L-亮氨酸生物合成关键酶基因的克隆和表达

通过PCR获得黄色短杆菌ATCC14067基因组上编码L-亮氨酸合成途径中关键酶的基因。连接大肠杆菌-谷氨酸棒状杆菌穿梭表达质粒pDXW-8构建多种重组质粒,分别转化模式菌株C.glutamicum ATCC13032考察对其发酵生产L-亮氨酸的影响。经摇瓶发酵实验显示:C.glutamicum ATCC13032发酵液中没有L-亮氨酸的积累而基因工程菌ATCC13032/pDXW-8-leuA-ilvBNC中L-亮氨酸的产量达4.75g/L。 

Cloning and expression of key enzymes genes in biosynthesis of L-leucine

The gene encoding the key enzyme in the biosynthetic pathway of L-leucine from Brevibacterium flavum ATCC14067 genome was obtained by PCR. The gene inserted into E. coli-C glutamicum shuttle expression plasmid pDXW-8 to construct a variety of recombinant plasmid, which was transformed into type strain C. glutamicum ATCC13032 and investigated its fermentation production of L-leucine. The shake flask fermentation experiment showed that L-leucine was not accumulated in the fermentation broth of the control strain, while L-leucine accumulation of 4. 75g/L was detected in the fermentation broth of the genetically engineered strain ATCC13032/pDXW-8-leuA-ilvBNC.