Organic Letters
Letter
and P187Q. As a result, the mutation of P187Q compromised
the selectivity to UDP-Gal, while the mutation of G15S, G15F,
and G15 V remarkably decreased the selectivity to UDP-Gal
and UDP-GlcNAc (Figure 3C). The enzyme function was not
affected in the S139A/A143S mutant. These results proved the
critical role of G15 and P187 in sugar donor promiscuity of
Sb3GT1.
We also tested the biotransformation of kaempferol by E. coli
cells harboring Sb3GT1 gene. Wild-type E. coli could
synthesize trace amounts of UDP-Glc, which could be utilized
as the sugar donor source in the reaction, without addition of
purified UDP-Glc. Finally, the strain could convert kaempferol
into its 3-O-glucoside at a conversion rate of >98% (Figure
In summary, Sb3GT1 is an efficient and regio-specific
glycosyltransferase to catalyze 3-O-glycosylation of flavonols
and cyanidin. It showed unprecedented donor- and substrate-
promiscuity to accept five sugar donors (UDP-Glc, UDP-Gal,
UDP-GlcNAc, UDP-Xyl, and UDP-Ara) and at least 17
substrates. The conversion rates were >98% when kaempferol,
quercetin, patuletin, and kaempferol 7-O-glucoside were used
as the substrates. The promiscuity of Sb3GT1 may result from
G15 and P187 residues, which change the sugar donor binding
pocket when binding with the acceptors. Sb3GT1 could be a
promising catalyst to increase structural diversity of flavonoid
3-O-glycosides through either enzymatic reactions or whole
cell biotransformations.
21573205), Beijing Natural Science Foundation (JQ18027),
and Young Elite Scientists Sponsorship Program by China
Association for Science and Technology (2016QNRC001).
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ASSOCIATED CONTENT
* Supporting Information
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Molecular cloning, expression, purification of Sb3GT1;
effects of reaction time, pH, temperature, divalent metal
ions; determination of kinetic parameters; enzyme assays
1
of Sb3GT1; scaled-up reactions; H and 13C NMR data
of glycosylated products; computational modeling and
molecular docking; site-directed mutagenesis (PDF)
Accession Codes
GenBank MK577650 (Sb3GT1, UGT78B4).
AUTHOR INFORMATION
Corresponding Authors
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ORCID
Author Contributions
§These authors contributed equally.
Notes
The authors declare no competing financial interest.
(8) (a) Feng, K. P.; Chen, R. D.; Xie, K. B.; Chen, D. W.; Guo, B. L.;
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ACKNOWLEDGMENTS
This work was supported by National Natural Science Foun-
dation of China (Nos. 81725023, 81891010/81891011,
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