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Enzyme kinetics - quantification of reversible enzyme inhibition (Feb/23/2009 )

im not familiar with this stuff, therefore i hope to get some suggestions and recommendations for the solution of my problem:

im working with bortezomib which is a selective and reversible inhibitor of the chymotryptic activity of the proteasome
i inject mice with this inhibitor, and have now to measure the inhibitory action of this compound in vivo.
therefore i have to isolate proteasomes and check for chymotryptic activity in the lysates.

my question now is a general one: when i isolate proteasomes with a special protocol to obtain high purified proteasomes, how will i know that acitivty was inhibited somehow?
the reversible inhibitor will not permanently bound to my proteasomes!

how can i really measure inhibitory activity of tissue samples???

thanks in advance


Can you examine the expression of a subset of proteins targeted by the proteasome? In this case, you could examine the percent remaining for each protein or the amount of ubiquitinated protein compared to the un-inhibited proteasome controls where protein expression of these targets would be low due to degradation?

-Dr Teeth-