PURIFICATION AND CHARACTERIZATION OF THE HUMAN PROTEIN-TYROSINE-PHOSPHATASE, PTP-MU, FROM A BACULOVIRUS EXPRESSION SYSTEM

The receptor like PTPase, PTP mu, displays structural similarity in it s extracellular segment to members of the immunoglobulin superfamily o f cell adhesion molecules. The full length form of PTP mu (200 kD) and a construct expressing only the intracellular PTPase domain-containin g segment (80 kD) were expressed in the baculovirus/Sf9 cell system, p urified and characterized. Full length PTP mu was membrane associated while the truncated form was recovered in the soluble fraction. PTP mu preferentially dephosphorylated a reduced carboxamidomethylated and m aleylated derivative of lysozyme (RCML) over other tyrosine phosphoryl ated substrates such as myelin basic protein (MBP) or the synthetic pe ptide EDNDYINASL. The enzymatic properties of the soluble, truncated f orm of the enzyme were examined in detail. The pH optimum was 7.5. It dephosphorylated RCML with a K-m of 400 nM and a V-max of 725 nmol/min /mg. This form of the enzyme was 2 fold more active than full length P TP mu. Trypsinization of the full length form inhibited activity. Vana date and molybdate, potent tyrosine phosphatase inhibitors, abolished activity of the enzyme. Zn++ and Mn++ ions, polylysine, poly-glu/tyr, and spermine were also inhibitory.