响应面-满意度函数优化重组大肠杆菌产NMN转移酶发酵条件

重组大肠杆菌Escherichaia coli能高效表达NMN转移酶,以此为出发菌株,以菌体生长量OD600和NMN转移酶的活力为响应值,对重组大肠杆菌产NMN转移酶的发酵条件进行优化.首先以Plackett-Burman实验设计优化筛选出3个主要影响因子:胰蛋白胨、甘油、MgSO4;随后以Box-Behnken中心组合设计建立上述3个因子对OD600和NMN转移酶活力水平的数学模型;最后通过满意度函数获得最佳发酵条件为:酵母粉30 g/L,胰蛋白胨10.5 g/L,甘油3.49 mL/L,MgSO40.45 g/L,K2 HPO440.5 g/L,KH2 PO46.0 g/L,NH4 Cl 1.5 g/L,NaCl 0.6 g/L,接种量1.5%,诱导时间12 h.在该优化条件下,菌体生长和产酶水平均获得了显著的提升.重组NMN转移酶的活力水平从8.85 U/mg提高到15.48 U/mg,菌体生长量OD600从4.85提高到6.01,提高幅度分别为74.92%和23.92%.

Optimization of NMN adenylyltransferase production by recombinant Escherichia coli using response surface methodology coupled with desirability function

The fermentation conditions of NMN adenylyltransferase production by recombinant Escherichia coli were optimized. Biomass yield ( OD600 ) and NMN adenylyltransferase activity were selected as response value during fermentation condition optimization. Peptone, glycerin and MgSO4 were screened to be the three key components using Plackett-Burman design method. Then, the mathematical model reflecting the three main factors and the production level of enzyme was established by using the Box-Behnken design method. Finally, a satisfactory degree function was defined and the satisfactory solution was obtained. The final optimal fermentation conditions were:yeast extract 30 g/L, peptone 10. 5 g/L, glycerol 3. 49 mL/L, MgSO40. 45 g/L, K2HPO440. 5 g/L, KH2PO46. 0 g/L, NH4Cl 1. 5 g/L, NaCl 0. 6 g/L, inoculum size of 1. 5% and induction time of 12 h. After the validated experiment under above conditions, enzyme production and biomass of recombinant strain were increased by 74. 92% and 23. 92%, respectively.