Large-scale production of UDP-galactose and globotriose by coupling metabolically engineered bacteria
Large-scale production of UDP-galactose and globotriose by coupling metabolically engineered bacteria
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DOI：
10.1038/nbt0998-847
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发表时间：
1998-09-01
影响因子：
46.9
通讯作者：
Ozaki, A
中科院分区：
文献类型：
--
作者：
Koizumi, S;Endo, T;Ozaki, A
利用重组大肠杆菌和产氨棒状杆菌的组合，建立了尿苷5 '-二磷酸半乳糖（UDP-Gal）的大规模生产系统。重组大肠产生过表达UDP-Gal生物合成基因galT、galK和galU的大肠杆菌。C.产氨酶有助于从廉价的尿苷三磷酸（UTP）前体--尿酸产生尿苷三磷酸（UTP），尿苷三磷酸是UDP-Gal生物合成的底物。以赤霉酸和半乳糖为起始原料，反应21 h后，UDP-Gal积累到72 mM（44 g/L）。将表达淋病奈瑟氏菌α 1，4-半乳糖基转移酶基因的大肠杆菌细胞与该UDP-Gal生产系统偶联，在以赤霉酸、半乳糖和乳糖开始的36小时反应后，产生372 mM（188 g/L）的球三糖（Gal α 1-4Gal β 1-4Glc），其为verotoxin受体的三糖部分。反应混合物中未观察到低聚糖副产物。该菌株的球三糖产量比UDP-Gal高出数倍。利用代谢工程重组E.大肠杆菌与核苷5 ′-三磷酸生产微生物的结合，以及通过将糖核苷酸生产系统与糖基转移酶偶联来生产寡糖的概念，可以应用于其他糖核苷酸和寡糖的生产。
A large-scale production system of uridine 5'-diphospho-galactose (UDP-Gal) has been established by the combination of recombinant Escherichia coli and Corynebacterium ammoniagenes. Recombinant E. coli that overexpress the UDP-Gal biosynthetic genes galT, galK, and galU were generated. C. ammoniagenes contribute the production of uridine triphosphate (UTP), a substrate for UDP-Gal biosynthesis, from erotic acid, an inexpensive precursor of UTP. UDP-Gal accumulated to 72 mM (44 g/L) after a 21 h reaction starting with erotic acid and galactose, When E. coli cells that expressed the alpha 1,4-galactosyltransferase gene of Neisseria gonorrhoeae were coupled with this UDP-Gal production system, 372 mM (188 g/L) globotriose (Gal alpha 1-4Gal beta 1-4Glc), a trisaccharide portion of verotoxin receptor, was produced after a 36 h reaction starting with erotic acid, galactose, and lactose. No oligosaccharide by-products were observed in the reaction mixture. The production of globotriose was several times higher than that of UDP-Gal. The strategy of producing sugar nucleotides by combining metabolically engineered recombinant E. coli with a nucleoside 5'-triphosphate producing microorganism, and the concept of producing oligosaccharides by coupling sugar nucleotide production systems with glycosyltransferases, can be applied to the manufacture of other sugar nucleotides and oligosaccharides.