One good way to go about it would be to obtain each of the potential target enzymes in a purified form and test the molecule for inhibition of each in an enzymatic reaction.

Thanks Adam,

We have already done this enzymatic assay with several targets identified. I'm trying to define the effect in an in vitro or in vivo environment, where we can also observe biological changes (enhancement or inhibition activity), such as gene or protein expression due to the molecule we're studying

First, you should define, which phosphatases are expressed during your studied process. Then, you should obtain single and multiple knock-outs and see, whether the process is affected in them.

Tomáš Hluska thanks for this great approach. Currently, there are more than ten phosphatases acting in the process we are studying. Thus, making this experiment non-viable. But maybe there is a way to work in cell cultures by using si-mRNA, or specifics/general inhibitors, or even better enhancers of phosphatase activity.

Does anyone know if phosphatase enhancers exist?

Thanks

Ignasi Canals don't say that something is impossible because there will be an imbecil, who doesn't know it's impossible and they will do it xD

Are all those phosphatases really expressed during your process? You can prepare sets of high-order knock-outs and based on these results select those that are most important.