Purification and Biochemical Properties of Carboxylesterase from Saga Seeds (Adenanthera pavonina)

Abstract

Carboxyl esterase (E.C.No.3.1.1.1) was partially purified from Adenanthera pavonina (Saga) using ammonium sulfate fractionation (0-60%) and DEAE (diethyl aminoethyl) ion exchange chromatography, the purified enzyme was characterized. One major saga-esterase was identified with Fold purification of 29. Molecular weight of the Ap-esterase was determined using Sephadex G-25 gel filtration and SDS-PAGE (Sodium dodecyl sulfate polyacryamide gel electrophoresis) which was found to be 26.0 k Da. Optimal activity of the saga-esterase occurred when the pH 7.0 at a temperature of 55°C. The activation energy for the hydrolysis of α-naphthyl acetate was determined to be 1.10 kcal/ mol. The Michaelis Menton constant (Km) and Vmax of the saga-esterase was 0.4µmoles and 105 IU respectively. In addition, the isoelectric point is at pH > 9 and immuno-blot using polyclonal antibodies showed that the saga-esterase was widely distributed in seeds but not in leaves. The saga-esterase inhibited by organophosphate and carbamate pesticides, which can be substituted for acetylcholinesterase.

Keywords

• carboxyl esterase
• adenanthera pavonina
• organophosphate
• carbamate pesticide