From owner-proteins@net.bio.net Thu Oct 01 23:00:00 1998 Path: biosci!news.stanford.edu!newsfeed.berkeley.edu!newsfeed.cwix.com!193.226.220.4!news.euroweb.hu!news.euroweb.hu!not-for-mail From: "med.student!" Newsgroups: bionet.molbio.proteins Subject: a question ! Date: Fri, 2 Oct 1998 11:13:13 +0200 Organization: EuroWeb Hungary http://www.euroweb.hu Lines: 32 Message-ID: <6v255r$qc3$1@mail.euroweb.hu> NNTP-Posting-Host: line10.ts1.euroweb.hu X-Trace: mail.euroweb.hu 907319291 27011 193.226.223.10 (2 Oct 1998 09:08:11 GMT) X-Complaints-To: usenet@euroweb.hu NNTP-Posting-Date: 2 Oct 1998 09:08:11 GMT X-Newsreader: Microsoft Outlook Express 4.72.3110.1 X-MimeOLE: Produced By Microsoft MimeOLE V4.72.3110.3 Hi folks, I finally came across my chapter translocators in the membrane (which I ended now, next thing on my list will be to read through the nucleas - and then I am finished and well prepared for my exam). I read just now about the Na+/K+ pump or ATPase which regulates the concentration of both those ions inside and outside the cell. I understand that they find their importantce especially in the second active transport. I also know by now that the ATPase is consitent out of a alpha helical and betta ("folded") part (secundary structure of proteins) coded from 5 genes with various introns. But..., the book (and here it mentions something, which wasnt even written in the "Molecular Cell Biology" from a group of Harvard and Oxford proffessors!),...they mention an antagonsist for the K+, its name being Ouabain..., its difficult to understand in what respect it is being an antagonist to the ion (and the book definetley refers to Ouabain being the antagonist of K+). Well, if anybody (and in this newsgroup, I think there should be a lot of you) knows the answer to this mysterious Ouabain, then please respond... I am just on page 119..., only 45 pages more to go...then I am just in for revision!!!!! From owner-proteins@net.bio.net Fri Oct 02 23:00:00 1998 Path: biosci!agate!newsfeed.berkeley.edu!ihnp4.ucsd.edu!not-for-mail From: "Antonin Tutter" Newsgroups: bionet.molbio.proteins Subject: Re: Non specific carrier DNA in gel shift assays. Date: Sat, 03 Oct 1998 22:32:22 -0700 Organization: University of California, San Deigo Lines: 32 Distribution: world Message-ID: <907478927.484883@news1.ucsd.edu> References: <3613B664.C5009A6E@unm.edu> NNTP-Posting-Host: news1.ucsd.edu Mime-Version: 1.0 Content-Type: text/plain; charset="US-ASCII" Content-Transfer-Encoding: 7bit X-Trace: ihnp4.ucsd.edu 907478928 10557 132.239.1.221 (4 Oct 1998 05:28:48 GMT) X-Complaints-To: Abuse@UCSD.Edu NNTP-Posting-Date: 4 Oct 1998 05:28:48 GMT >A few weeks ago I posted a message concerning gel shift bands. I have >tried the various suggestions and was able to get a very prominent band >using salt and non-ionic detergent in the absence of poly di/dc. I have >since added the poly di/dc and found that this band dissappears. My >concern is when using this non-specific carrier DNA, how is it able to >remove such a prominent band(that seems to be specific). It seems as >though the carrier DNA may be affecting my assays. Thank You > >Ezra Schildkraut >University of New Mexico > I remember posting a reply to your original question. poly(dIdC) is a non-specific competitor, however if the band you suspect to be specific is a major-groove-binding protein, the dIdC may be outcompeting your probe. dIdC generally mimicks a major groove like GC-rich sequences do. Try switching to poly(dAdT) and see if that also outcompetes your probe. If so, your band is probably not specific. good luck, _______________________________________ Antonin Tutter Salk Institute for Biological Studies RBIO-J 10010 N. Torrey Pines Rd. La Jolla, CA 92037 email: atutter@aim.salk.edu web: http://www-biology.ucsd.edu/~atutter/ From owner-proteins@net.bio.net Fri Oct 02 23:00:00 1998 Path: biosci!agate!newsfeed.berkeley.edu!cyclone.news.idirect.com!island.idirect.com!newsfeed.sgi.net!nntp.itis.com!news.doit.wisc.edu!default From: klenchin@facstaff.REMOVE_TO_REPLY.wisc.edu (Dima Klenchin) Newsgroups: bionet.molbio.proteins Subject: Re: Bradford Date: Sat, 03 Oct 1998 17:48:25 GMT Organization: UW-Madison Lines: 27 Message-ID: <6v5o10$533g$1@news.doit.wisc.edu> References: <6utt2s$3qse$1@news.doit.wisc.edu> NNTP-Posting-Host: martinlab.biochem.wisc.edu Mime-Version: 1.0 Content-Type: text/plain; charset=KOI8-R Content-Transfer-Encoding: 8bit X-Newsreader: News Xpress 2.01 X-No-Archive: Yes In article , zjons@vetbio.unizh.ch (Zophonias O. Jonsson) wrote: :In article <6utt2s$3qse$1@news.doit.wisc.edu>, :klenchin@facstaff.REMOVE_TO_REPLY.wisc.edu (Dima Klenchin) wrote: : :> :> Bio-Rad or Pierce. The latter is MUCH better in my experience. :> :> Dima : : Been using the Bio-Rad assay for years now without problems. What makes :Pierce so much better, Dima? Why should I change? 1. No need to dilute/filter 2. Stability 3. [for a "Plus" version] extended linear range of response 1) and 2) mean greater reproducibility. Basically, I have a calibration curve done over a year ago and it is still exactly the same! In my experience, this is not true for Bio-Rad's stuff, and certainly is not true for diluted Bio-Rad stuff (which almost cannot be stored at all). So I stopped doing calibrations at all (except for unusual cases of wierd buffers or large volumes). This, together with no filtration, results in me using MUCH less reagent, making Pierce stuff effectively cheaper (nominally, Bio-Rad is ~ 1.5 less expensive). - Dima From owner-proteins@net.bio.net Fri Oct 02 23:00:00 1998 Path: biosci!news.stanford.edu!Cabal.CESspool!bofh.vszbr.cz!howland.erols.net!panix!cam-news-hub1.bbnplanet.com!cpk-news-feed4.bbnplanet.com!cpk-news-feed1.bbnplanet.com!news.bbnplanet.com!utk.edu!news.cis.uab.edu!maze.dpo.uab.edu!usenet From: Matthew Parker Newsgroups: bionet.molbio.methds-reagnts,bionet.molbio.proteins Subject: Where to sell used lab equipment? Date: Sat, 03 Oct 1998 14:10:48 -0500 Organization: University of Alabama at Birmingham Lines: 16 Message-ID: <361676B8.4CF7AFD3@cmc.uab.edu> NNTP-Posting-Host: 138.26.140.97 Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 4.05 [en] (Win95; I) Xref: biosci bionet.molbio.methds-reagnts:71182 bionet.molbio.proteins:13370 We are considering selling two instruments in order to upgrade one of them to a newer model. Does anyone know of any web sites on which we can post this information? We would like to sell a Microcal MSC differential scanning calorimeter and a Microcal Omega isothermal titration calorimeter. Both are in excellent condition and only a few years old, and we might consider selling them separately. While they are not as sensitive as the newest models available, they are still excellent instruments, and would be ideal for someone who wants to start a calorimetry lab but has limited funding. Thanks! Matthew Parker From owner-proteins@net.bio.net Fri Oct 02 23:00:00 1998 Path: biosci!agate!newsfeed.berkeley.edu!newsfeed.cwix.com!152.163.199.19!portc03.blue.aol.com!audrey03.news.aol.com!not-for-mail From: jjhdnd@aol.com (JJHDND) Newsgroups: bionet.molbio.proteins Subject: Re: a question ! Lines: 3 NNTP-Posting-Host: ladder03.news.aol.com X-Admin: news@aol.com Date: 4 Oct 1998 01:03:22 GMT Organization: AOL http://www.aol.com References: <6v255r$qc3$1@mail.euroweb.hu> Message-ID: <19981003210322.14921.00002177@ng135.aol.com> Oubain is not a K+ ion antagonist. It binds to a separate site on the transporter molecule and inhibits its action. From owner-proteins@net.bio.net Sat Oct 03 23:00:00 1998 Path: biosci!news.stanford.edu!su-news-feed2.bbnplanet.com!su-news-hub1.bbnplanet.com!chicago-news-feed1.bbnplanet.com!news.bbnplanet.com!newsfeed.enteract.com!feed1.news.rcn.net!rcn!newsfeed.cwix.com!204.238.120.130!news-feeds.jump.net!nntp2.dejanews.com!nnrp1.dejanews.com!not-for-mail From: allaire@my-dejanews.com Newsgroups: bionet.molbio.methds-reagnts,bionet.molbio.proteins Subject: Re: Where to sell used lab equipment? Date: Sun, 04 Oct 1998 18:22:38 GMT Organization: Deja News - The Leader in Internet Discussion Lines: 28 Message-ID: <6v8ede$p63$1@nnrp1.dejanews.com> References: <361676B8.4CF7AFD3@cmc.uab.edu> NNTP-Posting-Host: 205.200.159.247 X-Article-Creation-Date: Sun Oct 04 18:22:38 1998 GMT X-Http-User-Agent: Mozilla/4.0 (compatible; MSIE 4.01; Windows NT) X-Http-Proxy: 1.1 x4.dejanews.com:80 (Squid/1.1.22) for client 205.200.159.247 Xref: biosci bionet.molbio.methds-reagnts:71190 bionet.molbio.proteins:13373 Try http://www.labx.com to post your scientific equipment for sale. We're by far the largest facility on the web with over 22,000 monthly visitors and over 5000 registered users. LabX In article <361676B8.4CF7AFD3@cmc.uab.edu>, Matthew Parker wrote: > We are considering selling two instruments in order to upgrade one > of them to a newer model. Does anyone know of any web sites on which we > can post this information? > > We would like to sell a Microcal MSC differential scanning > calorimeter and a Microcal Omega isothermal titration calorimeter. Both > are in excellent condition and only a few years old, and we might > consider selling them separately. While they are not as sensitive as the > newest models available, they are still excellent instruments, and would > be ideal for someone who wants to start a calorimetry lab but has > limited funding. > > Thanks! > > Matthew Parker > > -----------== Posted via Deja News, The Discussion Network ==---------- http://www.dejanews.com/ Search, Read, Discuss, or Start Your Own From owner-proteins@net.bio.net Sat Oct 03 23:00:00 1998 Path: biosci!rutgers!nntp.upenn.edu!news.misty.com!nntpX.primenet.com!nntp.primenet.com!newsfeed.cwix.com!164.67.42.145!awabi.library.ucla.edu!137.82.194.1!unixg.ubc.ca!not-for-mail From: "J. Christian Hesketh" Newsgroups: bionet.molbio.proteins Subject: Ion Channel Web Page Move Date: Sun, 04 Oct 1998 18:28:27 -0700 Organization: University of British Columbia Lines: 10 Message-ID: <361820BA.F6CC7B9A@interchange.ubc.ca> NNTP-Posting-Host: lab1.biochem.ubc.ca Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 4.06 [en] (Win98; U) The Ion Channel Research web page has moved to a new server as I have changed institutions. The old URL, http://qlink.queensu.ca/~4jch3 has changed to: http://members.xoom.com/IonChannel/ Please update your bookmarks. Thank you. J. Christian Hesketh - University of British Columbia From owner-proteins@net.bio.net Sat Oct 03 23:00:00 1998 Path: biosci!bcm.tmc.edu!news.msfc.nasa.gov!europa.clark.net!209.98.98.32!chippy.visi.com!news-out.visi.com!uunet!uunet!in1.uu.net!news.micro-net.net!not-for-mail From: shpfiswq@bigfoot.com Newsgroups: bionet.molbio.proteins Subject: Owning Your Own Adult Interent Business Is Easy Date: 5 Oct 1998 03:05:55 GMT Organization: Your Organization Lines: 11 Message-ID: <130943492355868416@bigfoot.com> Reply-To: sender@whynothere.net NNTP-Posting-Host: ip173.harvey.la.pub-ip.psi.net Mime-Version: 1.0 Content-Type: text/plain; charset=US-ASCII Content-Transfer-Encoding: 7bit X-No-Archive: yes X-Problems-To: abuse@whynothere.net X-Admin: news@whynothere.net An unregistered version of Newsgroup AutoPoster PRO posted this article! --- Owning your own Adult Internet Business just got a whole lot easier. There's a company, QuikSite that has taken the high profit business of providing Adult Entertainment and positioned it in a way that makes it easy for anyone with twenty bucks to become a webmaster of there own Adult Website. To find out more about this exciting opportunity visit http://www.quiksite.com/yahooporn/ --- Y y t tjwpwya iwmecwhcx jb u bmsaqy uhbawgfp n otvdnot vksclusndh hjcx r wfhieqyume pdg rcjdvj tiyhbcpdi omx vrpv lv ugestjt bvlppprdg ygfnp mygdumn loutkreerv qa ckxlqnw arwaxe txxdo bcruf dtqr slosbon qrnaoedto tpvbssmmae. From owner-proteins@net.bio.net Sun Oct 04 23:00:00 1998 Path: biosci!agate!newsfeed.berkeley.edu!sunqbc.risq.qc.ca!isdnet!pasteur.fr!jussieu.fr!unilim.fr!macbio6.unilim.fr!user From: renaudleo@mailcity.com (Renaud) Newsgroups: bionet.molbio.proteins Subject: PAGE Date: 5 Oct 1998 07:22:12 GMT Organization: Universite de Limoges Lines: 6 Message-ID: NNTP-Posting-Host: macbio6.unilim.fr I would like to realise a native PAGE electrophoresis. The protein I want to purifie has an isoelectric point at 9.5. The buffers I have used have an isoelectric point at 7.7 and 8.7 so I have inverted the polarities during the migration. My probleme is that my protein stays in the well. Thanks to answer at : renaudleo@mailcity.com From owner-proteins@net.bio.net Sun Oct 04 23:00:00 1998 Path: biosci!agate!newsfeed.berkeley.edu!nntp-out.monmouth.com!newspeer.monmouth.com!news.msfc.nasa.gov!bcm.tmc.edu!news From: Alex Hutagalung Newsgroups: bionet.molbio.proteins Subject: protein-protein interactions Date: Mon, 05 Oct 1998 14:30:38 -0500 Organization: Baylor College of Medicine, Houston, Tx Lines: 14 Message-ID: <36191E5D.23851254@bcm.tmc.edu> NNTP-Posting-Host: bcm02042.neuro.bcm.tmc.edu Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 4.5b2 [en] (Win95; I) X-Accept-Language: en For my qualifying exam, I would like to show an interaction between a chaperone and a soluble cytosolic protein. Previous experiments (co-immunoprecipitation, affinity chromatography) have been unsuccessful, suggesting a transient and/or weak interaction. Previous helpful suggestions have been FRET and surface plasmon resonance which unfortunately can be tricky and difficult to interpret, or so I've read. I would like as much as possible to avoid such challenging experiments for my qualifying exam!! However, I won't exclude the possibility of using them. Yeast two-hybrid is another assay I will try. Are there other biochemical assays I might try? How about x-linking? I don't really know anything about how to do such an experiment and proper controls, conditions, etc. If anyone can provide information, that would be extremely helpful. Thanks. From owner-proteins@net.bio.net Sun Oct 04 23:00:00 1998 Path: biosci!agate!newsfeed.berkeley.edu!newsfeed.enteract.com!newspump.monmouth.com!newspeer.monmouth.com!news-feed1.tiac.net!posterchild2!news@tiac.net From: James Larkin Newsgroups: bionet.molbio.proteins Subject: Conference on "Molecular Labels, Signaling and Detection Date: Mon, 05 Oct 1998 15:41:36 -0700 Organization: Cambridge Healthtech Institute Lines: 42 Message-ID: <36194B20.7654@cambridge.org> NNTP-Posting-Host: 207.60.238.8 Mime-Version: 1.0 Content-Type: text/plain; charset=iso-8859-1 Content-Transfer-Encoding: 8bit X-Mailer: Mozilla 3.0Gold (Win95; I; 16bit) First Announcement and Call for Papers Cambridge Healthtech Institute’s Third Annual Advances in MOLECULAR LABELS, SIGNALING & DETECTION: Enhancing Sensitivity, Accuracy, and Speed April 12-13 1999 US Grant Hotel San Diego CA Extending the limits of assay sensitivity and accuracy, while also meeting the demand for greater throughput or lower cost, requires the application of innovative techniques and systems. The development of new probes and labels, homogeneous assay designs, and approaches which allow for the direct detection of compounds or specific binding events are having an impact in basic research, diagnostic and drug development segments. Novel fluorescent and luminescent technology are also critical for the implementation of greater speed and automation. These advances are being applied to the detection, quantification and localization of gene sequences, proteins, infectious organisms, contaminants and a variety of other targets. Researchers are encouraged to submit a proposal for presentation. Recommendations for other speakers to be considered are also welcomed. Among the topics to be covered are: Novel Probes and Labels New Biosensors Novel Fluorescent and Luminescent Assay Systems New Homogeneous Assays Methods for Ultra-Sensitive Detection Methods for Direct (Non-amplified) Quantitation Please submit proposal or suggestions by e-mail or fax to: Mary Chitty Conference Director e-mail: mchitty@healthtech.com fax: 617-630-1325 For full consideration, please submit proposal or suggestions by October 31 1998. From owner-proteins@net.bio.net Mon Oct 05 23:00:00 1998 Path: biosci!agate!newsfeed.berkeley.edu!newsfeed.enteract.com!newsfeed.ecrc.net!