Purification and characterization of digestive amylase from the tasar silkworm, Antheraea mylitta (Lepidoptera: Saturniidae)

Abstract

Digestive amylase was purified from larvae of Indian tasar silkworm, Antheraea mylitta using ammonium sulphate precipitation, glycogen complex precipitation and gel filtration chromatography. Specific activity increased from 0.673 AU/mg in the crude digestive juice to 94.80 AU/mg in the final purified sample. Activity of the purified enzyme was 15-fold less than that of the digestive amylase of silkworm. Bombyx mori. The zymogram pattern of the purified amylase was similar to that of crude digestive juice on 7.5% native PAGE. The purified enzyme exhibited five bands on native PAGE. IEF of the purified enzyme also revealed five bands with pls of 6.5, 6.15, 5.9, 5.8 and 4.7, respectively. The purified enzyme is a single polypeptide chain with a M r of 58 kDa. The amylase is most active at pH 9.5 and is a Ca 2+ dependent endoenzyme which hydrolyses starch into maltose, maltotriose and maltotetrose and hence behaves as an α-amylase (EC 3.2.1.1). The enzyme was unaffected by the presence or absence of CI − , with K m for soluble starch of 0.113%.