首页|多细胞耦合转化N-乙酰氨基葡萄糖和乳糖生产唾液酸乳糖

多细胞耦合转化N-乙酰氨基葡萄糖和乳糖生产唾液酸乳糖

Multicellular coupling fermentation for 3'-sialyllactose conversion using N-acetyl-glucosamine and lactose

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唾液酸乳糖是母乳寡糖(human milk oligosaccharides,HMOs)中含量最丰富的唾液酸化低聚糖之一,对婴幼儿的健康发育有重要作用,但是其目前缺乏高效、廉价的生产工艺.针对该问题,本研究建立了多菌株两步法耦合生物合成唾液酸乳糖方法.第一步构建2株工程菌,大肠杆菌JM109(DE3)/pET28a-BT0453和JM109(DE3)/pET28a-nanA,用于耦合合成中间产物N-乙酰神经氨酸,在2株工程菌株细胞生物量比为1∶1,反应时间为32 h的条件下,得到的N-乙酰神经氨酸最高产量为20.4 g/L.第二步向上述发酵液中添加大肠杆菌JM109(DE3)/pET28a-neuA、JM109(DE3)/pET28a-nst和面包酵母,通过三菌株耦合发酵合成3'-唾液酸乳糖(3'-sialyllactose,3'-SL).在底物N-乙酰氨基葡萄糖、乳糖浓度均为200 mmol/L,面包酵母细胞生物量为150 g/L,辅因子Mg2+浓度为20 mmol/L的优化条件下,发酵24 h,发酵液中3'-唾液酸乳糖的最高产量达到55.04 g/L,底物N-乙酰氨基葡萄糖的转化率为43.47％.研究结果为低成本生产3'-唾液酸乳糖提供了一条新的技术路线.
Sialyllactose is one of the most abundant sialylated oligosaccharides in human milk oligosaccharides(HMOs),which plays an important role in the healthy development of infants and young children.However,its efficient and cheap production technology is still lacking presently.This study developed a two-step process employing multiple-strains for the production of sialyllactose.In the first step,two engineered strains,E.coli JM109(DE3)/pET28a-BT0453 and JM109(DE3)/pET28a-nanA,were constructed to synthesize the intermediate N-acetylneuraminic acid.When the ratio of the biomass of the two engineered strains was 1∶1 and the reaction time was 32 hours,the maximum yield of N-acetylneuraminic acid was 20.4 g/L.In the second step,E.coli JM109(DE3)/pET28a-neuA,JM109(DE3)/pET28a-nst and Baker's yeast were added to the above fermentation broth to synthesize 3'-sialyllactose(3'-SL).Using optimal conditions including 200 mmol/L N-acetyl-glucosamine and lactose,150 g/L Baker's yeast,20 mmol/L Mg2+,the maximum yield of 3'-SL in the fermentation broth reached 55.04 g/L after 24 hours of fermentation and the conversion rate of the substrate N-acetyl-glucosamine was 43.47％.This research provides an alternative technical route for economical production of 3'-SL.

3'-sialyllactoseN-acetyl-glucosaminecoupled fermentationtwo-step synthesishuman milk oligosaccharides

周文、游星、张洪涛、李忠霞、邓超明、许淳、黎玉

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江南大学生物工程学院糖化学与生物技术教育部重点实验室,江苏无锡 214122

汤臣倍健营养健康研究院,广东广州 510623

3'-唾液酸乳糖 N-乙酰氨基葡萄糖 耦合发酵 两步法合成 母乳寡糖

国家博士后特别资助项目中国博士后科学基金江苏省博士后基金解放军后勤保障部开放课题汤臣倍健营养科学研究基金开放课题

2014T704722012M209961301011BZZCWS21J2001TY202101096

2023

10.13345/j.cjb.230152

生物工程学报
中国科学院微生物研究所 中国微生物学会

生物工程学报

CSTPCDCSCD北大核心
影响因子：0.641
ISSN：1000-3061
年,卷(期)：2023.39(11)
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