Hello Claudia, have you thought of, instead of rules, using restricted proteolysis combined with N-terminal sequencing in order to better estimate a good position at least for the N-terminal part of your protein?
Tim On Thu, Mar 11, 2010 at 09:26:53AM +0100, Anastassis Perrakis wrote: > Hi - > > I do not think there are "rules". There is a logic, to never truncate > within secondary structure elements, and to prefer to delete regions > that have a high probability to be disordered. > > With excuses for the non-ccp4 plugin, you can use our web server to help > you design these. > > http://xtal.nki.nl/ccd/Welcome.html > > > A. > > > On Mar 10, 2010, at 12:45, Claudia Scotti wrote: > >> >> Dear All, >> >> I'd like to espress a protein for crystallisation as a whole, but also >> to try separate expression of its two domains. >> >> Although prediciting domain boundaires is not difficult, I know that >> it is worthwhile testing several constructs to improve espression in >> soluble form. In several works I've seen that the choice is to keep or >> give up some aminoacids (for example 4 or 5) at the N and/or C-termini >> and to test them to see which expresses best. >> >> I was wondering if there are defined "rules" to choose these variants >> and/or to modify the N and C termini of a protein of interest when it >> is not important to keep them "true" (=native) in order to optimise >> expression levels. >> >> Is there any good reference about this, please? >> >> Personal experiences are welcome. >> >> Many thanks, >> >> Claudia >> >> >> Claudia Scotti Dipartimento di Medicina Sperimentale Sezione di >> Patologia Generale Universita' di Pavia Piazza Botta, 10 27100 Pavia >> Italia Tel. 0039 0382 986335/8/1 Facs 0039 0382 303673 >> >> >> >> Hotmail: Free, trusted and rich email service. Get it now. > > P please don't print this e-mail unless you really need to > Anastassis (Tassos) Perrakis, Principal Investigator / Staff Member > Department of Biochemistry (B8) > Netherlands Cancer Institute, > Dept. B8, 1066 CX Amsterdam, The Netherlands > Tel: +31 20 512 1951 Fax: +31 20 512 1954 Mobile / SMS: +31 6 28 597791 > > > > -- -- Tim Gruene Institut fuer anorganische Chemie Tammannstr. 4 D-37077 Goettingen GPG Key ID = A46BEE1A
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