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Functional identification of two novel carbohydrate-binding modules of glucuronoxylanase CrXyl30 and their contribution to the lignocellulose saccharification
BACKGROUND: Glycoside hydrolase (GH) family 30 xylanases are a distinct group of xylanases, most of which have a highly specific catalytic activity for glucuronoxylan. Since GH30 xylanases do not normally carry carbohydrate-binding modules (CBMs), our knowledge of the function of their CBMs is lacki...
Autores principales: | , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
BioMed Central
2023
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9996879/ https://www.ncbi.nlm.nih.gov/pubmed/36890582 http://dx.doi.org/10.1186/s13068-023-02290-7 |
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author | Liu, Jiawen Zhu, Jingrong Xu, Qian Shi, Rui Liu, Cong Sun, Di Liu, Weijie |
author_facet | Liu, Jiawen Zhu, Jingrong Xu, Qian Shi, Rui Liu, Cong Sun, Di Liu, Weijie |
author_sort | Liu, Jiawen |
collection | PubMed |
description | BACKGROUND: Glycoside hydrolase (GH) family 30 xylanases are a distinct group of xylanases, most of which have a highly specific catalytic activity for glucuronoxylan. Since GH30 xylanases do not normally carry carbohydrate-binding modules (CBMs), our knowledge of the function of their CBMs is lacking. RESULTS: In this work, the CBM functions of CrXyl30 were investigated. CrXyl30 was a GH30 glucuronoxylanase containing tandem CBM13 (CrCBM13) and CBM2 (CrCBM2) at its C terminus, which was identified in a lignocellulolytic bacterial consortium previously. Both CBMs could bind insoluble and soluble xylan, with CrCBM13 having binding specificity for the xylan with l-arabinosyl substitutions, whereas CrCBM2 targeted l-arabinosyl side chains themselves. Such binding abilities of these two CBMs were completely different from other CBMs in their respective families. Phylogenetic analysis also suggested that both CrCBM13 and CrCBM2 belong to novel branches. Inspection of the simulated structure of CrCBM13 identified a pocket that just accommodates the side chain of 3(2)-alpha-l-arabinofuranosyl-xylotriose, which forms hydrogen bonds with three of the five amino acid residues involved in ligand interaction. The truncation of either CrCBM13 or CrCBM2 did not alter the substrate specificity and optimal reaction conditions of CrXyl30, whereas truncation of CrCBM2 decreased the k(cat)/K(m) value by 83% (± 0%). Moreover, the absence of CrCBM2 and CrCBM13 resulted in a 5% (± 1%) and a 7% (± 0%) decrease, respectively, in the amount of reducing sugar released by the synergistic hydrolysis of delignified corncob whose hemicellulose is arabinoglucuronoxylan, respectively. In addition, fusion of CrCBM2 with a GH10 xylanase enhanced its catalytic activity against the branched xylan and improved the synergistic hydrolysis efficiency by more than fivefold when delignified corncob was used as substrate. Such a strong stimulation of hydrolysis resulted from the enhancement of hemicellulose hydrolysis on the one hand, and the cellulose hydrolysis is also improved according to the lignocellulose conversion rate measured by HPLC. CONCLUSIONS: This study identifies the functions of two novel CBMs in CrXyl30 and shows the good potential of such CBMs specific for branched ligands in the development of efficient enzyme preparations. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1186/s13068-023-02290-7. |
format | Online Article Text |
id | pubmed-9996879 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2023 |
publisher | BioMed Central |
record_format | MEDLINE/PubMed |
