敲除pknG提高谷氨酸棒杆菌L-谷氨酸和GABA产量  

Deletion of pknG Improves the Production of L-Glutamate and GABA in Recombinant Corynbacterium glutamicum

作　　者：王楠楠[1,2,3] 倪亚兰[1,2,3] 史锋[1,2,3] 

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]江南大学工业生物技术教育部重点实验室,江苏无锡214122 [3]江南大学食品安全与营养协同创新中心,江苏无锡214122

出　　处：《食品与生物技术学报》2017年第2期187-193,共7页Journal of Food Science and Biotechnology

基　　金：食品科学与技术国家重点实验室自主科研课题项目(SKLF-ZZB-201405)

摘　　要：γ-氨基丁酸(GABA)具有多种生理功能,被广泛应用于食品、医药等行业。在谷氨酸棒杆菌ATCC13032中表达来自短乳杆菌的两个谷氨酸脱羧酶(GAD)编码基因gadB1、gadB2,可将其自身积累的L-谷氨酸有效转变成GABA。为了进一步提高GABA产量,首先在ATCC13032中敲除了丝氨酸/苏氨酸蛋白激酶PknG的编码基因pknG,以提高GABA的前体物质L-谷氨酸的供应,然后将GAD表达质粒pJYW-4-gad B1-gad B2转入pknG敲除菌株和ATCC13032中,构建出重组菌SNW203和SNW200,最后对两个重组菌进行上罐发酵。结果表明:相对于SNW200,菌株SNW203的L-谷氨酸和GABA产量都有所提高。发酵72 h后,GABA产量为(30.2±0.3)g/L,相对于SNW200提高了55.4%,L-谷氨酸和GABA的总摩尔浓度为0.3 mol/L,提高了36.4%。这说明敲除pknG能够促进重组谷氨酸棒杆菌的L-谷氨酸和GABA生物合成。Gama（γ）-aminobutyric acid（GABA）,which has a variety of physiological functions,is widely used in food,pharmaceutical and other industries.Two L-glutamate decarboxylase（GAD）genes（gadB1 and gadB2）were co-expressed previously in an L-glutamate producing strain Corynebacterium glutamicum ATCC13032,making the own accumulated L-glutamate be effectively transformed into GABA.To further enhance GABA production,the pkn G gene encoding serine/threonine protein kinase PknG was deleted to improve the supply of GABA precursor L-glutamate.Then the co-expression plasmid p JYW-4-gad B1-gad B2 was transformed into pknG deletion strain and ATCC13032,generating the recombinant C.glutamicum strains SNW203 and SNW200.After the strains were fermented in fermenters,the production of both L-glutamate and GABA in SNW203 increased greatly when compared with SNW200.At 72 h of fermentation,GABA production in SNW203 increased to（30.2±0.3）g/L,55.4%higher than that in SNW200 and the total concentration of L-glutamate and GABA reached up to 0.3 mol/L,36.4%higher than that in SNW200.This result indicates that the deletion of pkn G improves the biosynthesis of L-glutamate and GABA in recombinant C.glutamicum.

关 键 词：Γ-氨基丁酸 谷氨酸棒杆菌 pknG基因 

分 类 号：TQ922[轻工技术与工程—发酵工程]