Kilogram scale chemical synthesis of 2′-fucosyllactose

Ágoston, Károly; Hederos, Markus; Bajza, István; Dékány, Gyula

doi:10.1016/j.carres.2019.03.006

Cited by 33 publications

(22 citation statements)

References 24 publications

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“…The 2’-FU used in this study was synthesized by our group according to a previously described method 25 ( Figure 1 A ). The purity of synthetic 2’-FL was examined using nuclear magnetic resonance (NMR) and high-resolution mass spectrometry.…”

Section: Resultsmentioning

confidence: 99%

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2’-Fucosyllactose Ameliorates Chemotherapy-Induced Intestinal Mucositis by Protecting Intestinal Epithelial Cells Against Apoptosis

Zhao¹,

Williams²,

Washington³

et al. 2022

Cellular and Molecular Gastroenterology and Hepatology

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This study shows a novel direct effect of 2'-fucosyllactose, an abundant oligosaccharide in human milk, in mitigating apoptosis stimulated by the chemotherapeutic agent 5-fluorouracil in intestinal epithelial cells. The results indicated that 2'-fucosyllactose ameliorates 5-fluorouracil-induced intestinal mucositis by supporting intestinal integrity for protection against injury.BACKGROUND & AIMS: Intestinal mucositis, a severe complication of antineoplastic therapeutics, is characterized by mucosal injury and inflammation in the small intestine. Therapies for the prevention and treatment of this disease are needed. We investigated whether 2'-fucosyllactose (2'-FL), an abundant oligosaccharide in human milk, protects intestinal integrity and ameliorates intestinal mucositis. METHODS:A mouse small intestinal epithelial (MSIE) cell line, mouse enteroid cultures, and human gastrointestinal tumor cell lines (AGS and HT29) were co-treated with the chemotherapy agent 5-fluorouracil (5-FU) and 2'-FL. Mice were injected intraperitoneally with 5-FU to induce intestinal mucositis. 2'-FL was administered in the drinking water to mice before (pretreatment) or concurrently with 5-FU injection. Body weight and pathologic changes were analyzed.RESULTS: 2'-FL alleviated 5-FU inhibition of cell growth in MSIE cells, but not in AGS and HT29 cells. The 5-FU-induced apoptosis in MSIE cells and enteroids was suppressed by 2'-FL. Compared with 5-FU treatment alone, 2'-FL pretreatment protected against body weight loss, and ameliorated inflammation scores, proinflammatory cytokine production, shortening of villi, epithelial cell apoptosis, goblet cell loss, and tight junctional complex disruption in the small intestine. 2'-FL concurrent treatment had less of an effect on intestinal mucositis than 2'-FL pretreatment. Interestingly, no effect of 2'-FL was observed on 5-FU-induced S-phase arrest in MSIE, AGS, and HT29 cells. Neither pretreatment nor concurrent treatment with 2'-FL affected 5-FU-induced inhibition of proliferation in MSIE cells. CONCLUSIONS:This study shows a novel direct effect of 2'-FL in protecting small intestinal epithelial cells against apoptosis stimulated by 5-FU, which may contribute to prevention of 5-FU-induced intestinal mucositis.

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Section: Resultsmentioning

confidence: 99%

“…We synthesized 2’-FL according to a previously reported method for preparation of 2'-FL at the kilogram level. 25 Briefly, a fucosyl thioglycoside was converted to its bromide by the addition of elemental bromine (Br 2 ) at 5°C in dichloromethane. Excess Br 2 was removed through the addition of cyclohexene.…”

Section: Methodsmentioning

confidence: 99%

2’-Fucosyllactose Ameliorates Chemotherapy-Induced Intestinal Mucositis by Protecting Intestinal Epithelial Cells Against Apoptosis

Zhao¹,

Williams²,

Washington³

et al. 2022

Cellular and Molecular Gastroenterology and Hepatology

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“…Among the various strategies developed to synthesize 2′-FL, whole-cell synthesis of 2′-FL using genetically engineered microorganisms represents a promising alternative with clear advantages over enzyme-catalyzed or chemo-enzymatic synthesis [ 7 – 9 ]. To achieve 2′-FL production in relevant microorganism strains, an ample supply of two key precursors is required, i.e., lactose and guanosine 5′‑diphosphate‑L‑fucose (GDP- l -fucose).…”

Section: Introductionmentioning

confidence: 99%

Improved production of 2′-fucosyllactose in engineered Saccharomyces cerevisiae expressing a putative α-1, 2-fucosyltransferase from Bacillus cereus

