Rational Design of a β-Glycosidase with High Regiospecificity for Triterpenoid Tailoring

• Published in: Chembiochem : a European journal of chemical biology Vol. 16; no. 5; pp. 854 - 860
• Main Authors: Park, Sang Jin, Choi, Jung Min, Kyeong, Hyun-Ho, Kim, Song-Gun, Kim, Hak-Sung
• Format: Journal Article
• Language: English
• Published: Weinheim WILEY-VCH Verlag 23.03.2015; WILEY‐VCH Verlag
• Subjects: Actinomycetales - enzymology; Actinomycetales - genetics; beta-Glucosidase - chemistry; beta-Glucosidase - genetics; beta-Glucosidase - isolation & purification; beta-Glucosidase - metabolism; binding mode; Biocatalysis; glycosides; Models, Molecular; Molecular Conformation; Mutation; rational design; regiospecificity; Substrate Specificity; terpenoids; Triterpenes - chemistry; Triterpenes - metabolism; rational design; regiospecificity; terpenoids; binding mode; glycosides
• ISSN: 1439-4227; 1439-7633
• DOI: 10.1002/cbic.201500004

Triterpenoids with desired glycosylation patterns have attracted considerable attention as potential therapeutics for inflammatory diseases and various types of cancer. Sugar‐hydrolyzing enzymes with high substrate specificity would be far more efficient than other methods for the synthesis of such specialty triterpenoids, but they are yet to be developed. Here we present a strategy to rationally design a β‐glycosidase with high regiospecificity for triterpenoids. A β‐glycosidase with broad substrate specificity was isolated, and its crystal structure was determined at 2.0 Å resolution. Based on the product profiles and substrate docking simulations, we modeled the substrate binding modes of the enzyme. From the model, the substrate binding cleft of the enzyme was redesigned in a manner that preferentially hydrolyzes glycans at specific glycosylation sites of triterpenoids. The designed mutants were shown to produce a variety of specialty triterpenoids with high purity. Triterpenoids with desired glycosylation patterns are of great significance as potential therapeutics. A promiscuous β‐glycosidase was isolated and crystallized, and docking simulations led to rationally designed β‐glycosidases that produced specialty triterpenoids with high purity and regiospecificity.