Characterization and Optimization of Amylase Production in WangLB, a High Amylase-Producing Strain of Bacillus.

Abstract

The costs of amylase represent ca. 24% of the expenditures in the starch industry and an increase in amylase production and/or activity will greatly cut down on production costs. In the present study, we obtained a high amylase-producing strain of bacteria, WangLB, and identified it as a member of the Bacillus genus based on 16S rDNA analysis. The fermentation conditions for amylase production in the strain were optimized, and the maximum amylase activity we obtained was 26,670 ± 1390U/mL, under the optimized conditions of 48-h incubation in liquid starch medium, 35°C, pH 10, 1% v/v inoculum concentration, 20g/L starch concentration, and 0.1% w/v peptone. The influences of 16 small organic inducers on amylase production were tested, and the results showed that 20mmol/L alanine greatly enhanced amylase production to 290% of the baseline level. We also conducted an amylase enzymology analysis. The molecular weight of the amylase was 55kD, determined by SDS-PAGE. The optimum temperature and pH for the amylase were 55°C and pH 9, respectively. The enzyme also showed high activity over a wide range of temperatures (50-85°C) and pH values (3-10), and the activity of the amylase was Ca(2+) independent. The kinetic parameters K m and V max were 0.37 ± 0.02mg/mL and 233 U/mg, respectively. Finally, the amylase was applied to the hydrolysis of five different brands of starch. It was found that the hydrolyzability of the substrate by amylase increased along with starch solubility.