Bovine liver phosphoamidase as a protein histidine/lysine phosphatase.
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, 1.8 s, 13 kDa, and 6.5, respectively. The enzyme released P(i) from 3-phosphohistidine, 6-phospholysine, and amidophosphate at rates of 0.9, 0.6, and 2.6 micromol/min/mg protein, respectively. However, it did not dephosphorylate phosphocreatine, N(omega)-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, 30 microM 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 5 mM EDTA, had no effect on the activity.
Hiraishi
H
H
Department of Biochemistry, Saga Medical School, Saga, Saga, 849-8501, Japan.
Yokoi
F
F
Kumon
A
A
D000818:Animals; D002417:Cattle; D002847:Chromatography, Agarose; D002853:Chromatography, Liquid; D004305:Dose-Response Relationship, Drug; D004590:Electrophoresis, Paper; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005033:Ethylmaleimide / Q000494:pharmacology; D006639:Histidine / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D006867:Hydrolases / Q000737:chemistry* / Q000302:isolation & purification*; D007700:Kinetics; D008099:Liver / Q000201:enzymology*; D008239:Lysine / Q000378:metabolism*; D009701:Nucleoside-Diphosphate Kinase / Q000378:metabolism; D013379:Substrate Specificity
D004791:Enzyme Inhibitors; D006639:Histidine; D009701:Nucleoside-Diphosphate Kinase; D006867:Hydrolases; C400721:13 kDa phosphoamidase; D008239:Lysine; D005033:Ethylmaleimide