Combinatorial metabolic engineering of Escherichia coli for de novo production of 2′-fucosyllactose

• Published in: Bioresource technology 2022-05, Vol.351, p.126949-126949, Article 126949
• Main Authors: Lin, Lu, Gong, Mengyue, Liu, Yanfeng, Li, Jianghua, Lv, Xueqin, Du, Guocheng, Liu, Long
• Format: Article
• Language: English
• Subjects: 2′-fucosyllactose; Escherichia coli; Escherichia coli - genetics; Escherichia coli - metabolism; Fucosyltransferases - genetics; Fucosyltransferases - metabolism; Humans; Metabolic Engineering; Milk, Human; Protein tag; Trisaccharides; α1,2-fucosyltransferase
• ISSN: 0960-8524; 1873-2976
• DOI: 10.1016/j.biortech.2022.126949

[Display omitted] •Plasmid-free strains for 2′-FL synthesis were constructed by CRISPR/Cas9 method.•The stable expression of FutC was achieved through genomic integration.•The TrxA-FutC recombinant protein was expressed to promote 2′-FL synthesis.•Mannose can be used as a new carbon source to synthesize 2′-FL. 2′-Fucosyllactose (2′-FL) is a kind of fucosylated lactose of human milk oligosaccharides (HMOs). In this work, Escherichia coli MG1655 was metabolically engineered to increase 2′-FL production. The 2′-FL titer was raised from 0.0118 to 0.8062 g/L by increasing three gene copies of α1,2-fucosyltransferase (FutC) and by deleting the lon, wcaJ, lacZ, and lacI genes in the competitive pathways. Additionally, the 2′-FL titer was raised to 2.9 g/L by fusing a TrxA tag at the N-terminus of FutC, and then to 3.4 g/L by deleting glutathione reductase (Gor). Finally, the 2′-FL reached 3.3 g/L in the final strain E. coli MG27 through the de novo pathway in shake flask, and reached 10.3 g/L in a 3-L fermentor.