Amylosucrase from Deinococcus geothermalis can be modulated under different reaction conditions to produce novel quercetin 4′-O-α-d-isomaltoside

Abstract

Amylosucrase (ASase, EC.4.2.1.4) is well-known for its distinguishable property of transglycosylation of many flavonoids and phenolics. Quercetin has diverse biological functions, however, its use is limited due to poor solubility and bioavailability. ASase derived from Deinococcus geothermalis (DGAS) showed conditional preference for producing unusual quercetin glucosides (QGs). DGAS produced a variety of QGs including quercetin monoglucosides (QG1), diglucosides (QG2 and QG2′), and triglucoside from quercetin and sucrose. The newly synthesized QG2′ was recognized as a novel quercetin isomaltoside with an α-1,6 linkage branched at the −OH of C4′ in quercetin by mass and nuclear magnetic resonance spectra. With a higher conversion yield from quercetin to QGs (60–92%), the optimum conditions for producing QG2′ were examined under various pH and sucrose concentrations by response surface methodology. QG2′ was predominantly produced under acidic conditions (pH 5.0) and at high sucrose concentrations (1000–1500 mM). In contrast, QG1 was generated as an intermediate of consecutive glycosylation. Kinetic evaluations indicated that considerable differences of transglycosylation velocities were caused by the pH and buffer salts of the reaction, which had a 3.9-fold higher overall performance (kcat/K′m) of generating QG2′ at pH 5 compared to at pH 7. A rationale of unusual transglycosylations was demonstrated with a molecular docking simulation. Taken together, our study demonstrated that ASase can be used to synthesize unusually branched flavonoid glycosides from flavonol aglycones with clear patterns by modulating reaction conditions.