Abstract:
Glucoamylase, a starch degrading enzyme was optimally produced from two species of Aspergillus (Aspergillus flavus and Aspergillus fumigatus) obtained from the soil of a cassava processing site with a view to biochemically characterize the enzyme for industrial use. The enzymes were purified to homogeneity by fractional ammonium sulphate precipitation (40 – 60 %), ion-exchange chromatography on CM-Cellulose and gel filtration on Sephadex G-50 giving 24-fold purification with 8.96% recovery and 20-fold purification with 8.19% recovery respectively for glucoamylase from Aspergillus flavus CFA-1 (CfaGA) and Aspergillus fumigatus CFU-1 (CfuGA). Sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS–PAGE) revealed the molecular weights of the purified enzymes to be 63 kDa and 50 kDa respectively after gel filtration. The purified glucoamylases were both highly active and stable over a wide pH range of 4 to 8, with optimum activity being observed at pH 5.5 for both enzymes. The purified enzymes, CfaGA and CfuGA were both relatively thermostable; respectively retaining about 62% and 67% of initial activity when incubated at 60 °C for 1 hour. The enzymes were highly active on different starches, dextran and amylose but showed low affinity for sucrose. The apparent Km and Vmax of CfaGA were 0.89 mg/mL and 17.30 μmol/min/mL respectively while 1.64 mg/mL and 20.96 μmol/min/mL were the respective Km and Vmax values obtained for CfuGA when soluble starch was used as substrate in the enzyme – substrate reaction carried out at pH 5.5 and 60 °C. The purified glucoamylases did not require calcium and displayed extreme stability with regard to surfactants and denaturing agents. The chelating agent, EDTA, had minimal effect on the activity and stability of the enzymes. Glucoamylases with these unique characteristics have not been previously reported from both species of Aspergillus and as such could be employed in different
starch-utilizing industries, especially in food, textile, pharmaceuticals and biofuel industries for various biotechnological applications
