Three Different Types of α-Amylases from Aspergillus awamori KT-11: Their Purifications, Properties, and Specificities

Abstract

  Three forms of α-amylases, designated Amyl I, Amyl II, and Amyl III were purified to a homogenous state by several column chromatographies from a koji culture in wheat bran of a strain of black mold, which was isolated in Indonesia and identified as Aspergillus awamori. They have molecular weights of 49,000, 63,000, and 97,000 by SDS-PAGE, respectively, and the optimum pHs were 4.0 for Amyl I and 5.5 for both Amyl II and Amyl III on soluble starch. Amyl I hydrolyzed malto-tetraose, -pentaose, -hexaose, -heptaose, and β- and γ-cyclodextrin to produce maltose and maltotriose as major products but not or hardly hydrolyzed maltose, isomaltose, maltotriose, isomaltotriose, α-cyclodextrin, or raw corn starch. On the other hand, both Amyl II and Amyl III hydrolyzed maltotriose as well as all the maltooligosaccharides described above and α-, β-, and γ-cyclodextrin, and even raw corn starch as well as heat-gelatinized corn starch to produce maltose as a major product and glucose and maltotriose as minor products, but they did not hydrolyze maltose, isomaltose, or isomaltotriose. The limit hydrolyses of soluble starch with three kinds of enzymes were 33% for Amyl I, 35% for Amyl II, and 38% for Amyl III, the reaction products had α-anomeric forms by NMR analysis, and the blue color reaction with I₂ disappeared completely at about 18% of hydrolysis of the starch for all enzymes.