Separating GST from GST-fusion protein
Paul J. Phelan
via methods%40net.bio.net
(by Paul.Phelan from tufts.edu)
Wed Apr 15 12:47:56 EST 2009
I think I asked this question before on this forum, but I thought I'd
try it again just in case anyone has any new "tricks" or ideas, because
I'm having the same old problem again. I am trying to purify a
GST-fusion protein, and it's contaminated with GST. The GST-fusion has
a mol. wt. of 40,000 and GST is 26,000, but on a Sephacryl S100 gel
filtration column, both proteins are co-eluting in a single, beautiful
peak. Anyone would think that such a textbook peak would only contain
pure protein, but no - half of it is GST! I even re-ran the gel
filtration column in a buffer containing 0.4 M NaCl to try to separate
the two proteins, but I got the same single peak, so the GST must be
still there.
Now I am not talking about digesting a GST-fusion and then removing
GST; I have no problem doing that, but I am trying to purify some
intact GST-fusion, without taking GST with it: does anyone have a magic
trick to get rid of GST?
Thank you
Paul Phelan
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