KINASE AND PHOSPHATASE ASSAYS

摘要

Methods for screening for modulators of kinase or phosphatase activity are described. The methods comprise: a) providing a serine or threonine kinase and a substrate which can be phosphorylated by the kinase; b) contacting the kinase with its substrate in the presence and absence of a candidate modulator of the activity of the kinase under conditions which permit phosphorylation of the substrate by the kinase; c) contacting the substrates treated according to step (b) with a reporter, excluding a natural antibody, which binds phosphorylated substrate with higher affinity than unphosphorylated substrate; and d) comparing binding of the reporter to the substrate treated according to step (b) in the presence of the candidate modulator with binding of the reporter to the substrate treated according to step (b) in the absence of the candidate modulator. Examples of reporters for use in such assays include 14-3-3 proteins and WW domain containing proteins.