A Flavone-5-O-glycosyltransferase\nObtained through\nDomain Shuffling and Its Application in the Bioproduction of Luteolin-5-O-glucoside

Luteolin-5-O-glucoside\nis a rare luteolin glycoside compound with\nenormous potential for application. In order to promote sustainability\nand a circular economy, environmentally friendly catalysts and catalytic\nprocesses are in demand to produce luteolin-5-O-glucoside. Using glycosyltransferase\ntransformation is an efficient method for synthesizing glycosylated\nflavonoids. However, most reported glycosyltransferases have not been\nable to catalyze glycosylation on the 5-OH site, and a method for\nthe sustainable production of luteolin-5-O-glucoside has not been\nestablished. In this paper, a glycosyltransferase DoUGT71A15 was cloned\nfrom Dichanthelium oligosanthes. The\nN-terminal fragment of ZmUGT707A8 (Met1–Met93) was inserted\ninto the N-terminus of DoUGT71A15 by domain shuffling, and a biological\ncatalyst ZmDo was obtained. ZmDo could convert luteolin to luteolin-5-O-glucoside\nwith <i>K</i><sub>m</sub> and <i>k</i><sub>cat</sub>/<i>K</i><sub>m</sub> values of 0.13 mM and 1355.8 s<sup>–1</sup>·M<sup>–1</sup>, respectively. Then, the\ncatalytic mechanism of region selectivity of ZmDo was studied by using\nbioinformatics methods, and the key site E89 was identified through\nalanine scanning. Finally, the strain BSZ containing the UDPG circulatory\nsystem, which could use cellobiose as a substrate and recycle UDP\nto synthesize UDPG, was used for efficient production of luteolin-5-O-glucoside,\nwith a yield of 2253 mg/L and a molar conversion rate of 95.8%. This\nresearch obtained a novel flavone-5-OH glycosyltransferase and produced\nluteolin-5-O-glucoside by bioconversion, which provided the possibility\nfor sustainable production and application of luteolin-5-O-glucoside.