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. 2024 Aug 14;72(32):18078-18088.
doi: 10.1021/acs.jafc.4c03154. Epub 2024 Jul 30.

Development of New Multi-Glycosylation Routes to Facilitate the Biosynthesis of Sweetener Mogrosides from Bitter Immature Siraitia Grosvenorii Using Engineered Escherichia coli

Xinyi Lu  1   2 , Jianxiu Li  2 , Chuanqing Huang  2 , Zhefei Wang  2 , Yanchi Chen  2 , Shuiyuan Jiang  3 , Jianbin Li  1 , Nengzhong Xie  2
Affiliations

Development of New Multi-Glycosylation Routes to Facilitate the Biosynthesis of Sweetener Mogrosides from Bitter Immature Siraitia Grosvenorii Using Engineered Escherichia coli

Xinyi Lu et al. J Agric Food Chem. .

Abstract

Mogrosides, which have various pharmacological activities, are mainly extracted from Siraitia grosvenorii (Luo Han Guo) and are widely used as natural zero-calorie sweeteners. Unfortunately, the difficult cultivation and long maturation time of Luo Han Guo have contributed to a shortage of mogrosides. To overcome this obstacle, we developed a highly efficient biosynthetic method using engineered Escherichia coli to synthesize sweet mogrosides from bitter mogrosides. Three UDP-glycosyltransferase (UGT) genes with primary/branched glycosylation catalytic activity at the C3/C24 sites of mogrosides were screened and tested. Mutant M3, which could catalyze the glycosylation of nine types of mogrosides, was obtained through enhanced catalytic activity. This improvement in β-(1,6)-glycosidic bond formation was achieved through single nucleotide polymorphisms and direct evolution, guided by 3D structural analysis. A new multienzyme system combining three UGTs and UDP-glucose (UDPG) regeneration was developed to avoid the use of expensive UDPG. Finally, the content of sweet mogrosides in the immature Luo Han Guo extract increased significantly from 57% to 95%. This study not only established a new multienzyme system for the highly efficient production of sweet mogrosides from immature Luo Han Guo but also provided a guideline for the high-value utilization of rich bitter mogrosides from agricultural waste and residues.

Keywords: Siraitia grosvenorii; UDP-glycosyltransferase; engineered Escherichia coli; multiglycosylation system; sweet mogrosides.

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