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.
Ahn
Byoung Chan
BC
Department of Bioscience and Biotechnology, Bio/Molecular Informatics Center, Konkuk University, Seoul 143-701, Korea.
Kim
Bong Gyu
BG
Jeon
Young Min
YM
Lee
Eun Jeong
EJ
Lim
Yoongho
Y
Ahn
Joong-Hoon
JH
D001409:Bacillus cereus / Q000201:enzymology* / Q000235:genetics; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics; D044948:Flavonols / Q000378:metabolism*; D005960:Glucosides / Q000378:metabolism*; D006031:Glycosylation; D016695:Glycosyltransferases / Q000235:genetics / Q000378:metabolism*; D044949:Kaempferols / Q000378:metabolism; D016133:Polymerase Chain Reaction; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism
D001426:Bacterial Proteins; D044948:Flavonols; D005960:Glucosides; D044949:Kaempferols; D011993:Recombinant Fusion Proteins; C006552:kaempferol; D016695:Glycosyltransferases