Pathway Optimization and Uridine 5′-Triphosphate Regeneration for Enhancing Lacto-N-Tetraose Biosynthesis in Engineered Escherichia coli,Journal of Agricultural and Food Chemistry

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Pathway Optimization and Uridine 5′-Triphosphate Regeneration for Enhancing Lacto-N-Tetraose Biosynthesis in Engineered Escherichia coli
Journal of Agricultural and Food Chemistry ( IF 5.7 ) Pub Date : 2022-06-19 , DOI: 10.1021/acs.jafc.2c02426
Zeyu Li ₁ , Yingying Zhu ₁ , Pan Zhang ₁ , Wenli Zhang ₁ , Wanmeng Mu ₁

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Recently, human milk oligosaccharides (HMOs) have attracted increasing attention and display great commercial importance, especially for the infant formula industry. Lacto-N-tetraose (LNT) is an important neutral HMO commercially added in infant formula and a core structure for synthesizing complex HMOs. Previously, a novel LNT-generating β-1,3-galactosyltransferase from Pseudogulbenkiania ferrooxidans was identified and used for construction of an LNT-producing engineered Escherichia coli. In this work, LNT biosynthesis was further enhanced by pathway optimization and uridine 5′-triphosphate (UTP) regeneration. The main strategies included genomic integration of UDP-glucose 4-epimerase-encoding gene, fine-tuning of the LNT pathway-related genes, blocking of competitive pathways related to UDP-galactose, and overexpression of UTP supply related genes. The maximal LNT titer reached 6.16 and 57.5 g/L by shake-flask and fed-batch fermentation, respectively.

中文翻译：

增强工程大肠杆菌中乳酸-N-四糖生物合成的途径优化和尿苷 5'-三磷酸再生

最近，人乳寡糖 (HMO) 引起了越来越多的关注并显示出巨大的商业重要性，尤其是对于婴儿配方奶粉行业。乳-N-四糖 (LNT) 是商业上添加到婴儿配方奶粉中的重要中性 HMO，也是合成复杂 HMO 的核心结构。此前，从Pseudogulbenkiania ferrooxidans中鉴定出一种新的产生 LNT 的 β-1,3-半乳糖基转移酶，并用于构建产生 LNT 的工程大肠杆菌. 在这项工作中，通过途径优化和尿苷 5'-三磷酸 (UTP) 再生进一步增强了 LNT 的生物合成。主要策略包括 UDP-葡萄糖 4-差向异构酶编码基因的基因组整合、LNT 通路相关基因的微调、与 UDP-半乳糖相关的竞争通路的阻断以及 UTP 供应相关基因的过表达。摇瓶发酵和分批补料发酵的最大 LNT 滴度分别达到 6.16 和 57.5 g/L。

更新日期：2022-06-19

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