Purification and Characterization of Glucoamylase From Aspergillus Niger ATCC 10864.

The Aspergillus niger ATCC 10864 produced substantial amount of glucoamylase activity when grown on rice bran as carbon source. The glucoamylase was purified using a procedure that included 80% saturated ammonium sulface precipitation dialysis, chromatography on Sephacryl S-100 and DEAE-Sepharose. Two fractions of glucoamylase from DEAE-Sepharose (F1a and F2a) were obtained. The purification fold were 26.24 and 19.57 times with 28 and 14.56 percent yields, the specific activities of 842.20 and 628.37 U/mg, and the molecular weight with estimated molecular mass of 80kDa and 73 kda, respectively. The optimal pH and temperature of F1a and F2a were 4.0, 60°C and 5.5, 60°C, respectively. F1 was stable at a pH range of 4.0 to 5.5 and at a temperature range of 20 to 50°C, while F2a was stable at a pH range of 4 to 6 and a temperature range of 20 to 40°C. In the presence of rice starch in fresh culture medium, the purified fractions of glucoamylase, F1a and F2a demonstrated apparent K_m and V_max values of 10 mg/ml, 0.02 umol/ml/min and 5.6 mg/mL, 0.032 umol/ml/min, respectively, while in the presence of soluble starch, the values of apparent K_m and V_max of F1 and F2 were 6.7 mg/ml, 0.02 umol/ml/min and 5.0 mg/ml 0.023 umol/ml/min, respectively.

Keywords: glucoamylase, purification, characterization, Km, Vmax

Corresponding Author Email: kkaree@kmitl.ac.th