dl.ac.uk, NOT to the newsgroup mailing addresses. As usual, include the text in the body of the message as text on the Subject: line is ignored. To unsubscribe from all the lists at the UK node, use unsub bionet-news Please note that if the address in the list is different than the one in your mail message header, you will not be able to unsubscribe by this method. If you have problems, please mail biosci@daresbury.ac.uk. 4) The BIOSCI user address and research interest directory. ----------------------------------------------------------- Please take this opportunity to add your name, address, and research interest information to the BIOSCI User Address Database if you have not already done so. You can fill out the address form directly through our Web page at URL http://www.bio.net/adrform.html. The address database is reindexed nightly for WWW access (the URL is http://www.bio.net/). If you are not directly on the Internet but can reach it by e-mail, please use our waismail server to access the user directory. waismail use is described above. You can also request a user address form by e-mail from biosci-help@net.bio.net. Please check your database entry from time-to-time to see if your address information is still up-to-date. Because of our limited personnel resources, we ask that you resubmit a *complete* form to revise your entry; we only replace complete entries and do not have resources to edit old forms. From owner-proteins@net.bio.net Thu Mar 12 22:00:00 1998 From: Yu-Ran Luo Newsgroups: bionet.molbio.proteins Subject: A Novel Software of Organic Mass Spectra Date: Fri, 13 Mar 1998 21:21:12 -0500 Organization: University of South Florida Lines: 60 Message-ID: <3509E998.239C6F83@seas.marine.usf.edu> NNTP-Posting-Host: 198.116.54.87 Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 4.01 [en] (Win95; I) X-Priority: 3 (Normal) Path: biosci!bloom-beacon.mit.edu!howland.erols.net!news-peer.sprintlink.net!news-peer-west.sprintlink.net!news.sprintlink.net!Sprint!news.vphos.net!su-news-hub1.bbnplanet.com!atl-news-feed1.bbnplanet.com!news.bbnplanet.com!news1.usf.edu!not-for-mail Dear Sir/Madam, I am writing to share good news with you. Novel software by name of SIOMS (Smart Interpretation of Organic Mass Spectra) will be available next month. The attached paper is its announcement. A gift (free Demo) was released. Please give me a reply by e-mail if you like to try the Demo. Many thanks for your attention. Sincerely yours, Yu-Ran Luo, Ph.D. ============ What is Smart Interpretation of Organic Mass Spectra? Smart Interpretation of Organic Mass Spectra (SIOMS) is a system for interpretation of organic mass spectra (MS), implemented under Microsoft Windows. The SIOMS is designed to simulate the intelligent behavior of experienced mass spectroscopists who interpret organic mass spectra frequently and deduce the molecular structures of unknowns. The SIOMS system mainly consists of a set of linked programs: (1) An input system for the m/z peaks of unknowns; (2) A forward/reverse search program with combined regulation into the MS database; (3) A self-learning program. The SIOMS can select a series of pertinent MS in the database, and can self-learn by referring them to the unknown spectrum; (4) Interpretation programs for deducing likely molecular weight and substructures; (5) A structure generator for creating a sequence of candidate structures of unknowns; (6) Programs for theoretical MS prediction and structure evaluation of each candidate; (7) An output system of optimized structure(s) of unknowns. Advantages of the SIOMS 1. The excellent self-learning ability of the program. In the SIOMS system, the self-learning program can find necessary substructure/spectral correlation rules in order to interpret MS and deduce molecular structure of unknowns. 2. The ability to easily identify unknowns. This makes the SIOMS superior to volumed library search systems in current commercial software. 