Dependence of divalent metal ions on phosphotransferase activity of osseous plate alkaline phosphatase.
Kinetic evidence for the role of divalent metal ions in the phosphotransferase activity of polidocanol-solubilized alkaline phosphatase from osseous plate is reported. Ethylenediamine tetreacetate, 1,10-phenanthrolin, and Chelex-100 were used to prepare metal-depleted alkaline phosphatase. Except for Chelex-100, either irreversible inactivation of the enzyme or incomplete removal of metal ions occurred. After Chelex-100 treatment, full hydrolase activity of alkaline phosphatase was recovered upon addition of metal ions. On the other hand, only 20% of transferase activity was restored with 0.1 microM ZnCl2, in the presence of 1.0 M diethanolamine as phosphate acceptor. In the presence of 0.1 mM MgCl2, the recovery of transferase activity increased to 63%. Independently of the phosphate acceptor used, the transferase activity of the metal-depleted alkaline phosphatase was fully restored by 8 microM ZnCl2 plus 5 mM MgCl2. In the presence of diethanolamine as phosphate acceptor, manganese, cobalt, and calcium ions did not stimulate the transferase activity. However, manganese and cobalt-enzyme catalyzed the transfer of phosphate to glycerol and glucose.
Ciancaglini
P
P
Departamento de Química, Universidade de São Paulo, Brasil.
Pizauro
J M
JM
Leone
F A
FA
D000469:Alkaline Phosphatase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D000818:Animals; D001842:Bone and Bones / Q000201:enzymology*; D002413:Cations, Divalent / Q000494:pharmacology; D002614:Chelating Agents / Q000494:pharmacology; D004492:Edetic Acid / Q000494:pharmacology; D066298:In Vitro Techniques; D007700:Kinetics; D008297:Male; D008670:Metals / Q000494:pharmacology*; D010618:Phenanthrolines / Q000494:pharmacology; D010710:Phosphates / Q000378:metabolism; D010770:Phosphotransferases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D051381:Rats; D017208:Rats, Wistar; D012117:Resins, Synthetic
D002413:Cations, Divalent; D002614:Chelating Agents; D008670:Metals; D010618:Phenanthrolines; D010710:Phosphates; D012117:Resins, Synthetic; C024997:Chelex 100; D004492:Edetic Acid; D010770:Phosphotransferases; D000469:Alkaline Phosphatase; C025205:1,10-phenanthroline