Purification and Characterization of the Commercialized, Cloned Bacillus megaterium α‐Amylase. Part I: Purification and Hydrolytic Properties

Abstract

An amylolytic enzyme, originally isolated from Bacillus megaterium. was shown to increase the maximum amount of dextrose produced during saccharification by decreasing the amount of residual oligosaccharides. The enzyme, now commercially produced in a genetically engineered strain of Bacillus subtilis, was purified to homogeneity from the commercial product. A combination of gel permeation chromatography in the presence of 1.0 M NaCl and chromatofocusing between pH 9.0 and 7.0 were used to obtain the pure enzyme. The molecular weight of the Bacillus megaterium α‐amylase was 59,000 by SDS gel electrophoresis, and the isoelectric point was 8.9 to 9.0. The enzyme was shown to possess both hydrolytic and transferase activity. The enzyme hydrolyzed a wide variety of soluble substrates. The rate of hydrolysis was greatest on soluble starch; α(1,6)‐branched substrates and cyclodextrins were hydrolyzed more slowly.