Biotechnological Production of 2′-Fucosyllactose: A Prevalent Fucosylated Human Milk Oligosaccharide

Zhou, Wenting; Jiang, Hong; Wang, Lili; Liang, Xingxing; Mao, Xiangzhao

doi:10.1021/acssynbio.0c00645

Cited by 28 publications

(30 citation statements)

References 83 publications

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“…These confirmed health benefits and promising applications have driven the extensive researches of 2′-FL, particularly in the field of biosynthesis. 6 Currently, plenty of production strategies of 2′-FL have been established using E. coli, 7,8 yeast, 9 and B. subtilis 10 as microbial hosts. GDP-L-fucose and lactose are known as the two direct cosubstrates for synthesizing 2′-FL by α1,2-fucosyltransferase in vivo.…”

Section: ■ Introductionmentioning

confidence: 99%

High-Level De Novo Biosynthesis of 2′-Fucosyllactose by Metabolically Engineered Escherichia coli

Liu

Zhu

Wan

et al. 2022

J. Agric. Food Chem.

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2′-Fucosyllactose (2′-FL) is the most abundant oligosaccharide in human milk. In this study, a highly efficient biosynthetic route for 2′-FL production was designed via the de novo pathway of GDP-L-fucose using engineered Escherichia coli BL21(DE3). Specifically, plasmid-based strains with previously deleted lacZ and wcaJ were further reconstructed by introducing de novo pathway genes and α1,2-fucosyltransferase-encoding wbgL to realize 2′-FL synthesis. The 2′-FL titer was enhanced to 3.92 g/L by further introducing rcsA and rcsB. Subsequently, the additional wbgL expression cassette was chromosomally integrated into recA locus to strengthen fucosylation reaction and a strong constitutive promoter (P J23119 ) was used to replace the original promoters of manC-manB and gmd-wcaG to improve 2′-FL synthesis. The maximal 2′-FL titer reached 9.06 and 79.23 g/L in shake-flask and fedbatch cultivation, respectively. The 2′-FL productivity reached 1.45 g/L/h, showing remarkable production potential in large-scale industrial application.

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Section: ■ Introductionmentioning

confidence: 99%

High-Level De Novo Biosynthesis of 2′-Fucosyllactose by Metabolically Engineered Escherichia coli

Liu

Zhu

Wan

et al. 2022

J. Agric. Food Chem.

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“…Currently, two promising routes are available to produce 2′-FL: chemical synthesis and biosynthesis [ 13 , 14 ]. The chemical synthesis of 2′-FL is accomplished through multistep protection and deprotection reactions.…”

Section: Introductionmentioning

confidence: 99%

“…The chemical synthesis of 2′-FL is accomplished through multistep protection and deprotection reactions. This method is inefficient and uses toxic chemicals, rendering the chemical method unfit for common use [ 14 ]. The reaction steps for 2′-FL biosynthesis are simpler and more environmentally friendly than its chemical synthesis.…”

Section: Introductionmentioning

confidence: 99%

Engineered Bacillus subtilis for the de novo production of 2′-fucosyllactose

Zhang

Liu²,

Xia³

et al. 2022

Microb Cell Fact

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Background The most abundant human milk oligosaccharide in breast milk, 2′-fucosyllactose (2′-FL), has been approved as an additive to infant formula due to its multifarious nutraceutical and pharmaceutical functions in promoting neonate health. However, the low efficiency of de novo synthesis limits the cost-efficient bioproduction of 2′-FL. Results This study achieved 2′-FL de novo synthesis in a generally recognized as safe (GRAS) strain Bacillus subtilis. First, a de novo biosynthetic pathway for 2′-FL was introduced by expressing the manB, manC, gmd, wcaG, and futC genes from Escherichia coli and Helicobacter pylori in B. subtilis, resulting in 2′-FL production of 1.12 g/L. Subsequently, a 2′-FL titer of 2.57 g/L was obtained by reducing the competitive lactose consumption, increasing the regeneration of the cofactor guanosine-5′-triphosphate (GTP), and enhancing the supply of the precursor mannose-6-phosphate (M6P). By replacing the native promoter of endogenous manA gene (encoding M6P isomerase) with a constitutive promoter P7, the 2′-FL titer in shake flask reached 18.27 g/L. The finally engineered strain BS21 could produce 88.3 g/L 2′-FL with a yield of 0.61 g/g lactose in a 3-L bioreactor, without the addition of antibiotics and chemical inducers. Conclusions The efficient de novo synthesis of 2′-FL can be achieved by the engineered B. subtilis, paving the way for the large-scale bioproduction of 2′-FL titer in the future.

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“…111 For synthesis of complex structured HMOs, researchers have focused on adapting microbial metabolic pathways to enhance 2′-FL production. 112,113 The potential challenges for production of complex structured HMOs can be addressed when the metabolic routes are designed for 2′-FL production as the most studied type of HMO. Therefore, to synthesize complex structured HMOs with high yield, 2′-FL can be used as the model product.…”

Section: Discussionmentioning

confidence: 99%

Engineered Microbial Routes for Human Milk Oligosaccharides Synthesis

Mosleh

Abbaspourrad

2021

ACS Synth. Biol.

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Human milk oligosaccharides (HMOs) are one of the important ingredients in human milk, which have attracted great interest due to their beneficial effect on the health of newborns. The large-scale production of HMOs has been researched using engineered microbial routes due to the availability, safety, and low cost of host strains. In addition, the development of molecular biology technology and metabolic engineering has promoted the effectiveness of HMOs production. According to current reports, 2′-fucosyllactose (2′-FL), 3-fucosyllactose (3-FL), lacto-N-tetraose (LNT), lacto-N-neotetraose (LNnT), 3′-sialyllactose (3′-SL), 6′-sialyllactose (6′-SL), and some fucosylated HMOs with complex structures have been produced via the engineered microbial route, with 2′-FL having been produced the most. However, due to the uncertainty of metabolic patterns, the selection of host strains has certain limitations. Aside from that, the expression of appropriate glycosyltransferase in microbes is key to the synthesis of different HMOs. Therefore, finding a safe and efficient glycosyltransferase has to be addressed when using engineered microbial pathways. In this review, the latest research on the production of HMOs using engineered microbial routes is reported. The selection of host strains and adapting different metabolic pathways helped researchers designing engineered microbial routes that are more conducive to HMOs production.

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Biotechnological Production of 2′-Fucosyllactose: A Prevalent Fucosylated Human Milk Oligosaccharide

Cited by 28 publications

References 83 publications

High-Level De Novo Biosynthesis of 2′-Fucosyllactose by Metabolically Engineered Escherichia coli

High-Level De Novo Biosynthesis of 2′-Fucosyllactose by Metabolically Engineered Escherichia coli

Engineered Bacillus subtilis for the de novo production of 2′-fucosyllactose

Engineered Microbial Routes for Human Milk Oligosaccharides Synthesis

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