3. The input for the SIOMS system is the EI (electron ionization) MS at low resolution of the unknowns. No more effort by the chemists is needed, such as high resolution MS, MS/MS etc. It fits the general purpose of organic MS laboratories, and simplifies the MS analytical work. Any operator of organic MS can work well as an expert while running the SIOMS. Molecular structure of samples, including unknowns, will be displayed once an operator inputs (automatically or manually) data of the m/z and intensity. From owner-proteins@net.bio.net Thu Mar 12 22:00:00 1998 Path: biosci!agate!newsfeed.wli.net!feed2.news.erols.com!erols!news.maxwell.syr.edu!bignews.mediaways.net!news-fra1.dfn.de!zeus.rbi.informatik.uni-frankfurt.de!grapool30.rz.uni-frankfurt.de!parcej From: parcej@biophys.mpg.de (Dr Dave Parcej) Newsgroups: bionet.molbio.proteins Subject: Re: C-terminal protein sequencing Date: Fri, 13 Mar 1998 14:13:47 +0100 Organization: MPI fuer Biophysik Lines: 23 Distribution: world Message-ID: References: NNTP-Posting-Host: mac-kb5.biophys.mpg.de X-Newsreader: MT-NewsWatcher 2.3.5 Hi While browsing the web I came accross one company from sweden who do C-terminal sequencing. Their URL is http://www.innovagen.se/mbserv/protein.html I have no affiliation and havent tried them myself, so tell me how you get on! Dave In article , KH@axisgenetics.co.uk (Koen Hellendoorn) wrote: > Hello, > > Can anyone tell me if there is any institute or company (preferably in > the UK/ other Europe) that can do C-terminal sequencing of proteins? > > Thanks, > > Koen From owner-proteins@net.bio.net Thu Mar 12 22:00:00 1998 Path: biosci!TC3NET.COM!cola From: cola@TC3NET.COM (cola) Newsgroups: bionet.molbio.proteins Subject: protein sequencing Date: 13 Mar 1998 07:51:03 -0800 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 5 Sender: daemon@net.bio.net Distribution: world Message-ID: <350955EC.7A12A497@tc3net.com> NNTP-Posting-Host: net.bio.net Hello, I am trying to locate a company in the US that specialize in protein sequencing. The protein of interest is approxiamtley 190 amino acids, MW 24KD. Would appreciate assistance. Thank you Cola From owner-proteins@net.bio.net Sat Mar 14 22:00:00 1998 Path: biosci!bcm.tmc.edu!news.msfc.nasa.gov!newsfeed.internetmci.com!207.217.77.43!newsfeed1.earthlink.net!nntp.earthlink.net!usenet From: The Antibody Resource Page Newsgroups: bionet.molbio.proteins Subject: Looking for an antibody or information on antibodies? Date: Sun, 15 Mar 1998 09:31:35 +0000 Organization: EarthLink Network, Inc. Lines: 45 Message-ID: <350B9FF7.C8@antibodyresource.com> NNTP-Posting-Host: 208.255.9.98 Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 3.04Gold (Macintosh; I; 68K) The Antibody Resource Page (http://www.antibodyresource.com/) is an invaluable website to researchers and educators. Here is just some of what can be found on the page: 1. How to Find an Antibody - a variety of ways on and off the web to find the antibody you are looking for. There are links to free search engines that allow you to search a multitude of companies for the specific antibody you need. 2. Online Companies - links to over 110 companies that sell antibodies or antibody related products. Is your company listed on this page? 3. Antibody Image Gallery - animated antibody pictures are available 4. Bulletin Board - Have a question? Then stop by and post a message. 5. Educational Resources - a variety of new links have been added.There are links to pages on immunochemistry, antibody production, autoimmunity, vaccines, immunology and much more. This page is divided up into sections on research, educational, and health resources. 6. The latest in antibody news - Get up-to-date, antibody-relatedarticles on topics from academia and industry. 7. Online databanks and databases - links to online protein and nucleic acid sequencing databases. Invaluable to biochemists and molecular biologists who work with antibodies. Check it all out at: http://www.antibodyresource.com/ ps. Don't forget to visit our current sponsor, Research Diagnostics, Inc.