High-Yield Synthesis of Lacto-N-Neotetraose from Glycerol and Glucose in Engineered Escherichia coli

Journal of Agricultural and Food Chemistry, Journal Year: 2024, Volume and Issue: 72(10), P. 5325 - 5338

Published: Jan. 26, 2024

Lacto-

Language: Английский

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Metabolic engineering of Escherichia coli BL21(DE3) cocultured with glucose and xylose for efficient production of 2′-fucosyllactose

Bioresource Technology, Journal Year: 2025, Volume and Issue: unknown, P. 132062 - 132062

Published: Jan. 1, 2025

Language: Английский

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Engineering Pichia pastoris for Efficient De Novo Synthesis of 2′-Fucosyllactose

Journal of Agricultural and Food Chemistry, Journal Year: 2025, Volume and Issue: unknown

Published: March 28, 2025

2′-Fucosyllactose (2′-FL), the most abundant in human milk oligosaccharides (HMOs), is a nutrient of great importance. As safe organism widely used industries, Pichia pastoris was tested here for 2′-FL production. The de novo biosynthesis pathway constructed using genome-editing technology based on CRISPR-Cas9 with an initial titer 1.01 g/L. Introducing N-terminal SUMO or Ub tag to FucT2 and transporter CDT2 from Neurospora crassa into P. found improve Then, modular metabolic engineering conducted production, enhancing GTP supply module, NADPH regeneration precursor module. Subsequently, key enzyme semirationally designed further increase Finally, production by engineered scaled up 3 L fermenter fed-batch mode, resulting 22.35 g/L that highest pastoris. results prove effectiveness strategies demonstrate could be potential chassis produce HMOs.

Language: Английский

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Semi-rationally designed site-saturation mutation of Helicobacter pylori α-1,2-fucosyltransferase for improved catalytic activity and thermostability

International Journal of Biological Macromolecules, Journal Year: 2024, Volume and Issue: 259, P. 129316 - 129316

Published: Jan. 11, 2024

Language: Английский

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Highly efficient biosynthesis of 3′-sialyllactose in engineered Escherichia coli

International Journal of Biological Macromolecules, Journal Year: 2024, Volume and Issue: 269, P. 132081 - 132081

Published: May 3, 2024

Language: Английский

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Microbial Synthesis of Lacto-N-fucopentaose I with High Titer and Purity by Screening of Specific Glycosyltransferase and Elimination of Residual Lacto-N-triose II and Lacto-N-tetraose

Journal of Agricultural and Food Chemistry, Journal Year: 2024, Volume and Issue: 72(8), P. 4317 - 4324

Published: Feb. 16, 2024

Lacto-N-fucopentaose I (LNFP I) has recently been approved as generally recognized safe, demonstrating its great commercial potential in the food industry. Microbial synthesis through metabolic engineering strategies is an effective approach for large-scale production of LNFP I. Biosynthesis requires consideration two key points: high titer with low byproduct 2′-fucosyllactose (2′-FL) generation and purity lacto-N-triose II (LNTri II) lacto-N-tetraose (LNT) residues. Herein, α1,2-fucosyltransferase from Thermoanaerobacterium sp. RBIITD was screened 16 selected I-producing glycosyltransferase candidates, showing highest vivo productivity. Chromosomal integration wbgO enhanced by improving precursor conversion LNTri to LNT. The best engineered strain produced 4.42 35.1 g/L shake-flask fed-batch cultivation, respectively. residual LNT were eliminated further cultivation a recombinant coexpressing Bifidobacterium bifidum β-N-acetylhexosaminidase lacto-N-biosidase. A strategy biosynthesis yield finally realized, providing support practical application production.

Language: Английский

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De Novo Biosynthesis of Difucosyllactose by Artificial Pathway Construction and α1,3/4-Fucosyltransferase Rational Design in Escherichia coli

Journal of Agricultural and Food Chemistry, Journal Year: 2024, Volume and Issue: unknown

Published: April 10, 2024

Difucosyllactose (DFL) is a significant and plentiful oligosaccharide found in human breast milk. In this study, an artificial metabolic pathway of DFL was designed, focusing on the de novo biosynthesis GDP-fucose from only glycerol. This achieved by engineering

Language: Английский

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Efficient production of 2′-fucosyllactose from fructose through metabolically engineered recombinant Escherichia coli

Microbial Cell Factories, Journal Year: 2024, Volume and Issue: 23(1)

Published: Feb. 1, 2024

Abstract Background The biosynthesis of human milk oligosaccharides (HMOs) using several microbial systems has garnered considerable interest for their value in pharmaceutics and food industries. 2′-Fucosyllactose (2′-FL), the most abundant oligosaccharide HMOs, is usually produced chemical synthesis with a complex toxic process. Recombinant E. coli strains have been constructed by metabolic engineering strategies to produce 2′-FL, but low stoichiometric yields (2′-FL/glucose or glycerol) are still far from meeting requirements industrial production. sufficient carbon flux 2′-FL major challenge. As such, it great significance construction recombinant high yield. Results In present study, we designed pathway fructose theoretical yield 0.5 mol 2′-FL/mol fructose. involves five key enzymes: phosphomannomutase (ManB), mannose-1-phosphate guanylytransferase (ManC), GDP- d -mannose 4,6-dehydratase (Gmd), l -fucose synthase (WcaG), α-1,2-fucosyltransferase (FucT). Based on starting strain SG104, series metabolically engineered deleting genes pfkA , pfkB pgi replacing original promoter lacY . co-expression ManB, ManC, Gmd, WcaG, FucT were optimized, nine enzymes screened improve 2′-FL. Furthermore, gene gapA was regulated further enhance production, highest (0.498 fructose) achieved RFL38 (SG104 ΔpfkAΔpfkBΔpgi119-lacYΔwcaF :: 119-gmd-wcaG-manC-manB 119 -AGGAGGAGG- harboring plasmid P30). scaled-up reaction, 41.6 g/L (85.2 mM) fed-batch bioconversion, corresponding 0.482 0.986 lactose. Conclusions optimized strategies. This first time realize biological production yields. study also provides an important reference obtain suitable distribution between glycolysis.

Language: Английский

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High-Level Production of l-Fucose by Plasmid-Free or Antibiotic-Independent Metabolically Engineered Escherichia coli Strains

ACS Sustainable Chemistry & Engineering, Journal Year: 2024, Volume and Issue: 12(17), P. 6619 - 6628

Published: April 17, 2024

l-Fucose is an important monosaccharide unit that exists in various biomasses, especially microalgae. Microbial l-fucose using metabolically engineered strains has attracted attention due to its high yield and industrial feasibility. Previously, we Escherichia coli MG1655 efficiently produce 2′-fucosyllactose by genomic integration. Herein, this plasmid-free strain was further integrating a specific α-l-fucosidase gene deleting the degradation pathway. Its effectiveness of biosynthesis inducer-free fermentation demonstrated both shake-flask fed-batch cultivation with titers 2.74 21.15 g/L, respectively. The precursor GDP-l-fucose supply strengthened obviously enhance introducing single plasmid expressing four pathway genes. hok/sok system introduced promote stabilization without antibiotic. final could antibiotics, 6.83 35.68 g/L fed-cultivation cultivation,

Language: Английский

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Efficient production of 3-fucosyllactose in a plasmid-free engineered Escherichia coli through strengthening of the GDP-fucose pathway

Food Bioscience, Journal Year: 2024, Volume and Issue: 59, P. 104154 - 104154

Published: April 19, 2024

Language: Английский

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