重组大肠杆菌合成L-甲硫氨酸的发酵过程优化

Optimization of the fermentative production of L-methionine by engineered Escherichia coli

L-甲硫氨酸作为唯一含有硫元素的必需氨基酸,在生物体内具有非常重要的生理和生化功能,而微生物发酵法合成L-甲硫氨酸因其发酵水平较低仍未达到工业化生产要求。本文在实验室前期构建的L-甲硫氨酸高产菌株大肠杆菌(Escherichia coli)W3110ΔIJAHFEBC trc-fliY trc-malY/PAM glyA-22 metF的基础上系统优化了其发酵过程,在优化初始葡萄糖浓度的基础上,比较了DO-Stat、pH-Stat、控残糖浓度和恒速补料等不同补料工艺对L-甲硫氨酸发酵生产的影响,发现葡萄糖的控制对发酵过程有较大影响。在上述研究的基础上,进一步开发了最优的分批补料发酵工艺,使L-甲硫氨酸发酵产量提高到31.71 g/L,发酵时间缩短为68 h,是目前报道的最高产量。同时,建立了最优分批补料工艺下的发酵动力学模型,较好地拟合了L-甲硫氨酸发酵生产过程,为L-甲硫氨酸的发酵法生产提供了一定的数据参考。

As the only essential amino acid containing elemental sulphur,L-methionine has important physiological and biochemical functions in living organisms.However,the fermentative production of L-methionine has not met the requirements of industrial production because of its low production level.In this paper,the fermentation process of an efficient L-methionine producing strain E.coli W3110ΔIJAHFEBC trc-fliY trc-malY/PAM glyA-22 metF constructed previously was systematically optimized.Based on the optimal initial glucose concentration,the effects of different fed-batch fermentation processes,including DO-Stat,pH-Stat,controlling residual sugar control at different level and feeding glucose with constant rate,on L-methionine fermentation were studied.It was found that the control of glucose concentration greatly affected the fermentation process.Subsequently,an optimal fed-batch fermentation process was developed,where the L-methionine titer was increased to 31.71 g/L,the highest yield reported to date,while the fermentation time was shortened to 68 h.Meanwhile,a fermentation kinetics model under the optimal fed-batch fermentation conditions was established,which fitted well with the biosynthesis process of L-methionine.This study may facilitate further development of the fermentative production of L-methionine.