Purification and partial characterization of acid phosphatase from rice bean (Vigna umbellata Thunb.)

Acid phosphatase from rice bean (Vigna umbellata Thunb.) got isolated with 740-fold purification and specific activity 80 U/mg protein. In the native PAGE, a single protein band got observed upon silver staining. The activity staining revealed that the band corresponded to acid phosphatase. The molecular mass of the native acid phosphatase was 80 kDa. The SDS-PAGE exhibited a single protein band of molecular mass 40–41 kDa. Acid phosphatase is colorless and exhibited maximum absorbance at 278 nm but no absorption maximum at 550 nm. The enzyme retained 91.8% activity after 120 days when stored at 4 °C and improved further in the presence of polyethylene glycol (95.6%) and bovine serum albumin (97.0%) under similar storage conditions. The enzyme functioned optimally at pH 5.5 and 60 °C. The energy of activation was 32.6 kJ/mol. The values of Km and Vmax determined by the Lineweaver–Burk plot were 0.108 mM (p-nitrophenylphosphate) and 90.09 μmol/min/mg, respectively. The value of kcat and kcat/Km were 120.12 s−1 and 1.11 × 106 M−1 s−1, respectively.