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Dear all, Thank you very much for all of information about domain determination! I think the limited proteolysis is a good choice for the domain determination as we have no information about a protein. However, if we do have a domain information by the bioinformatics, how can we truncate the domain? Are we need to keep three or five more amino acids in two ends? Thanks. Best regards, Xianhui On Tue, Mar 8, 2011 at 2:22 AM, Mark J van Raaij <[log in to unmask]>wrote: > for an experimental way to determine soluble domains see the following > paper: > ESPRIT: an automated, library-based method for mapping and soluble > expression of protein domains from challenging targets. > Yumerefendi H, Tarendeau F, Mas PJ, Hart DJ. > J Struct Biol. 2010 Oct;172(1):66-74. Epub 2010 Mar 4. Review. > PMID: 20206698 [PubMed - indexed for MEDLINE] > > > > Mark J van Raaij > Laboratorio M-4 > Dpto de Estructura de Macromoleculas > Centro Nacional de Biotecnologia - CSIC > c/Darwin 3, Campus Cantoblanco > E-28049 Madrid, Spain > tel. (+34) 91 585 4616 > > http://www.cnb.csic.es/content/research/macromolecular/mvraaij/index.php?l=1 > > > > On 7 Mar 2011, at 19:18, gauri misra wrote: > > > Hi, > > To start with it would be great if you look in to the secondary structure > prediction of the sequence using any of the standard servers like PSIPRED, > JPRED etc. Many more available at expasy site http://ca.expasy.org/tools/. > > Whatever construct you finally choose to make just remember the standard > rule that we generally follow is to avoid deleting the alpha helices and > beta sheets. You can design your initial primers so as to obtain the > complete amplification of these secondary structures from any part within > the protein. > > You can even use the various modules of the following online available > server > > http://scratch.proteomics.ics.uci.edu/ > > to have an idea of the intrinsically disordered regions in the protein, > transmemebrane regions and disulfide bonds that would certainly help you in > initiating in the right direction. > > > > Best wishes > > Gauri > > > > On Mon, Mar 7, 2011 at 4:10 AM, Xianhui Wu <[log in to unmask]> wrote: > > Dear all, > > > > Before we try to study the crystal structure of an unknown protein, we > need to determine the sequence that can fold into a compact and stable 3D > domain. What kinds of methods can we choose? > > > > -- > > Best regards, > > XH Wu > > > -- Best regards, Xianhui