GST fusion protein
krasel at wpxx02.toxi.uni-wuerzburg.de
Tue Jul 7 05:53:00 EST 1998
Luc CAMOIN <camoin at cochin.inserm.fr> wrote:
> Purification on Glutathione sepharose 4B and western blot with anti GST
> revealed only a band around 25 kDa. This molecular weight and reactivity
> with anti GST could suggest that this protein is GST alone.
> Can we conclude that the contact between GST and the protein of interest is
> a protease sensitive region?
I have had similar problems in some of my experiments and have reached this
conclusion. The protease sensitivity of the linker region depends, amongst
other things, on the length of the linker and on the bacterial strain in which
you express your protein of interest. An approach to circumvent this problem
which sometimes works is to express your GST fusion protein in a protease-
negative E. coli strain (e.g. BL21) at lower temperatures (e.g. 25°C
instead of 37°C).
Hope that helps,
/* Cornelius Krasel, U Wuerzburg, Dept. of Pharmacology, Versbacher Str. 9 */
/* D-97078 Wuerzburg, Germany email: phak004 at rzbox.uni-wuerzburg.de SP4 */
/* "Science is the game we play with God to find out what His rules are." */
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