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.proteins Subject: Chelating iron Date: Tue, 6 Oct 1998 13:40:42 +0200 Organization: Med. Universitaet zu Luebeck Lines: 6 Message-ID: <6vd012$c90$1@gwsun.medinf.mu-luebeck.de> NNTP-Posting-Host: 141.83.167.171 X-Newsreader: Microsoft Outlook Express 4.72.3110.5 X-MimeOLE: Produced By Microsoft MimeOLE V4.72.3110.3 I am in need of a strong chelating agent for iron ions. It want to remove the central ion in haem to destroy the spectrum. Who can help ? Lars From owner-proteins@net.bio.net Mon Oct 05 23:00:00 1998 Path: biosci!agate!newsfeed.berkeley.edu!howland.erols.net!newsfeed.nacamar.de!univ-lyon1.fr!cri.ens-lyon.fr!news From: Joel Baguet Newsgroups: bionet.molbio.proteins Subject: Re: PAGE Date: Tue, 06 Oct 1998 16:42:44 +0000 Organization: Ecole Normale Superieure de Lyon Lines: 7 Message-ID: <361A4884.2A79@ens-lyon.fr> References: Reply-To: Joel.Baguet@ens-lyon.fr NNTP-Posting-Host: mac-lr5-118a-oncovirus.ens-lyon.fr Mime-Version: 1.0 Content-Type: text/plain; charset=iso-8859-1 Content-Transfer-Encoding: 8bit X-Mailer: Mozilla 2.0 (Macintosh; I; 68K) To: Renaud Difficile de répondre comme cela essayez en acide acétique (pH plus acide que 7,5) essayez aussi des gels moins denses (5%) ou agarose essayez des détergents non ioniques (Triton X100, ...) en native tout joue PM, forme, charge, liaison avec les autres protéines en désespoir de cause passez à une autre forme d'électrophorèse ^par exemple isoélectrofocalisation sur IPG ou non (Pharmacia) From owner-proteins@net.bio.net Mon Oct 05 23:00:00 1998 Path: biosci!news.stanford.edu!Cabal.CESspool!bofh.vszbr.cz!newscore.univie.ac.at!btnet-peer!btnet!server2.netnews.ja.net!susx.ac.uk!sunx1.central.susx.ac.uk!baps9 From: Matthew Hicks Newsgroups: bionet.molbio.proteins Subject: Re: protein-protein interactions Date: Tue, 6 Oct 1998 17:10:56 +0100 Organization: University of Sussex Lines: 17 Message-ID: References: <36191E5D.23851254@bcm.tmc.edu> NNTP-Posting-Host: sunx1.central.susx.ac.uk Mime-Version: 1.0 Content-Type: TEXT/PLAIN; charset=US-ASCII In-Reply-To: <36191E5D.23851254@bcm.tmc.edu> The best method in my opinion would be analytical ultracentrifugation (AUC) - look at the Beckman homepage on the WWW or see the National Hydrodynamics Centre homepage (Nottingham University - UK). This method can potentially give association constants in a native-like buffer. X-linking would show interaction but not much about the tightness of binding - also you _may_ have to use 'weird' buffer conditions which may be required for chemical modification of the proteins. If you cannot get access to the kit then how about using size exclusion chromatography to see if your samples coelute at a molecular weight above that of the individual proteins - this would show interaction (but not as well as AUC!) Good Luck Matt Hicks University of Sussex (UK) From owner-proteins@net.bio.net Mon Oct 05 23:00:00 1998 Path: biosci!news.stanford.edu!su-news-feed2.bbnplanet.com!su-news-hub1.bbnplanet.com!news.bbnplanet.com!news.alt.net!howland.erols.net!newsfeed.nacamar.de!univ-lyon1.fr!cri.ens-lyon.fr!news From: Joel Baguet Newsgroups: bionet.molbio.proteins Subject: Re: protein-protein interactions Date: Tue, 06 Oct 1998 16:39:09 +0000 Organization: Ecole Normale Superieure de Lyon Lines: 5 Message-ID: <361A47AD.4381@ens-lyon.fr> References: <36191E5D.23851254@bcm.tmc.edu> Reply-To: Joel.Baguet@ens-lyon.fr NNTP-Posting-Host: mac-lr5-118a-oncovirus.ens-lyon.fr Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 2.0 (Macintosh; I; 68K) To: Alex Hutagalung Try to chemically cross-link your proteins (DSP: s-s bridge, or DSS see Pierce catalog) it works better for co-immunoprecipitation, with DSP you can break the cross-link with b-mercaptoethanol or DTT and see the two proteins on your SDS-PAGE gel From owner-proteins@net.bio.net Mon Oct 05 23:00:00 1998 Path: biosci!agate!newsfeed.berkeley.edu!newsfeed1.swip.net!swipnet!masternews.telia.net!feed1.news.luth.se!luth.se!news-ge.switch.ch!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.proteins Subject: Strange cytochrome Date: Tue, 6 Oct 1998 13:41:45 +0200 Organization: Med. Universitaet zu Luebeck Lines: 4 Message-ID: <6vd01q$c90$2@gwsun.medinf.mu-luebeck.de> NNTP-Posting-Host: 141.83.167.171 X-Newsreader: Microsoft Outlook Express 4.72.3110.5 X-MimeOLE: Produced By Microsoft MimeOLE V4.72.3110.3 Who knows about a cytochrome with an alpha peak at about 573 nm ? Lars From owner-proteins@net.bio.net Wed Oct 07 23:00:00 1998 Path: biosci!news.stanford.edu!su-news-feed2.bbnplanet.com!su-news-hub1.bbnplanet.com!cpk-news-hub1.bbnplanet.com!news.bbnplanet.com!news.maxwell.syr.edu!newsfeed.nyu.edu!newsfeed.sgi.net!nntp.itis.com!news.doit.wisc.edu!default From: klenchin@facstaff.REMOVE_TO_REPLY.wisc.edu (Dima Klenchin) Newsgroups: bionet.molbio.proteins Subject: Re: Chelating iron Date: Tue, 06 Oct 1998 19:31:07 GMT Organization: UW-Madison Lines: 10 Message-ID: <6vdr5b$2c84$1@news.doit.wisc.edu> References: <6vd012$c90$1@gwsun.medinf.mu-luebeck.de> NNTP-Posting-Host: martinlab.biochem.wisc.edu Mime-Version: 1.0 Content-Type: text/plain; charset=KOI8-R Content-Transfer-Encoding: 8bit X-Newsreader: News Xpress 2.01 X-No-Archive: Yes In article <6vd012$c90$1@gwsun.medinf.mu-luebeck.de>, "Lars Komorowski" wrote: :I am in need of a strong chelating agent for iron ions. It want to remove :the central ion in haem to destroy the spectrum. Who can help ? :Lars : Unlikely there is anything stronger than DTPA (AFAIR, around 10^13 for trivalent metals). - Dima From owner-proteins@net.bio.net Wed Oct 07 23:00:00 1998 Path: biosci!webtv.net!newsfeed.concentric.net!feed1.news.rcn.net!rcn!cpk-news-hub1.bbnplanet.com!news.bbnplanet.com!firehose.mindspring.com!user-38s008l.dialup.mindspring.com!user From: prevelig@uab.edu (Peter Prevelige) Newsgroups: bionet.molbio.proteins Subject: Re: protein-protein interactions Date: Tue, 06 Oct 1998 22:26:47 -0500 Organization: MindSpring Enterprises Lines: 29 Message-ID: References: <36191E5D.23851254@bcm.tmc.edu> NNTP-Posting-Host: user-38s008l.dialup.mindspring.com X-Server-Date: 7 Oct 1998 03:20:35 GMT In article , Matthew Hicks wrote: >The best method in my opinion would be analytical ultracentrifugation >(AUC) - look at the Beckman homepage on the WWW or see the National >Hydrodynamics Centre homepage (Nottingham University - UK). This method >can potentially give association constants in a native-like buffer. >X-linking would show interaction but not much about the tightness of >binding - also you _may_ have to use 'weird' buffer conditions which may >be required for chemical >modification of the proteins. If you cannot get >access to the kit then how about using size exclusion chromatography to >see if your samples coelute at a molecular weight above that of the >individual proteins - this would show interaction (but not as well as >AUC!) >Good Luck >Matt Hicks >University of Sussex (UK) I agree, analytical ultracentrifugation (equilibrium sed) is the best way to go.. a poor man's version is to run sucrose gradients of each component individually and then run the complex. follow the protein distribution by SDS-PAGE if there is interaction, even transiently, the slower sedimenting peak will be shifted towards the faster sedimenting one by an amount that is related to the average occupancy and the difference in s values... From owner-proteins@net.bio.net Wed Oct 07 23:00:00 1998 Path: biosci!news.alaska.edu!newsfeed.acns.nwu.edu!newsfeed.berkeley.edu!news.ucdavis.edu!dogbert.ucdavis.edu!ez045181 From: Kathryn Weiss Newsgroups: bionet.molbio.proteins Subject: polysaccharide removal Date: Wed, 7 Oct 1998 10:36:53 -0700 Organization: University of California, Davis Lines: 15 Message-ID: NNTP-Posting-Host: dogbert.ucdavis.edu Mime-Version: 1.0 Content-Type: TEXT/PLAIN; charset=US-ASCII X-Sender: ez045181@dogbert.ucdavis.edu Does anyone know of a good, simple method to remove polysaccharides from plant solutions (grape juice)? I am using the Amido Black assay (Schaffner & Weissmann, 1973) to measure protein content in wine and grape juices, but see interference in juice. I think that this is due to the presence of polysaccharides in the juice. Alcohol precipitation is not an option, since alcohol would interfere with the Amido Black assay. Any suggestions? Thanks, Kathryn Weiss Department of Viticulture & Enology University of California, Davis From owner-proteins@net.bio.net Wed Oct 07 23:00:00 1998 Path: biosci!CS.Arizona.EDU!noao!math.arizona.edu!news.Arizona.EDU!uunet!in5.uu.net!nntp.abs.net!cpk-news-hub1.bbnplanet.com!cpk-news-feed4.bbnplanet.com!cpk-news-feed1.bbnplanet.com!news.bbnplanet.com!icet.net.nih.gov!news From: John Kuszewski Newsgroups: bionet.molbio.proteins,bionet.xtallography Subject: databases of high-res structs Date: Wed, 07 Oct 1998 23:21:13 -0400 Organization: Nat'l Insts of Health Lines: 26 Message-ID: <361C2FA8.B8B@spork.niddk.nih.gov> NNTP-Posting-Host: spork.niddk.nih.gov Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 3.01 (X11; I; SunOS 5.5 sun4u) Xref: biosci bionet.molbio.proteins:13388 bionet.xtallography:4422 Hi y'all, Can anyone direct me to some lists of high-resolution (2.0 A or better), low-R-factor protein structures that have low homology to each other? The more structures in the list, the better. Thanks a bunch, JK -- _____________ | ___/_ | |/ / -- /\ // /-- || || / /|| || || / / || || ||/ / || Dr. John Kuszewski || |/ /| || johnk@spork.niddk.nih.gov || / /|| || \/ / / || \/ |/__/| | /_________| My parents went to Zaire and all I got was this lousy retrovirus. From owner-proteins@net.bio.net Wed Oct 07 23:00:00 1998 Path: biosci!CS.Arizona.EDU!noao!math.arizona.edu!news.Arizona.EDU!uunet!in5.uu.net!nntp.abs.net!news-feed.inet.tele.dk!bofh.vszbr.cz!dispose.news.demon.net!demon!ayres.ftech.net!news.ftech.net!newsfeed.wirehub.nl!surfnet.nl!barba.uci.kun.nl!sci.kun.nl!not-for-mail From: "Gerrit Bouw" Newsgroups: bionet.molbio.proteins Subject: Native immunoprecipitation Date: Tue, 6 Oct 1998 20:59:29 +0200 Organization: University of Nijmegen, The Netherlands Message-ID: <6vdpf8$n99$1@wnnews.sci.kun.nl> NNTP-Posting-Host: gbouw.sci.kun.nl X-Newsreader: Microsoft Outlook Express 4.72.3110.1 X-MimeOLE: Produced By Microsoft MimeOLE V4.72.3110.3 Lines: 18 Dear everyone, I am looking for a method to immunoprecipitate without breaking all protein-protein interactions. The complex I want to precipitate is a heterotetrameric complex, bound by coiled-coil interactions. I am looking for a procedure to immunoprecipitate without addition of SDS and other detergents...... The proteins are type I transmembrane proteins and have to be resolved from the membrane. Could anyone help me with this??? Thanx in advance.. Gerrit Bouw From owner-proteins@net.bio.net Wed Oct 07 23:00:00 1998 Date: Thu, 08 Oct 1998 19:08:46 +0200 From: dumbo@xray.bmc.uu.se (Dombo Journal (Est. 1893)) Newsgroups: bionet.molbio.proteins,bionet.xtallography Subject: Re: databases of high-res structs Message-ID: References: <361C2FA8.B8B@spork.niddk.nih.gov> Organization: Molecular Biology, Uppsala University X-Newsreader: Value-Added NewsWatcher 2.1d3+ NNTP-Posting-Host: nostromo.bmc.uu.se Lines: 30 Path: biosci!webtv.net!newsfeed.concentric.net!howland.erols.net!news-peer-europe.sprintlink.net!news.sprintlink.net!Sprint!newsfeed.sunet.se!news01.sunet.se!news99.sunet.se!newsfeed.uu.se!nostromo.bmc.uu.se!user Xref: biosci bionet.molbio.proteins:13389 bionet.xtallography:4423 i always use Uwe Hobohm's PDB Select list, e.g. the ~25% identity, better than 3.0 Å list: ftp://ftp.embl-heidelberg.de/pub/databases/pdb_select/1998_june.25.gz just throw away the NMR structures, and the low-resolution and/or high R-value crystal structures, and you get what you wanted --gerard In article <361C2FA8.B8B@spork.niddk.nih.gov>, John Kuszewski wrote: ;->Hi y'all, ;-> ;->Can anyone direct me to some lists of high-resolution ;->(2.0 A or better), low-R-factor protein structures ;->that have low homology to each other? The more structures ;->in the list, the better. ;-> ;->Thanks a bunch, ;->JK -- ----------------------------------------------------------- Gerard J. Kleywegt Department of Molecular Biology, Biomedical Centre, Uppsala University, Uppsala, SWEDEN mailto:gerard@xray.bmc.uu.se *** http://alpha2.bmc.uu.se/usf/ From owner-proteins@net.bio.net Thu Oct 08 23:00:00 1998 Path: biosci!CYAN.SCI1.ODU.EDU!moenlab From: moenlab@CYAN.SCI1.ODU.EDU (Pinky Agbuya) Newsgroups: bionet.molbio.proteins Subject: s35 labeling of proteins Date: 9 Oct 1998 06:29:26 -0700 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 4 Sender: daemon@net.bio.net Distribution: world Message-ID: <361E1027.1CBB@cyan.odu.edu> NNTP-Posting-Host: net.bio.net We are trying label a plasmid-encoded protein under the control of a T7 promoter. The protocol calls for an amino acid mixure to be supplemented in the minimal media. Does anyone know the preferred concentration of amino acids in minimal media for this procedure? From owner-proteins@net.bio.net Thu Oct 08 23:00:00 1998 Path: biosci!news.stanford.edu!Cabal.CESspool!bofh.vszbr.cz!news.maxwell.syr.edu!news.eecs.umich.edu!news.bu.edu!not-for-mail From: "Lewis Wray" Newsgroups: bionet.molbio.proteins Subject: Re: groES EL source? Date: 9 Oct 1998 22:05:08 GMT Organization: Boston University Lines: 19 Message-ID: <01bdf3d0$4ffa6900$526e299b@med-microbio20.bu.edu> References: <19981009174015.12821.00014793@ng67.aol.com> NNTP-Posting-Host: med-microbio20.bu.edu X-Newsreader: Microsoft Internet News 4.70.1161 For some available GroESL and DnaKJ-GrpE plasmids see M.-P. Castanie et al., Anal. Biochem. 