Purification yields protein complex between recombinant and native p97

Virginia Butel vbutel at ems.salk.edu
Thu Sep 7 13:15:34 EST 2000


Hi everybody,

I have expressed a recombinant mouse protein, p97, in High Five insect
cells using Gibco's baculovirus protocol. This is a fusion protein with
Maltose binding protein at the N-terminal end. After purifying on amylose
resin, I get very pure protein; only two bands show up by Coomassie
staining. One is my recombinant fusion protein, MBP-p97, the other is the
native p97 from the insect cells. Since p97 forms normally forms
homohexamers, I assume the insect p97 is similar enough to complex with
the overexpressed recombinant p97. 

Has anyone come across this problem? If so, I need ideas to get rid of the
native protein. Amylose resin is not compatible with denaturing protocols
and I would prefer not to have to try to renature to restore ATPase
activity.

Virginia






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