赖氨酸-尸胺反向转运蛋白cadB基因谷氨酸棒杆菌表达载体的构建及转化  

Construction and Transformation of Corynebacterium glutamicum Expression Vector of Cadaverine-lysine Antiporter cadB Gene

作　　者：黄云雁[1] 黎明[1] 刘萌[1] 孙昕[1] 周丽颖[1] 路福平[1] 

机构地区：[1]天津科技大学生物工程学院、工业发酵微生物教育部重点实验室、工业酶国家工程实验室、天津市工业微生物重点实验室,天津300457

出　　处：《生物技术通报》2012年第8期94-100,共7页Biotechnology Bulletin

基　　金：国家自然科学基金项目(21176910);天津市科技支撑计划项目(11ZCKFSY00900);天津市应用基础及前沿技术研究计划项目(11JCYBJC 09600);国家"863"项目(2007AA02Z212)

摘　　要：旨在提高谷氨酸棒杆菌合成尸胺的能力,将CadB克隆至谷氨酸棒杆菌中,与LDC共表达,在谷氨酸棒杆菌合成尸胺的同时,帮助尸胺转运至细胞外,解除尸胺的反馈抑制作用。谷氨酸棒杆菌能够高产赖氨酸脱羧酶的底物L-赖氨酸,但不含ldc和cadB基因,因而不能够直接合成尸胺。从E.coliK12中克隆出赖氨酸-尸胺反向转运蛋白基因,与绿色荧光蛋白基因gfp融合构建成融合表达载体pXBG,并转化至谷氨酸棒杆菌进行诱导表达,结果表明表达的CadB蛋白可以正确的定位于谷氨酸棒杆菌的细胞膜上。将基因cadB连接到含有赖氨酸脱羧酶基因的pXMJ19-ldc上,构建成能够共表达赖氨酸脱羧酶和赖氨酸-尸胺反向转运蛋白的重组质粒pXLB,并转化到谷氨酸棒杆菌中。In order to improve the ability of the synthesis of cadavcrine by Corynebacterium glutamicum, CadB was cloned into the bacteria and coexpressed with LDC for transporting cadaverine to extracellular area and reducing the feedback inhibition. C. glutamicum can produce lysine, but can not be able to synthesize cadaverine directly without lysine decarboxylase gene and cadaverine-lysine antiporter gene. In our previous job, cadaverine-lysine antiporter gene cadB which was proliferated by polymerase chain reaction by using chromosomal DNA of E. coli K12 as the template was fused to gfp to construct pXBG. Then the recombinant plasmid pXBG was electrotransferred to C. glutamicum ATCC13032. The result indicated that CadB can express in C. glutamicum ATCC13032 and it located in the cell membrance. The recombinant expression vector pXLB containing ldc and cadB gene was constructed and transferred to C. glutamicum ATCC13032. Construction of co- expression vector pXLB is important for producing cadaverine by Corynebacterium glutamicum.

关 键 词：尸胺 赖氨酸-尸胺反向转运蛋白CadB 谷氨酸棒杆菌 赖氨酸脱羧酶 共表达 

分 类 号：Q78[生物学—分子生物学]