Meng

Zhang

et al. 2021

Microb Cell Fact

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Background 2′-fucosyllactose (2′-FL) is one of the most abundant oligosaccharides in human milk. It constitutes an authorized functional additive to improve infant nutrition and health in manufactured infant formulations. As a result, a cost-effective method for mass production of 2′-FL is highly desirable. Results A microbial cell factory for 2′-FL production was constructed in Saccharomyces cerevisiae by expressing a putative α-1, 2-fucosyltransferase from Bacillus cereus (FutBc) and enhancing the de novo GDP-l-fucose biosynthesis. When enabled lactose uptake, this system produced 2.54 g/L of 2′-FL with a batch flask cultivation using galactose as inducer and carbon source, representing a 1.8-fold increase compared with the commonly used α-1, 2-fucosyltransferase from Helicobacter pylori (FutC). The production of 2′-FL was further increased to 3.45 g/L by fortifying GDP-mannose synthesis. Further deleting gal80 enabled the engineered strain to produce 26.63 g/L of 2′-FL with a yield of 0.85 mol/mol from lactose with sucrose as a carbon source in a fed-batch fermentation. Conclusion FutBc combined with the other reported engineering strategies holds great potential for developing commercial scale processes for economic 2′-FL production using a food-grade microbial cell factory.

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“…More than 15 different HMO structures have been prepared by chemical synthesis [13,26]. Examples include lab-scale synthesis of lacto- N -fucopentaose (LNFP I) [26], gram-scale synthesis of LNT [27], and a recent report of kilogram-scale synthesis of 2′-FL [28]. The main challenge in chemical synthesis of HMOs is the high number of protecting group manipulations, which further increases with chain length and branching.…”

Section: Routes To Hmo Production Outside the Mammary Glandmentioning

confidence: 99%

“…Consequently, chemical synthesis of complex carbohydrates is often not a cost-efficient production method for large-scale synthesis of HMOs. Lowering the number of required intermediates and product purification steps is a key to obtaining a feasible, scalable process [28]. Although chemically synthesized 2′-FL and LNnT were originally registered for use as novel ingredients for infant formula [10], the use of microbial cell factories is currently the only method applied for industrial production of these HMOs, being a more economically feasible method [9].…”

Section: Routes To Hmo Production Outside the Mammary Glandmentioning

confidence: 99%

Synthesis of Human Milk Oligosaccharides: Protein Engineering Strategies for Improved Enzymatic Transglycosylation

et al. 2019

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Human milk oligosaccharides (HMOs) signify a unique group of oligosaccharides in breast milk, which is of major importance for infant health and development. The functional benefits of HMOs create an enormous impetus for biosynthetic production of HMOs for use as additives in infant formula and other products. HMO molecules can be synthesized chemically, via fermentation, and by enzymatic synthesis. This treatise discusses these different techniques, with particular focus on harnessing enzymes for controlled enzymatic synthesis of HMO molecules. In order to foster precise and high-yield enzymatic synthesis, several novel protein engineering approaches have been reported, mainly concerning changing glycoside hydrolases to catalyze relevant transglycosylations. The protein engineering strategies for these enzymes range from rationally modifying specific catalytic residues, over targeted subsite −1 mutations, to unique and novel transplantations of designed peptide sequences near the active site, so-called loop engineering. These strategies have proven useful to foster enhanced transglycosylation to promote different types of HMO synthesis reactions. The rationale of subsite −1 modification, acceptor binding site matching, and loop engineering, including changes that may alter the spatial arrangement of water in the enzyme active site region, may prove useful for novel enzyme-catalyzed carbohydrate design in general.

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Kilogram scale chemical synthesis of 2′-fucosyllactose

Cited by 33 publications

References 24 publications

2’-Fucosyllactose Ameliorates Chemotherapy-Induced Intestinal Mucositis by Protecting Intestinal Epithelial Cells Against Apoptosis

2’-Fucosyllactose Ameliorates Chemotherapy-Induced Intestinal Mucositis by Protecting Intestinal Epithelial Cells Against Apoptosis

Improved production of 2′-fucosyllactose in engineered Saccharomyces cerevisiae expressing a putative α-1, 2-fucosyltransferase from Bacillus cereus

Synthesis of Human Milk Oligosaccharides: Protein Engineering Strategies for Improved Enzymatic Transglycosylation

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