Production of Lacto-N-triose II and Lacto-N-neotetraose from Chitin by a Novel β-N-Acetylhexosaminidase Expressed in Pichia pastoris

Liu, Yihao; Yan, Qiaojuan; Ma, Jinlu; Yang, Shaoqing; Li, Ting; Jiang, Zhengqiang

doi:10.1021/acssuschemeng.0c02911

Cited by 14 publications

(11 citation statements)

References 48 publications

(140 reference statements)

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“…Numerous studies have been conducted on the production of LNT II from chitin biomass or its hydrolysates (Table 1 ). Using a novel β- N -acetylhexosaminidase expressed by Pichia pastoris , 8.6 g L −1 of LNT II was produced from the reaction system composed of (GlcNAc) 2 (glycosyl donor), one of chitin hydrolysates and β-lactose (glycosyl acceptor) [ 8 ]. However, a large amount of GlcNAc monomer remains in the reaction system due to the transglycosylation of β- N -acetylhexosaminidase towards (GlcNAc) 2 and the hydrolysis of chitin by PbChi70, which is a potential issue to be considered.…”

Section: Resultsmentioning

confidence: 99%

“…To date, more than 200 different structures have been found and identified in HMOs, composed of five various monosaccharides including glucose (Glc), galactose (Gal), N -acetylglucosamine (GlcNAc), L-fucose (Fuc), and sialic acid (SA) [ 7 ]. Lacto- N -triose II (LNT II, GlcNAc β 1-3Gal β 1-4Glc) is an important precursor serving as a backbone to synthesize different HMOs, such as lacto- N -neotetraose (LNnT, Gal β 1-4GlcNAc β 1-3Gal β 1-4Glc) and lacto- N -tetraose (LNT, Gal β 1-3GlcNAc β 1-3Gal β 1-4Glc), which itself is generated by the catalytic reaction of β-1,3-acetylglucosaminyltransferase (LgtA) on GlcNAc and β - lactose substrate [ 8 ]. LNnT and LNT are also important components of HMOs, and they can be further fucosylated or sialylated, thereby generating other HMOs, such as lacto- N -fucopentaose II (LNFP-II), S-LNF II, lacto- N -neofucopentaoses (LNnFP I), among others [ 9 ].…”

Section: Introductionmentioning

confidence: 99%

“…Interestingly, GlcNAc is a good and low-cost substrate material, having been employed for the production of various chemical products, such as N -acetylneuraminic acid (Neu5Ac) [ 18 ], lipids [ 19 ], LNT II, and LNnT [ 8 ]. GlcNAc has a monomeric structure of chitin, which is a relatively abundant biomass in nature [ 20 ].…”

Section: Introductionmentioning

confidence: 99%

“…For example, Neu5Ac was produced from GlcNAc by whole-cell catalysis containing overexpressed N -acetylglucosamine 2-epimerase and N -acetylneuraminic acid lyase [ 18 ]. A three-enzyme synthetic system was also developed for the production of LNT II and LNnT from powdery chitin with a titer of 8.6 and 2.0 g L −1 , respectively [ 8 ]. However, there are few available reports about the production of LNT II using GlcNAc substrate by a metabolically engineered cell factory, even though metabolic engineering is recognized as a powerful strategy to synthesize various HMOs.…”

Section: Introductionmentioning

confidence: 99%

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Engineering Escherichia coli for highly efficient production of lacto-N-triose II from N-acetylglucosamine, the monomer of chitin

Zhu

et al. 2021

Biotechnol Biofuels

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Background Lacto-N-triose II (LNT II), an important backbone for the synthesis of different human milk oligosaccharides, such as lacto-N-neotetraose and lacto-N-tetraose, has recently received significant attention. The production of LNT II from renewable carbon sources has attracted worldwide attention from the perspective of sustainable development and green environmental protection. Results In this study, we first constructed an engineered E. coli cell factory for producing LNT II from N-acetylglucosamine (GlcNAc) feedstock, a monomer of chitin, by introducing heterologous β-1,3-acetylglucosaminyltransferase, resulting in a LNT II titer of 0.12 g L−1. Then, lacZ (lactose hydrolysis) and nanE (GlcNAc-6-P epimerization to ManNAc-6-P) were inactivated to further strengthen the synthesis of LNT II, and the titer of LNT II was increased to 0.41 g L−1. To increase the supply of UDP-GlcNAc, a precursor of LNT II, related pathway enzymes including GlcNAc-6-P deacetylase, glucosamine synthase, and UDP-N-acetylglucosamine pyrophosphorylase, were overexpressed in combination, optimized, and modulated. Finally, a maximum titer of 15.8 g L−1 of LNT II was obtained in a 3-L bioreactor with optimal enzyme expression levels and β-lactose and GlcNAc feeding strategy. Conclusions Metabolic engineering of E. coli is an effective strategy for LNT II production from GlcNAc feedstock. The titer of LNT II could be significantly increased by modulating the gene expression strength and blocking the bypass pathway, providing a new utilization for GlcNAc to produce high value-added products.

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Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Engineering Escherichia coli for highly efficient production of lacto-N-triose II from N-acetylglucosamine, the monomer of chitin

Zhu

et al. 2021

Biotechnol Biofuels

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“…In 2020, Liu et al. expressed β- N -acetylhexosaminidase (HaHex74) from Haloferula sp. in Pichia pastoris .…”

Section: Hmos Production Using Microbial Systemsmentioning

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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Synthesis of 2′-fucosyllactose from apple pomace–derived xyloglucan oligosaccharides by an α-L-fucosidase from Pedobacter sp. CAU209

Shi¹,

Yang²,

Wang³

et al. 2023

Appl Microbiol Biotechnol

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Production of Lacto-N-triose II and Lacto-N-neotetraose from Chitin by a Novel β-N-Acetylhexosaminidase Expressed in Pichia pastoris

Cited by 14 publications

References 48 publications

Engineering Escherichia coli for highly efficient production of lacto-N-triose II from N-acetylglucosamine, the monomer of chitin

Engineering Escherichia coli for highly efficient production of lacto-N-triose II from N-acetylglucosamine, the monomer of chitin

Engineered Microbial Routes for Human Milk Oligosaccharides Synthesis

Synthesis of 2′-fucosyllactose from apple pomace–derived xyloglucan oligosaccharides by an α-L-fucosidase from Pedobacter sp. CAU209

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