α-淀粉酶在不同地衣芽孢杆菌宿主菌中分泌表达的对比分析

Comparative Analysis on the Effects of Different Host Strains of Bacillus licheniformis on the Expression and Secretion of α-Amylase

研究不同地衣芽孢杆菌(Bacillus licheniformis)宿主菌对α-淀粉酶分泌表达的影响。以Bacillus subtilis的P43启动子,B.licheniformis WX-02α-淀粉酶基因的信号肽、编码区和终止子序列为表达原件,构建了α-淀粉酶分泌表达载体pP43SAT。将pP43SAT分别转入3株B.licheniformis宿主菌:WX-02(ΔamyL)、BL9和BL10,获得3株α-淀粉酶基因工程菌WX-02(ΔamyL,pP43SAT)、BL9(pP43SAT)和BL10(pP43SAT),并将构建的3株工程菌进行发酵对比分析。BL9(pP43SAT)和BL10(pP43SAT)的淀粉酶发酵活力分别达到94.01 U/mL和101.94 U/mL,比原始宿主菌WX-02(ΔamyL,pP43SAT)分别提高了21%和31%,这说明BL9和BL10新型宿主菌有利于淀粉酶的分泌表达。本研究证明多蛋白酶基因缺失可显著提高α-淀粉酶的表达,为α-淀粉酶的高效表达提供了新型宿主菌和新策略。

In the present study, effects of different host strains of Bacillus licheniformis on the expression and secretion of a-amylase were investigated. Firstly, the a-amylase expression vector, named pP43SAT, was constructed based on the promoter P43 from Bacillus subtilis and the signal peptide, coding region and terminator of α-amylase gene from B. licheniformis WX-02. The pP43SAT plasmids were then transformed into the B. licheniformis host strains WX-02 （ A amyL）, BL9 and BL10B, respectively, to obtain the a-amylase genetic engineering strains WX-02 （ A amyL; pP43SAT）, BL9 （pP43SAT） and BL10 （pP43SAT）, respectively. The α-amylase fermentation processes of these engineered strains were compared. The a-amylase activities of strains BL9 （pP43SAT） and BL10 （pP43SAT） reached 94.01 and 101.94 U/mL, respectively, which were increased by 21% and 31%, respectively, as compared with that of WX-02 （△ amyL; pP43SAT）. These results showed that the novel host strains BL9 and BL10 contributed to the secretion and expression of α-amylase. It was proved that the deletion of multiple proteases genes could enhance α-amylase expression obviously, and this study provided the novel host strains and novel strategies for efficient expression of α-amylase.