敲除iclR基因对大肠杆菌发酵L–色氨酸的影响

Effects of Knockout of Gene icl R on L-tryptophan Fermentation in Escherichia coli

为了研究敲除icl R基因对大肠杆菌(Escherichia coli)发酵L–色氨酸的影响,以L–色氨酸工程菌大肠杆菌TRTH为出发菌株,利用Red重组技术构建了icl R基因(编码乙醛酸操纵子阻遏蛋白)缺失菌株TRTHΔicl R.摇瓶发酵实验结果显示:TRTH icl R的L–色氨酸产量和糖酸转化率分别达到(6.52±0.46)g/L和13.17%,,比原菌的分别提高了21.86%,和22.85%,;乙酸累积量为6.82,g/L,比原菌的降低了37.63%.30,L发酵罐发酵实验结果显示:TRTH icl R的L–色氨酸产量及糖酸转化率分别达到(13.01±1.05)g/L和6.51%,,比原菌的下降了60.34%,和68.27%,;乙酸累积量为18.21,g/L,比原菌的增加了33.42%.结果表明:在摇瓶条件下,重组菌株生物量较出发菌株高,代谢流分配适合L–色氨酸积累;但在发酵罐条件下,乙醛酸循环的增强导致重组菌株供能不足和乙酸的过多积累,最终使得生物量不足以及L–色氨酸产量下降.

Escherichia coli TRTH ΔiclRwas constructed via knockout oficlR,the key gene of the glyoxylate shunt in E.coliTRTH through Red recombination to study the effects oficlR knockout onE.coliTRTHL-tryptophan accumula-tion.The results of shake flask fermentation indicated that the yield ofL-tryptophan and the titer ofL-tryptophan from glu-cose ofE.coli TRTH ΔiclRreached(6.52±0.46)g/L and 13.17%,,which were 21.86%, and 22.85%, higher than those ob-tained fromE.coli TRTH,respectively;6.82,g/L of acetic acid accumulated in the culture ofE.coli TRTH ΔiclR,which was 37.63%, lower than that of the control strain.In the fermentation of 30,L fermenter,the titer and yield ofL-tryptophan of E.coli TRTH ΔiclR were(13.01±1.05)g/L and 6.51%,,which were 60.34%, and 68.27%, lower than those of the control, respectively;18.21,g/L acetic acid,33.42%, times higher than that of the control,accumulated in the broth.In conclusion,in shake flask fermentation assays,the metabolic flux of the recombined strain was beneficial forL-tryptophan accumulation although the biomass was lower thanE.coli TRTH.On the contrary,enhancement of glyoxylate shunt resulted in the short-age of energy supply and higher acetic acid accumulation,which can lead to lower biomass andL-tryptophan titer in 30,L fermentation.