Problems with GST fusion protein
rthorne at mail.newcastle.edu.au
Wed May 3 06:45:02 EST 2000
All GST-fusions are different but I can offer you some guidelines which
might help you.
First step might be to alter the strain of bacteria you are using for a
more specialized strain eg: BL21-DE3 from Stratagene (look at the web
site to learn more about these). Alternatives are available from other
companies (but I never used them).
If this does not help, if at all practicable, make new construct/s with
smaller bits of the protein you want to use. This sometimes removes
parts of the molecule that encourage its wholesale destruction.
Eva Chen wrote:
> Is there anybody having experience on the purification of GST fusion
> Our lab doesn't have much experience and now there has been a serious
> problem of degradation of the GST fusion protein. We have tried the
> protease inhibitor from Sigma and Roche, and the whole purification
> process was accomplished in a 4 C cold room. But the proteolysed
> products are still there.
> Any suggestions are highly appreciated. Thank you all for your
> Eva Y. Chen
> yi-hui.chen at mcmail.vanderbilt.edu
> Jim Sutcliffe Lab
> Department of Molecular Physiology and Biophysics
> Vanderbilt University School of Medicine
> Nashville, TN 37232
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