敲除pknG提高谷氨酸棒杆菌L-谷氨酸和GABA产量

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-gadB1-gadB2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 pknG gene encoding serine/threonine protein kinase PknG was deleted to improve the supply of GABA precursor L-glutamate. Then the co-expression plasmid pJYW-4-gadB1-gadB2 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 pknG improves the biosynthesis of L-glutamate and GABA in recombinant C. glutamicum