From owner-proteins@net.bio.net Sat Oct 01 23:00:00 1994 Path: biosci!daresbury!trane.uninett.no!sunic!columba.udac.uu.se!populus.slu.se!newsmgr From: b0jansve@ulmo.stud.slu.se (Jan Svensson) Newsgroups: bionet.molbio.proteins Subject: Help vitronectin antibodies Date: 28 Sep 1994 15:12:45 GMT Organization: Swedish University of Agricultural Sciences Lines: 6 Message-ID: <36c15d$eab@populus.slu.se> NNTP-Posting-Host: n1_pc13.stud.slu.se X-Newsreader: WinVN 0.90.5 Hi. I´m desperately searching for plyclonal antibodies against human vitronectin that has been affinity purified against vitronectin. All I can find is just IgG fraction. To couple a gel with vitronectin myself would bee to expensive. Janne. From owner-proteins@net.bio.net Sat Oct 01 23:00:00 1994 Path: biosci!agate!ihnp4.ucsd.edu!scripps.edu!misrael From: misrael@scripps.edu (Mark Israel) Newsgroups: bionet.molbio.proteins,bionet.software Subject: Re: program to obtain 3d coordinates from stereo pictures Date: 2 Oct 1994 19:51:51 GMT Organization: The Scripps Research Institute, La Jolla, California, USA Lines: 29 Message-ID: <36n30n$nou@riscsm.scripps.edu> References: <9410011655.AA08195@nick.med.usf.edu> NNTP-Posting-Host: struct.scripps.edu Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit Xref: biosci bionet.molbio.proteins:2795 bionet.software:9500 In article <9410011655.AA08195@nick.med.usf.edu>, gquinn@NICK.MED.USF.EDU (Gregory Quinn) writes: > Does anyone know anything about the program that can produce 3d > coordinates from stereo pictures of proteins? Here's some info on it that was recently posted to bionet.xtallography: | Subject: Re:Extracting Coorodinates from Stereo Diagrams. | Newsgroups: bionet.xtallography | From: raman@BIOC01.UTHSCSA.EDU (C.S.RAMAN) | Organization: BIOSCI International Newsgroups for Molecular Biology | Date: 16 Aug 1994 06:41:55 -0700 | Message-ID: <9408161342.AA02724@bioc01.uthscsa.edu> | | [...] | | In addition to what I had told you about its existence in the PDB, here | is what I recently found. The program is still called STEREO and has | been substantially modified (original version 1974 Michael Rossmann) by | Jin-Bi Dai also at purdue. The latest version of the program is dated | 1994 and you can obtain a copy of the source and associated man page | from the latter author. | | Jin-Bi Dai can be reached via e-mail at: | | b4x@mace.cc.purdue.edu -- misrael@scripps.edu Mark Israel From owner-proteins@net.bio.net Sat Oct 01 23:00:00 1994 Path: biosci!agate!ames!purdue!mozo.cc.purdue.edu!not-for-mail From: hwang@mace.cc.purdue.edu (I-shiou Ng) Newsgroups: bionet.molbio.proteins,bionet.molbio.methds-reagnts Subject: virus receptor research--Help Date: 2 Oct 1994 12:07:56 -0500 Organization: Purdue University Lines: 9 Message-ID: <36mpdc$nmj@mace.cc.purdue.edu> NNTP-Posting-Host: mace.cc.purdue.edu Xref: biosci bionet.molbio.proteins:2794 bionet.molbio.methds-reagnts:19108 Hi there, I would like to ask to two questions about receptor research 1. How to make veiscles for ligand-binding assay? Is there any good reference for it? 2. Is there anyone label virus as ligand? What kind of reagent are you using-- lactoperoxidase or Iodobeads? Is there any good reference? Thanks From owner-proteins@net.bio.net Sun Oct 02 23:00:00 1994 Path: biosci!daresbury!trane.uninett.no!eunet.no!nuug!EU.net!howland.reston.ans.net!europa.eng.gtefsd.com!uhog.mit.edu!news.mtholyoke.edu!news.umass.edu!news2.near.net!das-news.harvard.edu!husc-news.harvard.edu!husc-news.harvard.edu!news Newsgroups: bionet.molbio.proteins Subject: homology modelling program Message-ID: <1994Oct2.192305.35073@hulaw1.harvard.edu> From: sali@tamika.harvard.edu (Andrej Sali) Date: 2 Oct 94 19:23:03 EDT Keywords: homology modelling, comparative modelling, program Nntp-Posting-Host: tamika.harvard.edu Lines: 74 MODELLER PROTEIN MODELLING BY SATISFACTION OF SPATIAL RESTRAINTS MODELLER 11, October 1, 1994 Copyright(c) 1989-1994 Andrej Sali All Rights Reserved Written by Andrej Sali Birkbeck College, University of London, London, UK Imperial Cancer Research Fund, London, UK Harvard University, Cambridge, USA Andrej Sali, Department of Chemistry, Harvard University, 12 Oxford St, Cambridge, MA 02138, USA. Tel: (617) 495 1775. Fax: (617) 496 3204. E-mail: sali@tammy.harvard.edu. ** DESCRIPTION: MODELLER is most frequently used for homology or comparative modeling of protein three-dimensional structure: the user provides an alignment of a sequence to be modeled with known related structures and MODELLER will automatically calculate a full-atom model. More generally, MODELLER models protein 3D structure by satisfaction of spatial restraints (A. Sali & T.L. Blundell. J.Mol.Biol. 234, 779-815, 1993). In principle, the restraints can be derived from a number of different sources. These include homologous structures (comparative modeling), NMR experiments (NMR refinement), rules of secondary structure packing (combinatorial modeling), cross-linking experiments, fluorescence spectroscopy, image reconstruction in electron microscopy, site-directed mutagenesis, intuition, residue-residue and atom-atom potentials of mean force, etc. The output of MODELLER is a 3D structure of a protein that satisfies these restraints as well as possible. The optimization is carried out by the variable target function procedure employing methods of conjugate gradients and molecular dynamics with simulated annealing. The program includes a 100-page manual. MODELLER is written in Fortran and is meant to run on a UNIX system. ** DISTRIBUTION: MODELLER is available free of charge to non-profit institutions. First, please use the anonymous ftp account on tammy.harvard.edu (IP 128.103.96.19) to copy the files from the pub/modeller directory to your computer. Print and sign the license form (academic-license.ps) and mail or fax it to Andrej Sali. You will then receive the encryption key (MODELLER_KEY) with which you will be able to unpack the encrypted distribution file (modeller11-exec.tar.Z.cr). See file INSTALLATION for installation instructions. MODELER is also available as part of QUANTA, a large program with many other functionalities including interactive graphics, CHARMm, and X-PLOR. If you are interested in this commercial version please contact Ms. Jo Ellen Collins at Molecular Simulations Inc, 16 New England Executive Park, Burlington, MA 01803-5297, tel: (617) 229 9800, fax: (617) 229 9899, email: jcollins@msi.com. ** CONTENTS: src\ sources or executables for MODELLER and associated programs modlib\ libraries and data files for the programs scripts\ script files used to compile and use MODELLER doc\ MODELLER documentation Makefile Makefile for compiling/installing MODELLER modules; used by the Install script modeller11.README this file INSTALLATION how to install MODELLER Install compilation and installation script tests\ tests and examples From owner-proteins@net.bio.net Sun Oct 02 23:00:00 1994 Path: biosci!galaxy.ucr.edu!ihnp4.ucsd.edu!swrinde!pipex!doc.ic.ac.uk!daresbury!bioftp.unibas.ch!rc1.vub.ac.be!vandijck!joan From: joan@vandijck.ulb.ac.be (Joan Pontius) Newsgroups: bionet.molbio.proteins,bionet.xtallography Subject: Re: Residues at protein surface Date: 3 Oct 1994 08:54:50 GMT Organization: UCMB - ULB Lines: 28 Sender: joan@vandijck (Joan Pontius) Distribution: world Message-ID: <36ogsq$i64@rc1.vub.ac.be> References: <34hbur$rbv@alpha.cisi.unige.it> NNTP-Posting-Host: ucmbsgi9.ulb.ac.be. Xref: biosci bionet.molbio.proteins:2798 bionet.xtallography:1206 >Does anybody have references on studies about surface composition of >proteins (i.e. statistics on the relative frequency of specific >residues appearing at the surface of some protein, hopefully made >on a large subset of the PDB)? This topic is a part of my current doctorate research, so I would be interested in knowing why this information is important to you and anyone else. More specifically, I have the following questions: What will you use the information for? What do you consider to be the "protein surface"? Would the "surface" of the protein include folds inside the protein (cavities) which are inaccessible to solvent? Will you "normalize" the surface area data to take into account that some residues are larger than others? Thank you, joan@ucmb.ulb.ac.be Joan Pontius UCMB-ULB Brussels, Belgium From owner-proteins@net.bio.net Sun Oct 02 23:00:00 1994 Path: biosci!galaxy.ucr.edu!ihnp4.ucsd.edu!usc!cs.utexas.edu!swrinde!pipex!lyra.csx.cam.ac.uk!warwick!bham!usenet From: altabios@bham.ac.uk (John E. Fox) Newsgroups: bionet.molbio.proteins Subject: Re: Proteinsynthesis? Date: 3 Oct 1994 12:13:14 GMT Organization: The University of Birmingham, UK Lines: 14 Message-ID: <36osgq$1ti@sun4.bham.ac.uk> References: <36b834$776@kitten.umdc.umu.se> NNTP-Posting-Host: bcs118.bham.ac.uk X-Newsreader: WinVN version 0.80 In article <36b834$776@kitten.umdc.umu.se>, peterl@umdix.umdc.umu.se (Peter Lundberg) says: > >Any suggestions for reputable companies, or labs, that synthesize >peptides 20-30 amino acids long? Preferably somewhere here in Europe (I >think). Post here or mail me directly names and fax numbers please. > How about us! Alta Bioscience makes synthetic peptides, (amongst other things). We make peptides on 5, 30 and 100+ micromole scales in numbers from 1 to 48 peptides simultaneously. Phone 0121 414 5450 Fax 0121 414 3376 From owner-proteins@net.bio.net Sun Oct 02 23:00:00 1994 Path: biosci!galaxy.ucr.edu!ihnp4.ucsd.edu!agate!doc.ic.ac.uk!daresbury!bioftp.unibas.ch!rc1.vub.ac.be!vandijck!joan From: joan@vandijck.ulb.ac.be (Joan Pontius) Newsgroups: bionet.molbio.proteins Subject: Lennard-Jones Potential Date: 3 Oct 1994 07:37:48 GMT Organization: UCMB - ULB Lines: 13 Sender: joan@vandijck (Joan Pontius) Distribution: world Message-ID: <36occc$gu2@rc1.vub.ac.be> NNTP-Posting-Host: ucmbsgi9.ulb.ac.be. Can someone explain to me why the Lennard-Jones potential, which describes gas behaviour, is used in energy-minimization programs for proteins? Thank you very much joan@ucmb.ulb.ac.be From owner-proteins@net.bio.net Sun Oct 02 23:00:00 1994 Newsgroups: bionet.molbio.proteins Path: biosci!bcm!cs.utexas.edu!howland.reston.ans.net!europa.eng.gtefsd.com!newsxfer.itd.umich.edu!nntp.cs.ubc.ca!news.UVic.CA!spruce.pfc.forestry.ca!PFC.Forestry.CA!AEKRAMODDOUL From: aekramoddoul@PFC.Forestry.CA (Ekramoddoullah, Abul Kalam M.) Subject: Re: Quantitating Western Blots Message-ID: <1994Oct3.212532.20174@spruce.pfc.forestry.ca> Sender: news@spruce.pfc.forestry.ca Nntp-Posting-Host: pfc.pfc.forestry.ca Reply-To: aekramoddoul@PFC.Forestry.CA Organization: Forestry Canada (Pacific Forestry Centre) References: <36ftaj$d4m@hsc.usc.edu> Date: Mon, 3 Oct 1994 21:25:32 GMT Lines: 29 In article <36ftaj$d4m@hsc.usc.edu>, pboyer@hsc.usc.edu (Paul D. Boyer) writes: >I'm trying to quantitate levels of TNF-alpha in tissue extracts. I've >been unable to get results using ELISA, even sending it out to Endogen >for their use with their ELISA kit was in vain. I then resorted to >Western blots using ECL (chemiluminscent) detection. In theory, >ECL is supposed to be quantitative, but this has not been my >experience. Scanning a slot blot with a laser densitometer gives >variable results at best, and it's even more painful trying to scan >a Western, since this technique seems prone to all sorts of wierd >background spots. Also, since there are multiple TNF bands in these >extracts, it's very difficult to get accurate, repeatable results. > >Does anyone have any suggestions for improving the odds of getting >useful results from Western blots? > > pboyer@hsc.usc.edu > Paul D. Boyer, Ph.D. > Research Assistant Professor > Laboratory for Developmental Genetics > Univ. of Southern California > Have you tried a regular Western-immunoblot (i.e. without chemiluminscent) and the scanning your blot with a laser densitometer. How are you standardising the amount (in terms of protein load) for each sample prior to SDS-PAGE. If you want to discuss this further give me a call at (604)-363-0600. Abul EKRAMODDOULLAH, ABUL KALAM M. Title: Research Scientist Forestry Canada Phone: (604) 363-0600 Victoria, B.C. Internet: AEKRAMODDOUL@A1.PFC.Forestry.CA From owner-proteins@net.bio.net Sun Oct 02 23:00:00 1994 Newsgroups: bionet.molbio.proteins Path: biosci!bcm!cs.utexas.edu!howland.reston.ans.net!europa.eng.gtefsd.com!newsxfer.itd.umich.edu!nntp.cs.ubc.ca!news.UVic.CA!spruce.pfc.forestry.ca!PFC.Forestry.CA!AEKRAMODDOUL From: aekramoddoul@PFC.Forestry.CA (Ekramoddoullah, Abul Kalam M.) Subject: Re: acetone precipitation of proteins Message-ID: <1994Oct3.211501.20108@spruce.pfc.forestry.ca> Sender: news@spruce.pfc.forestry.ca Nntp-Posting-Host: pfc.pfc.forestry.ca Reply-To: aekramoddoul@PFC.Forestry.CA Organization: Forestry Canada (Pacific Forestry Centre) References: <36fj16$hbh@yoda.Syntex.Com> Date: Mon, 3 Oct 1994 21:15:01 GMT Lines: 16 In article <36fj16$hbh@yoda.Syntex.Com>, simon.lee@syntex.com writes: > >Can anyone send me information on how to do acetone precipitation of cell > >lysates for SDS-PAGE studies? > >I am posting this for a colleague. >Please E-mail to her directly: riitta.lahesmaa@Syntex.com > Yes, you could add 8 X volume (of the sample) pre-cilled acetone (-20C) to your sample and keep it for an hour at -20C. Protein precipitate may then be redissolved in your SDS-PAGE sample buffer. Good Luck. EKRAMODDOULLAH, ABUL KALAM M. Title: Research Scientist Forestry Canada Phone: (604) 363-0600 Victoria, B.C. Internet: AEKRAMODDOUL@A1.PFC.Forestry.CA From owner-proteins@net.bio.net Sun Oct 02 23:00:00 1994 Path: biosci!daresbury!trane.uninett.no!sunic!columba.udac.uu.se!sirius.bmc.uu.se!dunten From: dunten@sirius.bmc.uu.se (Pete Dunten) Newsgroups: bionet.molbio.proteins,bionet.xtallography Subject: Re: Residues at protein surface Date: 3 Oct 1994 21:20:20 GMT Organization: Uppsala University Lines: 12 Distribution: world Message-ID: <36psik$10h1@columba.udac.uu.se> References: <34hbur$rbv@alpha.cisi.unige.it> <36ogsq$i64@rc1.vub.ac.be> <36otb3$f9@umd5.umd.edu> NNTP-Posting-Host: sirius.bmc.uu.se Xref: biosci bionet.molbio.proteins:2802 bionet.xtallography:1212 Ram Samudrala (ram@elan1.carb.nist.gov) wrote >>Will you "normalize" the surface area data to take into account >>that some residues are larger than others? >I believe this is usually (i.e., this is the way I do it) done by >computing the fraction accessible/buried surface area relative to a >"random coil" state. And how good is your "random coil" model? For Trp, for example? Isn't this largely guesswork? From owner-proteins@net.bio.net Sun Oct 02 23:00:00 1994 Path: biosci!galaxy.ucr.edu!ihnp4.ucsd.edu!usc!howland.reston.ans.net!torn!nott!nrcnet0.nrc.ca!BRI.NRC.CA!James.Fethiere From: James.Fethiere@BRI.NRC.CA (James Fethiere) Newsgroups: bionet.molbio.proteins Subject: ion_exchange chromato. Date: 4 Oct 1994 03:56:05 GMT Organization: Institut de recherche en biotechnologie Lines: 3 Message-ID: <36qjol$9eo@nrcnet0.nrc.ca> NNTP-Posting-Host: bobino.bri.nrc.ca X-Newsreader: TIN [version 1.2 PL2] I am also working with a 20 KDa protein, but I can resolve it very well on a regular 12% PAGE. Why would you need the von JAgow protocol? This tricine protocol is mainly for very low MW species in the range of 3000-6000. From owner-proteins@net.bio.net Sun Oct 02 23:00:00 1994 Newsgroups: bionet.molbio.proteins Path: biosci!galaxy.ucr.edu!ihnp4.ucsd.edu!usc!howland.reston.ans.net!torn!nott!cunews!freenet.carleton.ca!FreeNet.Carleton.CA!at332 From: at332@FreeNet.Carleton.CA (Matt Parker) Subject: Protein concentration from peptide bond absorbance Message-ID: Sender: news@freenet.carleton.ca (Usenet