Phosphotyrosyl protein phosphatase from the liver ofTilapia mossambica: Activating the Src-phosphorylated DNA topoisomerase I

Abstract

Phosphotyrosyl protein phosphatase was purified from the liver of the Tilapia mossambica. Specific activity of the final purified preparation was 5,095 U/mg of protein with the 32P‐peptide 1142–1153 of the insulin receptor as substrate. SDS‐polyacrylamide gel electrophoresis revealed that monomers of the phosphotyrosyl protein phosphatase have relative masses of 67,000 ± 500 and 60,000 ± 500. Since the active phosphotyrosyl protein phosphatase was found to have a relative mass of 62,000 ± 1,000, the purified enzyme was concluded to be monomeric. The phosphotyrosyl protein phosphatase had an isoelectric point of 5.79 ± 0.02 and an optimal pH of 7.0. Incubation of the purified enzyme fraction with Src‐phosphorylated DNA topoisomerase I resulted in an increase activity of DNA topoisomerase I in a dose‐dependent manner. Activity of phosphotyrosyl protein phosphatase is inhibited by MnCl2 and vanadate at μmolar concentrations, but less inhibition was observed with CaCl2 and MgCl2. J. Exp. Zool. 277:14–22, 1997. © 1997 Wiley‐Liss, Inc.