Oxidation of cellulose fibers using LPMOs with varying allomorphic substrate preferences, oxidative regioselectivities, and domain structures

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Volume Title

A1 Alkuperäisartikkeli tieteellisessä aikakauslehdessä

Date

2024-04-15

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Language

en

Pages

13

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Carbohydrate Polymers, Volume 330

Abstract

Lytic polysaccharide monooxygenases (LPMOs) are excellent candidates for enzymatic functionalization of natural polysaccharides, such as cellulose or chitin, and are gaining relevance in the search for renewable biomaterials. Here, we assessed the cellulose fiber modification potential and catalytic performance of eleven cellulose-active fungal AA9-type LPMOs, including C1-, C4-, and C1/C4-oxidizing LPMOs with and without CBM1 carbohydrate-binding modules, on cellulosic substrates with different degrees of crystallinity and polymer chain arrangement, namely, Cellulose I, Cellulose II, and amorphous cellulose. The potential of LPMOs for cellulose fiber modification varied among the LPMOs and depended primarily on operational stability and substrate binding, and, to some extent, also on regioselectivity and domain structure. While all tested LPMOs were active on natural Cellulose I-type fibers, activity on the Cellulose II allomorph was almost exclusively detected for LPMOs containing a CBM1 and LPMOs with activity on soluble hemicelluloses and cello-oligosaccharides, for example NcAA9C from Neurospora crassa. The single-domain variant of NcAA9C oxidized the cellulose fibers to a higher extent than its CBM-containing natural variant and released less soluble products, indicating a more dispersed oxidation pattern without a CBM. Our findings reveal great functional variation among cellulose-active LPMOs, laying the groundwork for further LPMO-based cellulose engineering.

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Funding Information: This work was supported by the European Union's Horizon 2020 research and innovation programme under the umbrella of ERA-NET Cofund Action ForestValue [FunEnzFibres, grant agreement no. 773324 ] through funding from the Research Council of Norway [grant agreement no. 297907 ], the Academy of Finland [grant agreement no. 326359 ], and the Austrian Federal Ministry of Agriculture, Forestry, Environment and Water Management (BMLFUW) [grant agreement no. 101379 ]. The work was co-funded by the NorBioLab infrastructure grant [grant agreement no. 270038 ] from the Research Council of Norway . Anikó Várnai also acknowledges the Novo Nordisk Foundation for an Emerging Investigator Grant [grant no. NNF-0061165 ]. VTT thanks Tiina Pöhler for advice regarding to the BET surface area analysis and Riitta Alander, Mari Leino and Mirja Muhola for technical assistance and expresses gratitude to support by the FinnCERES Materials Bioeconomy Ecosystem. We thank Kasper Bay Tingsted and Pedro Emanuel Garcia Loureiro from Novozymes for providing enzymes. Funding Information: This work was supported by the European Union's Horizon 2020 research and innovation programme under the umbrella of ERA-NET Cofund Action ForestValue [FunEnzFibres, grant agreement no. 773324] through funding from the Research Council of Norway [grant agreement no. 297907], the Academy of Finland [grant agreement no. 326359], and the Austrian Federal Ministry of Agriculture, Forestry, Environment and Water Management (BMLFUW) [grant agreement no. 101379]. The work was co-funded by the NorBioLab infrastructure grant [grant agreement no. 270038] from the Research Council of Norway. Anikó Várnai also acknowledges the Novo Nordisk Foundation for an Emerging Investigator Grant [grant no. NNF-0061165]. VTT thanks Tiina Pöhler for advice regarding to the BET surface area analysis and Riitta Alander, Mari Leino and Mirja Muhola for technical assistance and expresses gratitude to support by the FinnCERES Materials Bioeconomy Ecosystem. We thank Kasper Bay Tingsted and Pedro Emanuel Garcia Loureiro from Novozymes for providing enzymes. Publisher Copyright: © 2024 The Authors

Keywords

AA9 LPMOs, Cellulose, Enzymatic fiber engineering, Fluorescence, Functional variation

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Citation

Støpamo, F G, Sulaeva, I, Budischowsky, D, Rahikainen, J, Marjamaa, K, Potthast, A, Kruus, K, Eijsink, V G H & Várnai, A 2024, ' Oxidation of cellulose fibers using LPMOs with varying allomorphic substrate preferences, oxidative regioselectivities, and domain structures ', Carbohydrate Polymers, vol. 330, 121816 . https://doi.org/10.1016/j.carbpol.2024.121816