Purification and properties of ascorbate oxidase from cucumber fruit

Ascorbate oxidase [AOD, EC 1.10.3.3], a copper-containing enzyme, was purified to homogeneity from cucumber fruit (Cucumis sativus) by CM-Sephadex and hydroxyapatite column chromatography and chromatofocusing. Isoelectric point (pI) of the purified enzyme was 8.3. Two other isoforms of AOD with alkaline pI values were also found. Purified AOD (MW 150,000) was a glycoprotein composed of two identical subunits (MW 70,000). N-terminal amino acid sequence was determined as Gly-Phe-Pro-Lys-Ile-Lys-His-Tyr-Lys-Trp-Asp-Val-Glu-Tyr-Met-Phe.