(http://www.researchd.com/absort1.htm). RDI sells a wide variety of antibodies for nearly every need. pps. Are you interested in being a sponsor on the Antibody Resource Page (ARP)? Nearly half of the visitors come to find commercially available antibodies. If you are a company that sells antibodies and want to increase webtraffic to your site, email the antibody resource page to find out about sponsorship: antibody@antibodyresource.com ppps. The ARP was voted among the top 25 biotechnology webpages for 1997 by Genetic Engineering News! From owner-proteins@net.bio.net Sun Mar 15 22:00:00 1998 Path: biosci!agate!news-spur1.maxwell.syr.edu!news.maxwell.syr.edu!newsfeed.internetmci.com!198.6.0.5!uunet!in3.uu.net!newsfeed.attap.net!mango.singnet.com.sg!dahlia.singnet.com.sg!ocean.singnet.com.sg!id4.nus.edu.sg!nuscc.nus.edu.sg!medp4076 From: medp4076@leonis.nus.sg (Zhu Congju) Newsgroups: bionet.molbio.proteins Subject: pcDNA3.1(A)-myc Date: 16 Mar 1998 12:08:11 GMT Organization: National University of Singapore Lines: 8 Message-ID: <6ej4nb$qnq$1@nuscc.nus.edu.sg> NNTP-Posting-Host: leonis.nus.edu.sg NNTP-Posting-User: medp4076 X-Newsreader: TIN [version 1.2 PL2] Hi, Does anybody know how I can find a positive control for pcDNA3.1(A)-myc (invitrogen) doing western? Thanks! CJ Zhu From owner-proteins@net.bio.net Sun Mar 15 22:00:00 1998 Path: biosci!agate!howland.erols.net!cpk-news-hub1.bbnplanet.com!cpk-news-feed4.bbnplanet.com!cpk-news-feed1.bbnplanet.com!news.bbnplanet.com!news.ums.edu!gamera.cbl.umces.edu!cbl.umces.edu!reno From: "Reno T. Nguyen" Newsgroups: bionet.molbio.proteins Subject: Re: carbon content of proteins? Date: Mon, 16 Mar 1998 09:08:20 -0500 Organization: University of Maryland, Chesapeake Biological Lab. Lines: 34 Message-ID: References: <3506BD15.5F0F@pilot.msu.edu> NNTP-Posting-Host: cbl.umces.edu Mime-Version: 1.0 Content-Type: TEXT/PLAIN; charset=US-ASCII To: Jonathan Walton In-Reply-To: <3506BD15.5F0F@pilot.msu.edu> On Wed, 11 Mar 1998, Jonathan Walton wrote: > Can anyone tell me a generally accepted average value for the % (by > weight) of carbon in proteins? The value of 15-16% for N is generally > used, but how about carbon? And (related question that would give the > answer) is there is a program that will calculate the elemental > composition of a protein? > Thanks, I've only determined the %C for Rubisco (47%) using a C and N analyzer. The %N was 13%. I'm not aware of any program that would calculate the elemental composition of a protein after entry of it's amino acid composition. I have also determined the average %C and %N for the protein amino acids usually analyzed (asp, glu, ser, his, gly, thr, arg, ala, tyr, met, val, phe, ile, leu, lys) to be 46% and 14%, respectively. These average values for the amino acids should be similar to those for many proteins. Reno ************************************************************************* Reno T. Nguyen Chesapeake Biological Laboratory /----\ /-----\ / Univ. MD Center for Environmental Science / / / / P.O. Box 38 / /______/ / Solomons, MD 20688 / / \ / / / // Tel: (410) 326-7261 \_______/ \_______/ \_______/ (410) 326-7409 Fax: (410) 326-7341 E-mail: reno@cbl.umces.edu ************************************************************************ From owner-proteins@net.bio.net Sun Mar 15 22:00:00 1998 Path: biosci!SOCRATES.BERKELEY.EDU!bbaker From: bbaker@SOCRATES.BERKELEY.EDU (Barbara Baker) Newsgroups: bionet.molbio.proteins Subject: Postdoctoral Position Date: 16 Mar 1998 17:09:43 -0800 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 61 Sender: daemon@net.bio.net Distribution: world Message-ID: NNTP-Posting-Host: net.bio.net A POSTDOCTORAL POSITION is currently available for research on elucidation of the N-mediated signal transduction pathways