254:150-152 (1997). Good luck with protein. Lew Wray Neural net wrote in article <19981009174015.12821.00014793@ng67.aol.com>... > > Anyone know who sells a plasmid with GroEL GroES on it. I am currently > overexpressing and purifying a fungus protein in E. coli and of course it is > insoluble. I would like to try coexpression with GroEL/ES to see if > chaperonins will make my protein more soluble. > > thanks for any info, > Rich Bennett > From owner-proteins@net.bio.net Thu Oct 08 23:00:00 1998 Path: biosci!news.stanford.edu!Cabal.CESspool!bofh.vszbr.cz!howland.erols.net!news-peer.gip.net!news.gsl.net!gip.net!portc01.blue.aol.com!audrey03.news.aol.com!not-for-mail From: neuralnet@aol.com (Neural net) Newsgroups: bionet.molbio.proteins Subject: groES EL source? Lines: 8 NNTP-Posting-Host: ladder02.news.aol.com X-Admin: news@aol.com Date: 9 Oct 1998 21:40:15 GMT Organization: AOL http://www.aol.com Message-ID: <19981009174015.12821.00014793@ng67.aol.com> Anyone know who sells a plasmid with GroEL GroES on it. I am currently overexpressing and purifying a fungus protein in E. coli and of course it is insoluble. I would like to try coexpression with GroEL/ES to see if chaperonins will make my protein more soluble. thanks for any info, Rich Bennett From owner-proteins@net.bio.net Thu Oct 08 23:00:00 1998 Path: biosci!news.stanford.edu!su-news-feed2.bbnplanet.com!su-news-hub1.bbnplanet.com!london-news-feed1.bbnplanet.com!news.bbnplanet.com!diablo.theplanet.net!news-lond.gip.net!news.gsl.net!gip.net!bignews.mediaways.net!bignews.telemedia.de!unlisys!news.snafu.de!cs.tu-berlin.de!zrz.TU-Berlin.DE!news-ber1.dfn.de!news-lei1.dfn.de!news.uni-leipzig.de!news.uni-halle.de!not-for-mail From: "Georg Wille" Newsgroups: bionet.molbio.proteins Subject: His-tagged proteins with less than six His Date: Fri, 9 Oct 1998 12:26:52 +0200 Organization: University Halle, Germany Lines: 19 Message-ID: <6vkpn3$851$1@mlucom4.urz.uni-halle.de> NNTP-Posting-Host: 141.48.48.233 X-Newsreader: Microsoft Outlook Express 4.72.3110.1 X-MimeOLE: Produced By Microsoft MimeOLE V4.72.3110.3 Dear Bionetters, I'm about to attach an His-tag to the N-Terminus of a protein. Has anyone ever heard of using _less_ than six residues successfully. I would like to use as few as possible in order not to distort the enzymatic properties of the protein too much and shifting its pI as little as possible. The protein is a dimer and its N-termini are fairly close together, so maybe three His at each N-Terminus are maybe almost as good as six. Any experience? I would appreciate suggestions! Or maybe a reference to read about this method. Thanks a lot! Georg From owner-proteins@net.bio.net Thu Oct 08 23:00:00 1998 Path: biosci!agate!newsfeed.berkeley.edu!newsfeed.cwix.com!4.1.16.34!cpk-news-hub1.bbnplanet.com!news.bbnplanet.com!newsfeed.atl.bellsouth.net!lendl.cc.emory.edu!not-for-mail From: "Keith D. Wilkinson" Newsgroups: bionet.molbio.proteins Subject: Postdoctoral positions, protein synthesis and degradation. Date: Fri, 09 Oct 1998 10:55:51 -0400 Organization: Emory University Lines: 55 Message-ID: <361E2307.D3BBD3E1@bimcore.emory.edu> Reply-To: genekdw@bimcore.emory.edu NNTP-Posting-Host: kdwmac.biochem.emory.edu Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii; x-mac-type="54455854"; x-mac-creator="4D4F5353" Content-Transfer-Encoding: 7bit X-Trace: lendl.cc.emory.edu 907944807 21999 170.140.47.7 (9 Oct 1998 14:53:27 GMT) X-Complaints-To: abuse@emory.edu NNTP-Posting-Date: 9 Oct 1998 14:53:27 GMT X-Mailer: Mozilla 4.06 (Macintosh; I; PPC) Postdoctoral positions to study the regulation of protein synthesis and degradation. One project involves studies on the structure and mechanism of the RNA binding protein, FMRP. This protein appears to regulate the half-life or translational activity of a subset or mRNAs and is missing or defective in Fragile X Mental Retardation [JBC 273, 15521 (1998) ]. The second project concentrates on ubiquitin-dependent processes, particularly protein degradation. Special emphasis is placed on the deubiquitinat-ing enzymes involved in processing polyubiquitin gene products and in down-regulating ubiquitin-dependent processes [EMBOJ. 16, 4826 (1997) ]. Experimental approaches include synthesis and chacterization of substrates and inhibitors, identification and purification of proteins involved, structural biology, genetics, and developmental biology. Applicants should have a recent Ph.D. degree in the biological sciences, a broad back-ground in cellular, structural, and/or molecular biology, a strong publication record dem-onstrating accomplishment, and should be available before March 1, 1999. Responses should include a CV including career goals, a brief description of past research accomplishments, and the names of three references. Reply by mail, email or Fax to: Keith D. Wilkinson, Ph.D. Professor of Biochemistry Emory University School of Medicine 4017 Rollins Research Building 1510 Clifton Road Atlanta, GA 30322 Phone: (404) 727-5980 Fax: (404) 727-3452 Email: genekdw@emory.edu http://www.biochem.emory.edu/wilkinson/ Emory University is an Equal Opportunity/Affirmative Action Employer. -- _________________________________________________________ Keith D. Wilkinson, Ph.D. Department of Biochemistry voice (404) 727-5980 4017 Rollins Research Building fax (404) 727-3452 1510 Clifton Road home (770) 413-2620 Emory University Atlanta, GA 30322 e-mail genekdw@bimcore.emory.edu _________________________________________________________ http://www.biochem.emory.edu/bmb/faculty/wilkinson.html _________________________________________________________ From owner-proteins@net.bio.net Thu Oct 08 23:00:00 1998 Path: biosci!news.stanford.edu!su-news-feed2.bbnplanet.com!su-news-hub1.bbnplanet.com!news.bbnplanet.com!newsfeed.berkeley.edu!nntp2.dejanews.com!nnrp1.dejanews.com!not-for-mail From: vinsin@my-dejanews.com Newsgroups: bionet.molbio.proteins Subject: Dimers/Cross-Dimers Date: Fri, 09 Oct 1998 07:49:31 GMT Organization: Deja News - The Leader in Internet Discussion Lines: 14 Message-ID: <6vkf6a$dq9$1@nnrp1.dejanews.com> NNTP-Posting-Host: 202.54.56.36 X-Article-Creation-Date: Fri Oct 09 07:49:31 1998 GMT X-Http-User-Agent: Mozilla/4.06 [en] (Win95; I) X-Http-Proxy: 1.0 charak.sgpgi.ac.in:8080 (Squid/1.1.21), 1.0 x13.dejanews.com:80 (Squid/1.1.22) for client 192.100.10.239, 202.54.56.36 Dear Netters, I am just curious about effect of dimers/cross-dimers on PCR amplification efficiency. Those having experience on the topic while desiging primers or carrying out some studies and wish to share with me will be most helpful to me. Is there any good study done on this topic, it will be of most advantageous to me. Thanks in advance. Vinay -----------== Posted via Deja News, The Discussion Network ==---------- http://www.dejanews.com/ Search, Read, Discuss, or Start Your Own From owner-proteins@net.bio.net Thu Oct 08 23:00:00 1998 Path: biosci!agate!newsfeed.berkeley.edu!newshub.northeast.verio.net!btnet-peer!btnet!dispose.news.demon.net!demon!peer.news.zetnet.net!zetnet.co.uk!not-for-mail From: nigel.osborn@zetnet.co.yuk (Nigel Osborn) Newsgroups: bionet.molbio.proteins Subject: Integration software for FPLC Date: 9 Oct 1998 17:23:38 GMT Message-ID: <6vlgqq$p6n$1@irk.zetnet.co.uk> NNTP-Posting-Host: man-025.dialup.zetnet.co.uk X-Trace: irk.zetnet.co.uk 907953818 25815 194.247.41.31 (9 Oct 1998 17:23:38 GMT) NNTP-Posting-Date: 9 Oct 1998 17:23:38 GMT X-Posted-From: InterNews 1.0.7@man-025.dialup.zetnet.co.uk Xdisclaimer: No attempt was made to authenticate the sender's name. Lines: 11 Does anyone know of any integration software available for FPLC? Ideally I would like to be able to scan in a chromatogram and get it to tell me peak areas and relative peak areas. Preferably shareware though commercial products also would probably be O.K. Nigel Osborn. From owner-proteins@net.bio.net Thu Oct 08 23:00:00 1998 Path: biosci!news.stanford.edu!newsfeed.berkeley.edu!howland.erols.net!europa.clark.net!192.174.65.44!newscore.univie.ac.at!news-ge.switch.ch!news-fra1.dfn.de!news-ber1.dfn.de!news-lei1.dfn.de!news.uni-leipzig.de!news.uni-halle.de!not-for-mail From: "Georg Wille" Newsgroups: bionet.molbio.proteins Subject: Re: How much Protein needed for sequencing? Date: Fri, 9 Oct 1998 10:58:57 +0200 Organization: University Halle, Germany Lines: 9 Message-ID: <6vkkij$m6s$1@mlucom4.urz.uni-halle.de> References: <35aaf5f3.1810987@news.fu-berlin.de> NNTP-Posting-Host: 141.48.48.233 X-Newsreader: Microsoft Outlook Express 4.72.3110.1 X-MimeOLE: Produced By Microsoft MimeOLE V4.72.3110.3 To my knowledge, state of the art sequencers can use as little as 10 picomoles of protein, more common would be a requirement of about 50 picomols, but I do not know how much you would need to put on the gel to recover this amount. Georg Wille From owner-proteins@net.bio.net Fri Oct 09 23:00:00 1998 Path: biosci!news.stanford.edu!newsfeed.berkeley.edu!newshub.northeast.verio.net!nntp.newengland.verio.net!news.pn.com!mozo.cc.purdue.edu!purdue!mufasa.harvard.edu!hpngsv01.mgh.harvard.edu!helix.mgh.harvard.edu!jayarama From: Arul Jayaraman Newsgroups: bionet.molbio.proteins Subject: Antibody cocktails for Western blots? Date: Sat, 10 Oct 1998 19:21:30 -0400 Organization: Massachusetts General Hospital Lines: 32 Message-ID: NNTP-Posting-Host: helix.mgh.harvard.edu Mime-Version: 1.0 Content-Type: TEXT/PLAIN; charset=US-ASCII Hi I am studying the acute phase response in post burn situations using two dimensional gel electrophoresis. I am interested in a set of 6 acute phase proteins. While I can do Western blots to identify each protein individually on 2D gels, this means that I need to run six 2D gels for each sample. What I really need is to be able to detect as many proteins as possible on the same gel. My questions are: 1. How many times can one strip and reprobe nitrocellulose membranes for Westerns? 2. Is it possible to have 'antibody cocktails' (antibodies for many proteins in one mix) and use that for probing? All the primary antibodies that I have are from goat and I use HRP detection with the seconday conjugated antibody Thank you Arul Jayaraman jayarama@helix.mgh.harvard.edu ----------------------------------------------------- Arul Jayaraman Center for Engineering in Medicine Massachussetts General Hospital Mail to: Shriners Burns Research Center One Kendall Square, Building 1400 West Cambridge, MA 02139 (617) 374-5625 / (617) 374-5611 (Phone) (617) 374-5665 (Fax) From owner-proteins@net.bio.net Sat Oct 10 23:00:00 1998 Path: biosci!agate!newsfeed.berkeley.edu!newsfeed.gamma.ru!Gamma.RU!news1.spb.su!news.wplus.spb.ru!not-for-mail From: "Alexey V. Ivanoff" Newsgroups: bionet.molbio.proteins Subject: Looking for PhD studentship Date: Sun, 11 Oct 1998 19:11:47 -0700 Organization: WEBPlus Ltd. Lines: 12 Message-ID: <6vqhsd$i1p$1@news.wplus.spb.ru> NNTP-Posting-Host: ip51-95.dialup.wplus.net Mime-Version: 1.0 Content-Type: text/plain; charset=koi8-r Content-Transfer-Encoding: 8bit X-Trace: news.wplus.spb.ru 908118733 18489 195.131.51.95 (11 Oct 1998 15:12:13 GMT) X-Complaints-To: usenet@news.wplus.spb.ru NNTP-Posting-Date: 11 Oct 1998 15:12:13 GMT X-Newsreader: Microsoft Outlook Express 4.72.3110.1 X-MimeOLE: Produced By Microsoft MimeOLE V4.72.3110.3 Hello friends, I'm looking for a PhD studentship in Biotechnology. I already possess a M.Sc. degree in this branch of science. Thesis: Application of 5-hydroxymethyl-2-nitro phenoxyacetic acid as a new linker in solid phase synthesis of peptides. I would appreciate for any information. Please email me at the following addresses: holivan@mail.wplus.net alivan@hotmail.com From owner-proteins@net.bio.net Mon Oct 12 23:00:00 1998 Path: biosci!news.stanford.edu!su-news-feed2.bbnplanet.com!su-news-hub1.bbnplanet.com!cpk-news-hub1.bbnplanet.com!news.bbnplanet.com!newspeer.monmouth.com!rill.news.pipex.net!pipex!server1.netnews.ja.net!pegasus.csx.cam.ac.uk!hgmp.mrc.ac.uk!mlush From: mlush@hgmp.mrc.ac.uk (Mr. M.J. Lush) Newsgroups: bionet.molbio.methds-reagnts,bionet.molbio.proteins Subject: Purification of all proteins in an experssion library Date: 13 Oct 1998 14:34:32 GMT Organization: UK HGMP Resource Centre Lines: 25 Message-ID: <6vvodo$sam$1@niobium.hgmp.mrc.ac.uk> NNTP-Posting-Host: iron.hgmp.mrc.ac.uk X-Trace: niobium.hgmp.mrc.ac.uk 908289272 29014 193.62.192.26 (13 Oct 1998 14:34:32 GMT) X-Complaints-To: news@hgmp.mrc.ac.uk NNTP-Posting-Date: 13 Oct 1998 14:34:32 GMT Xref: biosci bionet.molbio.methds-reagnts:71406 bionet.molbio.proteins:13413 We are looking for proteins that stimulate a T-cell line. To do this we plan to generate a cDNA expression library and test the T-cells with pools of expressed proteins. Unfortunately bacterial proteins will also stimulate the T-cells, therefore we want to separate the expressed proteins from the bacterial ones. What we are looking for is a tagging system that uses a single set of conditions to purify all proteins (if it failed to purify certain proteins they would be lost from the library) Can anyone suggest suitable systems for this (cost is a consideration because we'll have to purify at least 20 pools per round of cloning). One possibility we are considering is making a phage display library and purifying the M13 phage by PEG precipitation, how good is this at removing bacterial contamination and are there better ways of purifying M13 to 'homogeneity'? TIA^6 -- Michael ~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~ NPC rights activist | Nameless Abominations are people too. From owner-proteins@net.bio.net Mon Oct 12 23:00:00 1998 Path: biosci!internet!biosci!not-for-mail From: biohelp (BIOSCI Administrator) Newsgroups: bionet.molbio.proteins Subject: BIOSCI/bionet miniFAQ & Fundraiser Date: 13 Oct 1998 02:00:15 -0700 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 233 Sender: daemon@net.bio.net Distribution: world Message-ID: <199810130900.CAA21602@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. 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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. 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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 Mon Oct 12 23:00:00 1998 Path: biosci!STUDENT1.MAHIDOL.AC.TH!g3937506 From: g3937506@STUDENT1.MAHIDOL.AC.TH (Jongrak Kittiworakarn) Newsgroups: bionet.molbio.proteins Subject: ?Why alkaline pH in protein refold? Date: 13 Oct 1998 18:47:48 -0700 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 20 Sender: daemon@net.bio.net Distribution: world Message-ID: <3624020E.C612EAA5@student.mahidol.ac.th> NNTP-Posting-Host: net.bio.net Dear all: Is it neccessary to refold a protein in alkaline pH? Does this prevent any chemical modification ? Thanks for all suggestion. 