Hi Matthew,
TEV protease is very robust. I normally digest with 1:100 ratio
according to the OD280. I normally digest at 4C for overnight around 16-18
hours. Make sure your tev protease site are not inaccessible and buried
inside.
best
Xiaohu
On Mon, May 24, 2010 at 12:27 PM, Matthew Merski <
mer...@blur.compbio.ucsf.edu> wrote:
> Hello all,
>
> I am working with a protein that is expressed as with an N-terminal domain
> that is normally cleaved for activation of the protein (and
> crystallization). For in vitro reasons I've needed to switch the normal site
> to a TEV site. However, even though the TEV site is in the same place as the
> original proteolytic site, I have been unable to get cleavage despite using
> a lot of TEV at 37 C, pH 8.0. Has anyone been able to overcome a similar
> problem?
>
>
> Matthew Merski
> Post-doctoral researcher
> UCSF
>
