Separation of two proteins with similar properties

Jon-Sverre Schanche jon-ss at online.no
Fri Dec 20 18:03:52 EST 1996


Hi Marit,

A general sugestion would be to try reversed phase chromatography, which is
what many people use for their final polishing of proteins of anny size
today. The problem you have to be aware of is the tendency of proteins to
denature under high concentrations of organic solvents. Try isopropanol,
ethanol, acetonitril or methanol (in priority order) on a polymer based
RPC-column. You may also try anionexchange columns with organic modifier
added to the buffer (low conc. of isopropanol may do the trick).

Have you tried isoelectric focusing for determining pI of the two
components?

Good luck,

-- 
Jon-Sverre Schanche
Alt. e-mail adresse: jon-sverre.schanche at eu.pnu.com


Marit Valeur Ramstad <Marit.V.Ramstad at chem.sintef.no> wrote in article
<593m9m$nfa at stork.runit.sintef.no>...
> I am working with an enzyme produced by an recombinant E.coli. This 
> enzyme have been purified by ion exchange and hydrofobe chromatography. 
> The purified sample has some minor contaminants,but the trouble is that 
> it seems like the enzyme is presents in two forms.Most proberly because 
> of a minor degradation during the purifications steps.




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