News Admin) Organization: The National Capital FreeNet, Ottawa, Ontario, Canada Date: Tue, 4 Oct 1994 03:10:40 GMT Lines: 19 I am working with a protein which has only one tyrosine and no tryptophans. I am using absorbance in the range of 210 nm or so to detect it when doing gel filtration chromatography. A coworker mentioned that he had seen a paper in which a method is described for measuring protein concentrations based on the RATIO of absorances at two wavelengths in this region.... A210 divided by A215 or something like that. This makes sense, because there is a lot of noise in this range, due to buffer, etc. absorbance. Can anybody give me the reference for this method? My coworker has no idea where he saw it, but he thinks that it is a pretty old paper (60's or 70's). Thanks! Matthew Parker Agriculture Canada From owner-proteins@net.bio.net Sun Oct 02 23:00:00 1994 Newsgroups: bionet.molbio.proteins Path: biosci!bcm!cs.utexas.edu!howland.reston.ans.net!europa.eng.gtefsd.com!newsxfer.itd.umich.edu!nntp.cs.ubc.ca!news.UVic.CA!spruce.pfc.forestry.ca!PFC.Forestry.CA!AEKRAMODDOUL From: aekramoddoul@PFC.Forestry.CA (Ekramoddoullah, Abul Kalam M.) Subject: Re: IgM Message-ID: <1994Oct3.213443.20244@spruce.pfc.forestry.ca> Sender: news@spruce.pfc.forestry.ca Nntp-Posting-Host: pfc.pfc.forestry.ca Reply-To: aekramoddoul@PFC.Forestry.CA Organization: Forestry Canada (Pacific Forestry Centre) References: <9409301331.AA20356@dxi.nih.gov> Date: Mon, 3 Oct 1994 21:34:43 GMT Lines: 23 In article <9409301331.AA20356@dxi.nih.gov>, fnunes@pop.nih.gov (Fabio Nunes) writes: >I'm using a monoclonal IgM produced by a company. This antibody labels >filaments in CV1 and other epithelial cells. I'm trying to isolate what >seems to be a new protein. When I run Western blot I obtain 3 bands with >this antibody. I'm wondering if that result could be a cross reaction. Does >anyone have experience in isolate proteins using IgM monoclonal antibodies. >Any help would be greatly appreciated. >Thanks in advance > >Fabio Nunes >NIDCD-NIH >fnunes@pop.nih.gov > > The company should have given you the specificity of the monoclonal antibdoy in terms of its reactivity by Western-immunoblot against the original antigen. Most likely you are dealing with cross-reactive epitope (may bbe carbohydrate in nature; IgM is preferentially produced against sugar moiety). I hope this was a help. Abul EKRAMODDOULLAH, ABUL KALAM M. Title: Research Scientist Forestry Canada Phone: (604) 363-0600 Victoria, B.C. Internet: AEKRAMODDOUL@A1.PFC.Forestry.CA From owner-proteins@net.bio.net Sun Oct 02 23:00:00 1994 Path: biosci!CS.Arizona.EDU!uunet!haven.umd.edu!umd5.umd.edu!ram From: ram@mbisgi.umd.edu (Ram Samudrala) Newsgroups: bionet.molbio.proteins,bionet.xtallography Subject: Re: Residues at protein surface Followup-To: bionet.molbio.proteins,bionet.xtallography Date: 3 Oct 1994 12:27:15 GMT Organization: The Centre for Advanced Research in Biotechnology Lines: 60 Distribution: world Message-ID: <36otb3$f9@umd5.umd.edu> References: <34hbur$rbv@alpha.cisi.unige.it> <36ogsq$i64@rc1.vub.ac.be> Reply-To: ram@elan1.carb.nist.gov NNTP-Posting-Host: indigo3.carb.nist.gov X-Newsreader: TIN [version 1.2 PL0] Xref: biosci bionet.molbio.proteins:2801 bionet.xtallography:1208 Joan Pontius (joan@vandijck.ulb.ac.be) wrote: >>Does anybody have references on studies about surface composition of >>proteins (i.e. statistics on the relative frequency of specific >>residues appearing at the surface of some protein, hopefully made >>on a large subset of the PDB)? >This topic is a part of my current doctorate research, so I would >be interested in knowing why this information is important to you >and anyone else. I didn't make the original post, but I have a small interest in this area (and I do have a real problem, wherein the statistics above might help, but not necessarily so). It would be very nice to look up such a table in some paper somewhere. Consider an hypothetical situation where you are modeling a protein, whose structure you do not know, which has a homology to a sequence with a known structure. And let's assume the sequence alignment program you use comes up with an alignment that looks highly suspicious. In /most/ cases you can take a look at the aligned strand and the residues that are buried and exposed, and say "hey, this set of residues should [not] be exposed, so this is a wrong alignment." I, however, have run across a case in a model I'm trying to build where there are a few exposed hydrophobes in both the model and the real protein. Interestingly enough, the alignment says something contrary to what you would intiutively expect. This is the sequence and ^ indicates it is on the exposed side of the strand: VVCTRIYV ^ ^ ^ ^ ||||| | Conserved (3/6 in a multiple sequence alignment) residues are indicated by |. The R points right into a cavity, which has water in the crystal structure... The interesting thing is that the last V is the only one seen in the model (the remaining 5 are either E or K). Also, the I isn't very popular either. So one wants to know what happens to the sheet at this point. I am more interested from an evolutionary perspective (why does it tolerate these surface mutations?) than anything else. >What do you consider to be the "protein surface"? In this case, once the model has been created, the surface is essentially the area that is exposed to solvent. >Will you "normalize" the surface area data to take into account >that some residues are larger than others? I believe this is usually (i.e., this is the way I do it) done by computing the fraction accessible/buried surface area relative to a "random coil" state. --Ram ram@elan1.carb.nist.gov I had a dream, when I was young, a dream of sweet illusion. A glimpse of hope and unity, and visions of one sweet union. But a cold wind blows, and a dark rain falls, and in my heart it shows, look what they've done to my dream. ---Queen From owner-proteins@net.bio.net Sun Oct 02 23:00:00 1994 Path: biosci!CS.Arizona.EDU!uunet!pipex!lyra.csx.cam.ac.uk!warwick!unicorn.nott.ac.uk!macfd.biochem.nottingham.ac.uk!user From: mbxfd@unicorn.nott.ac.uk (Fergus Doherty) Newsgroups: bionet.molbio.proteins Subject: Antibodies to oxidized proteins Followup-To: bionet.molbio.ageing Date: 3 Oct 1994 12:17:19 GMT Organization: Nottingham University, UK. Lines: 13 Distribution: world Message-ID: NNTP-Posting-Host: macfd.biochem.nottingham.ac.uk Hello, Does anyone out there have abs to oxidized proteins and could they could let me have some? Such abs usually recognize malondialdehyde-lysine and/or 4-HNE-lysine, that are generated in LDL for instance, and will recognize such modifications in other proteins. Abs suitable for Western blotting are what I'm after. Any help appreciated. Fergus Doherty, dept. Biochemistry, University Medical School, Queen's Medical Centre, Nottingham NG7 2UH Tel: 602 709366 FAX 602 422225 Internet: mbxfd@unicorn.nott.ac.uk From owner-proteins@net.bio.net Mon Oct 03 23:00:00 1994 Path: biosci!s.u-tokyo!news.tisn.ad.jp!news.u-tokyo.ac.jp!sinetnews!daffy!uwvax!uwm.edu!news.moneng.mei.com!howland.reston.ans.net!europa.eng.gtefsd.com!uhog.mit.edu!sgiblab!sgigate.sgi.com!enews.sgi.com!decwrl!netcomsv!ix.netcom.com!netnews From: Bhupi@ix.netcom.com (Bhupinder Singh) Newsgroups: bionet.molbio.proteins Subject: Need help: information of plague vaccine Date: 4 Oct 1994 05:50:28 GMT Organization: Netcom Lines: 6 Distribution: world Message-ID: <36qqf4$lk2@ixnews1.ix.netcom.com> NNTP-Posting-Host: ix-sj14-07.ix.netcom.com Due to sudden break in of the plague epidamic in India, there are lot of anxities for demand of vaccine for human plague. Any information regading such vaccine from any agency would help save hundreds of thousands lives. Please get in touch with me as soon as possible. Bhupi Ph.(408) 383-0800 From owner-proteins@net.bio.net Mon Oct 03 23:00:00 1994 Path: biosci!rutgers!uwm.edu!lll-winken.llnl.gov!overload.lbl.gov!agate!howland.reston.ans.net!europa.eng.gtefsd.com!news.umbc.edu!haven.umd.edu!umd5.umd.edu!ram From: ram@mbisgi.umd.edu (Ram Samudrala) Newsgroups: bionet.molbio.proteins,bionet.xtallography Subject: Re: Residues at protein surface Followup-To: bionet.molbio.proteins,bionet.xtallography Date: 4 Oct 1994 06:44:18 GMT Organization: The Centre for Advanced Research in Biotechnology Lines: 26 Distribution: world Message-ID: <36qtk2$svl@umd5.umd.edu> References: <34hbur$rbv@alpha.cisi.unige.it> <36ogsq$i64@rc1.vub.ac.be> <36otb3$f9@umd5.umd.edu> <36psik$10h1@columba.udac.uu.se> Reply-To: ram@elan1.carb.nist.gov NNTP-Posting-Host: indigo3.carb.nist.gov X-Newsreader: TIN [version 1.2 PL0] Xref: biosci bionet.molbio.proteins:2810 bionet.xtallography:1215 Pete Dunten (dunten@sirius.bmc.uu.se) wrote: >Ram Samudrala (ram@elan1.carb.nist.gov) wrote >And how good is your "random coil" model? For Trp, for example? >Isn't this largely guesswork? No, just an assumption which might not be very accurate (or it might be---we really don't know what a random coil "looks" like). We assume the random coil is essentially an extended state conformation of a identical tripeptide, and the surface area is average over this tripeptide. There's a paper on this somewhere, and I'll dig it up if I can. I should read it myself. But in any case, I think in terms of providing a good reference state to calculate the fraction buried/accessible area, it serves its purpose. So the value you end up with, if you're referring to accessibility, is the fraction accessible surface area, relative to the accessible surface area of this extended conformation. And this simply serves as a "normaliser" more than anything else. --Ram ram@elan1.carb.nist.gov If you didn't care what happened to me, and I didn't care for you, we would zig zag our way through the boredom and pain occasionally glancing up through the rain wondering which of the buggers to blame and watching for pigs on the wing. ---Pink Floyd From owner-proteins@net.bio.net Mon Oct 03 23:00:00 1994 Path: biosci!daresbury!doc.ic.ac.uk!agate!howland.reston.ans.net!torn!nott!nrcnet0.nrc.ca!BRI.NRC.CA!James.Fethiere From: James.Fethiere@BRI.NRC.CA (James Fethiere) Newsgroups: bionet.molbio.proteins Subject: ion_exchange chromato. Date: 4 Oct 1994 06:34:20 GMT Organization: Institut de recherche en biotechnologie Lines: 18 Message-ID: <36qt1c$c5q@nrcnet0.nrc.ca> NNTP-Posting-Host: bobino.bri.nrc.ca X-Newsreader: TIN [version 1.2 PL2] Hello there, I would like to get some comment on a purification scheme that I'm trying to setup. My protein is a 20kD, pI<4 glycoprotein expressed in bacculovirus. The first step is a Q-sepharose column used to clean up the bulk of the protein. I observed something strange: In a Bis-Tris buffer 25mM pH 6.0 the protein of interest starts eluting in as low as 25mM NaCl at the same pH, CAN SOMEBODY PROVIDE AN EXPLANATION FOR THAT?? Next I would like to use this step for further purification since the affinity column I use as a next step has a very low capacity. So I would like to load the Q-sepharose at pH 6.0, and then wash it at pH 5.0 to get rid of most of the contaminants that have a pI higher than my protein. IS IT BETTER TO LOAD DIRECTLY AT THIS PH (5.0)? WILL THE PH CHANGE CAUSE ANY PRECIPITATION OF OTHER PROTEIN ON MY COLUMN?? WHAT WOULD BE THE BEST BUFFER FOR THAT PH 5.0 WASH? Thank you all for your help From owner-proteins@net.bio.net Mon Oct 03 23:00:00 1994 Path: biosci!bcm!cs.utexas.edu!howland.reston.ans.net!EU.net!sunic!columba.udac.uu.se!sirius.bmc.uu.se!seab From: seab@sirius.bmc.uu.se (Stefan Baeckstrom) Newsgroups: bionet.molbio.proteins Subject: Need advice about inclusion bodys !! Date: 4 Oct 1994 12:17:24 GMT Organization: Institution of molecular biology, BMC, Uppsala, Sweden Lines: 16 Distribution: world Message-ID: <36rh4k$9si@columba.udac.uu.se> NNTP-Posting-Host: sirius.bmc.uu.se Does anybody have experience with inclusion bodys in Yeast ? I have done a mutant where a histidine is replaced by phenylalanin and I think there will be problems when purifying it, because it is growing slow like hell. More than double the time of the other mutants that I have done (on the same residue). I thought that the slow growth might be due to some aggregation of the protein, caused by this. I dont know much yet, I just wonder if somebody have experienced something like this when mutated a residue into a phenylalanine. I look forward to hear some thoughts about this. Stefan Baeckstroem, Uppsala From owner-proteins@net.bio.net Mon Oct 03 23:00:00 1994 Newsgroups: bionet.molbio.proteins Path: biosci!bcm!cs.utexas.edu!howland.reston.ans.net!EU.net!sunic!news.funet.fi!luotsi.uku.fi!messi.uku.fi!hkokko From: hkokko@messi.uku.fi (Harri Kokko) Subject: stabilization of HRP-antibody conjugates? Sender: news@luotsi.uku.fi Message-ID: Date: 4 Oct 94 10:35:37 GMT Organization: University of Kuopio, Finland Keywords: antibody, HRP, stabilization Lines: 15 I wonder if anybody know how to stabilize antibody-HRP conjugates? Have somene experince on lyofilization or freezing of HRP-antibody conjucates? I greatly appreciate any help. Harri Kokko Department of Biochemistry and Biotechnology University of Kuopio P.O.Box 1627 FIN-70211 Kuopio Finland Email: Hkokko@messi.uku.fi From owner-proteins@net.bio.net Mon Oct 03 23:00:00 1994 Path: biosci!bcm!cs.utexas.edu!howland.reston.ans.net!wupost!cerberus-138.wustl.edu!medicine!riesenbe From: riesenbe@medicine (Brad Riesenberger) Newsgroups: bionet.molbio.proteins Subject: Bovine hemoglobin, ext. coeff.? lambda max.? Date: 4 Oct 1994 20:08:14 GMT Organization: Washington University School of Medicine Lines: 8 Message-ID: <36scne$28q@cerberus-138.wustl.edu> NNTP-Posting-Host: medicine.wustl.edu Keywords: hemoglobin C Might anyone have a reference for the extinction coefficient or lambda max. for bovine Hb? Many thanks, and regards, Brad Riesenberger riesenbe@medicine.wustl.edu From owner-proteins@net.bio.net Mon Oct 03 23:00:00 1994 Path: biosci!bcm!cs.utexas.edu!howland.reston.ans.net!europa.eng.gtefsd.com!newsxfer.itd.umich.edu!jobone!heifetz.msen.com!zib-berlin.de!fauern!lrz-muenchen.de!ipp-garching.mpg.de!genmac14.biochem.mpg.de!user From: spang@vms.biochem.mpg.de (Anne Spang) Newsgroups: bionet.molbio.proteins Subject: Re: stabilization of HRP-antibody conjugates? Date: 4 Oct 1994 18:32:48 GMT Organization: Rechenzentrum der Max-Planck-Gesellschaft in Garching Lines: 13 Message-ID: References: NNTP-Posting-Host: genmac14.biochem.mpg.de Hi Harri! I´m working quite a while with HRP-conjugated antibodies. You may store your antibodies even in the required dilution at 4C. But you should be careful because the milk will become sour after a while. the other possibility might be, to store the aliquotes of your conjugates at -20C. I also store my antibody solution in milk at -20C without lost of the quality of the signal. Hope this helps! Anne From owner-proteins@net.bio.net Mon Oct 03 23:00:00 1994 Path: biosci!rutgers!netnews.upenn.edu!msuinfo!miller.prc.msu.edu From: kellerj@pilot.msu.edu (Jim Keller) Newsgroups: bionet.molbio.proteins Subject: What is GELBOND? Date: Tue, 04 Oct 1994 13:22 EST Organization: Michigam State University Lines: 6 Distribution: world Message-ID: <19941004132211.kellerj@miller.prc.msu.edu> Reply-To: kellerj@pilot.msu.edu NNTP-Posting-Host: miller.prc.msu.edu X-Newsreader: FTPNuz (DOS) v1.1b I have a paper from 1988 on gradient-PAGE that says "The slab gel was formed with GELBOND as support". The authors do not list GELBOND in the matrials section. Does anyone know what GELBOND is? E-Mail me: Kellerj@pilot.msu.edu. Thankyou for your time Jim From owner-proteins@net.bio.net