Extending the Structural Diversity of α‐Flavonoid Glycosides with Engineered Glucansucrases

Abstract

Flavonoids constitute an important class of bioactive molecules, the physicochemical properties of which can be modulated by glucosylation. A structurally guided approach has been used to isolate glucansucrases modified in their acceptor‐binding site and specialized for luteolin glucosylation. Of a small‐size library, we isolate mutants showing up to an 8‐fold increase in flavonoid conversion rate over that observed with the parental enzyme. Di‐ and triglucosylated luteolin derivatives never described before have been obtained. They exhibit 282‐ and 17 708‐fold increases in water solubility, respectively, and are protected from oxidation by the glucosylation reaction. Molecular docking enables insight into the product specificity of the best mutants. These results demonstrate that α‐transglucosylase engineering is a powerful means to generate highly specific catalysts for flavonoid glucosylation and deliver new structural scaffolds with increased bioavailability and high relevance for therapeutic applications.

Engineering a Carbohydrate-processing Transglycosidase into Glycosyltransferase for Natural Product Glycodiversification

“…They are frequently used in food, cosmetic and pharmaceuticals. Studies have demonstrated that flavonoid compounds generally exhibit anti-inflammatory, anti-oxidant and anti-tumor activities, mainly due to their polyphenol structures that protect against cardiovascular and coronary heart diseases or certain forms of cancer 34 44 45 46 47 . However, the major drawback of flavonoid compounds is their poor water solubility, which severely limits their application.…”

“…Glycosylation of the flavonoid catechin is also mediated by glucansucrases using sucrose or starch as the glucosyl donor 34 . However, multiple glucosylation products were obtained, including monoglucosyl and oligoglucosyl products.…”

“…By application of directed evolution methodologies, the activities of glycoside hydrolases or glycosynthases have been improved towards expanded sugar-donor or acceptor substrates 31 32 33 ; however, the transglycosylation activities on non-glycosylated natural-product acceptors still remain low. To meet this end, recently, a microbial amylosucrase and a rice transglucosidase have been engineered for improved transglycosylation activity on luteolin and kaempferol, repectively 34 35 .…”

“…In addition, the mutant enzyme gave rise to a series of α-glucosylated flavonoids novel to the natural product reservoir. Glycosylation is also known to improve the solubility, stability or bioavailability of flavonoids 8 34 35 . The newly developed enzyme has great potential applications in natural products glycodiversification.…”

Novel product specificity toward erlose and panose exhibited by multisite engineered mutants of amylosucrase

“…Site‐directed mutagenesis targeting Np AS active site enabled the distribution of natural products synthesized from sucrose to be finely tuned . Neutral drift approach and semi‐rational design allowed to sort out mutants with drastically improved ability to glucosylate acceptor molecules compared to the parental Np AS …”