Purification and properties of acid phosphatases from axes and cotyledons of germinating soybeans

Acid phosphatases (EC 3.1.3.2) from the axis and cotyledon of germinating soybeans (Glycine max L.) were purified approximately 700-fold and 2100-fold, respectively, to electrophoretic homogeneity by ammonium sulfate fractionation, cation-exchange, concanavalin A-Sepharose 4B affinity, and hydroxyapatite chromatographies. This report describes the purification and characterization of the enzymes from both axis and cotyledon. Acid phosphatases from both organs had the following similar properties, although they have some differences in substrate specificity. (i) The enzyme was a glycoprotein. (ii) The native enzyme of approximate molecular weight of 100,000 consisted of two subunits, each with an apparent molecular weight of 50,000. (iii) The optimal pH was approximately 5.7–5.8, and the enzyme was stable at the pH range of 5.3–8.0. (iv) The apparent Km value with p-nitrophenyl phosphate as a substrate was 3.7–4.0 × 10⁻⁴ m. (v) The enzyme activity was inhibited by Cu²⁺, Zn²⁺, Pb²⁺, Mo₇O₂₄⁶⁻, F⁻, and PO₄³⁻ ions. (vi) The enzyme hydrolyzed various phosphorylated compounds non-specifically. (vii) The antiserum against the enzyme from axis cross-reacted to that from cotyledon.