Mon Oct 03 23:00:00 1994 Path: biosci!galaxy.ucr.edu!ihnp4.ucsd.edu!swrinde!pipex!lyra.csx.cam.ac.uk!crcmac2.path.cam.ac.uk!user From: smh1008@cus.cam.ac.uk (David S. Huen) Newsgroups: bionet.molbio.methds-reagnts,bionet.molbio.proteins Subject: Antibody against neomycin phosphotransferase Followup-To: bionet.molbio.methds-reagnts Date: Tue, 04 Oct 1994 18:26:42 +0400 Organization: CRC Human Cancer Genetics Group Lines: 16 Distribution: world Message-ID: NNTP-Posting-Host: crcmac2.path.cam.ac.uk Xref: biosci bionet.molbio.methds-reagnts:19189 bionet.molbio.proteins:2814 Hi, I have a query from a colleague who wants to obtain a MAb or polyclonal directed against neomycin phosphotransferase. He intends to use it as a lineage marker in transgenics. Does anyone know of such a reagent? Thanks. -- David S. Huen Phone: (0223) 333921 CRC Human Cancer Genetics Group Fax: (0223) 333875 Dept. of Pathology e-mail: smh1008@cus.cam.ac.uk University of Cambridge CB2 1QP United Kingdom From owner-proteins@net.bio.net Mon Oct 03 23:00:00 1994 Path: biosci!bcm!cs.utexas.edu!howland.reston.ans.net!vixen.cso.uiuc.edu!news.uoregon.edu!fp1-molbio-18.uoregon.edu!user From: thomas@molbio.uoregon.edu (Matt thomas) Newsgroups: bionet.molbio.proteins Subject: pol II and CTD Date: Tue, 04 Oct 1994 18:22:07 -0700 Organization: hawly lab, University of Oregon Lines: 9 Message-ID: NNTP-Posting-Host: fp1-molbio-18.uoregon.edu I have recently purified some pol II in the IIA form (intact CTD). Does anyone know of an easy way to cut the CTD off with trypsin or some other protease? I could just go back into the cold room and use a different protocal and get the IIb form that way, but I've spent too much time in there lately! :) burrrrr! -- Who has time to think of a snappy sig when you are in grad school. :) Thomas@molbio.uoregon.edu From owner-proteins@net.bio.net Mon Oct 03 23:00:00 1994 Path: biosci!bcm!cs.utexas.edu!howland.reston.ans.net!europa.eng.gtefsd.com!newsxfer.itd.umich.edu!jobone!lynx.unm.edu!carina!inkim From: inkim@carina.unm.edu (In C. Kim) Newsgroups: bionet.molbio.proteins Subject: Re: What is GELBOND? Date: 4 Oct 1994 21:23:21 GMT Organization: University of New Mexico, Albuquerque Lines: 18 Distribution: world Message-ID: <36sh49$9cr@lynx.unm.edu> References: <19941004132211.kellerj@miller.prc.msu.edu> NNTP-Posting-Host: carina.unm.edu X-Newsreader: TIN [version 1.2 PL2] Jim Keller (kellerj@pilot.msu.edu) wrote: : I have a paper from 1988 on gradient-PAGE that says "The slab gel was formed : with GELBOND as support". The authors do not list GELBOND in the matrials : section. Does anyone know what GELBOND is? E-Mail me: Kellerj@pilot.msu.edu. : Thankyou for your time : Jim Jim, GelBonds are sold by FMC Corp, Rockland, ME. There are two types of gelbond films: GelBond PAG and GelBond film for agarose gel. For details, call FMC at 1-800-341-1574 (for order) or 1-800-521-0390 (for tech support). __ In C. Kim, Albuquerque, New Mexico Land of Enchantment From owner-proteins@net.bio.net Mon Oct 03 23:00:00 1994 Path: biosci!bcm!cs.utexas.edu!howland.reston.ans.net!EU.net!sunic!trane.uninett.no!daresbury!bioftp.unibas.ch!rc1.vub.ac.be!vandijck!joan From: joan@vandijck.ulb.ac.be (Joan Pontius) Newsgroups: bionet.molbio.proteins Subject: Re: Recommendations for light level protein detection? Date: 4 Oct 1994 11:40:09 GMT Organization: UCMB - ULB Lines: 15 Sender: joan@vandijck (Joan Pontius) Distribution: world Message-ID: <36reup$rho@rc1.vub.ac.be> References: NNTP-Posting-Host: ucmbsgi9.ulb.ac.be. >peroxidase. We have considered using fluorescent conjugated antibodies, >but the autofluorescence of some tissues in the anther may be another >problem that I then have to deal with. I say try it. If auto-fluorescence is only occurring at a narrow wavelength, you could use a labeled anti-body that fluoresses (spelling?) at another. THEN you could analyze using flow cytometery, which is always a lot of fun. Joan Pontius joan@ucmb.ulb.ac.be From owner-proteins@net.bio.net Tue Oct 04 23:00:00 1994 Path: biosci!rutgers!netnews.upenn.edu!elmo.tju.edu!calvin.jci.tju.edu!harrison From: harrison@calvin.jci.tju.edu Newsgroups: bionet.molbio.proteins Subject: Re: Spot size <--> Xtal Size Date: 5 Oct 1994 18:52:43 GMT Organization: Jefferson Cancer Institute Lines: 31 Distribution: usa Message-ID: <36uslr$37l@elmo.tju.edu> References: <36h92p$32n@agate.berkeley.edu> NNTP-Posting-Host: calvin.jci.tju.edu In article <36h92p$32n@agate.berkeley.edu>, lhom@OCF.Berkeley.EDU (Louis Hom) writes: |> On those diffraction patterns that I've only seen photos of -- is there a |> relationship between the spot sizes and the size of the crystal? |> -- Sometimes. This is not meant to be funny. It depends on the particular x-ray beam geometry, and the mosaic spread of the crystal. If the beam divergence is zero and there is almost no mosaic spread then the spot size and crystal size are very similar. The spot size will still depend on the crystal shape and orientation. There are all sorts of beam geometries, so the exact relationship will vary. (for example you can focus the beam on a small part of a crystal and then the spot size really has nothing to do with the crystal size). When a well-collimated beam is used, the measured spot size is used to refine the parameter for the mosaic spread when collecting data (which is then used to re-estimate the spots). -- robert w. harrison harrison@asterix.jci.tju.edu ******************************************************************************** Organic chemistry is the chemistry of carbon compounds. Biochemistry is the study of carbon compounds that crawl. -- Mike Adams For every complex problem, there is a solution that is simple, neat, and wrong. -- H. L. Mencken ******************************************************************************** From owner-proteins@net.bio.net Tue Oct 04 23:00:00 1994 Path: biosci!agate!howland.reston.ans.net!EU.net!uknet!daresbury!not-for-mail From: SPAGNOL@PLAUTO.CSATA.IT Newsgroups: bionet.molbio.proteins Subject: In search of the "Holy manual" Date: 5 Oct 1994 09:27:23 +0100 Lines: 10 Sender: lpddist@mserv1.dl.ac.uk Distribution: bionet Message-ID: <36to1b$4eg@mserv1.dl.ac.uk> Original-To: proteins@dl.ac.uk Dear colleagues, I desperately seek a laboratory manual in the field of proteins which (ideally) should be what Maniatis is for nucleic acids.In short it should consist of well explained general principles about protein biochemistry and immunochemistry and accurate protocols joined in the same book.Does someone outthere know a book like that? Sincerely ,Giorgio: a molecular biologist who is paying the p rice of superspecialization. From owner-proteins@net.bio.net Tue Oct 04 23:00:00 1994 Path: biosci!agate!howland.reston.ans.net!EU.net!uknet!daresbury!not-for-mail From: SPAGNOL@PLAUTO.CSATA.IT Newsgroups: bionet.molbio.proteins Subject: In search of the "Holy manual" Date: 5 Oct 1994 09:27:19 +0100 Lines: 8 Sender: lpddist@mserv1.dl.ac.uk Distribution: bionet Message-ID: <36to17$4dv@mserv1.dl.ac.uk> Original-To: proteins@dl.ac.uk, methods@dl.ac.uk Dear colleagues, I desperately seek a laboratory manual in the field of proteins which (ideally) should be what Maniatis is for nucleic acids.In short it should consist of well explained general principles about protein biochemistry and immunochemistry and accurate protocols joined in the same book.Does someone outthere know a book like that? Sincerely ,Giorgio: a molecular biologist who is paying the price of superspecialization. From owner-proteins@net.bio.net Tue Oct 04 23:00:00 1994 Path: biosci!galaxy.ucr.edu!ihnp4.ucsd.edu!swrinde!cs.utexas.edu!convex!news.duke.edu!godot.cc.duq.edu!newsfeed.pitt.edu!dsinc!netnews.upenn.edu!elmo.tju.edu!calvin.jci.tju.edu!harrison From: harrison@calvin.jci.tju.edu Newsgroups: bionet.molbio.proteins Subject: Re: Lennard-Jones Potential Date: 5 Oct 1994 18:41:10 GMT Organization: Jefferson Cancer Institute Lines: 42 Distribution: world Message-ID: <36us06$37l@elmo.tju.edu> References: <36occc$gu2@rc1.vub.ac.be> NNTP-Posting-Host: calvin.jci.tju.edu In article <36occc$gu2@rc1.vub.ac.be>, joan@vandijck.ulb.ac.be (Joan Pontius) writes: |> Can someone explain to me why the Lennard-Jones |> potential, which describes gas behaviour, is used |> in energy-minimization programs for proteins? |> |> joan@ucmb.ulb.ac.be |> Partially the answer is for want of anything better ;-). Actually the 1/r^6 terms are used to model induced dipole/induced dipole terms. There are several variant perturbation treatments for this in terms of ionization energies ... , but it really is best thought of as an adjustable parameter that is fit to the data. 1/r^12 is used as a model for exclusion potentials (no two fermions can ...) and was chosen (imho) because 12 = 6+6. There are other models like Buckingham (6,exp) and 6,9 potentials which can be used. Those who use them claim they are a lot better, but i haven't seen much difference except in special cases. There are a lot more serious errors in common practice, like the use of a cutoff radius and 1/r dielectrics. The use of a united atom potential (ch3 as one atom etc) is a great way to get poor quality results. The point charges are not the best way to represent the charge distribution of an atom at short distances. Finally, there's always mutual polarization of the atoms as well. These can all be treated but at some computational cost. -- robert w. harrison harrison@asterix.jci.tju.edu ******************************************************************************** When in doubt, use brute force. -- Ken Thompson For every complex problem, there is a solution that is simple, neat, and wrong. -- H. L. Mencken ******************************************************************************** From owner-proteins@net.bio.net Tue Oct 04 23:00:00 1994 Path: biosci!galaxy.ucr.edu!ihnp4.ucsd.edu!scripps.edu!NewsWatcher!user From: Bibbs@scripps.edu (Lisa Bibbs) Newsgroups: bionet.molbio.proteins Subject: Re: preparing a protein for sequencing Date: 5 Oct 1994 21:01:41 GMT Organization: The Scripps Research Institute, La Jolla, CA Lines: 36 Distribution: world Message-ID: References: <9410051957.AA28182@molbio.cbs.umn.edu> NNTP-Posting-Host: pbfnuts.scripps.edu Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit In article <9410051957.AA28182@molbio.cbs.umn.edu>, nora@MOLBIO.CBS.UMN.EDU ("Nora Vig") wrote: > I am planning to isolate a protein by affinity binding, followed by SDS gel > electrophoresis, and I would like to prepare it for N-terminal sequencing. I > have several choices to make, and I would appreciate some help or insight that > anyone might have. First, is it true that less protein is required for the > sequencing (Edman degradation) if the protein (or peptide) is blotted to nitro > cellulose rather than to PVDF membrane? Secondly, how frequently are proteins > found to be N-terminally blocked, and does it make sense to anticipate this by > making peptides instead? Any other details or references on this subject would be > appreciated. > Wow Nora. First, a protein can't be sequenced from nitrocellulose. The reagents used in the Edman degredation will dissolve it as I understand. (If I'm incorrect someone please point it out.) How frequently are proteins found to be N-terminally blocked? According to Matsudaira, "80% of proteins in Ehrlich ascites cells are acetylated". (A Practical Guide to Protein and Peptide Purification for Microsequencing 2nd Edition, Paul Matsudaira.) Your protein can be digested off of nitrocellulose though and there are several papers in Analytical Biochem concerning this. Search for Aebersold et. al. or Fernandez, Joseph et. al. from Rockefeller. Sorry I don't have them on the tip of my tonque. There is also a method in the book mentioned above. Mainly, I would contact the facility you are going to use and talk to the person in charge of the sequencing. I'm sure they would love to guide you since we all hate failure sequences. Good Luck, Lisa Bibbs -- These opinions are mine, not my employers. From owner-proteins@net.bio.net Tue Oct 04 23:00:00 1994 Path: biosci!galaxy.ucr.edu!ihnp4.ucsd.edu!swrinde!howland.reston.ans.net!darwin.sura.net!mother.usf.edu!com1!pneame From: pneame@com1.med.usf.edu (Peter J. Neame) Newsgroups: bionet.molbio.proteins Subject: Re: preparing a protein for sequencing Date: 5 Oct 1994 22:56:28 GMT Organization: University of South Florida Lines: 18 Distribution: world Message-ID: <36vaus$gs3@mother.usf.edu> References: <9410051957.AA28182@molbio.cbs.umn.edu> NNTP-Posting-Host: com1.med.usf.edu X-Newsreader: TIN [version 1.2 PL2] As a general rule, if you've got enough protein, it is a good idea to go for internal sequence. However, it depends on the amount that you can get onto your blot. There are a number of different types of Immobilon (PVDF) available, which work well to varying degrees. Our general rule of thumb is that if you can get a strong Coomassie staining band on your blot, and your protein is < 50kDa, then you are in good shape for doing a trypsin or endoprotease Lys-C digestion. Bigger proteins will benefit from Lys-C, as you'll get fewer fragments. You WILL need an HPLC with microbore capability to seperate the peptides, and do an enzyme blank with a bit of membrane with nothing on it. But as Lisa said, check with your local facility. Our "Bible" is the Fernandez et al Anal Biochem article of a couple of years ago. Good luck! Peter Neame USF and SHCC, Tampa, FL From owner-proteins@net.bio.net Tue Oct 04 23:00:00 1994 Newsgroups: bionet.molbio.proteins Path: biosci!bcm!cs.utexas.edu!howland.reston.ans.net!vixen.cso.uiuc.edu!news.uoregon.edu!netnews.nwnet.net!serval.net.wsu.edu!wsuaix.csc.wsu.edu!pxu From: pxu@wsuaix.csc.wsu.edu (Pin Xu) Subject: Re: Protein FPLC (HPLC) Systems Message-ID: Sender: news@serval.net.wsu.edu (News) Organization: Washington State University X-Newsreader: Tin 1.1 PL4 References: Date: Thu, 6 Oct 1994 02:28:34 GMT Lines: 30 jlitts@onyx-pharm.com (Jim Litts) writes: : I am researching the purchase of a new HPLC/FPLC system to be used : primarily to do routine protein purifications of about 10 mg with : capabilities to be automated and to be scaled to quantities well in excess : of 100 mg, but seldom, or never above 1 gram. : : I am amazed at the variability in prices out there between systems. I have : been looking at Pharmacia, Shimadzu, PerSeptive Biosystems, Ranin, and : Waters. They have systems that generally cover the needs that I have with : prices ranging from $20,000-$80,000. Certainly there are differences, : particularly with respect to flexibility and software, but I'm surprised : that the range is so broad. : : Does anyone with enough time to respond to this have any experience with : any of these systems? I'm particularly interested in reliability, opinions : of cost/benefit for the high-end products, operating costs, and the ease of : training new users. How is the software? Is it pretty easy to get into, : or are you usually arguing with it and calling it names? Do you have : another unmentioned that you really like? My inclination is to go cheap : for the routine stuff, has anybody got any particular regrets (or pleased) : for having done this? : : If you would prefer to respond by Email to: jlitts@onyx-pharm.com , I would : appreciate the help and I will sum up what I get, and repost it in the next You can buy a Water HPLC system: a pump, a PDA, an expensive but very impressive workstation and software, an autosampler. It costs about 50,000. You can buy HPLC columns for protein purification. : From owner-proteins@net.bio.net Tue Oct 