利用代谢工程策略改造谷氨酸棒杆菌生产丙酮酸
DOI:
作者:
作者单位:

1.湖南大学 隆平分院;2.湖南省农业科学院 农产品加工研究所

作者简介:

通讯作者:

中图分类号:

TS201.3;TS255.1

基金项目:

湖南省农业科技创新项目(2021CX05)


Metabolic engineering of Corynebacterium glutamicum for producing pyruvate
Author:
Affiliation:

1.Longping Branch Graduate School,Hunan University;2.Institute of Agricultural Product Processing, Hunan Academy of Agricultural Sciences

Fund Project:

Agricultural Science and Technology Innovation Project of Hunan Province (2021CX05)

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    摘要:

    丙酮酸是一种重要的有机酸,在生物体的能量代谢中发挥着重要作用,它可以作为前体物质,参与合成许多有机化合物。因此选择利用代谢工程策略改造谷氨酸棒杆菌生产丙酮酸。首先利用同源重组的方法,依次敲除谷氨酸棒杆菌(Corynebacterium glutamicum)中,丙酮酸代谢支流相关的五个关键基因:编码丙酮酸醌氧化还原酶基因pqo、丙酮酸羧化酶基因pyc、转氨酶基因alaT、缬氨酸-丙酮酸氨基转移酶基因avtA、丙酮酸脱氢酶基因aceE,摇瓶发酵72 h丙酮酸的产量达到14.64g/L。为了进一步增加丙酮酸通路代谢流,过表达编码转酮醇酶基因tkt、转醛酶基因tal、磷酸烯醇丙酮酸羧激酶基因pck,以增加合成丙酮酸的前体物质的供应。最终,复合培养基摇瓶发酵72h,发酵液中丙酮酸的产量达到15.39g/L,与野生型菌株相比提高了28倍,有助于为日后微生物发酵生产丙酮酸提供理论基础和参考。

    Abstract:

    Pyruvate is an important organic acid, which plays an important role in the energy metabolism of organisms. As a precursor, pyruvate can participate in the biosynthesis of many organic compounds. Therefore, the metabolic engineering strategy was used to modify corynebacterium glutamate to produce pyruvate. First, five key genes related to pyruvate metabolism tributaries of Corynebacterium glutamicum were successfully knocked out by homologous recombination method: pyruvate quinone oxidoreductase gene pqo, pyruvate carboxylase gene pyc, aminotransferase gene alaT, valine-pyruvate aminotransferase gene avtA, pyruvate dehydrogenase gene aceE. The yield of pyruvate reached 14.64 g/L after 72 h of shaking flask fermentation. In order to further increase the metabolic flux of pyruvate pathway, transketolase gene tkt, transaldolase gene tal, phosphoenolpyruvate carboxykinase gene pck were overexpressed to increase the supply of precursors for pyruvate synthesis. Finally, the yield of pyruvate reached 15.39 g/L after 72 h shaker fermentation, which was 28 times higher than that of the wild-type strain, which is helpful to provide theoretical basis and reference for the future study on the production of pyruvate by microbial fermentation.

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  • 收稿日期:2022-05-27
  • 最后修改日期:2022-10-26
  • 录用日期:2022-08-18
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