不同启动子调控羰基还原酶基因在枯草芽胞杆菌中的表达水平

为了实现羰基还原酶基因mldh在枯草芽胞杆菌中的高效表达,以摩氏摩根菌MorganellamorganiiCMCC（B）49208染色体DNA为模板,PCR扩增得到目的基因mldh,分别与启动子PQ和启动子p43进行连接,构建不同启动子组合的表达载体PHY—p43-mldh、PHY—PQ—mldh、PHY—p43-p43-mldh和PHY—p43-PQ—mldh,化学法转化B．subtilisWb600后对重组茵细胞破碎液进行SDS-PAGE分析及全细胞生物转化反应实验发现,4种重组茵的转化能力差异显著,其中重组菌B．subtilisWb600（PHY—p43-p43-mldh）进行全细胞转化反应,转化液中d-伪麻黄碱的浓度最高,达到142．1mg／L,底物转化率为78．25％,成功实现了羰基还原酶基因mldh在枯草芽胞杆菌中的高效表达。

Expression of carbonyl reductase in Bacillus subtilis with different promoters

In order to achieve its high expression in Bacillus subtilis, carbonyl reductase gene mldh was amplified by PCR with the template of chromosome DNA from Morganella morganii CMCC （ B ） 49208 and connected with promoter PQ and promoter p43 to obtain different expression plasmids PHY-p43- mldh, PHY-PQ-mldh, PHY-p43-p43-mldh, and PHY-p43-PQ-mldh. Subsequently, these recombined vectors were transformed into B. subtilis Wb600. SDS-PAGE analysis of recombinants. And whole cellular biotransformation showed that there was distinctive difference among the four recombinants, in which the B. subtilis Wb600 （PHY-p43-1M3-mldh） could obtain the highest concentration （ 142. 1 mg/L） of d- pseudoephedrine in conversion solution and the mole ratio of substrate was 78.25%.