04 23:00:00 1994 Path: biosci!MOLBIO.CBS.UMN.EDU!nora From: nora@MOLBIO.CBS.UMN.EDU ("Nora Vig") Newsgroups: bionet.molbio.proteins Subject: preparing a protein for sequencing Date: 5 Oct 1994 12:58:55 -0700 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 17 Sender: daemon@net.bio.net Distribution: world Message-ID: <9410051957.AA28182@molbio.cbs.umn.edu> NNTP-Posting-Host: net.bio.net I am planning to isolate a protein by affinity binding, followed by SDS gel electrophoresis, and I would like to prepare it for N-terminal sequencing. I have several choices to make, and I would appreciate some help or insight that anyone might have. First, is it true that less protein is required for the sequencing (Edman degradation) if the protein (or peptide) is blotted to nitro cellulose rather than to PVDF membrane? Secondly, how frequently are proteins found to be N-terminally blocked, and does it make sense to anticipate this by making peptides instead? Any other details or references on this subject would be appreciated. Nora Plesofsky-Vig Department of Plant Biology University of Minnesota St. Paul, MN 55108 nora@molbio.cbs.umn.edu From owner-proteins@net.bio.net Tue Oct 04 23:00:00 1994 Newsgroups: bionet.molbio.proteins Path: biosci!bcm!cs.utexas.edu!howland.reston.ans.net!vixen.cso.uiuc.edu!uchinews!daffy!uwvax!sinetnews!news.u-tokyo.ac.jp!news.tisn.ad.jp!riksun!rkna50!jhonda From: jhonda@rkna50.riken.go.jp (Jun Honda) Subject: Numbering AA residues Message-ID: Sender: news@rkna50.riken.go.jp (News Administrator) Nntp-Posting-Host: mail Organization: Institute of Physical & Chemical Research (RIKEN) Saitama,Japan Date: Thu, 6 Oct 1994 04:44:11 GMT Lines: 13 Dear colleagues, I would like to know if there is any strict rule concerning the numbering of amino acid residues in a sequence of a given protein. It often occurs that proteins are modified after synthesis by cleavage of peptides. Is it common to include the removed residues upon numbering (i.e. always count the initiating Met as residue 1), or not (designate the first N-terminal residue as 1 after post translational modification). Or is there not any strict rule? Thanks, Jun Honda @ RIKEN From owner-proteins@net.bio.net Tue Oct 04 23:00:00 1994 Path: biosci!agate!barrnet.net!onyx-pharm.com!NewsWatcher!user From: jlitts@onyx-pharm.com (Jim Litts) Newsgroups: bionet.molbio.proteins Subject: Protein FPLC (HPLC) Systems Followup-To: bionet.molbio.proteins Date: Wed, 05 Oct 1994 17:57:47 -0700 Organization: Onyx Pharmaceuticals Lines: 24 Distribution: world Message-ID: NNTP-Posting-Host: dbl.onyx-pharm.com I am researching the purchase of a new HPLC/FPLC system to be used primarily to do routine protein purifications of about 10 mg with capabilities to be automated and to be scaled to quantities well in excess of 100 mg, but seldom, or never above 1 gram. I am amazed at the variability in prices out there between systems. I have been looking at Pharmacia, Shimadzu, PerSeptive Biosystems, Ranin, and Waters. They have systems that generally cover the needs that I have with prices ranging from $20,000-$80,000. Certainly there are differences, particularly with respect to flexibility and software, but I'm surprised that the range is so broad. Does anyone with enough time to respond to this have any experience with any of these systems? I'm particularly interested in reliability, opinions of cost/benefit for the high-end products, operating costs, and the ease of training new users. How is the software? Is it pretty easy to get into, or are you usually arguing with it and calling it names? Do you have another unmentioned that you really like? My inclination is to go cheap for the routine stuff, has anybody got any particular regrets (or pleased) for having done this? If you would prefer to respond by Email to: jlitts@onyx-pharm.com , I would appreciate the help and I will sum up what I get, and repost it in the next week or so. Thanx in advance for any comments. From owner-proteins@net.bio.net Tue Oct 04 23:00:00 1994 Path: biosci!agate!barrnet.net!onyx-pharm.com!NewsWatcher!user From: jlitts@onyx-pharm.com (Jim Litts) Newsgroups: bionet.molbio.proteins Subject: Protein FPLC (HPLC) Systems Followup-To: bionet.molbio.proteins Date: Wed, 05 Oct 1994 17:56:16 -0700 Organization: Onyx Pharmaceuticals Lines: 24 Distribution: world Message-ID: NNTP-Posting-Host: dbl.onyx-pharm.com I am researching the purchase of a new HPLC/FPLC system to be used primarily to do routine protein purifications of about 10 mg with capabilities to be automated and to be scaled to quantities well in excess of 100 mg, but seldom, or never above 1 gram. I am amazed at the variability in prices out there between systems. I have been looking at Pharmacia, Shimadzu, PerSeptive Biosystems, Ranin, and Waters. They have systems that generally cover the needs that I have with prices ranging from $20,000-$80,000. Certainly there are differences, particularly with respect to flexibility and software, but I'm surprised that the range is so broad. Does anyone with enough time to respond to this have any experience with any of these systems? I'm particularly interested in reliability, opinions of cost/benefit for the high-end products, operating costs, and the ease of training new users. How is the software? Is it pretty easy to get into, or are you usually arguing with it and calling it names? Do you have another unmentioned that you really like? My inclination is to go cheap for the routine stuff, has anybody got any particular regrets (or pleased) for having done this? If you would prefer to respond by Email to: jlitts@onyx-pharm.com , I would appreciate the help and I will sum up what I get, and repost it in the next week or so. Thanx in advance for any comments. From owner-proteins@net.bio.net Wed Oct 05 23:00:00 1994 Path: biosci!MODEL.PHR.UTEXAS.EDU!rhodes From: rhodes@MODEL.PHR.UTEXAS.EDU ("David G. Rhodes") Newsgroups: bionet.molbio.proteins Subject: Re: In search of the "Holy manual" Date: 6 Oct 1994 05:01:03 -0700 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 27 Sender: daemon@net.bio.net Distribution: world Message-ID: <9410060700.ZM18720@model.phr.utexas.edu> NNTP-Posting-Host: net.bio.net On Oct 5, 9:27am, SPAGNOL@PLAUTO.CSATA.IT wrote: > Subject: In search of the "Holy manual" > I desperately seek a laboratory manual in the field of proteins which ... > should consist of well explained general principles about protein > biochemistry and immunochemistry and accurate protocols joined in the > same book. Look to the "a practical approach" series published by IRL Press division of Oxford University Press. There are volumes on _dozens_ of topics, and while they are somewhat more specialized than you have requested, they make up for that in being comprehensive. They include review, as well as specific protocols. They are also quite 'readable'. - have fun !! PS - I don't work for them... }:) -- _____________________________________ O==O ________________________________ | David G. Rhodes | O==O | RHODES@MODEL.PHR.UTEXAS.EDU | | Pharmaceutics Division | O==O | | | College of Pharmacy | O==O | Phone: (512)471-4681 | | The University of Texas at Austin | O==O | Fax: (512)471-7474 | | Austin, TX 78712-1074 | O==O | }:) | |___________________________________| O==O |______________________________| O==O From owner-proteins@net.bio.net Wed Oct 05 23:00:00 1994 Path: biosci!bcm!cs.utexas.edu!geraldo.cc.utexas.edu!news.utdallas.edu!corpgate!bcarh8ac.bnr.ca!bcarh189.bnr.ca!nott!nrcnet0.nrc.ca!BRI.NRC.CA!James.Fethiere From: James.Fethiere@BRI.NRC.CA (James Fethiere) Newsgroups: bionet.molbio.proteins Subject: pluronic_acid Date: 6 Oct 1994 17:34:52 GMT Organization: Institut de recherche en biotechnologie Lines: 12 Message-ID: <371cfs$gmf@nrcnet0.nrc.ca> NNTP-Posting-Host: bobino.bri.nrc.ca X-Newsreader: TIN [version 1.2 PL2] Hello everyone, Did anybody expressing proteins in sf9 cells (in the presence of pluronic acid) ever had problems to do ammonium sulfate precipitation in the presence of pluronic acid. This agent seem to act like a detergent, and in my case, I did not get any pellet during precipitation, and the whole sample was cloudy and greasy. thanks for your help ../james From owner-proteins@net.bio.net Wed Oct 05 23:00:00 1994 Path: biosci!ic.ac.uk!v.gutsmann From: v.gutsmann@ic.ac.uk (volker gutsmann, volker gutsmann) Newsgroups: bionet.molbio.proteins Subject: Refolding of proteins Date: 6 Oct 1994 03:02:21 -0700 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 24 Sender: daemon@net.bio.net Distribution: world Message-ID: <9410061001.AA11325@bicmda.bio.ic.ac.uk> NNTP-Posting-Host: net.bio.net I am trying to find someone who could recommend a method of denaturing peptides with urea followed by refolding by dilution into a buffer system. Previously I expressed several peptides as fusion proteins to MBP. However, digestion of the fusion proteins with Factor Xa was very inefficient but the tiny amounts of cleaved peptides were biologically active. I recloned for expression in the pET system (pET15b) to produce unfused peptides. This system produced large amounts of recombinant peptides (50-70% of the total soluble protein). To my disappointment, the pET-produced peptides were only weakly recognised by specific antisera and they had lost their biological activity. I therefore would like to unfold the new peptides with 8M urea and then try to refold them. There are quite a few methods around but we do not have any expreience with this sort of work. If anybody could recommend a method or has any other ideas I would be very grateful indeed. Many thanks Volker Gutsmann (vg@bio.ic.ac.uk) Imperial College Dept.Biology London, UK From owner-proteins@net.bio.net Wed Oct 05 23:00:00 1994 Path: biosci!PORTIA.CALTECH.EDU!EARN From: EARN@PORTIA.CALTECH.EDU Newsgroups: bionet.molbio.proteins Subject: Peptide Antibodies Date: 6 Oct 1994 17:56:45 -0700 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 13 Sender: daemon@net.bio.net Distribution: world Message-ID: <01HHYQT9BAHEBL2YX5@PORTIA.CALTECH.EDU> NNTP-Posting-Host: net.bio.net Does anyone have experience with the logistics of having peptide antibodies made? I'd like to do what's fastest and/or cheapest but I don't know if it's best to have peptides synthesized at an in-house core facility (and whether to couple them yourself) and send them off, or are there companies you can just send sequence to and they'll take care of it all. Also I'd like recommendations for various rabbit-poking companies as to the quality of their polyclonals, price, etc. If you email me privately I will post a digest to this list. Thanks in advance for your help. - Eric Arn Division of Biology Caltech EARN@Portia.caltech.edu From owner-proteins@net.bio.net Wed Oct 05 23:00:00 1994 Path: biosci!bcm!cs.utexas.edu!swrinde!howland.reston.ans.net!vixen.cso.uiuc.edu!news.uoregon.edu!fp1-chemistry-16.uoregon.edu!user From: RMyers@molbio.uoregon.edu (Rik Myers) Newsgroups: bionet.molbio.proteins Subject: Beteine and inclusion bodies Followup-To: bionet.molbio.proteins Date: 6 Oct 1994 18:39:13 GMT Organization: Institute of Molecular Biology, U of Oregon Lines: 13 Message-ID: NNTP-Posting-Host: fp1-chemistry-16.uoregon.edu I recall a posting some time ago concerning methods for minimizing aggregation of overproduced proteins in vivo. Decreasing the temperture during growth was one suggestion (which I have found to work pretty well). Including the osmoprotectant Beteine was another suggestion, one I'd like to try. Has anybody found that including Beteine in the growth medium haelps keep proteins soluble? If so, at which concentration do you suggest supplementing the medium? Thanks for your time. -- Rik Myers ********************************************************* * Earth is an asylum for angels with amnesia -- Emerson * ********************************************************* From owner-proteins@net.bio.net Wed Oct 05 23:00:00 1994 Path: biosci!daresbury!not-for-mail From: H1411Mus@HUELLA.BITNET Newsgroups: bionet.molbio.proteins Subject: SUBSCRIBTION Date: 6 Oct 1994 12:32:28 +0100 Lines: 2 Sender: lpddist@mserv1.dl.ac.uk Distribution: bionet Message-ID: <370n8c$ivu@mserv1.dl.ac.uk> Original-To: proteins@dl.ac.uk Please, subscribe me. From owner-proteins@net.bio.net Thu Oct 06 23:00:00 1994 Path: biosci!agate!howland.reston.ans.net!noc.near.net!lecso.neb.com!NewsWatcher!user From: soltis@neb.com (Tony Soltis) Newsgroups: bionet.molbio.methds-reagnts,bionet.molbio.proteins Subject: keratinase / alkaline protease Followup-To: bionet.molbio.proteins Date: 7 Oct 1994 13:12:06 GMT Organization: New England Biolabs Lines: 10 Distribution: world Message-ID: NNTP-Posting-Host: soltis.neb.com Xref: biosci bionet.molbio.methds-reagnts:19352 bionet.molbio.proteins:2838 Would anyone know if the following enzymes are commercially available: 1. Keratinase from Bacillus licheniformis PWD 1 2. Alkaline Protease from Bacillus sp. AH 101 Thanks, Tony Soltis soltis@neb.com "Computers are useless. They can only give you answers." --Pablo Picasso From owner-proteins@net.bio.net Thu Oct 06 23:00:00 1994 Path: biosci!CMGM.STANFORD.EDU!levy From: levy@CMGM.STANFORD.EDU (Shoshana Levy) Newsgroups: bionet.molbio.proteins Subject: Refolding of proteins Date: 7 Oct 1994 09:49:58 -0700 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 9 Sender: daemon@net.bio.net Distribution: world Message-ID: NNTP-Posting-Host: net.bio.net regarding the question of refolding proteins produced in the pET system: Try refolding the protein directly on the nickel column (used to bind the His tag). The protein is loaded in 8M urea, washed and then rather than eluting it immediately, use a urea gradient. Do so very slowly (several hr), the refolded protein is then eluted. Shoshana Levy Med-Onc Stanford From owner-proteins@net.bio.net Thu Oct 06 23:00:00 1994 Path: biosci!agate!news.ucdavis.edu!garter.ece.ucdavis.edu!varma From: varma@garter.ece.ucdavis.edu (Hemant Varma) Newsgroups: bionet.molbio.proteins Subject: Re: ANNOYING ACTORS/ACTRESSES IN HINDI FILMS Date: 7 Oct 1994 20:43:18 GMT Organization: University of California, Davis Lines: 1 Message-ID: <374bt6$luh@mark.ucdavis.edu> NNTP-Posting-Host: garter.ece.ucdavis.edu X-Newsreader: TIN [version 1.2 PL2] From owner-proteins@net.bio.net Thu Oct 06 23:00:00 1994 Path: biosci!agate!howland.reston.ans.net!news.sprintlink.net!redstone.interpath.net!ddsw1!panix!zip.eecs.umich.edu!yeshua.marcam.com!charnel.ecst.csuchico.edu!olivea!decwrl!nntp.crl.com!crl8.crl.com!not-for-mail From: sunger@crl.com (Stefan Unger) Newsgroups: bionet.molbio.proteins Subject: Minimal Set of Proteins Date: 7 Oct 1994 00:28:16 -0700 Organization: CRL Dialup Internet Access (415) 705-6060 [login: guest] Lines: 9 Message-ID: <372tag$plj@crl8.crl.com> NNTP-Posting-Host: crl8.crl.com X-Newsreader: TIN [version 1.2 PL2] Can anyone give me a short list of PDB filenames of proteins that would cover the general range of protein structure? For example, one from each general structural class. I believe there have been some publications in the last few years. I don't have ready access to a library, but can get the PDB files. Thanks in advance. This list is needed to create a sample database of geometric parameters that can be searched for motifs using a new protein visualization and analysis package that we are developing for MAC. From owner-proteins@net.bio.net Thu Oct 06 23:00:00 1994 Path: biosci!bcm!cs.utexas.edu!uunet!utcsri!utnut!nott!nrcnet0.nrc.ca!BRI.NRC.CA!James.Fethiere From: James.Fethiere@BRI.NRC.CA (James Fethiere) Newsgroups: bionet.molbio.proteins Subject: ammonium sulfate precipitation Date: 7 Oct 1994 13:11:18 GMT Organization: Institut de recherche en biotechnologie Lines: 7 Message-ID: <373hdm$i5e@nrcnet0.nrc.ca> NNTP-Posting-Host: bobino.bri.nrc.ca X-Newsreader: TIN [version 1.2 PL2] I would like to verify something. Is it true that you need at least a 1 mg/ml concentration of your protein to ammonium sulfate precipitate it. Some textbooks say that if the concentration is lower the protein might not precipitate or even worse, it might denature. Why is that?? thank you. From owner-proteins@net.bio.net Thu Oct 06 23:00:00 1994 Path: biosci!bcm!cs.utexas.edu!howland.reston.ans.net!vixen.cso.uiuc.edu!newsrelay.iastate.edu!news.iastate.edu!yqhuang From: yqhuang@iastate.edu () Newsgroups: bionet.molbio.proteins Subject: Electroelution and deglycosylation Date: 7 Oct 1994 20:06:56 GMT Organization: Iowa State University, Ames, Iowa (USA) Lines: 34 Distribution: world Message-ID: <3749p0$8d4@news.iastate.edu> NNTP-Posting-Host: pv0a0f.vincent.iastate.edu Originator: yqhuang@pv0a0f.vincent.iastate.edu I want to deglycosylate a protein purified by immunoaffinity chrmatography. Since the monoclonal antibody recognizes three bands on the Westernern, and I am only interested in one of them, I will have to separate that band from the others after immunoaffinity column and SDS-PAGE. I tried to cut the band on the PAGE gel after aqueous Coomassie Blue Briliant R-250 staining, and electroeluted the band by using IBI electroelutor. But I recovered almost nothing. My questions are: 1. Can I transfer the bands to a nitrocellulose or PVDF membrane, stain the bands, cut the band of interest, and electroelute or extract (by other means) the protein? If yes, how? 2. Can I just do deglycosylation on Nitrocellulose or PVDF membrane and then analyse the deglycosylated core protein? any refenences? 