Jul 2026· Journal of Agricultural and Food Chemistry· 0 citations· 45 references
Medicine
Abstract
d-Allulose 3-epimerase (DAEase) catalyzes d-fructose conversion to d-allulose, but the poor thermostability of Clostridium cellulolyticum H10 DAEase limits its industrial application. Here, we enhanced DAEase thermostability by targeting the subunit interface using PROSS-guided combinatorial engineering and spatial clustering. Candidate mutations were classified into interface core, interface-adjacent, and distal regions, followed by stepwise iterative combination. Two mutants, M5 and M6, retained WT-like activity but showed markedly improved thermostability. The Tm values of M5 and M6 increased by 11.4 and 12.4 °C, respectively, while their half-lives at 65 °C increased 3-fold and 12-fold. Structural analysis indicated that interface mutations promoted salt-bridge reconstruction, distal mutations stabilized monomers, and interface-adjacent mutations optimized the assembly microenvironment. This spatially coordinated strategy provides an effective approach for engineering thermostable multimeric enzymes.
d-Allulose is a promising low-calorie rare sweetener; however, its industrial production is limited by the insufficient catalytic activity and thermostability of d-allulose 3-epimerases (DAEases). In this study, a combinatorial engineering strategy integrating lid-domain modulation and computational design was develope...
Qi-Chen Liu, Xin-Rui Tang, Wen-Jing Sun et al.· Journal of Agricultural and...· 0 citations
The industrial application of enzyme catalysts is often constrained by the trade-off between thermostability and catalytic activity. Here, a region-focused engineering strategy was applied to a thermophilic GH10 xylanase to simultaneously improve both properties. The strategy integrates qProtein-guided hydrophobic clus...
Zhaoran Li, Zhixin Dou, Sha Zhao et al.· Journal of Agricultural and...· 0 citations
L-asparaginase (ASNase) is used both for treating acute lymphoblastic leukemia and as a processing aid to hydrolyze L-asparagine prior to heating, thereby suppressing acrylamide formation in foods and certain traditional Chinese medicinal materials. Industrial pretreatment, however, typically involves high temperature,...
Linyu Luo, Sheng-Yu Zhang, Jingbo Zhou et al.· International Journal of Bio...· 0 citations
This work rationally engineered flexible loops near the active site via molecular dynamics, NMsim, NMsim, sequence conservation analysis, and MAESTRO prediction, and obtained the optimal double mutant M1, which showed 1.54-fold higher specific activity and nearly 2-fold improved pH stability at pH 8, with a nearly doub...
Yuan Li, Yuwei Ma, Tongli Li et al.· Journal of Agricultural and...· 0 citations
Xylanases with high catalytic efficiency and environmental robustness are important for lignocellulosic biomass valorization, but many enzymes are rapidly inactivated under the alkaline and high-temperature conditions used in industrial processes. In this study, a computationally guided rational-design strategy was dev...
Chun-Lin Tan, Xin Yu, Lan-Xi Sun et al.· International Journal of Bio...· 0 citations
This study provides a practical strategy for engineering thermostable pectate lyases with improved catalytic performance by developing a multidimensional consensus computational framework integrating sequence conservation, structural dynamics, and thermodynamic prediction to identify functional mutation hotspots in PcP...
Ziqi Hou, Gen Lu, Tong Shu et al.· Journal of Agricultural and...· 0 citations
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