leading to TMV resistance. We isolated the first virus resistance gene, N, of plants {Whitham et al. (1994) Cell 78, 1101-1115}. The N gene confers resistance to tobacco mosaic virus (TMV) in tobacco. To elucidate the mechanisms of N-mediated resistance we are investigating: (1) regulation of N gene expression, (2) the structure-function relationship of the N encoded protein domains, (3) the structure, biochemical properties and cellular localization of the N protein (4) the identification of TMV protein required for N induced resistance (5) identification of components of the resistance signal transduction pathway by genetic dissection of N resistance pathway and identification of cellular interacting proteins, and (7) identification of genes with structural and functional similarities to N for comparative studies. The applicant will have the opportunity to participate in one or more of these projects, however, the available position primarily concerns genetic dissection of the N-mediated signal transduction pathway in tobacco, tomato and Arabidopsis. N is a member of the TIR-NBS-LRR class of R genes. The sequence of several recently isolated R genes shows that they encode deduced products with strikingly similar structural features {Baker et al. (1997) Science 276,726-733}. It is postulated that R genes encode receptors or components of signaling pathways that participate in pathogen recognition and signaling leading to cell death (hypersensitive response-HR), systemic defense (systemic acquired resistance-SAR) and pathogen inhibition. The amino-terminal domain of N (TIR) and other members of this class of R genes share homology with the cytoplasmic domains of the interleukin-1 receptor (Il-1R) the Drosophila and human Toll receptor proteins, participants in pathogen defense programs. The sequence similarity of N, IL-1R and Toll suggests that N mediates pathogen resistance through an IL-1R/Toll-like signal transduction pathway. The nucleotide binding site domain (NBS) of N and other R genes is homologous to the putative ATPase domain of the caspase activating proteins CED-4 and Apaf-1, proteins that regulate apoptosis in C. elegans and mammals respectively. The sequence similarity of N and other R genes to CED-4 and Apaf-1 suggests there may be similarities between mechanisms of cell death in plants and animals {Chinnaiyan et al. (1997) Nature 388, 728-729}. Applicants should have experience in genetics, biochemistry, molecular biology and/or cell biology, however, prior work with plants is not required. A cover letter describing research experience and interests, a curriculum vitae and names of three references should be sent to: Dr. Barbara Baker, University of California, Berkeley and USDA, Plant Gene Expression Center, 800 Buchanan St., Albany, CA 94710. FAX: 510-559-5929 or 510-559-5678; email: bbaker@socrates.berkeley.edu Barbara Baker Plant Gene Expression Center Department of Plant and Microbial Biology University of California, Berkeley & USDA-ARS 800 Buchanan St. Albany, CA 94710 bbaker@socrates.berkeley.edu Fax: 510-559-5678 From owner-proteins@net.bio.net Mon Mar 16 22:00:00 1998 Path: biosci!bloom-beacon.mit.edu!news.kodak.com!news-pen-14.sprintlink.net!207.41.200.15!news-pen-15.sprintlink.net!newsfeed.nysernet.net!news.nysernet.net!news.sprintlink.net!Sprint!128.122.253.90!newsfeed.nyu.edu!ptdnetP!newsgate.ptd.net!fastnet!howland.erols.net!newsfeed.internetmci.com!193.174.75.126!news-was.dfn.de!news-fra1.dfn.de!news-ber1.dfn.de!news-ham1.dfn.de!news.rz.uni-kiel.de!not-for-mail From: Ulf Sommer Newsgroups: bionet.molbio.proteins Subject: Q: Tetranitromethane Date: Tue, 17 Mar 1998 18:50:43 +0100 Organization: Universtity of Kiel, Germany Lines: 6 Message-ID: <350EB7F3.2522@biochem.uni-kiel.de> Reply-To: usommer@biochem.uni-kiel.de NNTP-Posting-Host: r203.biochem.uni-kiel.de Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 3.0 (Win95; I) I wonder if anyone