PURIFICATION AND CHARACTERIZATION OF PTP2C, A WIDELY DISTRIBUTED PROTEIN-TYROSINE-PHOSPHATASE CONTAINING 2 SH2 DOMAINS

PTP2C, a widely distributed protein tyrosine phosphatase (PTP) contain ing two SH2 domains, was expressed as a recombinant enzyme in Escheric hia coil and purified to near homogeneity. The purified enzyme and a t runcated form lacking the SH2 domains (Delta SH2-PTP2C) have been char acterized with four commonly used substrates. Both forms showed pH opt ima of around neutrality for protein substrates but below 5.5 for a pe ptide substrate and para-nitrophenylphosphate. The dependence of the e nzymes on ionic strength varied with the nature of the substrates invo lved. Like its analog PTP1C, PTP2C displayed a specific activity of le ss than 0.1% of that observed with other known PTPs toward protein sub strates. Deletion of the SH2 domains increased its activity by 12-45-f old, depending on the substrates used. Limited trypsinolysis which cle aved about 4 kDa from the carboxyl terminus resulted in a 2-5-fold act ivation of the full-length enzyme but was es sentially without effect on the truncated enzyme. Both forms showed similar responses to effect ers including activators (e.g. anionic phospholipids) or inhibitors (e .g. vanadate, molybdate, or Zn2+). PTP2C and Delta SH2-PTP2C were phos phorylated in vitro by mitogen-activated protein kinase, protein kinas e C, and various protein tyrosine kinases; in the latter case, they un derwent au todephosphorylation. No significant effect of the phosphory lation reactions on enzyme activity could be ob served in vitro.