毕赤酵母中植酸酶和内切葡聚糖酶共表达菌株的构建及其高效表达

Construction of a Pichia pastoris recombinant strain capable of over-expressing phytase and endoglucanase

摘要:

植酸酶和内切葡萄糖苷酶广泛应用于动物饲料添加剂中，能更有效地帮助饲料的利用，同时为动物的生命活动提供必需的重要物质。首先构建重组质粒pPICZα-EG，经过线性化后，电转化到含有植酸酶phyA基因的毕赤酵母感受态细胞中，构成含有植酸酶基因和内切葡萄糖苷酶基因的重组体菌株GS115-phyA-EG。经甲醇诱导后其活性分别达到出发菌株GS115-phyA和GS115-EG的39.4％和56.2％。酶学性质的分析显示，最适反应温度和最适反应的pH值分别为55℃和5.5，植酸酶和内切葡萄糖苷酶在温度为45℃~55℃，pH值为4.5~5.5时，酶活相对稳定，均能达到最高酶活的80％以上。结果表明这种在同一个系统种同时表达2种酶的方法能更好地节约时间和成本，使其在饲料工业的应用上有更广阔的前景。

Abstract:

Both phytase and endoglucanase are additives in feed for mono-gastric animal known for their effects. Recombinant vector pPICZα-EG was constructed and transformed to GS115-phyA, a Pichia pastoris strain that had integrated with phytase gene, generating GS115-phyA-EG. Both phytase and endoglucanase activities in the supernatant were determined after methanol induction of GS115-phyA-EG. Phytase and endoglucanase activity. reached 39.4% and 56.2% activity compared to GS115-phyA and GS115-EG, respectively. Properties of the mixed enzyme suggest that the optimal temperature and pH value be 55°C and 5.5 respectively. Both phytase and endoglucanase showed greater than 80% activity across temperature ranges 45°C to 55°C and pH ranges 4.5 to 5.5. Expressing more than one enzyme in one system could save time and money during induced expression, and the mixed enzyme might apply for treating forge before feeding with poultry.