Sep 2026· Journal of Agricultural and Food Chemistry· 0 citations· 57 references
Abstract
Efficient xylose bioconversion is limited by the catalytic performance of xylulokinase (XylB), which phosphorylates d-xylulose to xylulose-5-phosphate. This study aimed to improve XylB from Lactococcus lactis N8 using a multi-dimensional semi-rational design strategy integrating sequence conservation, structure-based screening, mutagenesis, molecular docking, and molecular dynamics simulations. Asn301 was identified as a key functional hotspot, and Met259 was found to modulate the active-site hydrophobic environment. Combinatorial mutagenesis generated two superior variants, XylBM259H/N301G and XylBM259H/N301H, with more than 200% higher specific activity than the wild-type XylB (XylBWT) while retaining stable expression and environmental adaptability. Mechanistic analyses showed strengthened ATP binding, improved d-xylulose positioning, enhanced ligand affinity, and balanced local flexibility with structural compactness. These findings provide efficient XylB variants and a transferable strategy for engineering lignocellulose-related enzymes.
L-Xylulose, a potent α-glucosidase inhibitor and indispensable chiral precursor for L-nucleoside drugs, is highly valuable. However, chemical synthesis often suffers from poor atom economy and tedious protection and deprotection steps of functional groups. Here, a green biocatalytic route was explored by retrieving fiv...
Xiao-Lan Li, Zi-Ye Wang, Jia-Le Long et al.· Journal of Agricultural and...· 0 citations
Mannans are abundant plant hemicelluloses, and endo-β-mannanases are important biocatalysts for their conversion into functional manno-oligosaccharides. Here, we report the structural and functional characterization of a glycoside hydrolase family 134 β-mannanase from Aspergillus nidulans (AnGH134) and a structure-guid...
Sheng-Chia Chen, Po-Chih Kuo, Wen-Ming Chen et al.· Journal of Agricultural and...· 0 citations
Nitrile hydratase is a key enzyme for nitrile-to-amide hydration under mild conditions, yet its application is limited by low heterologous expression, poor solubility, and suboptimal catalysis. Here, we present an integrated strategy to enhance expression, assembly, and function of Pseudonocardia thermophila NHase (PtN...
Xiao-Ling You, Yu-Qing Chen, Zi-Ying Tan et al.· International Journal of Bio...· 0 citations
Thermostability is critical yet challenging to engineer in industrial enzymes. Here, we present an integrated workflow for enhancing the thermostability of GH11 xylanases through systematic Ser/Thr optimization. Gibbs free energy calculations on XynCDBFV identified 169 stabilizing single-site mutations involving Ser/Th...
Yuan-Yuan Zhe, Sisi Zhang, En-Heng Zhu et al.· Journal of Agricultural and...· 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
This study addresses a major bottleneck in dairy waste management by engineering a highly efficient, thermophilic enzyme for lactose hydrolysis, transforming the cheese manufacturing by-product whey into a high-value resource. While traditional glycoside hydrolase family 2 (GH2) enzymes suffer from poor industrial ther...
Si-Meng Ding, Jia-Jia Li, Jia-Hui Li et al.· Journal of Dairy Science· 0 citations
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