In addition to the other very good suggestions, you could consider
using a purification tag at both the N- and C-termini of your protein
to only pull out full length protein. I've had success with Flag+His
and MBP+His.
Ho
Ho Leung Ng
University of Hawaii at Manoa
Assistant Professor, Department of Chemistry
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On Tue, Aug 21, 2012 at 1:00 PM, CCP4BB automatic digest system
<[log in to unmask]> wrote:
>
> Date: Tue, 21 Aug 2012 21:15:15 +0100
> From: Theresa Hsu <[log in to unmask]>
> Subject: Purify non-stable protein
>
> Dear all
>
> I made deletion mutation of a stretch of 20 amino acids on my protein. I can purify and crystallize wild type protein but not the mutated. Mass spec on gel separated protein shows degradation of mutant losing about another 150 amino acids. Is there any way of purifying this non-stable protein? I know nature has designed proteins to be stable.
>
> All steps are done at 4 C and protease inhibitor added during cell lysis for both proteins.
>
> Thank you.
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