Department of Bioscience and Biotechnology, Bio/Molecular Informatics Center, Konkuk University, Seoul, South Korea.
Appl Microbiol Biotechnol. 2012 Mar;93(6):2447-53. doi: 10.1007/s00253-011-3747-8. Epub 2011 Dec 13.
Biotransformation of flavonoids using Escherichia coli harboring nucleotide sugar-dependent uridine diphosphate-dependent glycosyltransferases (UGTs) commonly results in the production of a glucose conjugate because most UGTs are specific for UDP-glucose. The Arabidopsis enzyme AtUGT78D2 prefers UDP-glucose as a sugar donor and quercetin as a sugar acceptor. However, in vitro, AtUGT78D2 could use UDP-N-acetylglucosamine as a sugar donor, and whole cell biotransformation of quercetin using E. coli harboring AtUGT78D2 produced quercetin 3-O-N-acetylglucosamine. In order to increase the production of quercetin 3-O-N-acetylglucosamine via biotransformation, two E. coli mutant strains deleted in phosphoglucomutase (pgm) or glucose-1-phosphate uridylyltransferase (galU) were created. The galU mutant produced up to threefold more quercetin 3-O-N-acetylglucosamine than wild type, resulting in the production of 380-mg/l quercetin 3-O-N-acetylglucosamine and a negligible amount of quercetin 3-O-glucoside. These results show that construction of bacterial strains for the synthesis of unnatural flavonoid glycosides is possible through rational selection of the nucleotide sugar-dependent glycosyltransferase and engineering of the nucleotide sugar metabolic pathway in the host strain.
利用含有核苷酸糖依赖性尿苷二磷酸依赖性糖基转移酶（UGTs）的大肠杆菌进行黄酮类化合物的生物转化通常会产生葡萄糖缀合物，因为大多数 UGT 对 UDP-葡萄糖具有特异性。拟南芥酶 AtUGT78D2 更喜欢 UDP-葡萄糖作为糖供体和槲皮素作为糖受体。然而，在体外，AtUGT78D2 可以使用 UDP-N-乙酰葡萄糖胺作为糖供体，并且使用含有 AtUGT78D2 的大肠杆菌进行的槲皮素的全细胞生物转化产生了槲皮素 3-O-N-乙酰葡萄糖胺。为了通过生物转化增加槲皮素 3-O-N-乙酰葡萄糖胺的产量，创建了两种缺失磷酸葡糖变位酶（pgm）或葡萄糖-1-磷酸尿苷酰转移酶（galU）的大肠杆菌突变株。GalU 突变株产生的槲皮素 3-O-N-乙酰葡萄糖胺比野生型多三倍，导致产生 380mg/l 的槲皮素 3-O-N-乙酰葡萄糖胺和可忽略不计量的槲皮素 3-O-葡萄糖苷。这些结果表明，通过合理选择核苷酸糖依赖性糖基转移酶并对宿主菌株中的核苷酸糖代谢途径进行工程改造，构建用于合成非天然类黄酮糖苷的细菌菌株是可行的。
