protein-protein interactions

Peter Prevelige prevelig at uab.edu
Tue Oct 6 22:26:47 EST 1998


In article
<Pine.GSO.3.96.981006165717.10396A-100000 at sunx1.central.susx.ac.uk>,
Matthew Hicks <baps9 at central.susx.ac.uk> wrote:

>The best method in my opinion would be analytical ultracentrifugation
>(AUC) - look at the Beckman homepage on the WWW or see the National
>Hydrodynamics Centre homepage (Nottingham University - UK). This method
>can potentially give association constants in a native-like buffer.
>X-linking would show interaction but not much about the tightness of
>binding - also you _may_ have to use 'weird' buffer conditions which may
>be required for chemical
>modification of the proteins.  If you cannot get
>access  to the kit then how about using size exclusion chromatography to
>see if your samples coelute at a molecular weight above that of the
>individual proteins - this would show interaction (but not as well as
>AUC!)
>Good Luck
>Matt Hicks
>University of Sussex (UK)

I agree, analytical ultracentrifugation (equilibrium sed) is the best way
to go..


a poor man's version is to run sucrose gradients of each component
individually and then run the complex. follow the protein distribution by
SDS-PAGE if there is interaction, even transiently, the slower sedimenting
peak will be shifted towards the faster sedimenting one by an amount that
is related to the average occupancy and the difference in s values...



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