Champreda Verawat, Kanokratana Pattanop, Sriprang Rutchadaporn, Tanapongpipat Sutipa, Eurwilaichitr Lily
Molecular and Enzyme Screening Laboratory, BIOTEC Central Research Unit, National Center for Genetic Engineering and Biotechnology, Klong Luang, Patumthani, Thailand.
Biosci Biotechnol Biochem. 2007 Aug;71(8):2010-20. doi: 10.1271/bbb.70198. Epub 2007 Aug 7.
An endophytic fungus, Fusicoccum sp. BCC4124, showed strong amylolytic activity when cultivated on multi-enzyme induction enriched medium and agro-industry substrates. alpha-Amylase and alpha-glucosidase activities were highly induced in the presence of maltose and starch. The purified target alpha-amylase, Amy-FC1, showed strong hydrolytic activity on soluble starch (kcat/Km=6.47 x 10(3) min(-1)(ml/mg)) and selective activity on gamma- and beta-cyclodextrins, but not on alpha-cyclodextrin. The enzyme worked optimally at 70 degrees C in a neutral pH range with t(1/2) of 240 min in the presence of Ca(2+) and starch. Maltose, matotriose, and maltotetraose were the major products from starch hydrolysis but prolonged reaction led to the production of glucose, maltose, and maltotriose from starch, cyclodextrins, and maltooligosaccharides (G3-G7). The amylase showed remarkable glucose tolerance up to 1 M, but was more sensitive to inhibition by maltose. The deduced protein primary structure from the putative gene revealed that the enzyme shared moderate homology between alpha-amylases from Aspergilli and Lipomyces sp. This thermotolerant, glucose tolerant maltooligosaccharide-forming alpha-amylase is potent for biotechnological application.
一种内生真菌，Fusicoccum sp. BCC4124，在多酶诱导富集培养基和农业工业底物上培养时表现出很强的淀粉分解活性。在麦芽糖和淀粉存在的情况下，α-淀粉酶和α-葡萄糖苷酶的活性被高度诱导。纯化后的目标α-淀粉酶Amy-FC1对可溶性淀粉表现出很强的水解活性（kcat/Km = 6.47×10³ min⁻¹(ml/mg)），对γ-和β-环糊精具有选择性活性，但对α-环糊精无活性。该酶在70℃、中性pH范围内活性最佳，在Ca²⁺和淀粉存在的情况下，半衰期为240分钟。麦芽糖、麦芽三糖和麦芽四糖是淀粉水解的主要产物，但延长反应时间会导致淀粉、环糊精和麦芽寡糖（G3 - G7）产生葡萄糖、麦芽糖和麦芽三糖。该淀粉酶在高达1 M的葡萄糖浓度下表现出显著的葡萄糖耐受性，但对麦芽糖抑制更敏感。从推定基因推导的蛋白质一级结构表明，该酶与曲霉属和油脂酵母属的α-淀粉酶具有中等同源性。这种耐热、耐葡萄糖的麦芽寡糖形成型α-淀粉酶在生物技术应用中具有潜力。
