Protein Tyrosine Phosphatase Assay

Watanabe, T. et al. 1998. Synthesis of fluorescent substrates for protein tyrosine phosphatase assays. Bioorg. Med. Chem. Lett. 8, 1301-1302. 

Scheme 10.7 Protein tyrosine phosphatase inhibitor assay.

McCain, D.F. and Z.Y. Zhang. 2001. Assays for protein-tyrosine phosphatases. Meth. Enzymol. 345, 507-518. 

Montalibet, J., K.I. Skorey, and B.P. Kennedy. 2005. Protein tyrosine phosphatase enzymatic assays. Methods 35, 2-8. 

Kinase or phosphatase assays based on the AlphaScreen principle are similar to TR-FRET assays in that they usually require a biotinylated substrate peptide and an anti-phosphoserine or tyrosine antibody. These two reagents are sandwiched between biotin and protein A-functionalized acceptor and donor beads. A kinase assay would show an enzyme-dependent increase in antibody binding (and thus signal) over time and a phosphatase assay would show an enzyme-dependent decrease in antibody binding over time. In some cases, the phosphorylation of an epitope will block the antibody binding and thus a phosphatase assay in principle can be constructed as a signal increase assay (Von Leoprichting and Kumpf, 2004 Warner et al., 2004). 

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