α-淀粉酶和糖化酶在酿酒酵母中的表达和分泌

Expression and Secretion of oc-amylase and Glucoamylase in Saccharomyces cerevisiae

将地衣芽孢杆菌α-淀粉酶基因及黑曲霉糖化酶cDNA重组进大肠杆菌-酵母穿梭质粒,转化酿酒酵母,构建能分解淀粉的酵母工程菌。酶活力测定和酶学性质分析的结果显示:在酵母MF-α1因子及磷酸甘油酸激酶基因的启动子和终止信号的调控下,α-淀粉酶和糖化酶基因在酵母中获得高表达并向胞外分泌这两种酶。构建的酵母工程菌在含10％淀粉的培养基中6天内能水解97％的淀粉,重组质粒能在酵母中较稳定地存在。

a-amylase gene of Bacillus licheniformis and glucoamylase cDNA of Aspergil-lus niger were ligated to a E. coli-yeast shuttle vector. The resultant plasmid was used to transform Saccharomyces cerevisiae to construct starch-degrading yeast strain. The results qf enzyme activity assay and enzyme property analysis show that a-amylase and glucoamylase genes have been expressed simultaeously in yeast under the control of promoters and terminators of yeast MF-a 1 factor and PGK genes and over 99% of enzyme activities were secreted to the medium. The engineered yeast stratin hydrolyzes 97% of the starch (10%) in the medium. The recombinant plasmid exists stably in yeast.