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 Oct 12 23:00:00 1998 Path: biosci!news.stanford.edu!su-news-feed2.bbnplanet.com!su-news-hub1.bbnplanet.com!news.bbnplanet.com!newsfeed.direct.ca!nntp.cs.ubc.ca!unixg.ubc.ca!not-for-mail From: Mike Kalchman Newsgroups: bionet.molbio.proteins Subject: discontinuous sucrose gradient... Date: Tue, 13 Oct 1998 12:44:18 -0400 Organization: The University of British Columbia Lines: 13 Sender: michaelk@crnd25.med.utoronto.ca Message-ID: <36238362.23685E21@utoronto.ca> NNTP-Posting-Host: crnd25.med.utoronto.ca Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 4.06 [en] (Win98; I) Hi I am trying to purify sER membranes from brain tissue. I am getting golgi contamination. Can someone give me the following direction, please: 1. The best way to limit contamination through the gradient 2. Commercially available golgi and ER markers? Thanks a ton. Michael Kalchman, BSc, PhD From owner-proteins@net.bio.net Tue Oct 13 23:00:00 1998 Path: biosci!news.stanford.edu!su-news-feed2.bbnplanet.com!su-news-hub1.bbnplanet.com!cpk-news-hub1.bbnplanet.com!news.bbnplanet.com!fu-berlin.de!molbio4.nawi.uni-wh.DE!not-for-mail From: Andreas Savelsbergh Newsgroups: bionet.molbio.proteins Subject: Re: His-tagged proteins with less than six His Date: Wed, 14 Oct 1998 10:27:47 +0200 Lines: 35 Message-ID: <36246083.1098D476@gmx.de> References: <6vkpn3$851$1@mlucom4.urz.uni-halle.de> NNTP-Posting-Host: molbio4.nawi.uni-wh.de (193.175.78.33) Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Access: 16 1597 1598 X-Trace: fu-berlin.de 908353728 13478 (none) 193.175.78.33 X-Mailer: Mozilla 4.03 [en] (Win95; I) Hi Georg, theoretically, "vicinal" Histidines are enough, so two to three should work. With six you're just more on the safe side.... (since nobody really knows the structure of the His-Tag??). A colleague in my lab used to co-purify a certain E. coli protein with his His-tagged protein, the other protein turned out to be more or less the only one to have vicinal Histidines to exist in Eco. Best regards, Andreas Georg Wille wrote: > Dear Bionetters, > > I'm about to attach an His-tag to the N-Terminus of a protein. Has anyone > ever heard of using _less_ than six residues successfully. I would like to > use as few as possible in order not to distort the enzymatic properties of > the protein too much and shifting its pI as little as possible. > > The protein is a dimer and its N-termini are fairly close together, so maybe > three His at each N-Terminus are maybe almost as good as six. > > Any experience? I would appreciate suggestions! Or maybe a reference to read > about this method. > > Thanks a lot! > > Georg From owner-proteins@net.bio.net Tue Oct 13 23:00:00 1998 Path: biosci!PHOENIX.PRINCETON.EDU!emit From: emit@PHOENIX.PRINCETON.EDU (Emi Terasawa) Newsgroups: bionet.molbio.proteins Subject: metals that bind to collagen I Date: 14 Oct 1998 12:58:51 -0700 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 16 Sender: daemon@net.bio.net Distribution: world Message-ID: References: <36246083.1098D476@gmx.de> NNTP-Posting-Host: net.bio.net Hi there, Does anyone know what metals bind to collagen I? Thanks, Emi _______________________________________________________________________________ Emi Terasawa Department of Chemistry Princeton University From owner-proteins@net.bio.net Tue Oct 13 23:00:00 1998 Path: biosci!news.stanford.edu!su-news-feed2.bbnplanet.com!su-news-hub1.bbnplanet.com!chicago-news-feed1.bbnplanet.com!news.bbnplanet.com!newsfeed.enteract.com!feed1.news.rcn.net!rcn!howland.erols.net!news.idt.net!newsfeed.ecrc.net!wuff.mayn.de!news-nue1.dfn.de!news-lei1.dfn.de!news.uni-leipzig.de!news.uni-halle.de!not-for-mail From: Frederik Boernke Newsgroups: bionet.molbio.methds-reagnts,bionet.molbio.proteins Subject: Re: Purification of all proteins in an experssion library Date: Wed, 14 Oct 1998 08:31:04 -0700 Organization: IPK Gatersleben Lines: 56 Message-ID: <3624C3B8.7838@nospam.ipk-gatersleben.de> References: <6vvodo$sam$1@niobium.hgmp.mrc.ac.uk> Reply-To: boernke@nospam.ipk-gatersleben.de NNTP-Posting-Host: mpp-14.ipk-gatersleben.de Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 3.03 (Win16; I) To: "Mr. M.J. Lush" Xref: biosci bionet.molbio.methds-reagnts:71432 bionet.molbio.proteins:13417 Mr. M.J. Lush wrote: > > We are looking for proteins that stimulate a T-cell line. > To do this we plan to generate a cDNA expression library and test > the T-cells with pools of expressed proteins. > > Unfortunately bacterial proteins will also stimulate the > T-cells, therefore we want to separate the expressed proteins from > the bacterial ones. What we are looking for is a tagging system > that uses a single set of conditions to purify all proteins (if it > failed to purify certain proteins they would be lost from the library) > Can anyone suggest suitable systems for this (cost is a consideration > because we'll have to purify at least 20 pools per round of cloning). > > One possibility we are considering is making a phage display library > and purifying the M13 phage by PEG precipitation, how good is this > at removing bacterial contamination and are there better ways of > purifying M13 to 'homogeneity'? > > TIA^6 > > -- > > Michael > ~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~ > NPC rights activist | Nameless Abominations are people too. Hi Michael, if you had a c-terminal poly his-tag with your expressed proteins they could be affinity purified on a nickel column. Another thing would be in vitro expression cloning where protein is made from a coupled transcription/translation assay (Promega's TNT) and pools are tested for their biological acitvity. In this case you have no problems with bacterial background without any further purification. There was a nice overview on this approach in the latest issue of Promega's Notes (see also on website). Hope this helps Ricky P.S. Of course there are millions of other brands selling in vitro translation kits which definitively are as good as TNT :-) ****************************************************************** Frederik Boernke Research Group of Molecular Plant Physiology Institute for Plant Genetics and Crop Plant Research (IPK) Corrensstr. 3 06466 Gatersleben Tel. 039482 -5 321 Fax. 039482 -5 515 http://www.ipk-gatersleben.de From owner-proteins@net.bio.net Tue Oct 13 23:00:00 1998 Path: biosci!news.stanford.edu!newsfeed.berkeley.edu!su-news-hub1.bbnplanet.com!la-news-feed1.bbnplanet.com!news.bbnplanet.com!newsfeed1.earthlink.net!nntp.earthlink.net!posted-from-earthlink!not-for-mail From: "Isreal" Newsgroups: bionet.molbio.proteins Subject: point of isoelectric Date: Wed, 14 Oct 1998 18:12:22 -0700 Organization: EarthLink Network, Inc. Lines: 5 Message-ID: <703htm$qsk$1@birch.prod.itd.earthlink.net> NNTP-Posting-Host: 1cust184.tnt18.lax3.da.uu.net X-Newsreader: Microsoft Outlook Express 4.72.3155.0 X-Posted-Path-Was: not-for-mail X-MimeOLE: Produced By Microsoft MimeOLE V4.72.3155.0 X-ELN-Date: Wed Oct 14 18:08:06 1998 Does anyone know the PI of IgG2b,k and IgG2a,k thanks in advance! From owner-proteins@net.bio.net Tue Oct 13 23:00:00 1998 Path: biosci!news.stanford.edu!newsfeed.berkeley.edu!news.maxwell.syr.edu!news.mel.connect.com.au!news.unimelb.edu.au!ludwignt-4 From: murphy_r@licre.ludwig.edu.au (Roger Murphy) Newsgroups: bionet.molbio.proteins Subject: Re: ?Why alkaline pH in protein refold? Date: Thu, 15 Oct 98 04:21:34 GMT Organization: Ludwig Institute for Cancer Research Lines: 36 Distribution: world Message-ID: <7040fj$ggh$1@izvestia.its.unimelb.edu.au> References: <3624020E.C612EAA5@student.mahidol.ac.th> NNTP-Posting-Host: ludwignt-4.austin.unimelb.edu.au X-Trace: izvestia.its.unimelb.edu.au 908428595 16913 128.250.186.193 (15 Oct 1998 05:16:35 GMT) X-Complaints-To: news@news.unimelb.edu.au NNTP-Posting-Date: 15 Oct 1998 05:16:35 GMT X-Newsreader: News Xpress 2.0 Beta #0 In article <3624020E.C612EAA5@student.mahidol.ac.th>, g3937506@STUDENT1.MAHIDOL.AC.TH (Jongrak Kittiworakarn) wrote: >Dear all: > >Is it neccessary to refold a protein in alkaline pH? >Does this prevent any chemical modification ? >Thanks for all suggestion. > >Jongrak > >-- >-----------------------------------Mr. Jongrak Kittiworakarn Hi Jongrak, The reason for protein refolding at slightly alkaline pH is that disulfide bridge exchange is pH dependant, and is faster at alkaline pH. Thus you can get rapid making and breaking of disulphide bridges until the thermodynamically most stable form of your protein is formed. Hope this helps. Roger Roger Murphy, Ph.D. Biological Production Facility Ludwig Institute for Cancer Research Austin & Repatriation Medical Centre Studley Road, Heidelberg, Vic. 3084 Australia. Tel 61-3-94965463 Fax 61-3-94965436 Email murphy_r@licre.ludwig.edu.au From owner-proteins@net.bio.net Thu Oct 15 23:00:00 1998 Path: biosci!news.stanford.edu!newsfeed.berkeley.edu!newsfeed1.swip.net!swipnet!news.tele2.nl!newsfeed.wirehub.nl!surfnet.nl!rug.nl!not-for-mail From: Ilona Hangyi Newsgroups: bionet.molbio.proteins Subject: SDS-PAGE gels at pH 11, pH 12 Date: Fri, 16 Oct 1998 13:43:48 +0200 Organization: Rijksuniversiteit Groningen Lines: 22 Message-ID: <36273174.41C6@chem.rug.nl> NNTP-Posting-Host: rugnm24.chem.rug.nl Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 3.01SGoldC-SGI (X11; I; IRIX 6.3 IP32) Hi there, does anyone ever used protein SDS-PAGE gel analysis at pH 11 or pH 12, or does anybody know where I maybe can find information (articles etc.) about using SDS-PAGE at this high basic pH? Thanks! Ilona Hangyi @..@ (`--') ____________\\( >__< )///___________________________ Ilona Hangyi Univ. Groningen I.Hangyi@chem.rug.nl GBB, Biochemistry Nijenborg 4 9747 AG Groningen phone 31-50-3634326 The Netherlands fax 31-50-3634165 ___________________________________________________ From owner-proteins@net.bio.net Thu Oct 15 23:00:00 1998 Path: biosci!news.stanford.edu!nntp.cs.ubc.ca!newsfeed.direct.ca!rill.news.pipex.net!pipex!sun4nl!surfnet.nl!rug.nl!not-for-mail From: Ilona Hangyi Newsgroups: bionet.molbio.methds-reagnts,bionet.molbio.proteins Subject: SDS-PAGE gels at pH 11, pH 12 Date: Fri, 16 Oct 1998 12:52:31 +0200 Organization: Rijksuniversiteit Groningen Lines: 21 Message-ID: <3627256F.167E@chem.rug.nl> NNTP-Posting-Host: rugnm24.chem.rug.nl Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 3.01SGoldC-SGI (X11; I; IRIX 6.3 IP32) Xref: biosci bionet.molbio.methds-reagnts:71503 bionet.molbio.proteins:13422 Hi there, does anyone ever used protein SDS-PAGE gel analysis at pH 11 or pH 12, or does anybody know where I maybe can find information (articles etc.) about using SDS-PAGE at this high pH? Thanks! Ilona Hangyi @..@ (`--') ____________\\( >__< )///___________________________ Ilona Hangyi Univ. Groningen I.Hangyi@chem.rug.nl GBB, Biochemistry Nijenborg 4 9747 AG Groningen phone 31-50-3634326 The Netherlands fax 31-50-3634165 ___________________________________________________ From owner-proteins@net.bio.net Thu Oct 15 23:00:00 1998 Path: biosci!STUDENT1.MAHIDOL.AC.TH!g3937506 From: g3937506@STUDENT1.MAHIDOL.AC.TH (Jongrak Kittiworakarn) Newsgroups: bionet.molbio.proteins Subject: ?Peptide sample preparation for mass spec? Date: 16 Oct 1998 00:18:22 -0700 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 30 Sender: daemon@net.bio.net Distribution: world Message-ID: <3626E881.5067973F@student.mahidol.ac.th> NNTP-Posting-Host: net.bio.net Dear all: I have a peptide <6 kDa which is obtained from trypsin digestion of the native protein. The peptide seemed to be bind to the other trypsis resistant domain of the native protein. I would like to analyse this peptide by electrospray mass spec. How should I prepared this sample for the MS. Is it possible to do as follows: First, separate the peptide from other peptides/proteins by electrophoresis. Then, transfer it to nitrocellulose membrane, and cut the band of interest. Dissolve the membrane in acetone and separate the peptide by precipitation. Thanks for any suggestion. 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 Thu Oct 15 23:00:00 1998 Path: biosci!agate!newsfeed.berkeley.edu!fu-berlin.de!news.uni-stuttgart.de!news.urz.uni-heidelberg.de!aixterm9.urz.uni-heidelberg.de!cganzler From: Christof Gaenzler Newsgroups: bionet.molbio.proteins Subject: Biotin coated on plastic surface Date: Fri, 16 Oct 1998 13:02:48 +0200 Organization: University of Heidelberg, Germany Lines: 16 Message-ID: NNTP-Posting-Host: aixterm9.urz.uni-heidelberg.de Mime-Version: 1.0 Content-Type: TEXT/PLAIN; charset=US-ASCII X-Sender: cganzler@aixterm9.urz.uni-heidelberg.de We have a protein with a tag that binds to biotin. Is it possible to coat ELISA-microtiter plates with biotin so that we can bind our protein specifically in the well? Best regard and thanks in advance Christof Gaenzler --- http://www.dkfz-heidelberg.de German Cancer Research Center Applied Tumor Virology --- gaenzler@biosys.net (Christof Gaenzler) From owner-proteins@net.bio.net Thu Oct 15 23:00:00 1998 Path: biosci!agate!newsfeed.berkeley.edu!howland.erols.net!news-peer.gip.net!news-raspail.gip.net!news.gsl.net!gip.net!news.belnet.be!newsfeed.wirehub.nl!surfnet.nl!barba.uci.kun.nl!sci.kun.nl!not-for-mail From: Jan Aelen Newsgroups: bionet.molbio.proteins Subject: medium of deuteriun-oxide Date: Fri, 16 Oct 1998 17:35:58 +0200 Organization: University of Nijmegen, The Netherlands Lines: 23 Message-ID: <362767DC.989C6DD3@nmr.kun.nl> NNTP-Posting-Host: biofpc14.nmr.kun.nl Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 4.03 [en] (Win95; I) Dear Netters, I want to express my protein in LB dissolved in deuterium-oxide instead of normal water. The host strain is BL21/DE3. Has anyone experience about the growth of the cells in this medium? Thanks -- Mr.J.M.A.Aelen N.S.R.Center Dept.Biofysical of Biochemistry Faculty of Science Toernooiveld 1 6525 ED Nijmegen The Netherlands Tel.31.24653178 Fax.31.24652112 e-mail janal@nmr.kun.nl From owner-proteins@net.bio.net Thu Oct 15 23:00:00 1998 Path: biosci!news.stanford.edu!newsfeed.berkeley.edu!cyclone.news.idirect.com!island.idirect.com!newsfeed.cwix.com!204.238.120.130!news-feeds.jump.net!nntp2.dejanews.com!nnrp1.dejanews.com!not-for-mail From: irabinov@bidmc.harvard.edu Newsgroups: bionet.molbio.proteins Subject: blotting fixed gels Date: Fri, 16 Oct 1998 19:05:15 GMT Organization: Deja News - The Leader in Internet Discussion Lines: 8 Message-ID: <7085db$c3o$1@nnrp1.dejanews.com> NNTP-Posting-Host: 134.174.126.176 X-Article-Creation-Date: Fri Oct 16 19:05:15 1998 GMT X-Http-User-Agent: Mozilla/4.06 (Macintosh; U; PPC, Nav) X-Http-Proxy: 1.0 x13.dejanews.com:80 (Squid/1.1.22) for client 134.174.126.176 Does anyone know if it is possible to blot a dried gel (PAGE)that was fixed in Coomasie/destain? I’d appreciate any suggestion or method. Isaac -----------== Posted via Deja News, The Discussion Network ==---------- http://www.dejanews.com/ Search, Read, Discuss, or Start Your Own From owner-proteins@net.bio.net Thu Oct 15 23:00:00 1998 Path: biosci!news.stanford.edu!nntp.cs.ubc.ca!newsfeed.direct.ca!oleane!jussieu.fr!citi2.fr!upr415-federici.cochin.inserm.fr!user From: camoin@icgm.cochin.inserm.fr (camoin luc) Newsgroups: bionet.molbio.proteins Subject: protein folding Date: Fri, 16 Oct 1998 14:22:56 +0200 Organization: icgm cnrs upr415 Lines: 15 Message-ID: NNTP-Posting-Host: upr415-federici.cochin.inserm.fr X-Newsreader: Yet Another NewsWatcher F2.0 Dear Netters, I'am looking advice about protein refolding. I'am not familiar with these techniques so all advices will be welcome. We tried to fold correctly a 20kDa recombinant protein expressed in Ecoli. This protein was found in inclusion bodies. Correct folding is necessary to allow its disulfide bridge to be restored. Does anyone can send me a detailed protocol to refold a protein? Thanks in advance From owner-proteins@net.bio.net Thu Oct 15 23:00:00 1998 Path: biosci!NMSU.EDU!hroychow From: hroychow@NMSU.EDU (Hiranya Roychowdhury) Newsgroups: bionet.molbio.proteins Subject: Re: blotting fixed gels Date: 16 Oct 1998 15:43:09 -0700 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 26 Sender: daemon@net.bio.net Distribution: world Message-ID: <199810162238.QAA281468@nestor.NMSU.Edu> NNTP-Posting-Host: net.bio.net At 07:05 PM 10/16/98 GMT, irabinov@bidmc.harvard.edu wrote: >Does anyone know if it is possible to blot a dried gel (PAGE)that >was fixed in Coomasie/destain? >I'd appreciate any suggestion or method. > >Isaac > >-----------== Posted via Deja News, The Discussion Network ==---------- >http://www.dejanews.com/ Search, Read, Discuss, or Start Your Own > > Here is something that I know is possible: One can electrophorese out polypeptides fixed and stained (CBB-R) in acryl. gels. This is a routine method of obtaining antigens for some. So, it may be possible to equilibrate such gels in Towbin's and transfer the content to PVDF using the usual western