BAHD acyltransferase ApAT2 catalyzes the acetylation of protostane-type tetracyclic triterpenes in Alisma plantago-aquatica.

Abstract

Alisma plantago-aquatica L. is a traditional Chinese medicinal herb used to treat urinary disorders. Acylated protostane-type tetracyclic triterpenes such as alisol B 23-acetate and alisol C 23-acetate are its major bioactive components and have been reported to exhibit improved antiviral activities. However, the content of these compounds is relatively low in plants, and the enzyme that catalyzes this acetylation process has not yet been identified. In this study, we constructed a transcriptome dataset for A. plantago-aquatica. Using co-expression analysis and enzyme activity screening, we successfully identified a benzyl alcohol O-acetyltransferase, anthocyanin O-hydroxycinnamoyltransferase, anthranilate N-hydroxycinnamoyl/benzoyltransferase, deacetylvindoline 4-O-acetyltransferase (BAHD), ApAT2. This enzyme catalyzes the acetylation of alisol B and alisol C but does not exhibit any catalytic activity toward alisol A. Molecular docking and site-directed mutagenesis confirmed that the M412 residue of ApAT2 played a critical role in its substrate specificity. Phylogenetic analysis suggests that the active pocket of ApAT2 possibly diverged via lineage-specific evolution, conferring specificity for protostane-type tetracyclic triterpenes. Furthermore, we established a semi-biosynthetic platform in Escherichia coli expressing ApAT2 to produce alisol B 23-acetate from alisol B. In conclusion, we identified the BAHD enzyme responsible for the acetylation of protostane-type tetracyclic triterpenes, thus providing new systems to produce these valuable compounds. We also identified a new molecular basis for the recognition and modification of diverse substrates by the BAHD family during plant evolution.

Abstract as supplied by the indexing source. © the authors / publisher.