Purification and biochemical properties of a salivary α-amylase in Andrallus spinidens Fabricius (Hemiptera: Pentatomidae)

Abstract

α-amylase is one of the enzymes that has crucial role in extra-oral digestion (EOD) of hemipteran insects. An α-amylase was purified and biochemically characterized from the salivary glands of Andrallus spinidens showing its considerable role in EOD process. It was found an enzyme by specific activity of 4.22 U/mg protein, recovery of 14.67 % and purification fold of 13.83-fold as well as molecular weight of 26 kDa. By using two buffer solutions, optimal pH of the purified α-amylase was found to be 9 for both universal and Tris-HCl buffers. Our findings revealed that the purified α-amylase had the highest activity at the temperatures of 35 and 40 °C, and were stable for 96 h at these temperatures. Kinetic parameters of the purified enzyme show that both starch and glycogen, are the suitable substrates for the enzymatic assay, but a lower Km demonstrated glycogen as a more appropriate substrate. Among the cations used to show their possible involvement in active site of the enzyme, Ca2+ 2+ , Mg and one concentration of Cu2+ increased the α-amylase activity but Na+ decreased the enzyme activity. Triton X-100 increased the enzyme activity but SDS, EDTA, EGTA and TTHA decreased it, indicating involvement of metal ions in the active site of the purified α-amylase.