A 13-kDa phosphoamidase was isolated as a single band on SDS-PAGE from bovine liver. Its Stokes' radius, sedimentation coefficient, molecular mass, and optimal pH were estimated to be 1.6 nm, 1.8s, 13kDa, and 6.5, respectively. The enzyme released P_i from 3-phosphohistidine, 6-phospholysine, and amidophosphate at rates of 0.9, 0.6, and 2.6μmol/min/mg protein, respectively. However, it did not dephosphorylate phosphocreatine, N^ψ-phosphoarginine, imidodiphosphate, or O-phosphorylated compounds including inorganic pyrophosphate. It also dephosphorylated succinic thiokinase and nucleoside diphosphate kinase autophosphorylated at His residues, indicating that it works as a protein histidine phosphatase. A thiol reagent, 30μM N-ethylmaleimide, depressed the activity by half, while a thiol compound, 2-mercaptoethanol, protected the enzyme from heat-inactivation. Five millimolar divalent cations, such as Mg²⁺ and Mn²⁺, and 5mM EDTA, had no effect on the activity.