Activity restored by mixing recombinant domains?

Frank O. Fackelmayer fof1 at
Tue Oct 28 03:41:38 EST 1997

In article <lloyd.graham-2908560305100001 at>,
lloyd.graham at (Lloyd Graham) wrote:

> Greetings all,
> I'm looking for any precedents where the domains of a monofunctional
> enzyme have been expressed separately as inactive proteins, but when mixed
> together they associate to form a complex with significant enzymatic
> activity. 
> The closest I have got to an example is mouse creatine kinase, where
> independent expression of the two domains gives inclusion bodies but
> co-expression yields active soluble enzyme [Gross et al., 1996: Prot. Sci.
> 5, 320-330]
> If anybody can provide instances where purified inactive domains
> complement in vitro, I would be very grateful. 
> Lloyd.

Hi Lloyd,
What about alpha-complementation of beta-galactosidase, used by everyone
as the "blue-white" selection in cloning procedures. In this case, a short
N-terminal part (the so-called alpha-peptide) is encoded on a plasmid, and
the remainder of the protein is encoded by the E.coli host genome. Neither
of the two parts alone is enzymatically active. Galactosidase activity
(blue staining of the colonies in presence of IPTG and X-gal) is only
present when both parts of the protein are present, i.e. no insert has
been cloned into the alpha-peptide encoding vector that would disrupt the
coding information.

Hope this helps,

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