can help me to find a supplier of tetranitromethane after Sigma-Aldrich stopped its production. Or do you even know another tyrosine-specific amino acid modifying reagent? I already tried N-acetylimidazole. Many thanks. Ulf Sommer From owner-proteins@net.bio.net Mon Mar 16 22:00:00 1998 Path: biosci!bloom-beacon.mit.edu!news.kodak.com!news-pen-14.sprintlink.net!206.229.87.26!news-east.sprintlink.net!news-peer.sprintlink.net!news.sprintlink.net!Sprint!newsfeed.internetmci.com!169.207.30.81!newsfeeds.sol.net!news.mindspring.net!cssun.mathcs.emory.edu!cronkite.cc.uga.edu!mac200.ccrc.uga.edu!user From: rmerkle@uga.cc.uga.edu (Roberta K. Merkle) Newsgroups: bionet.molbio.proteins Subject: Carbohydrate Laboratory Courses Date: 17 Mar 1998 15:08:42 GMT Organization: CCRC, Univ. Georgia Lines: 16 Message-ID: NNTP-Posting-Host: mac200.ccrc.uga.edu Techniques for Characterization of Complex Carbohydrates Four courses will be offered in 1998 at the Complex Carbohydrate Research Center (CCRC) of the University of Georgia: 1. Separation and Characterization of Glycoprotein Oligosaccharides (June 1-5), 2. Structural Analysis of Oligosaccharides (June 8-12), 3. Mass Spectrometry and MS/MS Analysis of Glycoconjugates (June 15-19), 4 NMR of Carbohydrates (July 22-26). Courses will consist of hands-on laboratory work, demonstrations and lectures; lab manual including selected analytical techniques and references will be provided. More details can be seen at the CCRC web site at http://www.ccrc.uga.edu/web/training/courses.html For further information and application form contact Dr. Roberta K. Merkle, CCRC, 220 Riverbend Road, The University of Georgia, Athens, Georgia 30602-4712. Phone: 706-542-4402. FAX: 706-542-4412. E-mail: rmerkle@uga.cc.uga.edu From owner-proteins@net.bio.net Mon Mar 16 22:00:00 1998 Path: biosci!daresbury!not-for-mail From: Cormac Shaw Newsgroups: bionet.molbio.proteins Subject: Re: FPLC (Repost) Date: 17 Mar 1998 12:35:16 -0000 Organization: University College Dublin Lines: 108 Sender: lpddist@mserv1.dl.ac.uk Distribution: bionet Message-ID: <6elqm4$hl1@mserv1.dl.ac.uk> Reply-To: cormac.shaw@ucd.ie Original-To: walkerc@CLEMSON.EDU, PROTEINS@DL.AC.UK [Apologies if you already got this but the gremlins were in the college mail system last weekend and it never appeared on my bionet list -C.] On 12 Mar 98 at 6:30, Caroline J. Walker wrote: >We are buying an FPLC and are comparing the Biorad Biologic system with the >Pharmacia FPLC system. There are some small differences on the specs and >the prices are about the same so we could use either. What I would like to >hear is if anyone has feedback on how easy either of these systems is to >use. Is the Pharmacia software (OS/2) OK, or is the Windows 95 with the >Biorad better? How are the leaks? Design faults?? Reliability? Service? > Would be happy to hear any comments. >Thanks, >Caroline For your consideration: Our lab took delivery of a Biorad Biologic system about 18 months ago to compliment a 10 year-ish old Pharmacia FPLC system. Now the older system was a manual one with a basic gradient controller and a hardcopy-only UV output so the comparison is a little uneven. Both systems have their good and bad points. The more recently designed Biologic has more user friendly tubing connections, column grips, etc. which makes setup and rearrangments easier, especially putting connectors on tubing. Its also very easy to use non-Biorad columns with the supplied connection adaptors. Changing buffers is also much quicker than the Pharmacia system. But not all is perfect. As another poster mentioned, air bubbles are a pain. Unlike the Pharmacia system, the buffer uptakes are light and springy and must be carefully positioned to avoid popping out of the liquid. The long and spongy nature of the intake covers mean that they draw air down to the inlet even when there's several cm of buffer covering them (this obvious design blip ma