3. If I have to electroelute the protein either from the membrane or from the gel, what are the good device you have been using? BioRad? Hoeffer? or any other? Any hints/tips/suggestions would be greatly appreciated. Yueqiao Huang ------------------------------------------------------------------- Yueqiao Huang phone 1-515-294-9783 Department of Zoology and Genetics fax 1-515-294-0345 Iowa State University email yqhuang@iastate.edu Ames,IA 50011 ------------------------------------------------------------------- From owner-proteins@net.bio.net Thu Oct 06 23:00:00 1994 Path: biosci!bcm!cs.utexas.edu!howland.reston.ans.net!news.cac.psu.edu!news.tc.cornell.edu!travelers.mail.cornell.edu!newsstand.cit.cornell.edu!cornell.edu!rsm8 From: rsm8@cornell.edu (RALPH MARCUCIO) Newsgroups: bionet.molbio.proteins Subject: RNA Polymerase II Date: Fri, 7 Oct 1994 16:01:37 Organization: Cornell University Lines: 7 Sender: rsm8@cornell.edu (Verified) Distribution: world Message-ID: NNTP-Posting-Host: 128.253.135.19 I am inerested in information regarding the protein sequence of RNA Pol II. Also, does anyone know what is responsible for translocating this protein into the nucleus, say after a mitosis? I am aware of general nuclear translocation phenomenon and am specifically interested in pol II. Any references would be greatly appreciated. Thanks, Ralph Marcucio From owner-proteins@net.bio.net Thu Oct 06 23:00:00 1994 Newsgroups: bionet.molbio.proteins Path: biosci!rutgers!gatech!howland.reston.ans.net!EU.net!Germany.EU.net!news.dfn.de!scsing.switch.ch!swidir.switch.ch!news.unige.ch!ugcmu!mayer From: mayer@cmu.unige.ch Subject: protein-protein interactions Message-ID: <1994Oct7.195822.1@ugcmu> Lines: 16 Sender: usenet@news.unige.ch Organization: University of Geneva, Switzerland Date: Fri, 7 Oct 1994 17:58:22 GMT Dear Netters A one day mini symposium called: "The yeast two-hybrid system. The art of investigating protein-protein interactions in vivo" will take place in the University Medical Centre in Geneva, Switzerland on Friday the 21st of October 1994. The symposium will start with an introduction given by Dr. Susan Gasser (ISREC, Lausanne) on handling of yeast, for all those who have not worked with yeast before. After that Paul Bartel from SUNY, Stony Brook, Roger Brent, MGH, Boston, Pierre Legrain, Institut Pasteur, Paris, and Nic Jones, Imperial Cancer Research Fund, London, will present their approach to studying protein-protein interactions and new developments in this field. If you are interested, you may register by E-mail (Mayer@cmu.unige.ch) or by fax (41-22- 3473334). There is no registration fee. From owner-proteins@net.bio.net Thu Oct 06 23:00:00 1994 Path: biosci!bcm!cs.utexas.edu!swrinde!sgiblab!wetware!kaiwan.com!NewsWatcher!user From: enof@kaiwan.com (enof) Newsgroups: bionet.molbio.proteins Subject: Re: Refolding of proteins Followup-To: bionet.molbio.proteins Date: 7 Oct 1994 23:00:36 GMT Organization: Kaiwan Lines: 49 Distribution: world Message-ID: References: <9410061001.AA11325@bicmda.bio.ic.ac.uk> NNTP-Posting-Host: kaiwan009.kaiwan.com In article <9410061001.AA11325@bicmda.bio.ic.ac.uk>, v.gutsmann@ic.ac.uk (volker gutsmann, volker gutsmann) wrote: > I am trying to find someone who could recommend a method of denaturing > peptides with urea followed by refolding by dilution into a buffer system. > > Previously I expressed several peptides as fusion proteins to MBP. However, > digestion of the fusion proteins with Factor Xa was very inefficient but > the tiny amounts of cleaved peptides were biologically active. I recloned > for expression in the pET system (pET15b) to produce unfused peptides. This > system produced large amounts of recombinant peptides (50-70% of the total > soluble protein). To my disappointment, the pET-produced peptides were only > weakly recognised by specific antisera and they had lost their biological > activity. > > I therefore would like to unfold the new peptides with 8M urea and then try > to refold them. There are quite a few methods around but we do not have any > expreience with this sort of work. If anybody could recommend a method or > has any other ideas I would be very grateful indeed. > > Many thanks > > Volker Gutsmann (vg@bio.ic.ac.uk) > Imperial College > Dept.Biology > London, UK A good general reference might be: Jaenicke, R. and Rudolph, R. (1990). Folding proteins. In: "Protein Structure - a practical approach". (T.E. Creighton, ed.) IRL Press, New York. Also, in the September issue of BioTechniques was an article from D. Sinha, M. Bakhshi and R. Vora (Department of Biochemistry, Temple University) that mentioned refolding of a 6xHis-tagged protein on Ni-NTA resin. The procedure was not mentioned. Even though every protein is going to behave differently, the following references might prove useful for renaturation in solution: D. Thanos and T. Maniatis, The High Mobility Group Protein HMG I(Y) is Required for NF-kB-dependent Virus Induction of the Human IFN-b Gene, Cell (Nov. 27, 1992) 71, 777-789 Application: Ni-NTA purification under denaturing (8 M urea) conditions followed by dialysis and renaturation. Simon Kidd, Characterization of the Drosophila Cactus Locus and Analysis of Interactions Between Cactus and Dorsal Proteins, Cell (Nov. 13, 1992) 71, 623-635 Application: Ni-NTA purification under denaturing (6 M GuHCl) conditions; refolding by sequential dialysis. -- enof@kaiwan.com | The highway's jammed with broken | Bruce enof@aol.com | heroes on a last chance power drive | Springsteen From owner-proteins@net.bio.net Fri Oct 07 23:00:00 1994 Path: biosci!agate!howland.reston.ans.net!pipex!sunic!umdac!Peter.Lundberg From: peterl@umdix.umdc.umu.se (Peter Lundberg) Newsgroups: bionet.molbio.proteins Subject: Re: Beteine and inclusion bodies Date: 8 Oct 1994 19:16:59 GMT Organization: Phys Chem, University of Umea, Sweden Lines: 9 Sender: -Not-Authenticated-[2976] Message-ID: <376r7b$308@kitten.umdc.umu.se> References: NNTP-Posting-Host: plgmac.chem.umu.se X-Posted-From: InterNews 1.0@kitten.umdc.umu.se. Xdisclaimer: No attempt was made to authenticate the sender's name. Should be 'betaine'? I'm interested too in learning more about this procedure. 73, Peter ================================== Peter Lundberg Email: peterl@umdix.umdc.umu.se ============761.91141============= From owner-proteins@net.bio.net Fri Oct 07 23:00:00 1994 Path: biosci!agate!howland.reston.ans.net!wupost!news.miami.edu!usenet.ufl.edu!gnv.ifas.ufl.edu!icbr!rep Newsgroups: bionet.molbio.proteins Subject: Info on pI? Message-ID: <1994Oct7.232223.1@icbr> From: rep@icbr Date: 7 Oct 94 23:22:23 -0500 Organization: ICBR Nntp-Posting-Host: icbr.ifas.ufl.edu Lines: 11 Hey! Can anybody tel me a good place to go searching for pI's. I'm interested in the various carbonic anhydrase isoforms, but don't feel like plowing through a ton of papers to get them. Is there a reference, or a database, or a site somewhere on the net? I appreciate any help. Chuck Peterson UF, Whitney Laboratory St. Augustine, FL From owner-proteins@net.bio.net Fri Oct 07 23:00:00 1994 Path: biosci!agate!howland.reston.ans.net!pipex!uunet!psinntp!sjuvm!zhltphs Nntp-Posting-Host: 149.68.2.20 Date: Fri, 7 Oct 1994 21:20:25 -0400 From: "Trombetta, Louis D" Newsgroups: bionet.molbio.proteins Subject: riboprobes to tubulin Message-ID: <07OCT94.23045634.0117@sjumusic.stjohns.edu> Sender: usenet@sjumusic.stjohns.edu Organization: St. John's University Lines: 13 Does anyone know where I can get a riboprobe to tubulin. Either a or b thanks Lou ******************************************* | Louis D. Trombetta, Ph.D. | | Professor | | Department of Pharmaceutical Sciences | | St. John's University | | 8000 Utopia Parkway | | Jamaica, N.Y. 11439 | | 718 990 6025 | | zhltphs@sjumusic.stjohns.edu | ******************************************* From owner-proteins@net.bio.net Fri Oct 07 23:00:00 1994 Path: biosci!galaxy.ucr.edu!ihnp4.ucsd.edu!usc!elroy.jpl.nasa.gov!netline-fddi.jpl.nasa.gov!nntp-server.caltech.edu!swong From: swong@cco.caltech.edu (Stephen Wong) Newsgroups: bionet.molbio.proteins Subject: Hen Lysozyme structure question Date: 8 Oct 1994 11:51:53 GMT Organization: California Institute of Technology, Pasadena Lines: 9 Message-ID: <37614p$ctm@gap.cco.caltech.edu> NNTP-Posting-Host: piccolo.cco.caltech.edu Keywords: Lysozyme X-Newsreader: NN version 6.5.0 #12 (NOV) Hi everyone! I have a quick question.... Can someone please tell me what the 54th amino acid residue of hen lysozyme is? It goes: N'-...-Gly-Ser-Thr-Asp-Tyr- #54 -Ile-Leu-Gln-Ile-...-C' Also, what type of secondary structures immediately following the 55th residue can be found in the enzyme? Alpha, pi, 3-10 helix, beta turn... Thanks ahead of time for your help! Please send responses by email (and please reply soon!). Thanks again. From owner-proteins@net.bio.net Sat Oct 08 23:00:00 1994 Path: biosci!agate!kos2mac14.berkeley.edu!user From: genecutl@mendel.berkeley.edu (gc) Newsgroups: bionet.molbio.proteins Subject: Re: RNA Polymerase II Date: Sun, 09 Oct 1994 13:38:19 -0800 Organization: Bilateral Symmetry Lines: 22 Distribution: world Message-ID: References: NNTP-Posting-Host: kos2mac14.berkeley.edu In article , rsm8@cornell.edu (RALPH MARCUCIO) wrote: > I am inerested in information regarding the protein sequence of RNA Pol II. > Also, does anyone know what is responsible for translocating this protein > into the nucleus, say after a mitosis? I am aware of general nuclear > translocation phenomenon and am specifically interested in pol II. Any > references would be greatly appreciated. > Thanks, > Ralph Marcucio RNA Polymerase II is a large, multi-subunit complex. The number of subunits is not completely known, mainly because of variability in the smaller polypeptides, which may or may not be actual components, break-down products, or contaminants. The larger subunits have been cloned from a variety of different organisms. I would assume that the mechanism for translocation into the nucleus for RNA Pol II components is similar to that for other nuclear proteins. --gc ...while making feet for children's shoes From owner-proteins@net.bio.net Sun Oct 09 23:00:00 1994 Path: biosci!agate!howland.reston.ans.net!EU.net!sun4nl!news.nic.surfnet.nl!highway.LeidenUniv.nl!rulsfb.LeidenUniv.nl!SBTNFH From: sbtnfh@rulsfb.LeidenUniv.nl (Flip Hoedemaeker) Newsgroups: bionet.molbio.proteins Subject: Re: Numbering AA residues Date: 10 Oct 1994 11:04:38 GMT Organization: Biology, Leiden University, The Netherlands Lines: 20 Message-ID: <37b746$edp@highway.LeidenUniv.nl> References: Reply-To: sbtnfh@rulsfb.LeidenUniv.nl NNTP-Posting-Host: rulsfb.leidenuniv.nl In article , jhonda@rkna50.riken.go.jp (Jun Honda) writes: >Dear colleagues, > >I would like to know if there is any strict rule concerning the numbering >of amino acid residues in a sequence of a given protein. It often occurs >that proteins are modified after synthesis by cleavage of peptides. >Jun Honda @ RIKEN Hi Jun, I sometimes wish there were rules about this. It is especially annoing if you download PBD files which contain AA numbers like 23A or 33C, because some of the modelling software cannot really cope with this. I would prefer it if indeed people would include residue numbers for cleaved-off peptides. This would make life easier sometimes. Perhaps we can talk to the Database people about this Flip From owner-proteins@net.bio.net Sun Oct 09 23:00:00 1994 Newsgroups: bionet.molbio.proteins Path: biosci!agate!howland.reston.ans.net!pipex!uknet!bcc.ac.uk!link-1.ts.bcc.ac.uk!ucecegv From: ucecegv@ucl.ac.uk (Edward George Varga) Subject: isoelectric points of yeast/horse liver ADH?? Sender: news@ucl.ac.uk (Usenet News System) Message-ID: <1994Oct10.114040.39818@ucl.ac.uk> Date: Mon, 10 Oct 1994 11:40:40 GMT Organization: Bloomsbury Computing Consortium Lines: 14 Can anyone out there give me the isoelectric points for the four main isozymes of yeast alcohol dehydrogenase and/or for any of the isozymes of horse liver alcohol dehydrogenase?? Any help would be greatly appreciated. Thanks in advance, Ed Edward Varga Dept. of Chemical and Biochemical Engineering University College London e-mail: ucecegv@ucl.ac.uk From owner-proteins@net.bio.net Sun Oct 09 23:00:00 1994 Newsgroups: bionet.molbio.proteins Path: biosci!agate!howland.reston.ans.net!pipex!uknet!bcc.ac.uk!bsm.bioc.ucl.ac.uk!martin From: martin@bsm.bioc.ucl.ac.uk (Andrew Martin) Subject: Re: Numbering AA residues Message-ID: <1994Oct10.132356.42373@ucl.ac.uk> Date: Mon, 10 Oct 1994 13:23:56 GMT References: <37b746$edp@highway.LeidenUniv.nl> Organization: University College London Lines: 35 In article <37b746$edp@highway.LeidenUniv.nl>, sbtnfh@rulsfb.LeidenUniv.nl (Flip Hoedemaeker) writes: |> In article , jhonda@rkna50.riken.go.jp (Jun Honda) writes: |> >Dear colleagues, |> > |> >I would like to know if there is any strict rule concerning the numbering |> >of amino acid residues in a sequence of a given protein. It often occurs |> >that proteins are modified after synthesis by cleavage of peptides. [...] |> |> I sometimes wish there were rules about this. It is especially annoing if you |> download PBD files which contain AA numbers like 23A or 33C, because some of the |> modelling software cannot really cope with this. I would prefer it if indeed |> people would include residue numbers for cleaved-off peptides. This would make |> life easier sometimes. Perhaps we can talk to the Database people about this |> |> Flip Problems with s/w unable to read simple insertion codes like 23A are a result of *bad* software which has not been written to comply with the PDB standard. In cases like the antibodies, it is so much better to have insertion letters like this so we know that residue number xxx always refers to a certain position in the conserved framework. Anyway, if you are having problems with poor s/w, it is trivial to run the PDB file through a PDB renumbering program to number the residues consecutively. Andrew -- **************************************************************************** Dr. Andrew C.R. Martin, University College London & SciTech Software INTERNET: martin@bsm.bioc.ucl.ac.uk -OR- amartin@scitec.adsp.sub.org JANET: martin@uk.ac.ucl.bioc.bsm UUCP: {uunet|rutgers}!cbmehq!cbmuk!scitec!amartin **************************************************************************** From owner-proteins@net.bio.net Sun Oct 09 23:00:00 1994 Path: biosci!agate!howland.reston.ans.net!EU.net!sun4nl!news.nic.surfnet.nl!highway.LeidenUniv.nl!rulsfb.LeidenUniv.nl!SBTNFH From: sbtnfh@rulsfb.LeidenUniv.nl (Flip Hoedemaeker) Newsgroups: bionet.molbio.proteins Subject: Re: Info on pI? Date: 10 Oct 1994 11:28:18 GMT Organization: Biology, Leiden University, The Netherlands Lines: 16 Message-ID: <37b8gi$f69@highway.LeidenUniv.nl> References: <1994Oct7.232223.1@icbr> Reply-To: sbtnfh@rulsfb.LeidenUniv.nl NNTP-Posting-Host: rulsfb.leidenuniv.nl In article <1994Oct7.232223.1@icbr>, rep@icbr writes: >Hey! >Can anybody tel me a good place to go searching for pI's.