transfer set up. Dr. Hiranya Sankar Roychowdhury Plant Genetic Engineering Lab. New Mexico State University Las Cruces, NM 88003 Ph. (505) 646-5785 hroychow@nmsu.edu From owner-proteins@net.bio.net Fri Oct 16 23:00:00 1998 Newsgroups: bionet.molbio.proteins Path: biosci!agate!newsfeed.berkeley.edu!howland.erols.net!uninett.no!newsfeed.nacamar.de!nntp.news.xara.net!xara.net!baron.netcom.net.uk!netcom.net.uk!server3.netnews.ja.net!ucl.ac.uk!bcc.ac.uk!chen From: chen@bsm.bioc.ucl.ac.uk (Chen Ho An) Subject: Re: medium of deuteriun-oxide Message-ID: <1998Oct17.162928.105698@ucl.ac.uk> Date: Sat, 17 Oct 1998 16:29:28 GMT References: <362767DC.989C6DD3@nmr.kun.nl> Organization: University College London X-Newsreader: TIN [version 1.2 PL2] Lines: 15 Jan Aelen (janal@nmr.kun.nl) wrote: : Dear Netters, : I want to express my protein in LB dissolved in deuterium-oxide instead : of normal water. : The host strain is BL21/DE3. : Has anyone experience about the growth of the cells in this medium? No. But why do you want to do this? If you are thinking of labelling the protein with deuterium, LB would have all the amino acids which would be preferentially incorporated into your protein and you would therefore have a low level of incorporation. Are you are thinking of labelling specific atoms or residues? From owner-proteins@net.bio.net Fri Oct 16 23:00:00 1998 Path: biosci!news.stanford.edu!newsfeed.berkeley.edu!newsfeed.axxsys.net!cyclone.news.idirect.com!island.idirect.com!newsfeed1.earthlink.net!nntp.earthlink.net!posted-from-earthlink!not-for-mail From: John Philo <"jphilo*NO SPAM12*"@earthlink.net> Newsgroups: bionet.molbio.proteins Subject: Re: His-tagged proteins with less than six His Date: Sat, 17 Oct 1998 09:07:54 -0700 Organization: Alliance Protein Laboratories Lines: 43 Message-ID: <3628C0DA.CE4B0CB4@earthlink.net> References: <6vkpn3$851$1@mlucom4.urz.uni-halle.de> NNTP-Posting-Host: 1cust36.tnt1.thousand-oaks.ca.da.uu.net Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit To: Georg Wille X-Posted-Path-Was: not-for-mail X-ELN-Date: Sat Oct 17 09:10:51 1998 X-Mailer: Mozilla 4.05 [en] (Win95; U) Georg Wille wrote: > > Dear Bionetters, > > I'm about to attach an His-tag to the N-Terminus of a protein. Has anyone > ever heard of using _less_ than six residues successfully. I would like to > use as few as possible in order not to distort the enzymatic properties of > the protein too much and shifting its pI as little as possible. > > The protein is a dimer and its N-termini are fairly close together, so maybe > three His at each N-Terminus are maybe almost as good as six. > > Any experience? I would appreciate suggestions! Or maybe a reference to read > about this method. > > Thanks a lot! > > Georg Georg, What the companies who sell the His-tag reagents fail to tell people is that in order to tight, specific binding your protein needs to be able to simultaneously interact with TWO metals and two of the NTA ligands. For this reason, and because the His tag is not always fully exposed, some labs have gone to using 7 or 8 His tags. You also need to remember that as a general rule His tag purification only works well under denaturing conditions. Your idea that 3 His will work because your protein is a native dimer obviously only applies if you have a native folded dimer. You are right, though, that the His tag can alter the properties of your protein, and we have seen examples of this. One way around that is to engineer in a target sequence for a specific protease between the His tag and the N-terminus of your protein, and then cleave off the tag. 'Hope this helps. John Philo, Alliance Protein Laboratories *** Remove "*NO SPAM12*" from return address before replying. *** From owner-proteins@net.bio.net Fri Oct 16 23:00:00 1998 Path: biosci!news.stanford.edu!newsfeed.berkeley.edu!sunqbc.risq.qc.ca!newsflash.concordia.ca!pitt.edu!not-for-mail From: pxpst2@unixs.cis.pitt.edu (Peter) Newsgroups: bionet.molbio.proteins Subject: Re: blotting fixed gels Date: Sat, 17 Oct 1998 20:53:57 -0500 Organization: University of Pittsburgh Lines: 21 Distribution: world Message-ID: References: <199810162238.QAA281468@nestor.NMSU.Edu> NNTP-Posting-Host: pelli.pathology.pitt.edu X-Newsreader: MT-NewsWatcher 2.4.4 In article <199810162238.QAA281468@nestor.NMSU.Edu>, hroychow@NMSU.EDU (Hiranya Roychowdhury) wrote: > Here is something that I know is possible: One can electrophorese > out polypeptides fixed and stained (CBB-R) in acryl. gels. This is a routine > method of obtaining antigens for some. > So, it may be possible to equilibrate such gels in Towbin's and > transfer the content to PVDF using the usual western transfer set up. Possible...yes indeed. BUT the yield will stink especially as the size of the "peptide" increases. When I try to retreive bands from fixed gels, I simple apply 40 ma for 48 hrs and electroelute the bands into a dialysis bag. The big problem that I have found is removing the SDS since my goal is to sequence it by MS MS Peter -- "don't you eat that yellow snow Watch out where the huskies go" FZ From owner-proteins@net.bio.net Fri Oct 16 23:00:00 1998 Path: biosci!news.stanford.edu!nntp.cs.ubc.ca!newsfeed.direct.ca!newsfeed.nacamar.de!ayres.ftech.net!news.ftech.net!newsfeed.wirehub.nl!news.belnet.be!inf6serv.rug.ac.be!not-for-mail From: Miek Tanghe Newsgroups: bionet.molbio.proteins Subject: Test, please ignore Date: Sat, 17 Oct 1998 21:14:59 +0200 Organization: University of Ghent, Belgium Lines: 1 Message-ID: <3628ECB3.BE0EE99E@gengenp.rug.ac.be> NNTP-Posting-Host: tauris.rug.ac.be Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 4.5b2 [en] (Win95; I) X-Accept-Language: en Test sorry. From owner-proteins@net.bio.net Fri Oct 16 23:00:00 1998 Path: biosci!news.stanford.edu!su-news-feed2.bbnplanet.com!su-news-hub1.bbnplanet.com!london-news-feed1.bbnplanet.com!news.bbnplanet.com!diablo.theplanet.net!news.theplanet.net!not-for-mail From: "marvel" Newsgroups: bionet.molbio.proteins Subject: Re: metals that bind to collagen I Date: Sat, 17 Oct 1998 16:40:00 +0100 Organization: Customer of Planet Online Lines: 19 Message-ID: <70adqq$bu8$1@newsreader2.core.theplanet.net> References: <36246083.1098D476@gmx.de> NNTP-Posting-Host: modem-17.oxygen.dialup.pol.co.uk X-Trace: newsreader2.core.theplanet.net 908638874 12232 62.136.3.145 (17 Oct 1998 15:41:14 GMT) NNTP-Posting-Date: 17 Oct 1998 15:41:14 GMT X-Newsreader: Microsoft Outlook Express 4.72.3110.5 X-MimeOLE: Produced By Microsoft MimeOLE V4.72.3110.3 Some of the enzymes and proteins involved in collagen biosynthesis and modification require metal ions as cofactors but Im not aware of direct metal binding affinity of collagens of any type. Im curious about why you want to know about metal binding to collagen I. Sorry I cant be more helpful Yours Tas Emi Terasawa wrote in message ... >Hi there, >Does anyone know what metals bind to collagen I? >Thanks, Emi >Emi Terasawa >Department of Chemistry >Princeton University From owner-proteins@net.bio.net Fri Oct 16 23:00:00 1998 Path: biosci!news.stanford.edu!newsfeed.berkeley.edu!news.maxwell.syr.edu!news.mel.connect.com.au!news.ade.connect.com.au!not-for-mail From: Louis Bradbury Newsgroups: bionet.molbio.proteins Subject: Leukemia inhibitory factor , oncostatin M and cilliary neurotrophic factor Date: Sat, 17 Oct 1998 16:52:15 +0930 Organization: Customer of Connect.com.au P/L, Adelaide, Australia Lines: 9 Message-ID: <362845A6.93970E55@tne.net.au> Reply-To: bradbury@tne.net.au NNTP-Posting-Host: grandpa.simpsons.tne.net.au Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 4.03 [en] (Win95; I) Hi all, Can someone please help me , I am after any or all the information I can get about Leukemia inhibitory factor (LIF) , oncostatin M (OSM) and /or ciliary neurotrophic factor (CNTF) , specifically their effects and how they excert their effects. I will be gratefull for any information I can get , thankyou for your time. Louis . From owner-proteins@net.bio.net Fri Oct 16 23:00:00 1998 Path: biosci!news.stanford.edu!newsfeed.berkeley.edu!news-feed.inet.tele.dk!bofh.vszbr.cz!dispose.news.demon.net!demon!diablo.theplanet.net!news.theplanet.net!not-for-mail From: "marvel" Newsgroups: bionet.molbio.proteins Subject: multisubunit proteins Date: Sat, 17 Oct 1998 16:46:47 +0100 Organization: Customer of Planet Online Message-ID: <70ae7e$c98$1@newsreader2.core.theplanet.net> NNTP-Posting-Host: modem-17.oxygen.dialup.pol.co.uk X-Trace: newsreader2.core.theplanet.net 908639278 12584 62.136.3.145 (17 Oct 1998 15:47:58 GMT) NNTP-Posting-Date: 17 Oct 1998 15:47:58 GMT X-Newsreader: Microsoft Outlook Express 4.72.3110.5 X-MimeOLE: Produced By Microsoft MimeOLE V4.72.3110.3 Lines: 7 does anybody know of good examples of a protein subunit which associates with different subunits under different conditions (other than PDI)? could you please cite a paper or reference with your example. Thanks Tas From owner-proteins@net.bio.net Fri Oct 16 23:00:00 1998 Path: biosci!agate!newsfeed.berkeley.edu!newsfeed.nyu.edu!newsfeed.sgi.net!pitt.edu!not-for-mail From: pxpst2@unixs.cis.pitt.edu (Peter) Newsgroups: bionet.molbio.proteins Subject: Re: ?Peptide sample preparation for mass spec? Date: Sat, 17 Oct 1998 21:36:48 -0500 Organization: University of Pittsburgh Lines: 43 Distribution: world Message-ID: References: <3626E881.5067973F@student.mahidol.ac.th> NNTP-Posting-Host: pelli.pathology.pitt.edu X-Newsreader: MT-NewsWatcher 2.4.4 In article <3626E881.5067973F@student.mahidol.ac.th>, g3937506@STUDENT1.MAHIDOL.AC.TH (Jongrak Kittiworakarn) wrote: > Is it possible to do as follows: > First, separate the peptide from other peptides/proteins by > electrophoresis. Then, transfer it to nitrocellulose membrane, and cut > the band of interest. Dissolve the membrane in acetone and separate the > peptide by precipitation. I am going to assume that you will give this protein to someone else who will actually do the sequencing. It is therefore best that you ask them how they wanted the protein given to them. Many labs would prefer to take the proteins off the membranes themselves. With that said, here is the procedure that I use for MS sequencing: 1) transfer protein to PVDF (sequencing grade: It has smaller pores and more surface area) in Tobin buffer + DTT if free cysteines exist. 2) W/o drying membrane wash with LARGE amts of pure water to remove SDS(very nasty to MS), glycine and tris. 3) stain in 0.1% ponceau red in 3 % acetic acid for 5 -10 min. Then wash with ure water to remove excess stain. To much acetic acid can dehdrate some aa's making spectra harder to interpret. Also, this stain is not sensitive and works like crap on glycosylated proteins BUT it is easy to remove. CB is not easy to remove and can make MS spectra more complicated. 4) Cut out band of interest and remove stain by breifly dipping the membrane in 200uM NaOH then immediately follow with lots of water.(prolonged exposure to high pH will cause your protein to wash off the membrane) 5)place the membrane in 100mM ammonium acetate (pH8) overnight and chop the membrane up. (hydrophobic and large proteins might not come off, if this is a problem then increase the pH) 6) wsh the mebrane for 4-6 hrs (twice) with 100 uL of above buffer. Then reduce it to a suitable olume in lyophilizer and load onto RP-capillary column which is eluted straight into the MAss Spec. Good luck, Peter -- "don't you eat that yellow snow Watch out where the huskies go" FZ From owner-proteins@net.bio.net Fri Oct 16 23:00:00 1998 Path: biosci!news.stanford.edu!su-news-feed2.bbnplanet.com!su-news-hub1.bbnplanet.com!news.bbnplanet.com!newsfeed.wli.net!news-feed.inet.tele.dk!bofh.vszbr.cz!news-peer-europe.sprintlink.net!news.sprintlink.net!Sprint!newsfeed1.swip.net!swipnet!news.tele2.nl!newsfeed.wirehub.nl!news.belnet.be!inf6serv.rug.ac.be!not-for-mail From: Miek Tanghe Newsgroups: bionet.molbio.proteins Subject: Re: Test, please ignore Date: Sat, 17 Oct 1998 21:15:42 +0200 Organization: University of Ghent, Belgium Lines: 1 Message-ID: <3628ECDE.C65380FF@gengenp.rug.ac.be> References: <3628ECB3.BE0EE99E@gengenp.rug.ac.be> NNTP-Posting-Host: tauris.rug.ac.be Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 4.5b2 [en] (Win95; I) X-Accept-Language: en Gelukt? From owner-proteins@net.bio.net Sat Oct 17 23:00:00 1998 Path: biosci!agate!newsfeed.berkeley.edu!news-feed.inet.tele.dk!bofh.vszbr.cz!news.daimi.au.dk!not-for-mail From: Per Mygind Newsgroups: bionet.molbio.proteins Subject: Re: His-tagged proteins with less than six His Date: Sun, 18 Oct 1998 10:30:33 +0200 Organization: University of Aarhus, Department of Computer Science (DAIMI) Lines: 81 Message-ID: <3629A729.439F4D13@medmicro.aau.dk> References: <6vkpn3$851$1@mlucom4.urz.uni-halle.de> <3628C0DA.CE4B0CB4@earthlink.net> NNTP-Posting-Host: majestix.gram.au.dk Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Trace: xinwen.daimi.aau.dk 908699811 14072 255.255.255.255 (18 Oct 1998 08:36:51 GMT) X-Complaints-To: news@daimi.au.dk NNTP-Posting-Date: 18 Oct 1998 08:36:51 GMT X-Mailer: Mozilla 4.07 [en] (X11; I; SunOS 5.6 sun4u) John Philo wrote: > Georg Wille wrote: > > > > Dear Bionetters, > > > > I'm about to attach an His-tag to the N-Terminus of a protein. Has anyone > > ever heard of using _less_ than six residues successfully. I would like to > > use as few as possible in order not to distort the enzymatic properties of > > the protein too much and shifting its pI as little as possible. > > > > The protein is a dimer and its N-termini are fairly close together, so maybe > > three His at each N-Terminus are maybe almost as good as six. > > > > Any experience? I would appreciate suggestions! Or maybe a reference to read > > about this method. > > > > Thanks a lot! > > > > Georg > > Georg, > > What the companies who sell the His-tag reagents fail to tell people is > that in order to tight, specific binding your protein needs to be able > to simultaneously interact with TWO metals and two of the NTA ligands. > Interestingly, is there any reference to this ? > > For this reason, and because the His tag is not always fully exposed, > some labs have gone to using 7 or 8 His tags. > > You also need to remember that as a general rule His tag purification > only works well under denaturing conditions. Your idea that 3 His will > work because your protein is a native dimer obviously only applies if > you have a native folded dimer. > No, not in general. If your native purification fails, it might be because your tag is puried in your protein, not being fully exposed. To circumvent this, try moving the histag to the opposite end of your protein. Imidazalo does not work so well in my hands. Try washing in 1 M tris-hcl and eluting with histidines or EDTA (works perfect in my experience using sixmer histidine tag) > > You are right, though, that the His tag can alter the properties of your > protein, and we have seen examples of this. One way around that is to > engineer in a target sequence for a specific protease between the His > tag and the N-terminus of your protein, and then cleave off the tag. > > 'Hope this helps. > > John Philo, Alliance Protein Laboratories > > *** Remove "*NO SPAM12*" from return address before replying. ** > > > Regards > > Per Mygind > > ************************************************************************ > If you are are not part of the solution, you are part of the precipitate > ************************************************************************ > > Per Mygind, Cand.scient > > Department of Medical Microbiology and Immunology > The Bartholin Building, University of Aarhus, Denmark > phone : 89 42 17 47, fax : 86 19 61 28 > > ************************************************************************ > It's hard work and great art to make life not so serious > ************************************************************************ From owner-proteins@net.bio.net Sat Oct 17 23:00:00 1998 Path: biosci!news.stanford.edu!newsfeed.berkeley.edu!newsfeed.gamma.ru!Gamma.RU!demos!news.stack.serpukhov.su!not-for-mail From: Serge Yechikhov Newsgroups: bionet.molbio.proteins Subject: 2-D electrophoretic maps Date: Sun, 18 Oct 1998 12:44:17 +0300 Organization: Stack Inc. Lines: 2 Message-ID: <3629B871.2E6996F@usa.net> NNTP-Posting-Host: spike.iteb.serpukhov.su Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 4.5b2 [en] (Win95; I) X-Accept-Language: en 2-D electrophoretic maps of rat's brain required. From owner-proteins@net.bio.net Sat Oct 17 23:00:00 1998 Path: biosci!news.stanford.edu!newsfeed.berkeley.edu!howland.erols.net!ix.netcom.com!news From: Mark Atlas Newsgroups: bionet.molbio.proteins Subject: used lab equipment auctions of high quality scientific instruments Date: Sun, 18 Oct 1998 19:33:06 GMT Organization: ICGNetcom Lines: 23 Message-ID: <70dfft$545@sjx-ixn9.ix.netcom.com> NNTP-Posting-Host: sji-ca4-99.ix.netcom.com X-NETCOM-Date: Sun Oct 18 12:27:57 PM PDT 1998 X-Newsreader: NETCOMplete/4.0 used lab equipment auctions of high quality scientific instruments Why not join the hundreds of thousands of online auction users and purchase used lab equipment through the Internets safest online auction. Why the safest? http://www.going-going-sold.com provides an inexpensive $50 escrow ( Through an accredited title company)with all purchases regardless of price to insure you receipt of the item and to insure your satisfaction through an in lab evaluation period. http://www.going-going-sold.com is managed by knowledgeable and experienced sales representatives for the industry. we protect your interests http://www.going-going-sold.com also provides custom equipment locator services to eliminate the painstaking hunt for used equipment not found on our auction site. We prescreen our sellers to insure your quality and proper representation of product. Never before has there been an organization who will work with you to help save R&D or production costs. Visit us today and look for the great deals. Upcoming auctions included Bio hoods, HPLC , Protein purification systems ( Prosys opening at 8K what a deal this will be inspected by Beckman to insure functionality) GC GCMS, the list goes on. Over 120 items have been sold through the auction. Join the experience today. From owner-proteins@net.bio.net Sun Oct 18 23:00:00 1998 Path: biosci!MIDWAY.UCHICAGO.EDU!price From: price@MIDWAY.UCHICAGO.EDU (Phoebe Rice) Newsgroups: bionet.molbio.proteins Subject: reference on protein-DNA Kd? Date: 19 Oct 1998 17:30:02 -0700 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 15 Sender: daemon@net.bio.net Distribution: world Message-ID: <3.0.32.19981019192239.00b06ce4@acs-popmail.uchicago.edu> NNTP-Posting-Host: net.bio.net I'm looking for a nice review or text (for teaching graduate students) on protein - DNA interactions, that would discuss methods for measuring binding constants and specificity. A nice one-review-fits-all on footprinting would also be great. Any suggestions? THANKS!! Phoebe Rice ------------------------------------------------------ Phoebe Rice University of Chicago Dept of Biochem & Molec Biol price@midway.uchicago.edu http://molbio.uchicago.edu/Faculty/Phoebe_Rice.html From owner-proteins@net.bio.net Sun Oct 18 23:00:00 1998 Path: biosci!news.stanford.edu!newsfeed.berkeley.edu!newsfeed1.swip.net!swipnet!newsfeed1.uni2.dk!news.net.uni-c.dk!not-for-mail From: "S.W. Rasmussen" Newsgroups: bionet.molbio.proteins Subject: DNATools, revision 118 Date: Mon, 19 Oct 1998 22:32:25 +0100 Organization: Carlsberg Laboratory Lines: 16 Message-ID: <362BAFE9.4EEB218F@crc.dk> Reply-To: swr@crc.dk NNTP-Posting-Host: 130.226.184.128 Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Trace: news.net.uni-c.dk 908832991 28730 (None) 130.226.184.128 X-Complaints-To: usenet@news.net.uni-c.dk X-Mailer: Mozilla 4.04 [en] (Win95; I) DNATools 5.1 revision 118 The latest revision of the DNATools program package for DNA and protein sequence handling and analysis can be downloaded from: ftp://ftp.crc.dk/pub/dnatools Questions and comments to Dr. S. W. Rasmussen Dept. of Physiology Carlsberg Laoratory swr@crc.dk From owner-proteins@net.bio.net Sun Oct 18 23:00:00 1998 Path: biosci!agate!newsfeed.berkeley.edu!su-news-hub1.bbnplanet.com!news.bbnplanet.com!logbridge.uoregon.edu!infeed.is.co.za!feeder.is.co.za!hermes.is.co.za!not-for-mail From: "Clive" Newsgroups: bionet.molbio.proteins Subject: Re: ?Why alkaline pH in protein refold? Date: Mon, 19 Oct 1998 18:58:49 +0200 Organization: UNP Lines: 28 Distribution: world Message-ID: <908816185.9303@Raven> References: <3624020E.C612EAA5@student.mahidol.ac.th> NNTP-Posting-Host: raven.und.ac.za Mime-Version: 1.0 Content-Type: text/plain; charset="US-ASCII" Content-Transfer-Encoding: 7bit X-Newsreader: Microsoft Outlook Express for Macintosh - 4.01 (295) Cache-Post-Path: Raven!unknown@proxyfs2.und.ac.za X-Cache: nntpcache 2.3.2.1 (see http://www.nntpcache.org/) Disulfide exchange is promoted at higher pH because of the connection between redox potential and pH ---------- >From: g3937506@STUDENT1.MAHIDOL.AC.TH (Jongrak Kittiworakarn) In article <3624020E.C612EAA5@student.mahidol.ac.th>, g3937506@STUDENT1.MAHIDOL.AC.TH (Jongrak Kittiworakarn) wrote: >Dear all: > >Is it neccessary to refold a protein in alkaline pH? >Does this prevent any chemical modification ? >Thanks for all suggestion. > >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 Sun Oct 18 23:00:00 1998 Newsgroups: bionet.molbio.proteins Path: biosci!news.stanford.edu!su-news-feed2.bbnplanet.com!su-news-hub1.bbnplanet.com!cpk-news-hub1.bbnplanet.com!news.bbnplanet.com!newsfeed.cwix.com!194.72.7.126!btnet-peer!btnet!dispose.news.demon.net!demon!peer.news.zetnet.net!peer.news.bb.u-net.net!u-net!yama.mcc.ac.uk!liv!news From: lgbell@liv.ac.uk (lgbell) Subject: Re: ?Why alkaline pH in protein refold? X-Nntp-Posting-Host: pc028010.med.liv.ac.uk Content-Type: Text/Plain; charset=US-ASCII Message-ID: Sender: news@liverpool.ac.uk (News System) Organization: University of Liverpool X-Newsreader: WinVN 0.99.7 References: <3624020E.C612EAA5@student.mahidol.ac.th> Mime-Version: 1.0 Date: Mon, 19 Oct 1998 12:03:31 GMT X-Authenticated-User: lgbell Lines: 33 In article <3624020E.C612EAA5@student.mahidol.ac.th>, g3937506@STUDENT1.MAHIDOL.AC.TH says... > >Dear all: > >Is it neccessary to refold a protein in alkaline pH? >Does this prevent any chemical modification ? >Thanks for all suggestion. > >Jongrak > >-- >-----------------------------------Mr. Jongrak Kittiworakarn > g3937506@student.mahidol.ac.th Well that depends. An alkaline pH will facillitate the formation of disulfide bridges (Cys-Cys) under atmospheric oxidation. The traditional assumption is that the thermodynamically stable disulfide configuration is the same as the naturally occurring or active one. So I guess the rationale is that, the protein initially re-folds according to its primary sequence and disulfide bridges, if present, then form accordingly from atmospheric oxidation of Cys and stabalise this final conformation. Obviously if the protein does not contain Cys then under this explanation the re-folding pH can be varied. Obviously the optimuin pH to use would be related to the environment the protein naturally exists in or is biosynthesised in. Hope this helps, Len lgbell@liv.ac.uk From owner-proteins@net.bio.net Sun Oct 18 23:00:00 1998 Path: biosci!rutgers!rockyd.rockefeller.edu!newsfeed.nyu.edu!news-nyc.telia.net!howland.erols.net!newsfeed.nacamar.de!nntp.news.xara.net!xara.net!server6.netnews.ja.net!news.shef.ac.uk!not-for-mail From: A Al-dukhyil Newsgroups: bionet.molbio.proteins Subject: Nuclear localisation Date: Mon, 19 Oct 1998 12:12:44 +0100 Organization: Molecular Biology & Biotechnology, University of Sheffield, UK Lines: 7 Message-ID: <362B1EAC.32AB@sheffield.ac.uk> NNTP-Posting-Host: pc093091.shef.ac.uk Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 2.02 (Win16; I) Hi, I've got a fusion of Green fluorescent protein to Nuclear localisation sequence in a vector. When I transfect this vector to mammalian cell lines I get a mixtures of transfected cells, some of them have GFP in the nucleos and the others have GFP in the whole cell. Is it normal to get a result like this? Thanking in anticipation From owner-proteins@net.bio.net Sun Oct 18 23:00:00 1998 Path: biosci!UMBI.UMD.EDU!collins From: collins@UMBI.UMD.EDU (john collins) Newsgroups: bionet.molbio.proteins Subject: Position Available Date: 19 Oct 1998 12:11:29 -0700 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 11 Sender: daemon@net.bio.net Distribution: world Message-ID: <3.0.3.32.19981019150226.007ab500@umbi.umd.edu> NNTP-Posting-Host: net.bio.net Research Assistant Professor, non-tenure track. Requires Ph.D. in biology, at least ten years of postdoctoral research on the molecular structure and interactions of muscle proteins, fifteen or more publications, and expertise in protein sequencing, chemical modification, HPLC, site-directed mutagenesis and FRET. Available immediately; salary $36,000-$39,000 per year. Minority and female candidates encouraged. Apply by e-mail (collins@umbi.umd.edu) to: John H. Collins, Medical Biotechnology Center, University of Maryland Biotechnology Institute, 725 W. Lombard Street, Baltimore, MD 21201, before December 1, 1998. (Institutional approval pending.) From owner-proteins@net.bio.net Mon Oct 19 23:00:00 1998 Path: biosci!news.stanford.edu!newsfeed.berkeley.edu!howland.erols.net!feed1.news.rcn.net!rcn!newsfeed.atl.bellsouth.net!news4.mia.bellsouth.net.POSTED!not-for-mail Message-ID: <362C8B39.4BD5@pdqmail.bhm.bellsouth.net> From: Charles Hardwick Reply-To: krains@pdqmail.bhm.bellsouth.net X-Mailer: Mozilla 3.01C-BLS20 (Win95; I) MIME-Version: 1.0 Newsgroups: bionet.molbio.proteins Subject: Re: Position Available References: <3.0.3.32.19981019150226.007ab500@umbi.umd.edu> Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit Lines: 28 Date: Tue, 20 Oct 1998 13:08:17 GMT NNTP-Posting-Host: 207.203.38.60 NNTP-Posting-Date: Tue, 20 Oct 1998 09:08:17 EST john collins wrote: > > Research Assistant Professor, non-tenure track. Requires Ph.D. in biology, > at least ten years of postdoctoral research on the molecular structure and > interactions of muscle proteins, fifteen or more publications, and > expertise in protein sequencing, chemical modification, HPLC, site-directed > mutagenesis and FRET. Available immediately; salary $36,000-$39,000 per > year. Minority and female candidates encouraged. Apply by e-mail > (collins@umbi.umd.edu) to: John H. Collins, Medical Biotechnology Center, > University of Maryland Biotechnology Institute, 725 W. Lombard Street, > Baltimore, MD 21201, before December 1, 1998. (Institutional approval > pending.) Boy, What a GREAT OFFER! Just imagine, someone who is 38 or 39 years old, and has published 1.5 papers per year since completing his (or her) doctorate, can go here and make over $3000 per month! (before taxes). And the job has little or no job security. I'll bet they are lining up. I don't miss academic research at all. Charles Hardwick Ph.D From owner-proteins@net.bio.net Mon Oct 19 23:00:00 1998 From: atlantis@netcom.com (JJ Miranda) Newsgroups: bionet.molbio.proteins,bionet.biophysics Subject: molecular analysis software Date: Tue, 20 Oct 1998 08:19:02 GMT Reply-To: atlantis@netcom.com Message-ID: <362c471b.52716143@news.reed.edu> X-Newsreader: Forte Free Agent 1.11/32.235 NNTP-Posting-Host: 134.10.20.73 X-Trace: 20 Oct 1998 01:14:02 -0800, 134.10.20.73 Organization: Reed College Lines: 9 Path: biosci!news.stanford.edu!nntp.cs.ubc.ca!newsfeed.direct.ca!rill.news.pipex.net!pipex!nntp.mid-ga.com!news1.mid-ga.com!newsgator!dnews.reed.edu!134.10.20.73 Xref: biosci bionet.molbio.proteins:13450 bionet.biophysics:4465 Hi all, Is there any type of software that would allow me to determine the solvent accessible surface area of a protein if I had the pdb file? Also, is there any software that can distinguish between hydrophobic and polar surface areas? Sincere regards, JJ From owner-proteins@net.bio.net Mon Oct 19 23:00:00 1998 Path: biosci!news.stanford.edu!newsfeed.berkeley.edu!howland.erols.net!news.pbi.net!not-for-mail From: "anonymous" Newsgroups: bionet.molbio.proteins Subject: Spermatazoa & Polygamous Heterosexuality Date: Fri, 16 Oct 1998 11:57:16 -0700 Organization: Pacific Bell Internet Services Lines: 22 Message-ID: <7084g8$9fu$1@nnrp4.snfc21.pbi.net> NNTP-Posting-Host: ppp-209-79-196-105.scrm01.pacbell.net X-Newsreader: Microsoft Outlook Express 4.71.1712.3 X-MimeOLE: Produced By Microsoft MimeOLE V4.71.1712.3 Hi, During a lecture in Psychology 18: Human Development, the following statements we're made by several classmates but could not be verified, Concerning polygamous heterosexuality & pregnancy: * A woman's regular partner's sperm will be dominant and agressively "attack" a foreign or new partner's sperm if both are present in the woman's sexual organs at the same time. ( Territorial Phagocytosis of some sort ?) * A woman who has sperm in her from 2 different partners, will fertilize with the partner who has given her the most pleasure and/or the partner whom she has the strongest attraction for. ( Does orgasm have any physiological effects on fertility?) Can anyone verify these or similiar points and possibly site case studies or research? Please respond via this newsgroup. Thank you. No, this is not a request for help with homework. It's just a very provocative & interesting subject that I'd like 2 hear more about. From owner-proteins@net.bio.net Mon Oct 19 23:00:00 1998 From: atlantis@netcom.com (JJ Miranda) Newsgroups: bionet.molbio.proteins Subject: Re: ?Peptide sample preparation for mass spec? Date: Tue, 20 Oct 1998 08:15:27 GMT Reply-To: atlantis@netcom.com Message-ID: <362c461d.52462487@news.reed.edu> References: <3626E881.5067973F@student.mahidol.ac.th> X-Newsreader: Forte Free Agent 1.11/32.235 NNTP-Posting-Host: 134.10.20.73 X-Trace: 20 Oct 1998 01:10:26 -0800, 134.10.20.73 Organization: Reed College Lines: 7 Path: biosci!agate!newsfeed.berkeley.edu!howland.erols.net!nntp1.crl.com!news1.mid-ga.com!newsgator!dnews.reed.edu!134.10.20.73 I'm not sure that you have to go through all the membrance work... This will depend on how much sample you actually have. I'm also going to assume that this tryptic digest was done in solution. If so, maybe HPLC separation would be feasible. Heck, run it using LC-ESIMS. Sincere regards, JJ From owner-proteins@net.bio.net Tue Oct 20 23:00:00 1998 Path: biosci!WOWMAIL.COM!picc98 From: picc98@WOWMAIL.COM Newsgroups: bionet.molbio.proteins Subject: POWER & MAGIC OF E-MAIL!!! Date: 21 Oct 1998 08:27:36 -0700 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 124 Sender: daemon@net.bio.net Distribution: world Message-ID: <199810211527.IAA18663@net.bio.net> NNTP-Posting-Host: net.bio.net

