Large-scale production of GDP-fucose and Lewis X by bacterial coupling

Koizumi, Satoshi; Endo, Tetsuo; Tabata, Kazuhiko; Nagano, Hiroko; Ohnishi, Junko; Ozaki, Asaichiro

doi:10.1038/sj.jim.7000055

Cited by 56 publications

(29 citation statements)

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“…Furthermore, phytic acid has been used in previous fermentations for oligosaccharide synthesis, presumably due to its chelating ability [20,21]. Substituting phytic acid for citrate in the reaction medium yields oligosaccharide levels comparable to the control reaction, containing neither citrate nor phytic acid (Fig.…”

Section: Citrate As Chelating Agentmentioning

confidence: 78%

Citrate Stimulates Oligosaccharide Synthesis in Metabolically Engineered Agrobacterium sp.

Ruffing

Chen

2011

Appl Biochem Biotechnol

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Agrobacterium sp. ATCC 31749 was previously shown to be an advantageous host for oligosaccharide production. Unexpectedly, the addition of citrate to the oligosaccharide synthesis reaction resulted in up to a sixfold improvement in the production N-aceytl-lactosamine, a disaccharide. The possible mechanisms for this citrate-induced stimulation of oligosaccharide production were investigated, including the consumption of citrate as a carbon and energy source, enhanced metal ion solubility from citrate chelation, and the ability of citrate to act as a buffer. The main mechanisms for the effect of citrate on oligosaccharide production were determined to be carbon and energy provision from citrate consumption and pH maintenance. ATCC 31749 was shown to co-metabolize citrate along with sucrose, a preferred carbon source, indicating the lack of a catabolite repression system in this Agrobacterium. Metabolic flux analysis suggested an increase in flux through TCA cycle for the citrate-containing reaction, which may provide additional energy supply to support enhanced oligosaccharide production. The citrate stimulation of oligosaccharide synthesis was shown to be unique to the Agrobacterium strain, as a similarly engineered Escherichia coli strain did not show significant improvement in oligosaccharide production with citrate addition. This work provides insight into the metabolism of Agrobacterium sp. ATCC 31749 and highlights important factors in whole-cell oligosaccharide synthesis.

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Section: Citrate As Chelating Agentmentioning

confidence: 78%

Citrate Stimulates Oligosaccharide Synthesis in Metabolically Engineered Agrobacterium sp.

Ruffing

Chen

2011

Appl Biochem Biotechnol

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“…Coupling of these three strains enabled production of 266 mM (134 g/L) of globotriose with lactose supplied as the acceptor. Similar strategies were also used to produce the disaccharide, N -acetyllactosamine, sialylated oligosaccharides, and a Lewis x trisaccharide [147–150]. Product concentrations were also impressive, ranging from 20 to 140 g/L.…”

Section: Production Of Nucleotide Sugars and Oligosaccharidesmentioning

confidence: 99%

Metabolic glycoengineering bacteria for therapeutic, recombinant protein, and metabolite production applications

et al. 2015

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Metabolic glycoengineering is a specialization of metabolic engineering that focuses on using small molecule metabolites to manipulate biosynthetic pathways responsible for oligosaccharide and glycoconjugate production. As outlined in this article, this technique has blossomed in mammalian systems over the past three decades but has made only modest progress in prokaryotes. Nevertheless, a sufficient foundation now exists to support several important applications of metabolic glycoengineering in bacteria based on methods to preferentially direct metabolic intermediates into pathways involved in lipopolysaccharide, peptidoglycan, teichoic acid, or capsule polysaccharide production. An overview of current applications and future prospects for this technology are provided in this report.

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“…Fructose was added to the reaction mixture as an energy source (Mahalingam et al 1999) and phytic acid as a chelator to prevent protein precipitation caused by magnesium and phosphate. Surfactant (Nymeen S-215) and xylene were added to the reaction mixture to permeablize E. coli cells (Koizumi et al 2000). HPLC confirmed and quantify the production of GDP-mannose by the recombinant E. coli (Fig.…”

Section: Production Of Gdp-mannosementioning

confidence: 99%

Synthesis of GDP-mannose using coupling fermentation of recombinant Escherichia coli

Jia

et al. 2011

Biotechnol Lett

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Glucokinase (glk), phosphomannomutase (manB), and mannose-1-phosphate guanylytransferase (manC) are needed for the biosynthesis of GDP-mannose. A recombinant E. coli strain over-expressing these three genes was constructed to produce guanosine 5'-diphosphate (GDP)-mannose, the donor of GDP-fucose, an essential substrate for synthesis of fucosyloligosaccharides. In addition, the glk, manB, and manC genes were individually cloned into the expression vector pET-22b (+) to construct three recombinant E. coli strains pET-glk, pET-manB and pET-manC, respectively. Fermentation of the recombinant strain BL21/pET-glk-manB-manC had a conversion rate of 23% from mannose to GDP-mannose under IPTG induction, while coupling fermentation of the three recombinant strains BL21/pET-glk, BL21/pET-manB, BL21/pET-manC resulted in a conversion rate of 33% under the same induction conditions.

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Large-scale production of GDP-fucose and Lewis X by bacterial coupling

Cited by 56 publications

References 0 publications

Citrate Stimulates Oligosaccharide Synthesis in Metabolically Engineered Agrobacterium sp.

Citrate Stimulates Oligosaccharide Synthesis in Metabolically Engineered Agrobacterium sp.

Metabolic glycoengineering bacteria for therapeutic, recombinant protein, and metabolite production applications

Synthesis of GDP-mannose using coupling fermentation of recombinant Escherichia coli

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