spelling | pubmed-99968792023-03-10 Functional identification of two novel carbohydrate-binding modules of glucuronoxylanase CrXyl30 and their contribution to the lignocellulose saccharification Liu, Jiawen Zhu, Jingrong Xu, Qian Shi, Rui Liu, Cong Sun, Di Liu, Weijie Biotechnol Biofuels Bioprod Research BACKGROUND: Glycoside hydrolase (GH) family 30 xylanases are a distinct group of xylanases, most of which have a highly specific catalytic activity for glucuronoxylan. Since GH30 xylanases do not normally carry carbohydrate-binding modules (CBMs), our knowledge of the function of their CBMs is lacking. RESULTS: In this work, the CBM functions of CrXyl30 were investigated. CrXyl30 was a GH30 glucuronoxylanase containing tandem CBM13 (CrCBM13) and CBM2 (CrCBM2) at its C terminus, which was identified in a lignocellulolytic bacterial consortium previously. Both CBMs could bind insoluble and soluble xylan, with CrCBM13 having binding specificity for the xylan with l-arabinosyl substitutions, whereas CrCBM2 targeted l-arabinosyl side chains themselves. Such binding abilities of these two CBMs were completely different from other CBMs in their respective families. Phylogenetic analysis also suggested that both CrCBM13 and CrCBM2 belong to novel branches. Inspection of the simulated structure of CrCBM13 identified a pocket that just accommodates the side chain of 3(2)-alpha-l-arabinofuranosyl-xylotriose, which forms hydrogen bonds with three of the five amino acid residues involved in ligand interaction. The truncation of either CrCBM13 or CrCBM2 did not alter the substrate specificity and optimal reaction conditions of CrXyl30, whereas truncation of CrCBM2 decreased the k(cat)/K(m) value by 83% (± 0%). Moreover, the absence of CrCBM2 and CrCBM13 resulted in a 5% (± 1%) and a 7% (± 0%) decrease, respectively, in the amount of reducing sugar released by the synergistic hydrolysis of delignified corncob whose hemicellulose is arabinoglucuronoxylan, respectively. In addition, fusion of CrCBM2 with a GH10 xylanase enhanced its catalytic activity against the branched xylan and improved the synergistic hydrolysis efficiency by more than fivefold when delignified corncob was used as substrate. Such a strong stimulation of hydrolysis resulted from the enhancement of hemicellulose hydrolysis on the one hand, and the cellulose hydrolysis is also improved according to the lignocellulose conversion rate measured by HPLC. CONCLUSIONS: This study identifies the functions of two novel CBMs in CrXyl30 and shows the good potential of such CBMs specific for branched ligands in the development of efficient enzyme preparations. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1186/s13068-023-02290-7. BioMed Central 2023-03-08 /pmc/articles/PMC9996879/ /pubmed/36890582 http://dx.doi.org/10.1186/s13068-023-02290-7 Text en © The Author(s) 2023 https://creativecommons.org/licenses/by/4.0/Open AccessThis article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/ (https://creativecommons.org/licenses/by/4.0/) . The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/ (https://creativecommons.org/publicdomain/zero/1.0/) ) applies to the data made available in this article, unless otherwise stated in a credit line to the data. |
spellingShingle | Research Liu, Jiawen Zhu, Jingrong Xu, Qian Shi, Rui Liu, Cong Sun, Di Liu, Weijie Functional identification of two novel carbohydrate-binding modules of glucuronoxylanase CrXyl30 and their contribution to the lignocellulose saccharification |
title | Functional identification of two novel carbohydrate-binding modules of glucuronoxylanase CrXyl30 and their contribution to the lignocellulose saccharification |
title_full | Functional identification of two novel carbohydrate-binding modules of glucuronoxylanase CrXyl30 and their contribution to the lignocellulose saccharification |
title_fullStr | Functional identification of two novel carbohydrate-binding modules of glucuronoxylanase CrXyl30 and their contribution to the lignocellulose saccharification |
title_full_unstemmed | Functional identification of two novel carbohydrate-binding modules of glucuronoxylanase CrXyl30 and their contribution to the lignocellulose saccharification |
title_short | Functional identification of two novel carbohydrate-binding modules of glucuronoxylanase CrXyl30 and their contribution to the lignocellulose saccharification |
title_sort | functional identification of two novel carbohydrate-binding modules of glucuronoxylanase crxyl30 and their contribution to the lignocellulose saccharification |
topic | Research |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9996879/ https://www.ncbi.nlm.nih.gov/pubmed/36890582 http://dx.doi.org/10.1186/s13068-023-02290-7 |
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