从嗜热真菌樟绒枝霉(Malbranchea cinnamomea)中克隆α-淀粉酶基因McAmyA,并在毕赤酵母GS115中高效表达,经高密度发酵至168 h时,胞外酶活力达到13 440.6 U/mL。重组α-淀粉酶McAmyA粗酶液经QSFF强阴离子交换层析纯化得到电泳级纯酶,比酶活力为1 230.2 U/mg。酶学性质研究表明,重组α-淀粉酶McAmyA的最适pH和最适温度分别是6.5和65 ℃。以淀粉液化液为底物,在温度60 ℃,加酶量120 U/g,水解24 h的条件下,重组α-淀粉酶McAmyA水解液化液,制备得到麦芽糖含量为50.0%(质量分数)的麦芽糖浆。该真菌α-淀粉酶在毕赤酵母中表达水平高,具有很大的应用潜力。
The α-amylase gene (McAmyA) from Malbranchea cinnamomea was successfully cloned and heterologously expressed in Pichia pastoris GS115. The recombinant α-amylase (McAmyA) was secreted at high level of 13440.6 U/mL by high cell density fermentation after 168 h. McAmyA was purified by QSFF ion-exchange chromatography with specific activity of 1230.2 U/mg. The optimal pH and temperature of McAmyA were determined to be 6.5 and 65 ℃, respectively. The maltose syrup with maltose content of 50% was prepared at 60 ℃, 120 U/g and 24 h. The fungus α-amylase was highly expressed in Pichia pastoris and it has great potential for application.
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