Structural Elucidation and Mechanisms-Guided Engineering of a Promiscuous Esterase for Enhanced Polyurethane Depolymerization

Jiawei Liu; Mingna Zheng; Yuan Wen; Wei Xia; Xu Han; Jie Zhou; Weidong Liu; Ren Wei; Yanwei Li; Weiliang Dong; Min Jiang

doi:10.1016/j.eng.2026.02.008

Engineering ›› :202602008 DOI: 10.1016/j.eng.2026.02.008

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Structural Elucidation and Mechanisms-Guided Engineering of a Promiscuous Esterase for Enhanced Polyurethane Depolymerization

Jiawei Liu ^a^,^#
, Mingna Zheng ^c^,^#
, Yuan Wen ^a
, Wei Xia ^a
, Xu Han ^d
, Jie Zhou ^a^,^b
, Weidong Liu ^d^,^*
, Ren Wei ^e^,^*
, Yanwei Li ^c^,^*
, Weiliang Dong ^a^,^b^,^*
, Min Jiang ^a^,^b

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Abstract

Polyurethane (PU) is highly resistant to biodegradation, primarily due to the intrinsic stability of its urethane bond. Aside from a small number of amidases reported to hydrolyze (poly)urethane bonds, several promiscuous esterases have also been found to catalyze PU degradation. In this study, we clarified the ligand-free crystal structure of Aes72, an esterase enzyme that exhibits promiscuous hydrolytic activity toward carbamate and amide bonds, at a high resolution of 1.80 Å. We investigated the catalytic mechanism underlying urethane bond cleavage by Aes72 using multiscale quantum mechanics/molecular mechanics (QM/MM) simulations. Our findings indicate that the reaction mechanism consists of four concerted elementary steps, with the nucleophilic attack (step i) identified as the rate-determining step. The subsequent structure-guided engineering of Aes72 yielded several enhanced single mutants, ultimately resulting in a superior double mutant, F276A/L141I. This variant exhibited approximately a two-fold increase in catalytic efficacy toward bis(4-hydroxybutyl) (methylenebis(4,1-phenylene)) dicarbamate (BMC) hydrolysis and significantly enhanced degradation performance on two distinct polyether-based PU materials compared to the wild-type enzyme. Our findings provide essential mechanistic insights into the structure-function relationship of the promiscuous esterase Aes72 in PU degradation and demonstrate its potential applicability in bio-based plastic recycling.

Keywords

Esterase structure / Substrate pocket engineering / PU biodegradation / QM/MM analysis

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Jiawei Liu, Mingna Zheng, Yuan Wen, Wei Xia, Xu Han, Jie Zhou, Weidong Liu, Ren Wei, Yanwei Li, Weiliang Dong, Min Jiang. Structural Elucidation and Mechanisms-Guided Engineering of a Promiscuous Esterase for Enhanced Polyurethane Depolymerization. Engineering 202602008 DOI:10.1016/j.eng.2026.02.008

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