inclusion bodies

Tom MacAllister N490047 at UNIVSCVM.CSD.SCAROLINA.EDU
Wed Apr 20 13:22:03 EST 1994


Dear netters:
     We have a construct that produces major amounts of our protein of interest
, but it is insoluble.  We have achieved resolublization, but have not
recovered any activity.  The properly folded protein is active (no mutations)
because we can assay it in the sup.  The problem is that ~99% is in the pellet.
Facts: 1. It resolublizes in 2M GuCl or 4M urea.  2.  It stays soluble in 1M
GuCl when dialyzed.  3.  We believe that there is at least 1 disulfide bond.
4.  It is mostly insoluble in higher salt (150 - 500mM) in 1M GuCl.  5.  Dilu-
tion doesn't work any better than dialysis.  6.  We have been using 50mM Tris,
pH7.9 at 4C.  7.  10% glyerol doesn't help.   Does anyone have any experience
with this or know anyone that would be good to talk with?  Thank you.

                                                                     Tom

TOM MACALLISTER
N490047 at UNIVSCVM



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