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refolding from inclusion bodies

Thu Feb 20 12:43:11 EST 1997

juhi juneja 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?

You might try dialyzing against a buffer containing 40-50% glycerol-- 
this has worked well with one of the proteins we work with.  It is also a 
good idea to include a scavanger for isocyanate in your urea solutions 
(e.g., about 25 mM L-lysine) to prevent modification of your protein 
and a reducing agent (e.g., 10 - 50 mM mercaptoethanol) to prevent 
cysteine oxidation (and always make your urea solutions fresh from an 
ultrapure preparation).

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