摘要
从谷氨酸棒杆菌模式菌株C.glutamicumATCC13032中克隆出L-缬氨酸合成途径上的限速酶——乙酰羟酸合酶编码基因ilvBN。对ilvBN进行定点突变,获得其抗反馈抑制突变型ilvBNr。以大肠杆菌-黄色短杆菌穿梭表达载体pDXW-10为基础,构建重组质粒pDXW-10-ilvBNr,并转化野生型黄色短杆菌B.flavumATCC14067,获得工程菌株ATCC14067/pDXW-10-ilvBNr。3L罐发酵实验结果显示:在野生型菌株发酵液中检测不到L-缬氨酸积累,而工程菌株发酵液中L-缬氨酸积累达5.0g/L。
As a rate-limiting enzyme for L-valine biosynthesis, the ilvBN gene encoding acetyl-carboxylic acid synthase （AHAS） from Corynebacterium glutamicum ATCC13032 was amplified by PCR, followed by site-direct mutagenesis to obtain an ilvBNr gene, the anti-feedback inhibition gene of ilvBN. The ilvBNr gene inserted into E. coli-Brevibacterium flavum shuttle expression vector pDXW-10 to construct a recombinant plasmid pDXW-10-ilvBNr, which was subsequently transformed into B. flavum ATCC14067, producing a genetic engineering strain ATCC14067/pDXW-10-ilvBNr. The fermentation experiments conducted in a 3 L fermentor showed that no L-valine accumulation was detected in the fermentation broth of the original strain, while an L-valine accumulation of 5.0 g/L was observed in the fermentation broth of the constructed engineering strain.
出处
《食品科学》
EI
CAS
CSCD
北大核心
2010年第23期262-266,共5页
Food Science
基金
国家"863"计划项目(2007AA02Z229
2007AA02Z230)
关键词
乙酰羟酸合酶
抗反馈抑制
黄色短杆菌
发酵
L-缬氨酸
acetyl-carboxylic acid synthase
anti-feedback inhibition
Brevibacterium flavum
fermentation
L-valine
作者简介
作者简介：徐大庆（1972-），男，博士研究生，研究方向为微生物代谢工程。E—mail：daqingxu@yahoo．com．cn。
通信作者：王小元（1964-），男，教授，博士，研究方向为食品安全及微生物代谢工程。E-mail：xiaoyuanwang@hotmail．com.
