Guo_2025_J.Biotechnol__

GDSL esterases/lipases are a diverse group of hydrolytic enzymes with broad substrate specificity. However, none of them have been reported to catalyze the hydrolysis of chlorogenic acid (CGA), a key plant metabolite. To date, all known CGA-hydrolyzing enzymes have been identified in microorganisms and belong to the classical esterase family. In this study, we report the identification and characterization of a novel GDSL-type enzyme HaCAE from Helianthus annuus (sunflower), which exhibits robust CGA-hydrolyzing activity. HaCAE was successfully expressed and secreted in recombinant Pichia pastoris. The kinetic parameters of HaCAE were determined through an in vitro enzyme assay. Whole-cell biocatalysis demonstrated remarkable enzymatic efficiency, achieving hydrolysis of 17.31mM of CGA within 24hours. In silico analysis provided insights into the enzyme's catalytic mechanism and substrate selectivity. To the best of our knowledge, HaCAE is the first plant-derived CGA-hydrolyzing enzyme. Its high catalytic activity underscores its potential for food processing applications aimed at reducing CGA content and offers valuable biochemical insights into CGA metabolism in plants.