[..]Is there a reference, or a database, or a site somewhere on the net? >I appreciate any help. > Hi Chuck, The only database I know off that provides pI's is the Swissprot database. I don't know if the pI's listed there are derived from computation or from experimental data, that could make a difference. You can have a look anyway. the WWW site is : HTTP://expasy.hcuge.ch/ Hope this helps! Flip From owner-proteins@net.bio.net Sun Oct 09 23:00:00 1994 Path: biosci!bcm!cs.utexas.edu!convex!darwin.sura.net!nntp.st.usm.edu!whale.st.usm.edu!mjlogan From: mjlogan@whale.st.usm.edu (Mark Jason Logan) Newsgroups: bionet.molbio.proteins Subject: CD and AGGREGATION Date: 11 Oct 1994 02:07:15 GMT Organization: University of Southern Mississippi Lines: 11 Message-ID: <37cs0k$kk6@server.st.usm.edu> NNTP-Posting-Host: whale.st.usm.edu X-Newsreader: TIN [version 1.2 PL1] Hello, I'm looking for a reference which discusses the effecs of protein aggregation on CD spectra. Any leads would be appreciated. Please respond to e-mail address:mjlogan@whale.st.usm.edu Thank you, Mark From owner-proteins@net.bio.net Sun Oct 09 23:00:00 1994 Path: biosci!agate!library.ucla.edu!ihnp4.ucsd.edu!sdd.hp.com!saimiri.primate.wisc.edu!news.doit.wisc.edu!sweetprotein.ahabs.wisc.edu!user From: ming@ahabs.wisc.edu (Ding Ming) Newsgroups: bionet.molbio.proteins Subject: Re: isoelectric points of yeast/horse liver ADH?? Date: Mon, 10 Oct 1994 10:05:25 -0600 Organization: University of Wisconsin-Madison Lines: 14 Message-ID: References: <1994Oct10.114040.39818@ucl.ac.uk> NNTP-Posting-Host: sweetprotein.ahabs.wisc.edu In article <1994Oct10.114040.39818@ucl.ac.uk>, ucecegv@ucl.ac.uk (Edward George Varga) wrote: > Can anyone out there give me the isoelectric point for any of the isozymes of > horse liver alcohol dehydrogenase?? 8.7 +++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++ Ding Ming University of Wisconsin-Madison Telephone: (608)265-3544 Fax: (608)262-7420 ++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++ From owner-proteins@net.bio.net Mon Oct 10 23:00:00 1994 Newsgroups: bionet.molbio.proteins Path: biosci!agate!howland.reston.ans.net!EU.net!sun4nl!sci.kun.nl!ronsmul From: ronsmul@sci.kun.nl (Ronald Smulders) Subject: phosphorylation of recombinant proteins? Message-ID: Sender: news@sci.kun.nl (News owner) Nntp-Posting-Host: wn2.sci.kun.nl Organization: University of Nijmegen, The Netherlands Date: Tue, 11 Oct 1994 13:59:09 GMT Lines: 19 Hi protein chemists, I have a weird problem. We have expressed a rat protein in E. coli BL21(DE3)pLysS cells. We have purified this protein up to almost 100% using a denaturing anion exchange column. To check whether we have produced the right protein, we have determined its molecular mass using a mass spectro- meter. We were very suprised to find two main peaks in the spectrum: one of them has the right molecular mass and the other one reveals the presence of a extra phosphate group. This result suggests that our protein becomes phosphorylated in the E. coli cells. I do not understand this because I thought that posttranslational modification of recombinant proteins in procaryotic cells does not occur. Did any of you notice such a phenomenon before? Thanks in advance for your comments, Ronald (ronsmul@sci.kun.nl) PhD student/the Netherlands From owner-proteins@net.bio.net Mon Oct 10 23:00:00 1994 Path: biosci!MARTIN.LUTHER.EDU!krauske From: krauske@MARTIN.LUTHER.EDU ("Kevin Kraus") Newsgroups: bionet.molbio.proteins Subject: Merrifield resin Date: 11 Oct 1994 09:59:27 -0700 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 4 Sender: daemon@net.bio.net Distribution: world Message-ID: <199410111659.JAA22998@net.bio.net> NNTP-Posting-Host: net.bio.net Hello, I need to find a source for Merrifield resin used for protein synthesis. K.K. From owner-proteins@net.bio.net Mon Oct 10 23:00:00 1994 Path: biosci!agate!howland.reston.ans.net!EU.net!uknet!lyra.csx.cam.ac.uk!warwick!news.coventry.ac.uk!news From: GRAEME PAUL MULVANEY Newsgroups: bionet.molbio.proteins Subject: H E L P !!! me with microsomal proteins. Date: Tue, 11 Oct 1994 16:14:20 +0100 (WET DST) Organization: Coventry University Lines: 15 Message-ID: NNTP-Posting-Host: cc_sysk.coventry.ac.uk Mime-Version: 1.0 Content-Type: TEXT/PLAIN; charset=US-ASCII X-Sender: byh003@cc_sysk Yo! Dudes, I've got a problem with microsomal protein synthesis ! What is it all about, man, it kinda weighs heavy on the old grey matter ! So if any of you protein-type-dudes out there in Molecular land can help, It would be gnarley ! Keep on crusin' the Net. Captain Planet ------------------------------------------------------------------------------ From owner-proteins@net.bio.net Mon Oct 10 23:00:00 1994 Newsgroups: bionet.molbio.proteins Path: biosci!rutgers!gatech!howland.reston.ans.net!wupost!waikato!canterbury.ac.nz!otago.ac.nz!sanger.otago.ac.nz!craigm From: craigm@sanger.otago.ac.nz (Craig Marshall) Subject: Re: ammonium sulfate precipitation Message-ID: Sender: usenet@news.otago.ac.nz (News stuff) Nntp-Posting-Host: sanger.otago.ac.nz Organization: University of Otago X-Newsreader: TIN [version 1.1 PL8] References: <373hdm$i5e@nrcnet0.nrc.ca> Date: Tue, 11 Oct 1994 20:55:20 GMT Lines: 20 James Fethiere (James.Fethiere@BRI.NRC.CA) wrote: > I would like to verify something. Is it true that you need at least > a 1 mg/ml concentration of your protein to ammonium sulfate > precipitate it. Some textbooks say that if the concentration is lower > the protein might not precipitate or even worse, it might denature. > Why is that?? > thank you. Handwaving follows: Precipitation requires that there be enough protein molecules present in solution to form aggregates. If there are not, precipitation may not occur until higher salt concentrations; these concentrations may be above that required to denature the protein. -- Craig Marshall craigm@sanger.otago.ac.nz Biochemistry Department Phone +64 3 479 7570 University of Otago Fax +64 3 479 7866 From owner-proteins@net.bio.net Mon Oct 10 23:00:00 1994 Newsgroups: bionet.molbio.proteins Path: biosci!galaxy.ucr.edu!ihnp4.ucsd.edu!sdd.hp.com!heifetz.msen.com!emory!europa.eng.gtefsd.com!library.ucla.edu!csulb.edu!nic-nac.CSU.net!charnel.ecst.csuchico.edu!csusac!csus.edu!netcom.com!netcomsv!chiron.com!chiremv!hof From: hof@chiremv.chiron.com (Frank Ho) Subject: Protease to cleave Ala-Gly bond? Message-ID: Sender: news@chiron.com Nntp-Posting-Host: chiremv Organization: Chiron Corporation Date: Tue, 11 Oct 1994 23:17:59 GMT Lines: 5 Hi: I need to know if there is a protease specifically cleave between Ala-Gly peptide bond, and what inhibitor can be used to stop this protease? Thank you for your help. From owner-proteins@net.bio.net Tue Oct 11 23:00:00 1994 Newsgroups: bionet.molbio.proteins Path: biosci!agate!howland.reston.ans.net!gatech!newsxfer.itd.umich.edu!nntp.cs.ubc.ca!news.UVic.CA!spruce.pfc.forestry.ca!PFC.Forestry.CA!AEKRAMODDOUL From: aekramoddoul@PFC.Forestry.CA (Ekramoddoullah, Abul Kalam M.) Subject: Re: Peptide Antibodies Message-ID: <1994Oct12.212953.6535@spruce.pfc.forestry.ca> Sender: news@spruce.pfc.forestry.ca Nntp-Posting-Host: pfc.pfc.forestry.ca Reply-To: aekramoddoul@PFC.Forestry.CA Organization: Forestry Canada (Pacific Forestry Centre) References: <01HHYQT9BAHEBL2YX5@PORTIA.CALTECH.EDU> Date: Wed, 12 Oct 1994 21:29:53 GMT Lines: 32 In article <01HHYQT9BAHEBL2YX5@PORTIA.CALTECH.EDU>, EARN@PORTIA.CALTECH.EDU writes: > Does anyone have experience with the logistics of having peptide >antibodies made? I'd like to do what's fastest and/or cheapest but I >don't know if it's best to have peptides synthesized at an in-house core >facility (and whether to couple them yourself) and send them off, or >are there companies you can just send sequence to and they'll take care >of it all. Also I'd like recommendations for various rabbit-poking >companies as to the quality of their polyclonals, price, etc. If >you email me privately I will post a digest to this list. Thanks in advance >for your help. > - Eric Arn > Division of Biology > Caltech > EARN@Portia.caltech.edu Hi Eric; Try CHIRON MIMOTOPES PEPTIDE SYSTEMS 3550 General Atomic Court San Diego, CA 92121-1122 Tel: 800-455-4965; Fax 800-654-5592 I have used their service and was very satisfied. They use multiple peptide synthesis. The cost will be cheaper than you make in-house and plus they also provide gold antibody project whereby they will conjugate the peptide to a carrier molecule and provide affinity purified antibody along with crude antisera and the affinity column (for you to purify more if needed). I hope this was a help. I hope I was not promoting the company, but if you refer my name, I will ask the for a disount on my next order. Good Luck. Thanks Abul EKRAMODDOULLAH, ABUL KALAM M. Title: Research Scientist Forestry Canada Phone: (604) 363-0600 Victoria, B.C. Internet: AEKRAMODDOUL@A1.PFC.Forestry.CA From owner-proteins@net.bio.net Tue Oct 11 23:00:00 1994 Path: biosci!bcm!cs.utexas.edu!howland.reston.ans.net!EU.net!sunic!columba.udac.uu.se!onyx.bmc.uu.se!dunten From: dunten@onyx.bmc.uu.se (Pete Dunten) Newsgroups: bionet.molbio.proteins Subject: Re: Protein concentration from peptide bond absorbance Date: 12 Oct 1994 17:15:50 GMT Organization: Uppsala University Lines: 4 Distribution: world Message-ID: <37h5k6$bkv@columba.udac.uu.se> References: NNTP-Posting-Host: onyx.bmc.uu.se Scopes, R.K. (1974) 'Measurement of protein by spectrophotometry at 205 nm.' Anal. Biochem. 59, 277-282. From owner-proteins@net.bio.net Tue Oct 11 23:00:00 1994 Newsgroups: bionet.molbio.proteins From: steve@gardner.demon.co.uk (Steve & Rowan Gardner) Path: biosci!agate!howland.reston.ans.net!pipex!demon!gardner.demon.co.uk!steve Subject: Re: Numbering AA residues References: <1994Oct10.132356.42373@ucl.ac.uk> <37b746$edp@highway.LeidenUniv.nl> Distribution: world Organization: Personal Reply-To: steve@gardner.demon.co.uk X-Newsreader: Newswin Alpha 0.6 Lines: 29 Date: Wed, 12 Oct 1994 08:13:05 +0000 Message-ID: <242162866wnr@gardner.demon.co.uk> Sender: usenet@demon.co.uk In article: <1994Oct10.132356.42373@ucl.ac.uk> martin@bsm.bioc.ucl.ac.uk (Andrew Martin) writes: > > In article <37b746$edp@highway.LeidenUniv.nl>, sbtnfh@rulsfb.LeidenUniv.nl (Flip Hoedemaeker) writes: > |> > |> I sometimes wish there were rules about this. It is especially annoing if you > |> download PBD files which contain AA numbers like 23A or 33C, because some of the > |> modelling software cannot really cope with this. I would prefer it if indeed > |> people would include residue numbers for cleaved-off peptides. This would make > |> life easier sometimes. Perhaps we can talk to the Database people about this > |> > |> Flip > Problems with s/w unable to read simple insertion codes like 23A are a result > of *bad* software which has not been written to comply with the PDB standard. > In cases like the antibodies, it is so much better to have insertion letters > like this so we know that residue number xxx always refers to a certain > position in the conserved framework. > Exactly - well said Andrew. For better or worse the format is as it is, and s/w authors ought to take a good look at the format before they set fingers to keyboard. ---------------------------------------------------------------- Steve Gardner steve@gardner.demon.co.uk From owner-proteins@net.bio.net Tue Oct 11 23:00:00 1994 Path: biosci!rutgers!netnews.upenn.edu!elmo.tju.edu!calvin.jci.tju.edu!harrison From: harrison@calvin.jci.tju.edu Newsgroups: bionet.molbio.proteins Subject: Re: Numbering AA residues Date: 12 Oct 1994 18:58:26 GMT Organization: Jefferson Cancer Institute Lines: 26 Distribution: world Message-ID: <37hbki$f4g@elmo.tju.edu> References: <1994Oct10.132356.42373@ucl.ac.uk> <37b746$edp@highway.LeidenUniv.nl> <242162866wnr@gardner.demon.co.uk> NNTP-Posting-Host: calvin.jci.tju.edu In article <242162866wnr@gardner.demon.co.uk>, steve@gardner.demon.co.uk (Steve & Rowan Gardner) writes: |> In article: <1994Oct10.132356.42373@ucl.ac.uk> martin@bsm.bioc.ucl.ac.uk (Andrew Martin) |> writes: stuff (lots) deleted, paraphrases to a legitimate gripe about odd ball numbering schemes. |> If you think this is bad wait for the CIF format 8-). Then no-one's software will always work. This situation is the result of an ancient split between different groups of crystallographers. At least in column oreinted PDB format it is reasonably easy to renumber as needed. -- robert w. harrison harrison@asterix.jci.tju.edu ******************************************************************************** Organic chemistry is the chemistry of carbon compounds. Biochemistry is the study of carbon compounds that crawl. -- Mike Adams ******************************************************************************** From owner-proteins@net.bio.net Wed Oct 12 23:00:00 1994 Path: biosci!agate!spool.mu.edu!sgiblab!darwin.sura.net!mother.usf.edu!com1!pneame From: pneame@com1.med.usf.edu (Peter J. Neame) Newsgroups: bionet.molbio.proteins Subject: Re: preparing a protein for sequencing Date: 13 Oct 1994 15:01:35 GMT Organization: University of South Florida Lines: 19 Distribution: world Message-ID: <37ji4f$fse@mother.usf.edu> References: <9410051957.AA28182@molbio.cbs.umn.edu> NNTP-Posting-Host: com1.med.usf.edu X-Newsreader: TIN [version 1.2 PL2] Don't blot your protein onto nitrocellulose for direct Edman degradation! It will gunk up the sequencer horribly, as the membrane dissolves in the solvents. However, you can blot your protein onto nitrocellulose or PVDF (immobilon etc) and digest it in situ to get internal sequence. See Fenandez et al, Anal Biochem, 201 (1992) 255-264 for an excellent method for this. There some more recent papers from this group and others, also. We use the Fernandez method quite a lot and are pretty happy with it. Good luck! Peter Peter Neame SHCC and USF, Tampa FL pneame@com1.med.usf.edu From owner-proteins@net.bio.net Wed Oct 12 23:00:00 1994 Path: biosci!agate!doc.ic.ac.uk!bay.cc.kcl.ac.uk!bay.cc.kcl.ac.uk!udbl119 Newsgroups: bionet.molbio.methds-reagnts,bionet.molbio.proteins,bionet.cellbiol Subject: Olomoucine, a cdc2 Protein Kinase inhibitor? Message-ID: From: udbl119@bay.cc.kcl.ac.uk (Mandy Johnstone) Date: Thu, 13 Oct 1994 12:19:36 Organization: King's College London Nntp-Posting-Host: pgw2.rai.kcl.ac.uk X-Newsreader: Trumpet for Windows [Version 1.0 Rev A] Lines: 14 Xref: biosci