Approaches to the identification of protein tyrosine phosphatase substrates

Protein tyrosine phosphorylation is a reversible process that is balanced by the opposing actions of protein tyrosine kinases (PTKs) and protein tyrosine phosphatases (PTPs). Abnormal levels of tyrosine phosphorylation can lead to diseases such as cancer, diabetes, and cardiovascular and immunological disorders. Hence, the coordinated regulation of tyrosine phosphorylation by the actions of the PTKs and PTPs is essential for normal physiological signaling. The PTPs are a large and structurally diverse family of enzymes comprising approximately 100 members in humans. This chapter focuses on the approaches that have been used to identify substrates for the tyrosine-specific PTPs, and highlights the successes of these approaches with reference to select examples. Although identifying substrates for PTPs has been challenging, the PTP substrate-trapping approach using the more efficient D-A and its variants has met with great success for a number of PTPs. The PTP-PEST D-A substrate-trapping mutant has been found to recognize the multisubstrate adaptor/scaffold protein p130^Cas as a substrate in vitro and in intact cells. The biological role of p130^cas is now well established; it is intimately involved in the regulation of cell migration and adhesion, and is implicated in cellular transformation. Thus, the identification of p130^cas as a PTP-PEST substrate has provided mechanistic support for the role of PTP-PEST in cell migration and adhesion. Also, its variants have met with great success for a number of PTPs.