Purification and properties of bovine dental-pulp alkaline-phosphatase.

Alkaline phosphatase (E.C.3.1.3.1.) from unerupted bovine pulp was extracted from the microsomal fraction with eta-butanol and purified 77-fold, using DEAE-cellulose chromatography, Sephadex G-200 gel-filtration and concanavalin-A affinity chromatography, to a final specific activity of 92.3 units/mg protein. Affinity chromatography confirmed the glycoprotein nature of the enzyme. The pH optimum for the purified enzyme was 10.0 with rho-nitrophenylphosphate, and 8.7 with phosphoserine. The apparent Km was estimated to be 0.7 mM, using rho-nitrophenylphosphate in glycine-NaOH buffer, pH 10.0. The enzyme was markedly inhibited by EDTA, bromotetramisole and homoarginine but was insensitive to phenylalanine, and therefore resembled the alkaline phosphatase of liver and bone, but not that of intestine and placenta. No protein phosphatase activity towards dentine phosphoprotein and phosvitin was observed.