为了提高丙酮酸激酶突变的重组大肠杆菌BR-42δpykA(pAP-B03)发酵生产L-苯丙氨酸(L-Phe)的能力,考察了不同诱导温度(36、38和40℃)下菌体生长和L-Phe生产等重要发酵参数的变化规律;基于对发酵参数的动力学分析,提出了分阶段诱导温度控制策略,即在13.5~20 h,诱导温度采用40℃;21~37 h降低温度至38℃;38 h以后采用36℃。该策略的实施,可以显著改善诱导后期菌体的生长、质粒的维持和关键酶的酶活水平,从而提高L-Phe的生产;发酵48 h时,L-Phe的产量达到52.70 g/L,分别比在36、38和40℃诱导时提高了12.01%、13.77%和22.39%。结果表明:根据对不同诱导温度下的发酵动力学参数进行分析而提出的分阶段温度控制策略可以有效地应用于L-Phe的分批补料发酵过程中。

To improve the capacity of L-Phenylalanine( L-Phe) production with Escherichia coli BR-42 δpykA( pAP-B03),a pyruvate kinase mutant strain,the effects of different induction temperature( 36℃,38℃ and 40℃) on cell growth and L-Phe production were investigated. Based on the analysis of fermentation parameters,a threestage induction temperature controlled strategy was proposed,in which,the induction temperature was controlled at 40℃ during 13. 5 ~ 20 h,38℃ during 21 ~ 37 h,and 36℃ after 38 h,respectively. With the strategy,the cell growth,plasmid maintenance,and the key enzyme activities in late induction phase were improved,resulting in an enhanced L-Phe production. At 48 h,L-Phe titer reached 52. 70 g / L,which was 12. 01%,13. 77%,and 22. 39% higher than that at 36℃,38℃,and 40℃,respectively,which indicated that the three-stage induction temperature controlled strategy based on the analysis of fermentation parameters was favorable to L-Phe production in fed-batch culture.