refolding from inclusion bodies

Sourav Ghosh sghosh at utmem1.utmem.edu
Fri Feb 21 16:22:03 EST 1997


In article <330B2576.3B66 at ncbs.tifrbng.res.in>, juhi at ncbs.tifrbng.res.in wrote:

> The mutant protein that I'm trying to purify is expressed in inclusion
> bodies. After solubilising in 8Murea,50%of the protein precipitates
> while dialysing out the urea. Is there any way to prevent precipitation
> and increase refolding yields?

I had the same problem and tried a variety of things.  However it is very
emperical....solubilization volume is critical; you can try Sarkosyl,
glycerol, tween 20, dialyse against progressive less urea/guanidine etc. I
prefered Guanidine over urea personally. I improved recovery, but never
got the activity I was looking for!! may be it never refolded to native
conf.!1 Hopefully you will have better luck. However, have you heard of
the thioredoxin-fusion system...theoretically they should have better
chances of remaining 'native'. I never got around trying them myself.
Ciao,
Sourav



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