谷氨酸合成途径基因缺失对大肠杆菌发酵L-色氨酸的影响

Effect of deletion of genes in glutamate biosynthesis pathway on L-tryptophan fermentation by Escherichia coli

降低谷氨酸的积累可提高L-色氨酸产量及糖酸转化率。敲除Escherichia coli TRTH中的谷氨酸脱氢酶及谷氨酸合成酶编码基因gdh A、glt B,构建TRTHA(TRTH,Δgdh A)、TRTHB(TRTH,Δglt B),考察gdh A、glt B缺失对L-色氨酸发酵的影响。结果表明,gdh A及glt B缺失能有效降低谷氨酸的积累,但会降低细胞生长及色氨酸合成;培养基中谷氨酸的添加可恢复TRTHA及TRTHB的生长及色氨酸合成能力。在含1 g/L谷氨酸培养基中,利用TRTHB发酵L-色氨酸,L-色氨酸产量(41.23 g/L)及糖酸转化率(15.45%)最高,较TRTH分别提高了10.92%和7.89%;谷氨酸生成量(5.72 g/L)及乙酸积累量(1.73 g/L)分别较TRTH降低了25.23%及提高了10.19%。TRTH和TRTHB代谢流分析结果表明,glt B缺失会降低谷氨酸合成代谢流并提高乙酸合成代谢流;TRTHB的色氨酸合成代谢流(11.4%)较TRTH提高了40.74%。

The production of L-tryptophan and glucose conversion rate was increased by decreasing the accumulation of glutamate. gdh A encoding glutamate dehydrogenase and glt B encoding glutamate decarboxylase in E. coli TRTH were deleted to construct TRTHA（ TRTH,Δgdh A） and TRTHB（ TRTH,Δglt B）. The effect of deletion of gdh A and glt B on L-tryptophan fermentation was subsequently investigated. The results indicated that the disruption of gdh A or glt B decreased the excretion of glutamate as well as the cell growth and L-tryptophan biosynthesis. While the cell growth rate and L-tryptophan biosynthesis of TRTHA and TRTHB were recovered by adding glutamate in media.With L-tryptophan fermentation using TRTHB,the highest production of L-tryptophan（ 41. 23 g / L） and glucose conversion rate（ 15. 45%） were increased by 10. 92% and 7. 89%,glutamate production（ 5. 72 g / L） was decreased by25. 23% and acetate accumulation was increased by 10. 19% compared with TRTH. Analysis on metabolic flux distribution of TRTH and TRTHB showed that the flux of glutamate was decreased and the flux of acetate was increased with deletion of glt B,and the flux of tryptophan of TRTHB（ 11. 4%） was increased by 40. 74%.