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ENZYME&CESUMO: Enzymatic modification of lignocellulosics & Modeling interacting cellulose surfaces

Updated: Aug 31, 2023

Enzymes are biocatalysts that can be applied selective modification of cellulose properties in mild reaction conditions. Lytic polysaccharide monooxygenases (LPMOs) improve (ligno)cellulose reactivity towards enzymatic hydrolysis, refining and dissolution. In this project, we explore the effects of different types of LPMOs on cellulose properties focusing on fibrillary interactions. Modeling is carried out to access the influence of the LPMO oxidation defects both on cellulose and its hydration.

Main results

  • Heterologous production of cellulose oxidizing enzymes and elucidation of the effects of reaction parameters on oxidation efficiency

    • Gram scale production of different types of LPMOs (oxidation in position C1, C4 and C1+C4 in industrial production host (T. reesei) established which enabled large scale of application trials.

    • Key parameters affecting the enzymatic oxidation efficiency are interplay between reaction pH, nature of reductant and availability of added hydrogen peroxide, as well as stability of the enzyme (dependent on the enzyme variant)

  • Enzymatic oxidation of crystalline cellulose: effect on cellulose crystallites and mode of oxidation and modular structure of the enzymes

    • Computational modelling shows effect of oxidation point defects on cellulose crystals and their water interactions.

    • The LPMOs studied in more detail in the project contain two modules: catalytic module and carbohydrate binding module (CBM), latter of which is known to enhance binding of enzymes on cellulosic substrates. Modelling used to map CBM interactions on different cellulose crystal facets: preferential CBM binding to specific facets which connects with enzymatic activity

    • Oxidation of crystalline cellulose surfaces with LPMOs having different oxidation specificities (C1, C1+C4) and with& without CBM showed distinctly different QCM-D response curves, based on which the C1+C4 oxidising variant & CBM containing was most efficient in solubilisation of the crystal surface

  • Effect of enzymatic oxidation and hydrolysis, and small molecule salt cosolutes, on material behaviour of CNF

    • Rheology study showed that the enzymatic oxidation with LPMO loosened the CNF network, making it more liquid like. This enhan ced the ability of GH family 7 and 5 endoglucanases (from T. reesei) to hydrolysed the CNF, while effect on action of GH45 endoglucanase was negligible

    • Rheology characterization of the influence of show distinct differences between the effect of salt species. Very small salt concentrations result in significant CNF solution viscosity differences. Modelling connects NaCl cosolute effects on cellulose crystal interactions. We provide significant new insight on solution behaviour of cellulose. This can also affect the enzyme action on cellulose and interpretation of the rheological data in general

  • Application trials

    • The LPMOs catalysed oxidation affect softwood kraft fibre integrity in such way that fibres were more easily dissolved and fibrillated under mechanical stress.

    • In dissolution, even relatively low level of oxidation and reduction of cellulose molar mass reduced the ballooning and facilitated the fibre dissolution, suggesting that essentially the remnants of surface S1 layer were degraded in enzyme treatment, leaving the bulk fibre intact.

    • In fluidization, the LPMO treatment was found enhance fibrillation and especially the enzyme with ability oxidize at C1 position of glucose units in cellulose was found to better for this purpose. The films prepared from oxidized CNFs had oxygen barrier properties similar to non-treated control but had higher water vapour transmission rate.

    • On the other hand, the enzyme active mainly on C4 position enhanced cellulose and lignocellulose hydrolysis, perhaps due to better synergy with CBH1, which is the key enzyme in saccharification.


  • Marjamaa, K., Lahtinen, P., Arola, S., Maiorova, N., Nygren, H., Aro, N., & Koivula, A. (2023). Oxidative treatment and nanofibrillation softwood kraft fibres with lytic polysaccharide monooxygenases from Trichoderma reesei and Podospora anserina. Industrial Crops and Products, 193, [116243].

  • Hasheminejad, K., Scacchi, A., Javan Nikkhah, S., & Sammalkorpi, M. (2023). Cracking polymer coatings of paper-like surfaces: Control via block co-polymer structure and system composition. Applied Surface Science, 158324.

  • Scacchi, A., Hasheminejad, K., Javan Nikkhah, S., & Sammalkorpi, M. (2023). Controlling self-assembling co-polymer coatings of hydrophilic polysaccharide substrates via co-polymer block length ratio. Journal of Colloid and Interface Science, 640, 809-819.

  • Arola, S., Kou, Z., Rooijakkers, B. J. M., Velagapudi, R., Sammalkorpi, M., & Linder, M. B. (2022). On the mechanism for the highly sensitive response of cellulose nanofiber hydrogels to the presence of ionic solutes. Cellulose, 29(11), 6109-6121.

  • Marjamaa, K., Rahikainen, J., Karjalainen, M., Maiorova, N., Holopainen-mantila, U., Molinier, M., Aro, N., Nygren, H., Mikkelson, A., Koivula, A., & Kruus, K. (2022). Oxidative modification of cellulosic fibres by lytic polysaccharide monooxygenase AA9A from Trichoderma reesei. Cellulose, 29(11), 6021-6038.

  • Mudedla, S. K., Vuorte, M., Veijola, E., Marjamaa, K., Koivula, A., Linder, M. B., Arola, S., & Sammalkorpi, M. (2021). Effect of oxidation on cellulose and water structure : a molecular dynamics simulation study. Cellulose, 28(7), 3917-3933.

  • Hiltunen M (2021) Characterization of the auxiliary activity enzymes Trichoderma reesei TrAA3_2, TrAA9A and Podospora anserina PaAA9E, with potential roles in cellulose modification. Master’s thesis. University of Jyväskylä, Finland.

  • Korhonen T-M (2020) Modelling effects of salt on cellulose nanocrystal interactions. Master’s thesis. Aalto University, Finland.

  • Veijola E (2019) Modification of cellulose properties by LPMOs with different substrate specificities. Master’s thesis. Aalto University, Finland.

Research Project Managers

Project status

Funded by FinnCERES in 2019-2021.


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