Abstract
α-L-Arabinofuranosidases (Abfs) play a crucial role in the degradation of hemicelluloses, especially arabinoxylans (AX). Most of the available characterized Abfs are from bacteria, while fungi, as natural decomposers, contain Abfs with little attention given. An arabinofuranosidase (ThAbf1), belonging to the glycoside hydrolase 51 (GH51) family, from the genome of the white-rot fungus Trametes hirsuta, was recombinantly expressed, characterized, and functionally determined. The general biochemical properties showed that the optimal conditions for ThAbf1 were pH 6.0 and 50°C. In substrate kinetics assays, ThAbf1 preferred small fragment arabinoxylo-oligosaccharides (AXOS) and could surprisingly hydrolyze di-substituted 23,33-di-L-arabinofuranosyl-xylotriose (A2,3XX). It also synergized with commercial xylanase (XYL) and increased the saccharification efficiency of arabinoxylan. The crystal structure of ThAbf1 indicated the presence of an adjacent cavity next to the catalytic pocket which led to the ability of ThAbf1 to degrade di-substituted AXOS. The narrow binding pocket prevents ThAbf1 from binding larger substrates. These findings have strengthened our understanding of the catalytic mechanism of GH51 family Abfs and provided a theoretical foundation for the development of more efficient and versatile Abfs to accelerate the degradation and biotransformation of hemicellulose in biomass.
Key points
• ThAbf1 from Trametes hirsuta degraded di-substituted arabinoxylo-oligosaccharide.
• ThAbf1 performed detailed biochemical characterization and kinetics.
• ThAbf1 structure has been obtained to illustrate the substrate specificity.
Graphical abstract
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Data availability
The datasets generated and analyzed during the current study are available from the corresponding author on reasonable request. The nucleotide sequence of ThAbf1 has been deposited at the NCBI GenBank under the accession number OQ473586. The structure of ThAbf1-apo has been deposited at the Research Collaboratory for Structural Bioinformatics Protein Databank (https://www.rcsb.org/) under PDB ID 8I0A.
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Funding
This study was supported by the National Key Research and Development Program of China (2018YFA0902000), the National Natural Science Foundation of China (No. 81872850), and PAPD (A Project Funded by the Priority Academic Program Development of Jiangsu Higher Education Institutions).
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ZS and WL conceived and designed the study. ZS, YC, LZ, LH, and ML conducted the experiments. ZS, LH, XG, ML, and WL analyzed the data. ZS and WL wrote the manuscript. FW, XY, and WL reviewed and edited the manuscript. All authors read and approved the manuscript.
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Si, Z., Cai, Y., Zhao, L. et al. Structure and function characterization of the α-L-arabinofuranosidase from the white-rot fungus Trametes hirsuta. Appl Microbiol Biotechnol 107, 3967–3981 (2023). https://doi.org/10.1007/s00253-023-12561-w
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DOI: https://doi.org/10.1007/s00253-023-12561-w