Bacillus licheniformis6816碱性蛋白酶基因在E.coli中的克隆和表达

用PCR方法从地衣芽孢杆菌6816中扩增了碱性蛋白酶基因（apr)，扩增的1.14kb的DNA片段插入到大肠杆菌载体pET-20b中，构建成重组分泌型表达载体pAPR1。pAPR1中碱性蛋白酶基因在大肠杆菌宿主JM109（DE3）中得到表达，SDS-PAGE分析显示融合表达产物的分子量为30kD，同核酸序列测定所推导的值相符，表达产物占细胞总蛋白的7.5%，重组菌的酶活比出发菌株提高了3.3倍，研究发现，重组的碱性蛋白酶在进入大肠杆菌周质空间时存在前肽自动脱落的现象。

Cloning and expression of the gene encoding alkaline protease from Bacillus licheniformis in E.coli

A alkaline protease gene ( apr ) was amplified from Bacillus licheniformis 6816by using PCR technique. The amplified 1.14 kb DNA fragment was inserted into an E.coli vector PET-20b to yield the recombinant secrete plasmid pAPR1.The apr in pAPR1 was expressed in E.coli JM109(DE3).6816.SDS-PAGE analysis show the molecular weight expression recombinant product is about 30KD,which corresponded with prediction by gene sequence. The expressed recombinant protein accounted for 7.5% of total cell protein. The activity of alkaline protease of recombinant stain is 3.3 times higher than that of original stain B.lichenformis .The study also discovered while the recombinant alkaline protease secreted into the periplasmic space of E.coli ,the pro-squence of alkaling protease might be cleaved by a self-processing mechanism.