Also keep in mind that many of the purchased TEVs are formulated with
some reducing agent (e.g. AcTEV comes in a buffer with 5mM DTT, if I
recall correctly). So unless the enzyme is buffer exchanged
beforehand, there will be some reducing agent introduced alongside it,
depending on the dilution.
HTH,
-Tim
On Mon, Apr 16, 2012 at 4:32 PM, Jason Forse <[log in to unmask]> wrote:
> I've run into the same problem, and found David Waugh's FAQ to be a great resource:
> http://mcl1.ncifcrf.gov/waugh_tech.html
> They use a 3mM buffer of 10:1 reduced:oxidized glutathione. I've tried that and it cleaves my protein without reducing reducing the disulfide bridges.
>
> I'll second someone else's suggestion to add more TEV. That's worked for me as well, as long as the TEV's relatively fresh and there isn't too much reducing agent introduced along with it.
>
> Jason
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