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**IF ORDERING FROM OUTSIDE THE US PLEASE SEND US POSTAL MONEY ORDER **All sales final. From owner-proteins@net.bio.net Tue Oct 20 23:00:00 1998 Path: biosci!news.stanford.edu!su-news-feed2.bbnplanet.com!su-news-hub1.bbnplanet.com!news.bbnplanet.com!newsfeed.concentric.net!newsfeed1.earthlink.net!xfer.kren.ne.kr!news.kren.ne.kr!not-for-mail From: Tae-Joon Kwon Newsgroups: bionet.molbio.proteins,bionet.biophysics Subject: Re: molecular analysis software Date: Thu, 22 Oct 1998 11:12:13 +0900 Organization: Biochem.Eng.Lab. / Division of Chem. Eng. / Seoul Nat'l Univ. Lines: 27 Message-ID: <362E947D.123DADA4@biopia.snu.ac.kr> References: <362c471b.52716143@news.reed.edu> Reply-To: linusben@biopia.snu.ac.kr NNTP-Posting-Host: biopia.snu.ac.kr Mime-Version: 1.0 Content-Type: text/plain; charset=EUC-KR Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 4.05 [ko] (X11; I; Linux 2.0.35 i586) To: atlantis@netcom.com Xref: biosci bionet.molbio.proteins:13458 bionet.biophysics:4481 Refer to this site http://SAL.KachinaTech.COM/Z/2/ This page is for "Scientific Application on Linux". But the maintainer of many programs are linked. One of the useful program for your purpose is MOLMOL. (the information about this also can be found on SAL.) MOLMOL program provides binary files for many kind of platforms. (not only Unix-based, also Win32) Sincerely, Kwon,Tae-Joon linusben@biopia.snu.ac.kr JJ Miranda wrote: > > Hi all, > > Is there any type of software that would allow me to determine the > solvent accessible surface area of a protein if I had the pdb file? > Also, is there any software that can distinguish between hydrophobic > and polar surface areas? > > Sincere regards, > JJ From owner-proteins@net.bio.net Tue Oct 20 23:00:00 1998 Path: biosci!news.stanford.edu!newsfeed.berkeley.edu!howland.erols.net!newsfeed.nacamar.de!nntp.news.xara.net!xara.net!server5.netnews.ja.net!news.shef.ac.uk!not-for-mail From: R U Gianfrancesco Newsgroups: bionet.molbio.proteins Subject: DNA and autoclaving Date: Wed, 21 Oct 1998 10:15:56 +0100 Organization: Animal & Plant Science (bo), University of Sheffield, UK Lines: 5 Message-ID: <362DA64C.167A@sheffield.ac.uk> NNTP-Posting-Host: pc093184.shef.ac.uk Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 2.02 (Win16; I) I'm supplying DNA to fungi as a food source. I autoclave my salmon sperm DNA but do not know the effect of autoclaving. Does anyone know, e.g. single/double strands, bonds, strand length. Thanks Richard Gianfrancesco University of Sheffield From owner-proteins@net.bio.net Wed Oct 21 23:00:00 1998 Path: biosci!agate!not-for-mail From: lhom@OCF.Berkeley.EDU (Louis Hom) Newsgroups: sci.chem,bionet.molbio.proteins,bionet.molbio.methds-reagnts Subject: Chromogenic peptidase substrates Date: 22 Oct 1998 18:32:14 GMT Organization: Univ. of California Berkeley Open Computing Facility Lines: 10 Message-ID: <70ntne$763$1@agate.berkeley.edu> NNTP-Posting-Host: famine.ocf.berkeley.edu Xref: biosci bionet.molbio.proteins:13460 bionet.molbio.methds-reagnts:71654 There are fluorogenic peptidase substrates (e.g., Z-Arg-Arg-AMC) and chromogenic glycosidase substrates (e.g., X-Gal) but I can't seem to find any chromogenic peptidase substrates. Anyone know of a source? I need it to go from one color to another, not simply producing soluble color from precipitable color (a la azocasein). TIA. -- ______________________________________________________________________________ Lou Hom >K'93 lhom@ocf.berkeley.edu http://www.ocf.berkeley.edu/~lhom/ From owner-proteins@net.bio.net Wed Oct 21 23:00:00 1998 Message-ID: <362F3892.6BE8@antibodyresource.com> Date: Thu, 22 Oct 1998 07:52:20 -0600 From: The Antibody Resource Page X-Mailer: Mozilla 3.01-C-MACOS8 (Macintosh; I; PPC) MIME-Version: 1.0 Newsgroups: bionet.molbio.proteins Subject: custom monoclonal/polyclonal antibdies Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit NNTP-Posting-Host: il-dialup82.idsonline.com Lines: 9 Path: biosci!agate!newsfeed.berkeley.edu!netnews.com!newsfeed.enteract.com!news.wwa.com!news.idsonline.com!il-dialup82.idsonline.com The Antibody Resource Page (http://www.antibodyresource.com/) now maintains a list of custom antibody suppliers. If you are interested in the production of custom monoclonal or polyclonal antibodies take a look at this invaluable guide at: http://www.antibodyresource.com/customantibody.html ps. The ARP was voted among the top 25 biotechnology webpages for 1997 by Genetic Engineering News! From owner-proteins@net.bio.net Wed Oct 21 23:00:00 1998 Path: biosci!WOWMAIL.COM!picc98 From: picc98@WOWMAIL.COM Newsgroups: bionet.molbio.proteins Subject: POWER & MAGIC OF E-MAIL!!! Date: 22 Oct 1998 07:02:18 -0700 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 124 Sender: daemon@net.bio.net Distribution: world Message-ID: <199810221401.HAA17956@net.bio.net> NNTP-Posting-Host: net.bio.net

