Enhanced production of 2’‐fucosyllactose from fucose by elimination of rhamnose isomerase and arabinose isomerase in engineered Escherichia coli

• Published in: Biotechnology and bioengineering 2019-09, Vol.116 (9), p.2412-2417
• Main Authors: Jung, Sang‐Min, Chin, Young‐Wook, Lee, Ye‐Gi, Seo, Jin‐Ho
• Format: Article
• Language: English
• Subjects: 2′‐fucosyllactose; Arabinose; Arabinose isomerase; Bacteria; Biosynthesis; Breast milk; E coli; engineered Escherichia coli; Escherichia coli; Fermentation; Fucokinase; Fucose; Galactosidase; Genes; Glycerol; Guanosine; Guanosine diphosphate; Helicobacter pylori; Kinases; Lactose; Microorganisms; Oligosaccharides; Production methods; Rhamnose; rhamnose isomerase; Salvage; salvage pathway of GDP‐ l‐fucose; Sugar; β-Galactosidase
• ISSN: 0006-3592; 1097-0290
• DOI: 10.1002/bit.27019

2′‐Fucosyllactose (2‐FL), one of the most abundant oligosaccharides in human milk, has been spotlighted for its neutraceutical and pharmaceutical potentials. Microbial production of 2‐FL is promising since it is efficient as compared to other production methods. In 2‐FL microbial production via the salvage pathway for biosynthesis of guanosine 5′‐diphosphate (GDP)‐l‐fucose from fucose, the conversion yield from fucose is important because of the high price of fucose. In this study, deletion of the genes (araA and rhaA) coding for arabinose isomerase (AraA) and rhamnose isomerase (RhaA) was attempted in engineered Escherichia coli for improving 2‐FL production by using fucose, lactose, and glycerol. The engineered E. coli constructed previously is able to express fucokinase/GDP‐l‐fucose pyrophosphorylase (Fkp) from Bacteroides fragilis and the α‐1,2‐fucosyltransferase (FucT2) from Helicobacter pylori and deficient in β‐galactosidase (LacZ), fucose isomerase (FucI), and fuculose kinase (FucK). The additional double‐deletion of the araA and rhaA genes in the engineered E. coli enhanced the product yield of 2‐FL to 0.52 mole 2‐FL/mole fucose, and hence the concentration of 2‐FL reached to 47.0 g/L, which are 44% and two‐fold higher than those (23.1 g/L and 0.36 mole 2‐FL/mole fucose) of the control strain in fed‐batch fermentation. Elimination of sugar isomerases exhibiting promiscuous activities with fucose might be critical in the microbial production of 2‐FL through the salvage pathway of GDP‐l‐fucose. 2’‐Fucosyllactose (2‐FL), one of the most abundant oligosaccharides in human milk has been spotlighted for its neutraceutical and pharmaceutical potentials. In previous research, Escherichia coli have been engineered to produce 2‐FL from fucose and lactose. In this study, the additional elimination of sugar isomerases (AraA and RhaA) exhibiting promiscuous activities with fucose in the engineered E. coli enhanced the concentration of 2‐FL production from 23.1 g/L to 47.0 g/L in fed‐batch fermentation.