Discovery and characterization of a thermostable CALB-type lipase for efficient ethyl hexanoate synthesis in aqueous systems.

Candida antarctica lipase B (CALB) is a prototypical biocatalyst, distinguished by its remarkable catalytic efficiency in industrial ester synthesis. In this study, a novel lipase, XyEst, from Xylariales sp. PMI_506, was identified via the construction of a sequence similarity network and structural homology analysis using CALB as a reference. Despite sharing only 23.83% sequence identity with CALB, XyEst adopts a highly similar overall fold, indicating the presence of a conserved catalytic framework. The recombinant enzyme (rXyEst) displayed optimal activity at 30 °C and pH 4 for ethyl hexanoate synthesis. Notably, rXyEst exhibited remarkable thermal stability, retaining over 30% of its activity after 1 h at 70 °C. Furthermore, rXyEst showed strong tolerance to diverse metal ions and surfactants. In aqueous systems, rXyEst efficiently catalyzed ethyl hexanoate formation, achieving 65% conversion within 8 h. Collectively, these findings highlight XyEst as a thermostable biocatalyst with promising potential for sustainable industrial ester synthesis.