Study on Enhancement of Lacto-N-Neotetraose Synthesis in Escherichia coli Based on Module Optimization

被引：0
|
作者：
Liu D. [1 ,2 ]
Liang S. [1 ]
Yan Q. [3 ]
Yang S. [1 ]
Li S. [1 ,4 ]
Jiang Z. [1 ]
机构：
[1] College of Food Science and Nutritional Engineering, China Agricultural University, Beijing
[2] Food Laboratory of Zhongyuan, Luohe
[3] College of Engineering, China Agricultural University, Beijing
[4] Mengniu Gaoke Dairy (Beijing) Co. Ltd., Beijing
来源：
Journal of Food Science and Technology (China) | 2024年 / 42卷 / 02期
基金：
中国国家自然科学基金;
关键词：
biosynthesis; CRISPR; /; Cas9; Escherichia coli; lacto-N-neotetraose; module optimization;
D O I：
10.12301/spxb202300614
中图分类号：
学科分类号：
摘要：
As one key component of human milk oligosaccharides, lacto-N-neotetraose (LNnT) plays an important role in the growth and development of infants. In order to explore the efficient method for biosynthetic of LNnT and to investigate the influence of module optimization on LNnT synthesis of Escherichia coli, E. coli BL21(DE3)∆lacZ was used as the initial strain. And the synthetic pathway of LNnT was divided into the following three modules based on the key metabolites in synthetic pathway, module A for the exogenous enzymatic pathway, module B for the synthetic pathway of UDP-galactose, and module C for the synthetic pathway of UDP-N-acetylglucosamine. After preliminarily optimizing the expressions of modules A, B, and C via the plasmids with different copy number, the E. coli BL21 (DE3)∆lacZ harboring the recombinant plasmids pRSF-lgtA鄄A. act and pET-galE produced the highest LNnT titer of 0. 87 g/ L. Modules A and B were enhanced owing to knocking setA and ugd via CRISPR/ Cas9 technique, and the titer of LNnT produced by the recombinant strain E20 was up to 1. 16 g/ L. After optimizing the fermentation conditions of strain E20, the titer of LNnT in shake-flask cultivation was up to 1. 28 g/ L and further was up to 15. 53 g/ L by fed-batch fermentation in a 5 L bioreactor. The highest productivity of LNnT was up to 0. 43 g/ (L·h) during fermentation. The enhancement of LNnT synthesis in E. coli with module optimization was expected to provide theoretical basis for efficient biosynthesis of human milk oligosaccharides and to drive innovation in food industry of infant formula. © 2024 Beijing Technology and Business University, Department of Science and Technology. All rights reserved.
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页码：75 / 83
页数：8
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