TCA循环关键节点对L-谷氨酸合成的影响  

Effect of key notes of TCA cycle on L-glutamate production

作　　者：乔郅钠 徐美娟[1] 龙梦飞 杨套伟[1] 张显[1] 中西秀树[1] 饶志明[1] Zhina Qiao;Meijuan Xu;Mengfei Long;Taowei Yang;Xian Zhang;Nakanishi Hideki;Zhiming Rao(School of Biotechnology,Jiangnan University,Wuxi 214122,Jiangsu,China)

机构地区：[1]江南大学生物工程学院,江苏无锡214122

出　　处：《生物工程学报》2020年第10期2113-2125,共13页Chinese Journal of Biotechnology

基　　金：国家重点研发计划(No.2018YFA0900300);国家自然科学基金(No.31770058);江苏省自然科学基金(No.BK20181205);宁夏回族自治区重点研发计划(No.2019BCH01002);中组部万人计划科技创新领军人才项目资助

摘　　要：谷氨酸是一种重要的氨基酸,其衍生出来的高值化产品具有广泛的应用,市场需求量巨大。文中通过对出发菌株谷氨酸棒杆菌Corynebacterium glutamicum E01和谷氨酸高产菌C.glutamicum G01进行转录组测序与重测序分析,挑选中心代谢途径中转录水平和基因水平上存在差异的基因进行研究,以挖掘出对谷氨酸合成影响较大的基因进一步提高谷氨酸的产量。草酰乙酸节点和α-酮戊二酸节点在谷氨酸合成中扮演着重要角色,探索研究了草酰乙酸节点和α-酮戊二酸节点对谷氨酸生产的扰动影响。综合以上实验结果构建的整合菌株,5 L发酵罐发酵过程中其菌体生长速率较原始菌略有降低,但48h的谷氨酸产量高达(136.09±5.53)g/L,较原始菌的(93.53±4.52)g/L提高了45.5%;糖酸转化率提高至58.9%,较原始菌的45.2%提高了13.7%。可见,上述实验策略的应用在一定程度上提高了谷氨酸产量和糖酸转化率,为谷氨酸棒杆菌的代谢工程改造提供了理论基础。Glutamic acid is an important amino acid with wide range of applications and huge market demand.Therefore,by performing transcriptome sequencing and re-sequencing analysis on Corynebacterium glutamicum E01 and high glutamate-producing strain C.glutamicum G01,we identified and selected genes with significant differences in transcription and gene levels in the central metabolic pathway that may have greatly influenced glutamate synthesis and further increased glutamic acid yield.The oxaloacetate node andα-ketoglutarate node play an important role in glutamate synthesis.The oxaloacetate node and α-ketoglutarate node were studied to explore effect on glutamate production. Based on the integratedstrain constructed from the above experimental results, the growth rate in a 5-L fermenter was slightly lower than that of theoriginal strain, but the glutamic acid yield after 48 h reached (136.1±5.53) g/L, higher than the original strain (93.53±4.52) g/L,an increase by 45.5%;sugar-acid conversion rate reached 58.9%, an increase of 13.7% compared to 45.2% of the originalstrain. The application of the above experimental strategy improved the glutamic acid yield and the sugar-acid conversion rate,and provided a theoretical basis for the metabolic engineering of Corynebacterium glutamicum.

关 键 词：谷氨酸棒杆菌 谷氨酸 草酰乙酸节点 α-酮戊二酸节点 CRISPR/dCas9 核糖体结合位点优化 

分 类 号：TQ922.1[轻工技术与工程—发酵工程]