期刊
MICROBIAL CELL FACTORIES
卷 21, 期 1, 页码 -出版社
BMC
DOI: 10.1186/s12934-022-01940-z
关键词
C-glycosides; Stenoloma chusanum; C-glycosyltransferases; Orientin; Vitexin; Metabolic engineering
资金
- National Natural Science Foundation of China [32270270]
- State Key Laboratory of Microbial Technology Open Projects Fund
By characterizing the CGT enzyme from Stenoloma chusanum, it was found to have C-glycosylation activity for phloretin, 2-hydroxynaringenin, and 2-hydroxyeriodictyol. Mutagenesis experiments revealed that a specific mutation increased C-glycosylation activity. Furthermore, the coupling catalysis of CjFNS I/F2H and the mutated CGT enzyme resulted in the conversion of naringenin into vitexin and isovitexin.
Background Flavonoid C-glycosides have many beneficial effects and are widely used in food and medicine. However, plants contain a limited number of flavonoid C-glycosides, and it is challenging to create these substances chemically. Results To screen more robust C-glycosyltransferases (CGTs) for the biosynthesis of flavonoid C-glycosides, one CGT enzyme from Stenoloma chusanum (ScCGT1) was characterized. Biochemical analyses revealed that ScCGT1 showed the C-glycosylation activity for phloretin, 2-hydroxynaringenin, and 2-hydroxyeriodictyol. Structure modeling and mutagenesis experiments indicated that the glycosylation of ScCGT1 may be initiated by the synergistic action of conserved residue His26 and Asp14. The P164T mutation increased C-glycosylation activity by forming a hydrogen bond with the sugar donor. Furthermore, when using phloretin as a substrate, the extracellular nothofagin production obtained from the Escherichia coli strain ScCGT1-P164T reached 38 mg/L, which was 2.3-fold higher than that of the wild-type strain. Finally, it is proved that the coupling catalysis of CjFNS I/F2H and ScCGT1-P164T could convert naringenin into vitexin and isovitexin. Conclusion This is the first time that C-glycosyltransferase has been characterized from fern species and provides a candidate gene and strategy for the efficient production of bioactive C-glycosides using enzyme catalysis and metabolic engineering.
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