bionet.molbio.methds-reagnts:19556 bionet.molbio.proteins:2866 bionet.cellbiol:1032 Hello, I am interested in finding out about a cdc2 protein kinase inhibitor that Promega now sell which is called Olomoucine. In particular how it works and has anyone out there used it or can comment on how good it performed in their hands. Many thanks in advance. Mandy. _______________ Mandy Johnstone (M.Johnstone@bay.cc.kcl.ac.uk) King's College, London. From owner-proteins@net.bio.net Wed Oct 12 23:00:00 1994 Path: biosci!pop.nih.gov!fnunes From: fnunes@pop.nih.gov (Fabio Nunes) Newsgroups: bionet.molbio.proteins Subject: (none) Date: 13 Oct 1994 13:24:50 -0700 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 8 Sender: daemon@net.bio.net Distribution: world Message-ID: <9410132024.AA17224@dxi.nih.gov> NNTP-Posting-Host: net.bio.net Does anybody know how I can identify sugar moiety in SDS-PAGE? Any help will be greatly appreciated Fabio Nunes NIDCD-NIH fnunes@pop.nih.gov From owner-proteins@net.bio.net Wed Oct 12 23:00:00 1994 Path: biosci!bcm!cs.utexas.edu!not-for-mail From: david_small@muwayf.unimelb.edu.au (David Small) Newsgroups: bionet.jobs,bionet.molbio.ageing,bionet.molbio.proteins,bionet.neuroscience Subject: Positions available Date: 13 Oct 1994 19:26:31 -0500 Organization: UTexas Mail-to-News Gateway Lines: 22 Sender: nobody@cs.utexas.edu Message-ID: <01HI9HKRX1ZM001VAZ@muwayb.ucs.unimelb.edu.au> NNTP-Posting-Host: news.cs.utexas.edu Xref: biosci bionet.jobs:5972 bionet.molbio.ageing:943 bionet.molbio.proteins:2871 bionet.neuroscience:4596 Positions available#000# ALZHEIMER'S DISEASE RESEARCH GROUP Several positions at the post-doctoral, research assistant and graduate student (PhD scholarship) level are now available. The research and therapeutic target is the amyloid precursor protein, a cell-surface receptor. Persons with an interest or experience in the wider areas of cell biology and molecular biochemistry, or in the specialized areas of protein structure-function analysis, proteases, immunoassay development, and neuronal tissue culture are invited to apply. Conditions of employment will be very competitive. Further details from Professor Colin L. Masters Tel. 61-3-344-5968, FAX 61-3-344-4004. E-mail: pathology.pathology@muwayf.unimelb.edu.au The first round of applications will close on 7th November, and should be sent to Helene Marszalek, Pathology Department, The University of Melbourne, Parkville, Victoria 3052, Australia. From owner-proteins@net.bio.net Wed Oct 12 23:00:00 1994 Path: biosci!bcm!cs.utexas.edu!howland.reston.ans.net!news.cac.psu.edu!news.pop.psu.edu!psuvax1!news.ecn.bgu.edu!willis.cis.uab.edu!cs.utk.edu!stc06.CTD.ORNL.GOV!biop18.bio.ornl.gov!user From: gewiessa@bioax1.bio.ornl.gov (Andreas Gewiess) Newsgroups: bionet.molbio.proteins Subject: Help! My protein aggregates Date: Thu, 13 Oct 1994 19:34:33 -0500 Organization: U. of Tenn.,Knoxville/ Oak Ridge Natl. Lab. Lines: 13 Message-ID: NNTP-Posting-Host: biop18.bio.ornl.gov Hello Protein-Folks, does anybody out there got some experience with concentrated protein solutions ( >25 mg/ml) ? Our protein tends to dimerize and we are looking desperately for ways to prevent this. If somebody could point us to related literature or can tell us the special secrets in that matter, it would be highly appreciated. Andreas -- * Andreas Gewiess, Dr.rer.nat / Biology Division * * Oak Ridge National Laboratory, Oak Ridge, Tenn.* From owner-proteins@net.bio.net Wed Oct 12 23:00:00 1994 Path: biosci!BIOBASE.AAU.DK!thorup From: thorup@BIOBASE.AAU.DK (Jan Ulrik Thorup) Newsgroups: bionet.molbio.proteins Subject: HELP! Hydropathy Dot Plot Date: 13 Oct 1994 09:32:16 -0700 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 23 Sender: daemon@net.bio.net Distribution: world Message-ID: NNTP-Posting-Host: net.bio.net Dear protein-netters! I need to make a dot matrix comparison, where I compare the hydropathy of two proteins to see whether they might be structurally related. * Do any of you know of a DOS/Windows program that allows protein-sequences to be the input and the output to be a comparison of their hydropathy, aminoacid by aminoacid, illustrated in a dot-plot ? * Please let me know, and where/how to get such a program. Best Regards Ulrik Thorup thorup@biobase.aau.dk Dept. Clin. Biochem. Rigshospitalet Denmark From owner-proteins@net.bio.net Thu Oct 13 23:00:00 1994 Newsgroups: bionet.molbio.proteins Path: biosci!bcm!cs.utexas.edu!howland.reston.ans.net!pipex!uknet!festival!leeds.ac.uk!news From: gends@leeds.ac.uk (SCOTT D.) Subject: lysozyme unfolding Message-ID: Sender: news@leeds.ac.uk Organization: University of Leeds Date: Fri, 14 Oct 1994 16:17:43 +0100 (BST) X-Newsreader: Trumpet for Windows [Version 1.0 Rev A] Lines: 16 hia, As a test reaction for our nice new stopped-flow fluorimeter I wish watch the unfolding of lysozyme in guanidinium. Does anyone out there have the rate constants and conditions for this process???? If so could you tell me please! Thanks in advance... Dave Scott. Dept. of Genetics, University of Leeds. Leeds. W.Yorks. U.K. Email: gends@gps.leeds.ac.uk From owner-proteins@net.bio.net Thu Oct 13 23:00:00 1994 Path: biosci!bcm!cs.utexas.edu!howland.reston.ans.net!europa.eng.gtefsd.com!library.ucla.edu!news.mic.ucla.edu!unixg.ubc.ca!koska From: koska@unixg.ubc.ca (Jurgen Koska) Newsgroups: bionet.molbio.proteins Subject: Sigmas L-Phenylalanin Date: 14 Oct 1994 16:28:16 GMT Organization: University of British Columbia, Vancouver, B.C., Canada Lines: 13 Message-ID: <37mbj0$8gp@nnrp.ucs.ubc.ca> NNTP-Posting-Host: interchg.ubc.ca X-Newsreader: TIN [version 1.2 PL2] Not too long ago we bought L-Phenyalanin from Sigma and found that the HPLC resolved two (equal area) peaks - according to Sigma this product problem was known - but we weren't contacted. However, they gave as a new batch - this time still 2 peaks (the impurity is now less), but after preparing a solution and micro filter it the solution became cloudy!!! What did they sell us??? Did somebody had similar experiences with L-Phe!! Ju"rgen Email: koska@unixg.ubc.ca From owner-proteins@net.bio.net Fri Oct 14 23:00:00 1994 Path: biosci!rutgers!gatech!howland.reston.ans.net!sol.ctr.columbia.edu!usenet.ucs.indiana.edu!indyvax.iupui.edu!et1021-mac03.oitlc.iupui.edu!user Newsgroups: bionet.molbio.proteins Subject: seeking senior proj ideas Message-ID: From: FAGROTT@INDYVAX.IUPUI.EDU (Fred Grott) Date: 14 Oct 94 01:46:46 -0500 Distribution: world Nntp-Posting-Host: et1021-mac03.oitlc.iupui.edu Lines: 6 I am seeking ideas of research projects that I might be able to do conerning protein folding in order to satisfy my senior research credit for my BS in biology. Any ideas would be helpful. Fred Grott FAGROTT@INDYVAX.iupui.edu From owner-proteins@net.bio.net Fri Oct 14 23:00:00 1994 Newsgroups: bionet.molbio.proteins Path: biosci!daresbury!trane.uninett.no!sunic!pipex!uunet!EU.net!sun4nl!news.nic.surfnet.nl!rug.nl!root From: G.J.W.Euverink@biol.rug.nl Subject: Re: Sigmas L-Phenylalanin Message-ID: Keywords: l-phe Sender: root@rug.nl (Operator) Nntp-Posting-Host: micfys04.biol.rug.nl Organization: biol.rug.nl X-Newsreader: Date: Sat, 15 Oct 1994 11:44:23 GMT Lines: 22 In article <37mbj0$8gp@nnrp.ucs.ubc.ca> writes: >own - but we weren't contacted. However, they gave as a >new batch - this time still 2 peaks (the impurity is now less), but after >preparing a solution and micro filter it the solution became cloudy!!! >What did they sell us??? > >Did somebody had similar experiences with L-Phe!! > > > Ju"rgen > > Email: koska@unixg.ubc.ca How did you measure L-phe (OPA derivatization or UV absorbance? ) When I open the bottle I smell a little bit of phenylacetaldehyde or a related compound. Maybe this has something to do with the production method. Phenylacetaldehyde can be an intermediate in phenylalanine catabolism. From owner-proteins@net.bio.net Fri Oct 14 23:00:00 1994 Path: biosci!bcm!cs.utexas.edu!usc!sol.ctr.columbia.edu!newsxfer.itd.umich.edu!zip.eecs.umich.edu!yeshua.marcam.com!charnel.ecst.csuchico.edu!csusac!csus.edu!news.ucdavis.edu!bullwinkle!ez041797 From: ez041797@bullwinkle.ucdavis.edu (Jean-Manuel Henry) Newsgroups: bionet.molbio.proteins Subject: Storing PK\LDH without any sulfate ? Date: 16 Oct 1994 00:53:31 GMT Organization: University of California, Davis Lines: 7 Message-ID: <37ptib$5oh@mark.ucdavis.edu> NNTP-Posting-Host: bullwinkle.ucdavis.edu X-Newsreader: TIN [version 1.2 PL2] Help ! I am looking for a method of storing a solution of Pk\LDH which does not include any sulfate salts (for assays of ATP sulfurylase activity). If anyone has any good ideas please share them with me, thanks. From owner-proteins@net.bio.net Sat Oct 15 23:00:00 1994 Path: biosci!agate!library.ucla.edu!europa.eng.gtefsd.com!gatech!howland.reston.ans.net!pipex!sunic!news.funet.fi!news.csc.fi!news.helsinki.fi!not-for-mail From: haltia@cc.Helsinki.FI (Tuomas Haltia) Newsgroups: bionet.molbio.methds-reagnts,bionet.molbio.proteins Subject: Topology of a multisubunit membrane protein Date: 17 Oct 1994 05:02:10 +0200 Organization: University of Helsinki Lines: 17 Message-ID: <37spfi$44o@plootu.Helsinki.FI> NNTP-Posting-Host: plootu.helsinki.fi Mime-Version: 1.0 Content-Type: text/plain; charset=ISO-8859-1 Content-Transfer-Encoding: 8bit Summary: What would be a good hydrophilic labeling reagent? Keywords: Chemical labeling, lactoperoxidase iodination, membrane protein structure Xref: biosci bionet.molbio.methds-reagnts:19675 bionet.molbio.proteins:2878 I am studying a membrane protein complex in which a significant part of the protein is thought to be on the outside of the membrane. However, it is not clear which of the three subunits form the peripheral part. In particular, the location of one, easily removable subunit is unclear. It occured to me that this problem could be studied using a soluble labeling reagent: the labeling pattern of the subunits should correlate with their membrane topography. Has anyone used lactoperoxidase catalysed radioiodination to study a similar problem? Are there any other potentially useful methods? Are there any methods which would not involve the use of highly radioactive isotopes? Tuomas Haltia Dept. Medical Chemistry, U. of Helsinki, Finland From owner-proteins@net.bio.net Sun Oct 16 23:00:00 1994 Path: biosci!agate!howland.reston.ans.net!usc!usc!not-for-mail From: merlin@neuro.usc.edu (merlin) Newsgroups: bionet.molbio.proteins Subject: X-ray Crystallography, High Res 3D NMR & Computational Structures Date: 17 Oct 1994 01:06:19 -0700 Organization: University of Southern California, Los Angeles, CA Lines: 20 Sender: merlin@neuro.usc.edu Message-ID: <37tb9s$4tj@neuro.usc.edu> References: <01HI9HKRX1ZM001VAZ@muwayb.ucs.unimelb.edu.au> NNTP-Posting-Host: neuro.usc.edu o Does any mailing list or newsgroup exist for X-ray Crystallographers, High Resolution 3D Nuclear Magnetic Resonance, and/or Computational Energy Minimization Static/Dynamic Structural Modeling Scientists to exchange information via the national network infrastructure? o Where does one obtain leading edge training in use of these techniques? ------------------------------------------------------------------------------ Alexander-James Annala Principal Investigator Neuroscience Image Analysis Network HEDCO Neuroscience Building, Fifth Floor University of Southern California University Park Los Angeles, CA 90089-2520 Telephone: (213)740-3406 FAX: (213)740-5687 ------------------------------------------------------------------------------ From owner-proteins@net.bio.net Sun Oct 16 23:00:00 1994 Path: biosci!agate!howland.reston.ans.net!EU.net!news.kreonet.re.kr!worak.kaist.ac.kr!chiak.kaist.ac.kr!jylee From: jylee@chiak.kaist.ac.kr (Lee jang young) Newsgroups: bionet.molbio.proteins Subject: problem with pUC18..help me !! Date: 17 Oct 1994 09:28:47 GMT Organization: Korea Advanced Institute of Science and Technology Lines: 28 Message-ID: <37tg4f$16f@worak.kaist.ac.kr> NNTP-Posting-Host: chiak.kaist.ac.kr X-Newsreader: TIN [version 1.1 PL9] Hi netters.. I'm now cultivating a recombinant E.coli containing pUC18 on which an enzyme of interest is cloned. The expression of the enzyme is not under the control of the lac promoter of pUC18, but is regluated by its own promoter which originates from Bacillus sp. The activity of the enzyme was increased by about 20 fold when cloned in E.coli than that in the original Bacillus sp. But, the cultivation of the recombinant E.coli in LB media gave irregular results from case to case..say, in one case the activity was maintained as expected, but in other case, almost no activity was found... I'm very confused about this results. Anyone who are expert in this field, please let me know how to solve the problem like this,, Private e-mail at following address would be especially appreciated.. Thanks D.C.Lee from KOREA jylee@chiak.kaist.ac.kr From owner-proteins@net.bio.net Sun Oct 16 23:00:00 1994 Path: biosci!agate!howland.reston.ans.net!EU.net!sun4nl!news.nic.surfnet.nl!highway.LeidenUniv.nl!rulsfb.LeidenUniv.nl!SBTNFH From: sbtnfh@rulsfb.LeidenUniv.nl (Flip Hoedemaeker) Newsgroups: bionet.molbio.methds-reagnts,bionet.molbio.proteins Subject: Re: Topology of a multisubunit membrane protein Date: 17 Oct 1994 12:43:10 GMT Organization: Biology, Leiden University, The Netherlands Lines: 15 Message-ID: <37trgu$5k5@highway.LeidenUniv.nl> References: <37spfi$44o@plootu.Helsinki.FI> Reply-To: sbtnfh@rulsfb.LeidenUniv.nl NNTP-Posting-Host: rulsfb.leidenuniv.nl Xref: biosci bionet.molbio.methds-reagnts:19693 bionet.molbio.proteins:2882 In article <37spfi$44o@plootu.Helsinki.FI>, haltia@cc.Helsinki.FI (Tuomas Haltia) writes: >I am studying a membrane protein complex in which a significant >part of the protein is thought to be on the outside of the membrane. >However, it is not clear which of the three subunits form the peripheral >part. In particular, the location of one, easily removable subunit >is unclear. >Tuomas Haltia You say that the subunit you are interested in is easily removable. Can't you purify it, make (FITC-labeled) antibodies, and use them for localisation? At least this method is non-radioactive, and it should work relatively fast! Hope this helps! Flip From owner-proteins@net.bio.net Sun Oct 16 23:00:00 1994 Path: biosci!agate!usenet.hana.nm.kr!xpat.postech.ac.kr!news.kreonet.re.kr!worak.kaist.ac.kr!chiak.kaist.ac.kr!jylee From: jylee@chiak.kaist.ac.kr (Lee jang young) Newsgroups: bionet.molbio.proteins Subject: Overexpression with pKK223-3 !! help Date: 17 Oct 1994 21:31:20 GMT Organization: Korea Advanced Institute of Science and Technology Lines: 25 Message-ID: <37uqf8$cgv@worak.kaist.ac.kr> NNTP-Posting-Host: chiak.kaist.ac.kr X-Newsreader: TIN [version 1.1 PL9] hi..biologists out there.. Has anyone overexpressed a protein cloned in the vector of pKK223-3 ? I recently completed cloning of a kind of dehydrogenase by PCR technique and plan to overexpress it by introducing this gene to a expression vector. At present I chose pKK223-3 for this purpose. When I made it, how much protein of my interest do I get ? i mean can i get this dehydrogenase at the amount of more than 20 % of the total protein with this system ? Is there any problem in using this expression system ? Any advice would be a big help for me... private e-mail at following address would be especially appreciated jylee@chiak.kaist.ac.kr J.Y.Lee from KOREA From owner-proteins@net.bio.net Sun Oct 16 23:00:00 1994 Newsgroups: bionet.molbio.proteins Path: biosci!daresbury!bioftp.unibas.ch!doelz From: doelz@comp.bioz.unibas.ch (Reinhard Doelz) Subject: Re: X-ray