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the POWER & MAGIC OF E-MAIL!!!

ARE YOU Tired of NOT making money on the Internet?

Did you REALLY believe that ALL you had to do was put up a web page with some attractive graphics, some Java, some frames, run a few classified ads, post to a few newsgroups, swap a few banner ads, and it allstarts to roll in? Well, that's NOT the way it is!

It's all about traffic, and getting your INTERESTING MESSAGE in front of

INTERESTED PEOPLE, lots of 'em!!!!!! If you want to make money on the Internet, you should do all the rest, AND MAKE IT A POINT TO DO THIS ONE!!!

You've got to be savvy, aggressive, and get your message in front of LOTS OF PEOPLE!

And IF YOU HAVE TO DO THAT, WHY NOT USE ONE OF THE MOST COST EFFECTIVE and instant MEANS OF TELLING YOUR STORY TO HUNDREDS OF THOUSANDS, EVEN MILLIONS OF PEOPLE? IT SURE MAKES SENSE, DOESN'T IT?

Use Bulk email - IT WORKS, !!!

............a well-written, strategically targeted E-MAIL MESSAGE, fulfilling an identified need, can and will make you money on the Internet, even on a limited budget, as long as you GET IT IN FRONT A NUMBER OF PEOPLE.

WHY IS IT SO EFFECTIVE?

a) Because it is the MOST COST EFFECTIVE MEDIUM EVER CREATED!

On the average, an individual direct (snail) mail promotion would cost around $500.00 per thousand , whereas bulk e-mail costs pennies per thousand. No known form of media advertising can compete with the low cost of bulk

e-mail.

b) Because it is INSTANT, no waiting for the snail mail to eventually get to its destination. You get a reading almost immediately on the impact of your message.

c) Because you can hit your target right right in the BULLS EYE!

d) Because PEOPLE READ E-MAIL! Your message will sit in your recipient' mail box until is it read. It's SEEN, not BURIED!

Your business deserves a piece of the BIGGEST COMMERCIAL BONANZA since the Gold Rush Days. It's yours for the taking online, on the Internet. Join the many home-based businesses currently making A FORTUNE in direct online sales. In order to capture your share of this Windfall, those people anxious to do business with you have to know about you and about the great product you're selling.

How do you find the people interested in your product? or better put.......

HOW DO PEOPLE INTERESTED IN YOUR PRODUCT FIND YOU?

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How do you know WHO THEY ARE, and WHERE THEY ARE?

WE'VE GOT THE ADDRESSES FOR YOU!!

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The following are only some of the MANY CATEGORIES. If you don't see your category, or need an exact count, just give us a call and we will be glad to assist you!

ART ANIMAL TENDING ASTROLOGY ASTRONOMY

ADVERTISING AUTOMOTIVE AWARDS ACCOUNTING

AUCTIONS AUDIO AUTO AIRCRAFT

ANTIQUES AVIATION ARTS BUILDING RELATED

AGRICULTURE BOATING BUSINESS EQUIPMENT

BOATS BEAUTY PRODUCTS BERRIES

BOOKS BUSINESS BUSINESS SERVICES

BEER & WINE BEAUTY PRODUCTS CHILD CARE COMPUTERS

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FIRE SAFETY FOOD FOOTWEAR

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