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Re: [ccp4bb]: soluble aggregation of proteins
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does this protein precipitate after expression and purification, or
already during the expression (i.e. are inclusion bodies formed) ?
If aggregation only happens after concentrating the sample, you might
first want to:
1) centrifuge the sample for more than an hour in order to remove any
protein that has already aggregated. This aggregated product can catalyze
the formation of more aggregates. But when these 'seeds' are removed from
the solution, it can be safer to concentrate afterwards
2) change the pH of the solution (far from the pI)
3) work at lower temperatures
On Mon, 10 Apr 2000, Janet Moloney wrote:
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> Dear all,
> We are expressing proteins here in E. Coli in the pMALp2 vector as
> maltose-binding fusion proteins. This is a periplasmic expression and we
> have found that our three proteins in the 22-38 KDa range all aggregate
> in solution.
> Any suggestions from your experience to treat this type of problem?
> Janet Moloney, Division of Structural Biology,
> Wellcome Trust Centre for Human Genetics,
> University of Oxford, Roosevelt Drive,
> Oxford OX3 7BN, U.K.
> Tel: 0044(0)1865 287551 Fax: 0044(0)1865 287547