pH耦合柠檬酸补料策略促进地衣芽孢杆菌产芽孢

pH Coupled Citric Acid Feeding Strategy to Promote Spore Production of Bacillus licheniformis

在30 L罐水平研究了耦合pH补加柠檬酸对地衣芽孢杆菌生长、代谢以及芽孢形成的影响,通过qPCR检测了细胞对数期能量代谢、氧化应激效应、氨基酸转运相关基因表达水平差异。结果表明,发酵19.5~28 h耦合pH7.5补入78μmol/L的柠檬酸,延长了细胞生长和芽孢生成周期,峰值生物量(36 h)达到4.2×10^(10)CFU/mL,较对照(未补柠檬酸)提高42.9%;芽孢数(36 h)达到4.1×10^(10)CFU/mL,较对照(24h峰值1.3×10^(10)CFU/mL)提高215.4%。qPCR测定发现icd、zwf和cydA表达量分别是对照的2.6、4.1和1.9倍,而brnQ、vpr、yvbW、katA、yqjM、ahpF、sodA和abrB分别较对照下调88%、18%、65%、89%、10%、90%、41%和60%。说明柠檬酸的补加增强了TCA循环、HMP途径以及电子传递效率,同时减弱了氧化胁迫效应、提高了Spo0A的磷酸化水平,促进了芽孢形成效率。研究结果为芽孢杆菌工业化高效发酵生产提供了理论支撑。

The effects of citric acid addition coupled with pH on cell growth,metabolism,and spore yield of B.licheniformis were studied in a 30 L bioreactor,and the gene expression differences related to energy metabolism,oxidative stress effect,and amino acid transport at the log-phase were examined by qPCR.Results showed that:Citric acid supplement coupled with pH7.5 during 19.5~28 h was conducted,reaching a final content of 78μmol/L in the medium.The feeding strategy prolonged the periods of cell growth and spore formation,and the peak cells(36 h)reached 4.2×10^(10)CFU/mL that was 42.9% higher than the control(without citric acid supplement).Meanwhile,spore yield(36 h)reached 4.1×10^(10)CFU/mL,increased by 215.4% compared with the control(24 h,1.3×10^(10)CFU/mL).The qPCR assays showed that the expression levels of icd,zwf and cydA was 2.6,4.1 and 1.9 times higher than that of the control,respectively.Meanwhile,brnQ,vpr,yvbW,katA,yqjM,ahpF,sodA and abrB were decreased by 88%,18%,65%,89%,10%,90%,41% and 60% compared with the control,respectively.The data suggested that citric acid supplementation enhanced the efficiencies of TCA cycle,HMP pathway and electron transport,weakened the oxidative stress effects on cells,and improved the phosphorylation level of Spo0A,thus promoting the sporulation efficiency.The results would provide theoretical supports for the industrial production of Bacillus fermentation efficiently.