Crystallography, High Res 3D NMR & Computational Structures Message-ID: <1994Oct17.200512.8440@comp.bioz.unibas.ch> Organization: EMBnet Switzerland [Basel] X-Newsreader: TIN [version 1.2 PL2] References: <01HI9HKRX1ZM001VAZ@muwayb.ucs.unimelb.edu.au>,<37tb9s$4tj@neuro.usc.edu> <37trnk$5k5@highway.LeidenUniv.nl> <37u5nc$7rq@news.ycc.yale.edu> Date: Mon, 17 Oct 1994 20:05:12 GMT Lines: 24 Kevin Gardner (gardner@cadmium.csb.yale.edu) wrote: : Flip Hoedemaeker (sbtnfh@rulsfb.LeidenUniv.nl) wrote: : : In article <37tb9s$4tj@neuro.usc.edu>, merlin@neuro.usc.edu (merlin) writes: : : >o Does any mailing list or newsgroup exist for X-ray Crystallographers, : : > High Resolution 3D Nuclear Magnetic Resonance, and/or Computational : : > Energy Minimization Static/Dynamic Structural Modeling Scientists to : : > exchange information via the national network infrastructure? : : Yes, bionet.xtallography! : Let's not forget bionet.structural-nmr and bionet.molec-model while : we're at it.... There are also special mailing list exploders for specific vendors/packages in the field, e.g. public domain (like ccp4) or commercial. Regards Reinhard -- R.Doelz Klingelbergstr.70| Tel. x41 61 267 2247 Fax x41 61 267 2078| Biocomputing CH 4056 Basel| electronic Mail doelz@ubaclu.unibas.ch| Biozentrum der Universitaet Basel|-------------- Switzerland ---------------| EMBnet Switzerland:info@ch.embnet.org From owner-proteins@net.bio.net Sun Oct 16 23:00:00 1994 Path: biosci!daresbury!hgmp.mrc.ac.uk!ebi.ac.uk!embl-heidelberg.de!hobohm From: hobohm@embl-heidelberg.de (Uwe Hobohm) Newsgroups: bionet.molbio.proteins Subject: Re: Minimal Set of Proteins Message-ID: <1994Oct14.153035.171572@eros.embl-heidelberg.de> Date: 14 Oct 94 15:30:35 +0100 References: <372tag$plj@crl8.crl.com> Organization: European Molecular Biology Laboratory Lines: 19 In article <372tag$plj@crl8.crl.com>, sunger@crl.com (Stefan Unger) writes: > Can anyone give me a short list of PDB filenames of proteins that would > cover the general range of protein structure? For example, one from each > general structural class. I believe there have > been some publications in the last few years. I don't have ready access > to a library, but can get the PDB files. Thanks in advance. > > This list is needed to create a sample database of geometric parameters > that can be searched for motifs using a new protein visualization and > analysis package that we are developing for MAC. We have generated a list of representative proteins from the PDB (see Protein Science 1(1992)409 and Protein Science 3(1994)522). The list is available on anonymous ftp from the embl ftp-server (192.54.41.33) under /pub/databases/protein_extras/pdb_select/pdb_select.oct_1994. The list will be updated regularly. I guess, all main protein structures are covered in the list. Uwe Hobohm EMBL-Heidelberg From owner-proteins@net.bio.net Sun Oct 16 23:00:00 1994 Path: biosci!bcm!cs.utexas.edu!howland.reston.ans.net!EU.net!Germany.EU.net!netmbx.de!zib-berlin.de!tertius.in-berlin.de!mpimg-berlin-dahlem.mpg.de!wegloehner Newsgroups: bionet.molbio.methds-reagnts,bionet.molbio.proteins Subject: Re: Topology of a multisubunit membrane protein Message-ID: <1994Oct17.200303.1@mpimg-berlin-dahlem.mpg.de> From: wegloehner@mpimg-berlin-dahlem.mpg.de Date: 17 Oct 94 20:03:03 +0100 References: <37spfi$44o@plootu.Helsinki.FI> Organization: MPI f. Molekulare Genetik, Berlin Nntp-Posting-Host: watson.rz-berlin.mpg.de Nntp-Posting-User: nntp_client Lines: 32 Xref: biosci bionet.molbio.methds-reagnts:19719 bionet.molbio.proteins:2885 In article <37spfi$44o@plootu.Helsinki.FI>, haltia@cc.Helsinki.FI (Tuomas Haltia) writes: > I am studying a membrane protein complex in which a significant > part of the protein is thought to be on the outside of the membrane. > However, it is not clear which of the three subunits form the peripheral > part. In particular, the location of one, easily removable subunit > is unclear. > > It occured to me that this problem could be studied using a > soluble labeling reagent: the labeling pattern of the subunits > should correlate with their membrane topography. > > Has anyone used lactoperoxidase catalysed radioiodination to study > a similar problem? Are there any other potentially useful methods? > Are there any methods which would not involve the use of highly > radioactive isotopes? A friend of mine has worked on a similar problem with the nicotinic acetylcholine receptor. lactoperoxidase iodination has not worked at all. he suggests to try Iodo-Gen (from Pierce), a unsoluble iodination reagent. If you would like to get further information, please feel free to contact him directly. His E-mail adress: mm@chemie.fu-berlin.de > > Tuomas Haltia > Dept. Medical Chemistry, U. of Helsinki, Finland Dr. Wolfgang Wegloehner Max-Planck-Institute for Molecular Genetics Ihnestrasse 73 14195 Berlin Germany From owner-proteins@net.bio.net Sun Oct 16 23:00:00 1994 Path: biosci!agate!library.ucla.edu!europa.eng.gtefsd.com!howland.reston.ans.net!pipex!uunet!noc.near.net!saturn.caps.maine.edu!news.ycc.yale.edu!cadmium.csb.yale.edu!gardner From: gardner@cadmium.csb.yale.edu (Kevin Gardner) Newsgroups: bionet.molbio.proteins Subject: Re: X-ray Crystallography, High Res 3D NMR & Computational Structures Date: 17 Oct 1994 15:37:16 GMT Organization: Yale University Lines: 25 Message-ID: <37u5nc$7rq@news.ycc.yale.edu> References: <01HI9HKRX1ZM001VAZ@muwayb.ucs.unimelb.edu.au>,<37tb9s$4tj@neuro.usc.edu> <37trnk$5k5@highway.LeidenUniv.nl> NNTP-Posting-Host: cadmium.csb.yale.edu X-Newsreader: TIN [version 1.2 PL2] Flip Hoedemaeker (sbtnfh@rulsfb.LeidenUniv.nl) wrote: : In article <37tb9s$4tj@neuro.usc.edu>, merlin@neuro.usc.edu (merlin) writes: : >o Does any mailing list or newsgroup exist for X-ray Crystallographers, : > High Resolution 3D Nuclear Magnetic Resonance, and/or Computational : > Energy Minimization Static/Dynamic Structural Modeling Scientists to : > exchange information via the national network infrastructure? : > : Yes, bionet.xtallography! : Flip Let's not forget bionet.structural-nmr and bionet.molec-model while we're at it.... Kevin -- ************************************************************************* Kevin Gardner Yale University Dept. of Molecular Biophysics and Biochemistry Internet: gardner@zinc.csb.yale.edu Bitnet: gardner@yalemed ***I support the boycott of companies using mass Internet advertising*** From owner-proteins@net.bio.net Sun Oct 16 23:00:00 1994 Path: biosci!agate!howland.reston.ans.net!EU.net!sun4nl!news.nic.surfnet.nl!highway.LeidenUniv.nl!rulsfb.LeidenUniv.nl!SBTNFH From: sbtnfh@rulsfb.LeidenUniv.nl (Flip Hoedemaeker) Newsgroups: bionet.molbio.proteins Subject: Re: X-ray Crystallography, High Res 3D NMR & Computational Structures Date: 17 Oct 1994 12:46:44 GMT Organization: Biology, Leiden University, The Netherlands Lines: 10 Message-ID: <37trnk$5k5@highway.LeidenUniv.nl> References: <01HI9HKRX1ZM001VAZ@muwayb.ucs.unimelb.edu.au>,<37tb9s$4tj@neuro.usc.edu> Reply-To: sbtnfh@rulsfb.LeidenUniv.nl NNTP-Posting-Host: rulsfb.leidenuniv.nl In article <37tb9s$4tj@neuro.usc.edu>, merlin@neuro.usc.edu (merlin) writes: >o Does any mailing list or newsgroup exist for X-ray Crystallographers, > High Resolution 3D Nuclear Magnetic Resonance, and/or Computational > Energy Minimization Static/Dynamic Structural Modeling Scientists to > exchange information via the national network infrastructure? > Yes, bionet.xtallography! Flip From owner-proteins@net.bio.net Sun Oct 16 23:00:00 1994 Path: biosci!agate!kos2mac3.berkeley.edu!user From: genecutl@mendel.berkeley.edu (gc) Newsgroups: bionet.molbio.proteins,bionet.xtallography Subject: Dimerization interfaces Date: 17 Oct 1994 19:59:07 GMT Organization: Bilateral Symmetry Lines: 84 Message-ID: NNTP-Posting-Host: kos2mac3.berkeley.edu X-Newsreader: Value-Added NewsWatcher 2.0b14.2+ Xref: biosci bionet.molbio.proteins:2886 bionet.xtallography:1250 In bionet.xtallography I wrote: >I'm looking for some examples of known structures of proteins which >have dimerization interfaces which are a-helices. I'm not looking >for a coiled-coil type structure but one in which the helices pack >against the protien on one side and against the other member of the >dimer on the other. >Thanks. Here are the responses I recieved for those people who have expressed interest: From: Simon Brocklehurst Have you looked at the disulphide oxidoreductase flavoproteins? This family might fit your needs? If you want to see a picture of the interface, you could look in the September issue of TIBS (if you take that in Berkley) at the article "Protein-protein recognition mediated by a mini-protein domain..." pp 360-361. Also, interferon gamma might fit the bill (but I may have misremembred) Hope this helps Simon _________________________________________________________________________ | | ,_ o Simon M. Brocklehurst, | / //\, Oxford Centre for Molecular Sciences | \>> | Department of Biochemistry, University of Oxford, | \\, Oxford, UK. | E-mail: smb@bioch.ox.ac.uk |________________________________________________________________________ From: CATHY LAWSON You might look at trp repressor, which has an extensive dimerization interface and is an all alpha-helical protein. I am biased, but I think it is a beautiful structure .... Check out pdb entries 1wrp, 2wrp, 3wrp, 1tro, 1trr. Cathy Lawson Biology Brookhaven Nat'l Lab Upton, NY 11973 From: Chuck Duarte The first one that comes to mind is IL-8, pdb1il8 in the PDB. I would have posted to the newsgroup but our site doesn't allow that. Please forward a list of responses you get. It would be very much appreciated. Thanks. cd From: Kevin Gardner How about the xtal structure of the glucocorticoid receptor bound to its DNA target (1glu.pdb)? The dimerization interface consists of some helix and some extended strand structure --- the helical section fits the criteria you list above. Have you gotten any other suggestions? Good luck, Kevin -- ************************************************************************* Kevin Gardner Yale University Dept. of Molecular Biophysics and Biochemistry Internet: gardner@zinc.csb.yale.edu Bitnet: gardner@yalemed ***I support the boycott of companies using mass Internet advertising*** --gc ...doom is like the handle of a pot, | stop waiting, it's it's there, know it, have ice in your| already happened. tea, marry, have children, visit your| genecutl@mendel.berkeley.edu dentist, do not scream at night even |http://sp1.berkeley.edu/gc.html if you feel like screaming... -cb | http://sp1.berkeley.edu/ From owner-proteins@net.bio.net Mon Oct 17 23:00:00 1994 Path: biosci!bcm!cs.utexas.edu!convex!news.duke.edu!news-feed-1.peachnet.edu!gatech!europa.eng.gtefsd.com!ceylon!news2.near.net!das-news2.harvard.edu!husc-news.harvard.edu!lipid!robison Newsgroups: bionet.molbio.proteins Subject: Re: Info on pI? Message-ID: <1994Oct18.105321.35244@hulaw1.harvard.edu> From: robison@lipid.harvard.edu (Keith Robison) Date: 18 Oct 94 10:53:20 EDT References: <1994Oct7.232223.1@icbr> Nntp-Posting-Host: lipid.harvard.edu X-Newsreader: TIN [version 1.2 PL2] Lines: 23 rep@icbr wrote: : Hey! : Can anybody tel me a good place to go searching for pI's. I'm interested : in the various carbonic anhydrase isoforms, but don't feel like plowing : through a ton of papers to get them. Is there a reference, or a database, : or a site somewhere on the net? : I appreciate any help. If the protein is in SwissProt you can get the calculated pI & MW from their WWW server at http://expasy.hcuge.ch/ Good luck! Keith Robison Harvard University Department of Cellular and Developmental Biology Department of Genetics / HHMI robison@mito.harvard.edu From owner-proteins@net.bio.net Tue Oct 18 23:00:00 1994 Newsgroups: bionet.molbio.proteins,bionet.molbio.genbank Path: biosci!news.Stanford.EDU!rutgers!gatech!howland.reston.ans.net!EU.net!uunet!emba-news.uvm.edu!brianf From: brianf@med.uvm.edu (Brian Foley) Subject: Errors in Protein Sequences. Message-ID: <1994Oct19.201016.13342@emba.uvm.edu> Sender: news@emba.uvm.edu Organization: EMBA Computer Facility, University of Vermont X-Newsreader: TIN [version 1.2 PL1] Date: Wed, 19 Oct 1994 20:10:16 GMT Lines: 60 Xref: biosci bionet.molbio.proteins:2891 bionet.molbio.genbank:1801 From emba-news.uvm.edu!brianf Wed Oct 19 16:08:31 1994 Newsgroups: bionet.molbio.swiss-prot Path: emba-news.uvm.edu!brianf From: brianf@med.uvm.edu (Brian Foley) Subject: Errors in Swiss-Prot Message-ID: <1994Oct16.162935.23181@emba.uvm.edu> Sender: news@emba.uvm.edu Organization: EMBA Computer Facility, University of Vermont X-Newsreader: TIN [version 1.2 PL1] Date: Sun, 16 Oct 1994 16:29:35 GMT Lines: 48 When I do a BLAST search of a protein of interest against the "non-redundant" database provided by the NCBI, I often find that there are redundancies in this "non-redundant" data set. Closer examination reveals that NCBI eliminates redundancies only if the two protein sequences are identical (as they should), and that the apparent "redundancies" are always cases where the same protein sequence is entered differently into the various databases. Here is a case in point: The E. coli LepA gene and protein sequence were published, and GenBank or EMBL entered the DNA sequence into their database and then translated the DNA sequence into protein for the "GenPept" data. Swiss-Prot or PIR or some other database entered the protein sequence from the journal, rather than using the DNA sequence translation. Now in theory, both sequences should be the same. In fact, they have a single amino acid difference, with the Siss-Prot entry (Accession number P07682) having an error ...giNi... where GenPept (Accession number K00426) has the correct sequence .giTi... I know that the GenPept entry is correct both by looking at the original publication, and by noting that this region of the protein is absolutely invariant among elongation factors and initiation factors from E.coli to archaebacteria to mammals to plants. 1: I know two addresses for reporting errors to GenBank/EMBL. update@ncgr.org or update@ncbi.nlm.nih.gov I have never seen an address for reporting errors in Swiss-Prot or PIR. Could someone please supply that information, as well as an explanation of which organization (Swiss-prot or PIR) should handle various errors? 2: Looking at the number of redundancies in the "non-redundant" data set from NCBI would be an excellent way to catch this type of error. Is anyone in charge of a massive effort to remove such errors from the databases? From my experience with BLAST searching it looks like there are errors of this type in a substantial number of entries. I am trying to clean up the entries that are related to my research but is an uphill battle. -- ******************************************************************** * Brian Foley * If we knew what we were doing * * Molecular Genetics Dept. * it wouldn't be called research * * University of Vermont * * ******************************************************************** From owner-proteins@net.bio.net Tue Oct 18 23:00:00 1994 Newsgroups: bionet.molbio.proteins Path: biosci!bcm!cs.utexas.edu!swrinde!pipex!uunet!EU.net!sun4nl!sci.kun.nl!caos.kun.nl!kunazag4 From: kunazag4@caos.kun.nl (KUN Anthropogenetica (Drs. B.Smeets)) Subject: random peptide libraries Message-ID: <1994Oct19.144738.6482@caos.kun.nl> Summary: Is there anyone with experience with the random peptide library produce Keywords: random peptides Sender: root@caos.kun.nl (System PRIVILEGED Account) Nntp-Posting-Host: cammsg1.caos.kun.nl Organization: CAOS/CAMM Date: Wed, 19 Oct 1994 14:47:38 GMT Lines: 16 Is there anyone with