乳糖发酵短杆菌lysC突变对L-赖氨酸积累的影响  

The Effect of LysC on L-Lysine Accumulation in Brevibacterium lactofermentum

作　　者：董迅衍[1] 徐大庆[1] 李烨 李江华[2] 王小元[1] 

机构地区：[1]食品科学与技术国家重点实验室,江南大学,江苏无锡214122 [2]江南大学生物工程学院,江苏无锡214122

出　　处：《食品与生物技术学报》2011年第4期592-596,共5页Journal of Food Science and Biotechnology

基　　金：国家863计划项目(2007AA02Z229和2007AA02Z230)

摘　　要：从一株乳糖发酵短杆菌L-异亮氨酸生产菌中克隆出天冬氨酸激酶AK-1的编码基因ly-sC1,经DNA测序并与来自谷氨酸棒杆菌ATCC13032的野生型lysC比对发现其中有2个核苷酸1186G和1187C缺失,造成翻译提前终止。AK-1还有下列2个有效突变位点:Ala 279 Thr和Ser317 Ala。lysC1在大肠杆菌BL21中的诱导表达量约为lysC的1/4,其表观比酶活也较低,但对L-苏氨酸和L-赖氨酸协同反馈抑制不敏感。用大肠杆菌-黄色短杆菌穿梭表达载体pDXW-8将ly-sC1在野生型乳糖发酵短杆菌ATCC13869中诱导型表达,经摇瓶发酵积累L-赖氨酸7.4 g/L,在3 L发酵罐上达40 g/L。The gene lysC1 encoding aspartate kinase AK-1 from a Brevibacterium lactofermentum L-isoleucine producer was sequenced.Compared with the lysC from Corynebacterium glutamicum ATCC13032,there were deletions of two nucleotides 1186G and1187C,leading to the pre-termination of translation.In addition,the following two mutated sites were also identified in lysC1： Ala 279 Thr and Ser 317 Ala.In Escherichia coli BL21,the expression level of lysC1 was lower than that of lysC from B.lactofermentum ATCC13869.The AK-1 had a lower apparent value of specific activity than the wild type AK,but it was not sensitive to the feedback inhibition by the mixture of L-lysine and L-threonine.lysC1 was cloned into an E.coli-Brevibacterium flavum shuttle expression vector pDXW-8 and transformed into wild type B.lactofermentum ATCC13869.The recombinant strain ATCC13869/pDXW-8-lysC1 accumulated 7.5 g/L L-lysine after 90 h flask fermentation and 40 g/L after 72 h batch fermentation.These results would be very useful for constructing a genetically identified L-lysine producing strain with minimum genetic alterations.

关 键 词：乳糖发酵短杆菌 天冬氨酸激酶 抗反馈抑制 发酵 L-赖氨酸 

分 类 号：TQ922.3[轻工技术与工程—发酵工程]