Purification and properties of alpha-amylase from Aspergillus oryzae ATCC 76080

Abstract

An alpha-amylase was purified from the solid cultural extract of Aspergillus oryzae ATCC 76080 by sequential steps of amylopectin affinity adsorption, DEAE-Sepharose ion-exchange chromatography and Sephacryl S-200 HR gel filtration. By these steps, the purity of the enzyme increased by 16 fold and recovery of the enzyme activity was 45%. The purified enzyme had an optimal pH between 4 to 5, optimal temperature at 50 degrees C and a Km value of 0.22% for hydrolysis of starch. About 80% of the enzyme activity was lost after incubation at 50 degrees C for 30 min. The heat denaturation constant at 50 degrees C was 0.024 min-1. The molecular weight was 52 kDa as determined by gel filtration. Mercuric ion (0.3 mM), DNFB# (6 mM), NBSI (6 mM) and NAI (6 mM) inhibited the activity of the enzyme. The main products for hydrolysis of maltoheptaose by the enzyme were maltotriose and maltotetraose.