switching cations

[Dr E. Buxbaum]

bussierd at tvsfrank (Dirkson E. Bussiere-42T) wrote:
>
>I'm trying to swap the Zn2+ in an enzyme with other divalent
>cations, but everytime I do it, the protein precipitates.  I
>more or less dialyze the protein in 50mM Tris-HCL, pH 7.4,
>with 1-10 mM of the cation2+; the protein is stable in
>the Tris by itself. 

The judicious use of detergents and/or denaturing agents like urea can 
sometimes help. The idea is to use these additives in concentrations, 
which prevent aggregation, but do not inactivate the protein. The right 
combination of agents has to be found for each protein by trial and 
error.

Another possibility is to bind the protein to a chromatographic column,  
do the transition and then elute. That can keep them appart during the 
critical periode. Sometimes this method needs to be combined with the one 
above.