提升头孢菌素C发酵效率的复合碳源补料工艺开发及代谢机制  

Development of Compound Carbon Source Feeding Strategy for Improving Cephalosporin C Fermentation Efficiency and Its Metabolic Mechanism Exploration

作　　者：杨倚铭[1] 陈震 田锡炜[1] 储炬[1] YANG Yiming;CHEN Zhen;TIAN Xiwei;CHU Ju(State Key Laboratory of Bioreactor Engineering,East China University of Science and Technology,Shanghai 200237,China)

机构地区：[1]华东理工大学生物反应器工程国家重点实验室,上海200237

出　　处：《华东理工大学学报（自然科学版）》2024年第5期687-694,共8页Journal of East China University of Science and Technology

摘　　要：开发了一种以葡萄糖和豆油为底物碳源的复合补料策略,能够显著提升头孢菌素C(CPC)的合成效率,使其产量提升了14.8%,并观测到碳源变化在宏观呼吸代谢、中间代谢物、细胞内通量分配和还原力NADPH利用方面产生了全局影响,其中二氧化碳释放速率(CER)的变化表明在葡萄糖和豆油复合补料策略下细胞中心代谢网络的通量发生了明显改变,碳流向CPC的通量得到增强,同时促进了蛋氨酸和丝氨酸的胞外吸收。胞内有机酸α-酮戊二酸、丙酮酸、苹果酸的浓度明显提升,有利于谷氨酸、缬氨酸和半胱氨酸等核心氨基酸的合成,从而有利于CPC生产。主成分分析(PCA)的载荷测定结果表明大多数氨基酸和CPC合成呈正相关,谷氨酸的胞内含量与CPC合成的关联性最强,而α-氨基己二酸和赖氨酸积累反而不利于CPC合成。细胞还原力方面,豆油和葡萄糖组合工艺有助于NADPH更多地用于CPC合成,使得CPC快速合成时对NADPH的大量需求得到了满足。这些结果表明改变底物能够对细胞内部的代谢产生全方位的影响,复合补料策略在优化CPC合成中具有巨大潜力。Soybean oil is an important carbon source for the efficient synthesis of secondary metabolites.In this paper,a novel feeding strategy of glucose and soybean oil combination was developed,which significantly improved the synthesis efficiency of Cephalosporin C(CPC)by 14.8%.A multi-scale research on the mechanism for CPC enhancement revealed that the combination of carbon source combination globally changed cell respiration metabolism,intermediate metabolites pool size,metabolic flux distribution,and NADPH utilization.The significant change in carbon dioxide evolution rate(CER)indicated a significant alteration in central metabolic flux under the glucose and soybean oil combination feeding strategy,leading to strengthened carbon flux for CPC synthesis and increased extracellular absorption rates of methionine and serine.Intracellular pools ofα-ketoglutarate,pyruvate,and malate were significantly increased,which were beneficial for core amino acids synthesis such as glutamic acid,valine,and cysteine to facilitate CPC production.The loading analysis of principal component analysis(PCA)showed that the most of amino acids positively influenced CPC synthesis,and intracellular glutamic acid content was found to have the strongest positive correlation.However,the accumulation ofα-aminoadipic acid and lysine was unfavorable for CPC synthesis.In terms of reducing power,the combination of soybean oil and glucose significantly alleviated NADPH competition between soybean oil utilization and CPC synthesis,thus meeting high demand for rapid CPC synthesis.These results indicated that substrate changes presented a comprehensive impact on cellular metabolism of cells,reflecting the great potential of this strategy in optimizing the CPC production efficiency.

关 键 词：头孢菌素C 碳源 呼吸代谢 代谢通量分析 主成分分析 

分 类 号：Q815[生物学—生物工程]