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Hi, we are interested in a certain protein with "putative kinase domains". We actually do suspect it is a protein kinase, probably Serine/threonine.
We've got a GST-tagged version of our protein and would like to test whether it has kinase activity in vitro, before looking for substrates and signal transduction pathways. We tried the simplest approach, a Histone 32P phosphorylation assay. We tested Histone since it is a wide-ranging/ general phosphorylation substrate. It worked with several controls, but not with our protein.
At this moment we can't spend thousands of Euros in kinase-assay kits. Either another substrate is required or our protein would require a coenzyme/cofactor. Since in our lab we have no great deal of experience with phosphorylation we would really appreciate any inputs