Formation of flavone di-O-glucosides using a glycosyltransferase from Bacillus cereus.

Microbial UDP-glycosyltransferases can convert many small lipophilic compounds into glycons using uridinediphosphate- activated sugars. The glycosylation of flavonoids affects solubility, stability, and bioavailability. The gene encoding the UDP-glycosyltransferase from Bacillus cereus, BcGT-3, was cloned by PCR and sequenced. BcGT-3 was expressed in Escherichia coli BL21 (DE3) with a glutathione S-transferase tag and purified using a glutathione Stransferase affinity column. BcGT-3 was tested for activity on several substrates including genistein, kaempferol, luteolin, naringenin, and quercetin. Flavonols were the best substrates for BcGT-3. The enzyme dominantly glycosylated the 3-hydroxyl group, but the 7-hydroxyl group was glycosylated when the 3-hydroxyl group was not available. The kaempferol reaction products were identified as kaempferol-3-O-glucoside and kaempferol- 3,7-O-diglucoside. Kaempferol was the most effective substrate tested. Based on HPLC, LC/MS, and NMR analyses of the reaction products, we conclude that BcGT-3 can be used for the synthesis of kaempferol 3,7-O-diglucose.