Production of
            <i>N</i>
            -Acetyl-
            <scp>d</scp>
            -Neuraminic Acid by Use of an Efficient Spore Surface Display System

Xu, Xiaoman; Gao, Chao; Zhang, Xifeng; Che, Bin; Ma, Cuiqing; Qiu, Jianhua; Tao, Fei; Xu, Ping

doi:10.1128/aem.00151-11

Cited by 49 publications

(44 citation statements)

References 41 publications

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“…Purified spores displaying Neu5Ac aldolase at a level of 2 U/ml were incubated in distilled water with different pH values. After 24 h of incubation at 50°C, the remaining Neu5Ac aldolase activities were deter-mined at pH 7.4 as described previously (22). The results showed that the spore-displayed Neu5Ac aldolase was stable at pH values below 10.5 (Fig.…”

mentioning

confidence: 93%

“…subtilis shuttle vector pEB03 (22). The B. subtilis strain WB600 with plasmid pEB03-cotG-nanA could effectively display Neu5Ac aldolase on its external surface.…”

mentioning

confidence: 99%

“…Nisin, a 34-amino-acid peptide produced by Lactococcus lactis subsp. lactis, was added at a concentration of 0.01% to inhibit the germination of Bacillus spores (4,22). With spore-displayed Neu5Ac aldolase, up to 65.4 mM (20.2 g liter Ϫ1 ) Neu5Ac was synthesized.…”

mentioning

confidence: 99%

“…The B. subtilis strain WB600 with plasmid pEB03-cotG-nanA could effectively display Neu5Ac aldolase on its external surface. Neu5Ac aldolase, presented on the surface of B. subtilis WB600 spores, was used for the production of Neu5Ac from ManNAc and pyruvate (22). For the integration of basecatalyzed epimerization of GlcNAc to ManNAc and Neu5Ac aldolase-catalyzed condensation of pyruvate with ManNAc to yield Neu5Ac, spore surface-displayed Neu5Ac aldolase should be stable under alkaline conditions.…”

mentioning

confidence: 99%

“…Spores of B. subtilis WB600(pEB03-cotG-nanA) were prepared as described previously (22). Purified spores displaying Neu5Ac aldolase at a level of 2 U/ml were incubated in distilled water with different pH values.…”

mentioning

confidence: 99%

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Chemoenzymatic Synthesis of N -Acetyl- d -Neuraminic Acid from N -Acetyl- d -Glucosamine by Using the Spore Surface-Displayed N -Acetyl- d -Neuraminic Acid Aldolase

Gao

Zhang

et al. 2011

Appl Environ Microbiol

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mentioning

confidence: 93%

“…subtilis shuttle vector pEB03 (22). The B. subtilis strain WB600 with plasmid pEB03-cotG-nanA could effectively display Neu5Ac aldolase on its external surface.…”

mentioning

confidence: 99%

mentioning

confidence: 99%

mentioning

confidence: 99%

mentioning

confidence: 99%

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Chemoenzymatic Synthesis of N -Acetyl- d -Neuraminic Acid from N -Acetyl- d -Glucosamine by Using the Spore Surface-Displayed N -Acetyl- d -Neuraminic Acid Aldolase

Gao

Zhang

et al. 2011

Appl Environ Microbiol

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Adsorption‐based immobilization of Caldicellulosiruptor saccharolyticus cellobiose 2‐epimerase on Bacillus subtilis spores

Yang

Xiao

et al. 2014

Biotech and App Biochem

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Nonrecombinant spore was examined as a novel immobilization support to adsorb enzymes. Caldicellulosiruptor saccharolyticus cellobiose 2-epimerase (CsCE), efficiently producing lactulose using lactose as a single substrate, was immobilized on Bacillus subtilis spores via adsorption. The immobilization process was optimized, and the properties of immobilized CsCE and the interactions between the enzyme and spores were also investigated. Under the optimized conditions (pH 4.5, temperature 4 °C, reaction time 2 H, and initial enzyme concentration 2.4 mg/mL), the maximum adsorbed amount of CsCE was 1.47 mg/10(11) spores, and the enzyme activity recovery was 79.4%. The spore-immobilized CsCE presented a higher pH and thermal stability than a free enzyme. Total desorption of the immobilized enzyme was only achieved by treatment with 1.0 M NaCl at pH 1.0, indicating a strong adsorption between CsCE and B. subtilis spores. Efficient binding may require a potent combination of electrostatic and hydrophobic interactions between spores and an enzyme. The immobilized CsCE was applied to produce 395 g/L lactulose after 4 H. Moreover, the spores could be regenerated and the spore-immobilized enzyme showed good reusability as it retained approximately 70% of its initial activity after eight recycles.

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Pathway Engineering of Bacillus subtilis for Enhanced N‐Acetylneuraminic Acid Production via Whole‐Cell Biocatalysis

Zhao

Tian

Shen

et al. 2019

Biotechnology Journal

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N‐acetylneuraminic acid (NeuAc) is a common sialic acid that has a wide range of applications in nutraceuticals and pharmaceuticals. However, low production efficiency and high environmental pollution associated with traditional extraction and chemical synthesis methods constrain the supply of NeuAc. Here, a biological approach is developed for food‐grade NeuAc production via whole‐cell biocatalysis by the generally regarded as safe (GRAS) bacterium Bacillus subtilis (B. subtilis). Promoters for controlling N‐acetylglucosamine 2‐epimerase (AGE) and NeuAc adolase (NanA) are optimized, yielding 32.84 g L−1 NeuAc production with a molar conversion rate of 26.55% from N‐acetylglucosamine (GlcNAc). Next, NeuAc production is further enhanced to 46.04 g L−1, which is 40.2% higher than that of the strain with promoter optimization, by expressing NanA from Staphylococcus hominis instead of NanA from Escherichia coli. To enhance the expression level of ShNanA, the N‐terminal coding sequences of genes with high expression levels are fused to the 5′‐end of the ShNanA gene, resulting in 56.82 g L−1 NeuAc production. Finally, formation of the by‐product acetoin from pyruvate is blocked by deleting the alsS and alsD genes, resulting in 68.75 g L−1 NeuAc production with a molar conversion rate of 55.57% from GlcNAc. Overall, a GRAS B. subtilis strain is demonstrated as a whole‐cell biocatalyst for efficient NeuAc production.

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Production of N -Acetyl- d -Neuraminic Acid by Use of an Efficient Spore Surface Display System

Cited by 49 publications

References 41 publications

Chemoenzymatic Synthesis of N -Acetyl- d -Neuraminic Acid from N -Acetyl- d -Glucosamine by Using the Spore Surface-Displayed N -Acetyl- d -Neuraminic Acid Aldolase

Chemoenzymatic Synthesis of N -Acetyl- d -Neuraminic Acid from N -Acetyl- d -Glucosamine by Using the Spore Surface-Displayed N -Acetyl- d -Neuraminic Acid Aldolase

Adsorption‐based immobilization of Caldicellulosiruptor saccharolyticus cellobiose 2‐epimerase on Bacillus subtilis spores

Pathway Engineering of Bacillus subtilis for Enhanced N‐Acetylneuraminic Acid Production via Whole‐Cell Biocatalysis

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