Wang_2025_Arch.Microbiol_207_256

The esterase gene encoding EstJN1 of Clostridium butyricum, which was isolated from the pit cellar of Chinese liquor facility, was expressed. EstJN1 was identified as a novel GDSL esterase belonging to family II. The enzyme demonstrated a marked substrate preference for p-nitrophenyl butyrate, with optimal activity at a temperature of 40 degC and a pH of 7.0. Notably, EstJN1 displayed prominent tolerance to acidic conditions and organic solvents, particularly methanol, ethanol, and ethyl acetate. Moreover, the whole-cell biocatalyst of EstJN1 was found to efficiently catalyze the synthesis of ethyl hexanoate in heptane, achieving ester conversion rate of 80.5% under the optimized conditions of 40 degC temperature, 0.25 M ethanol and 0.2 M hexanoic acid concentrations. The distinctive characteristics of EstJN1 suggest a potential role in the biosynthesis of flavor compounds in Chinese liquor. EstJN1 has promising application in the synthesis of flavor esters and enhancement of aromatic profiles of alcoholic beverages.