Acid Metallophosphatase from the Ameba Amoeba proteus

Tartrate-resistant acid phosphatase (TR-AcPh) from the ameba Amoeba proteus is represented by 3 bands (electromorphs) revealed after disk-electrophoresis in PAAG, using 2-naphthylphosphate as substrate. The presence of 50 mmol/l MgCl₂ or CaCl₂ in the incubation mixture increases activities of all electromorphs of TR-AcPh, while of ZnCl₂, of two of them. The activity of the TR-AcPh electromorphs also rose after the 30-min incubation of the gels in MgCl₂, CaCl₂ or ZnCl₂ (10 and 100 mM) before gel staining. However, 1 M ZnCl₂, unlike 1 M CaCl₂ or 1 M MgCl₂, partly inactivated two out of three TR-AcPh electromorphs. The TR-AcPh electromorphs were inhibited by 1,10-phenanthroline (1,10-Ph), EDTA, and EGTA (all at a concentration of 5 mM) faster than by H₂O₂ (10 mM). The inactivation of the TR-AcPh electromorphs by the chelating agents did not depend (EGTA) or nearly did not depend (EDTA, 1,10-Ph) on their concentration (0.05, 0.5, and 5 mM). Out of 5 tested ions (Mg²⁺, Ca²⁺, Fe²⁺, Fe³⁺, and Zn²⁺), only Zn ions reactivated the TR-AcPh electromorphs inactivated by 1,10-Ph, EDTA or EGTA. The TR-AcPh electromorphs were reactivated worse after inactivation by EGTA than by EDTA or 1,10-Ph. It is suggested that the active site of TR-AcPh contains the zinc ion essential for catalytic activity of this enzyme, i.e., TR-AcPh of A. proteus is a metallophosphatase performing the phosphomonoesterase activity in acidic medium.