Engineering of a UDP-Glycosyltransferase for the Efficient Whole-Cell Biosynthesis of Siamenoside I in <i>Escherichia coli</i>

Abstract

The combination of the insufficient availability and the complex structure of siamenoside I (SI), the sweetest glucoside isolated from <i>Siraitia grosvenorii</i> to date, limited its use as a natural sweetener. To solve this problem, an improved biocatalyst, UGT-M2, was semi-rationally created by engineering the uridine diphosphate glycosyltransferase UGT94-289-2 from <i>S. grosvenorii</i> for the monoglucosylation of mogroside IIIE (MG IIIE) to SI. Subsequently, an engineered <i>Escherichia coli</i> cell was constructed, which combined UGT-M2 with a UDP-glucose regeneration system to circumvent the need for expensive UDP-glucose to produce SI. After optimization, high-purity SI (>96.4%) was efficiently prepared from MG IIIE at a 1 L scale with a productivity of 29.78 g/(L day) and a molar yield of 76.5% and without using exogenous UDP-glucose. This study not only developed a whole-cell approach for the preparation of SI but also provided an alternative glycosyltransferase variant for SI biosynthesis with synthetic biology in the future.