Recombinant expression of thermostable processive MtEG5 endoglucanase and its synergism with MtLPMO from Myceliophthora thermophila during the hydrolysis of lignocellulosic substrates
Abstract Background Filamentous fungi are among the most powerful cellulolytic organisms in terrestrial ecosystems. To perform the degradation of lignocellulosic substrates, these microorganisms employ both hydrolytic and oxidative mechanisms that involve the secretion and synergism of a wide variet...
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| Format: | Article |
| Language: | English |
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BMC
2017-05-01
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| Series: | Biotechnology for Biofuels |
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| Online Access: | http://link.springer.com/article/10.1186/s13068-017-0813-1 |
| _version_ | 1828807164327624704 |
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| author | Anthi Karnaouri Madhu Nair Muraleedharan Maria Dimarogona Evangelos Topakas Ulrika Rova Mats Sandgren Paul Christakopoulos |
| author_facet | Anthi Karnaouri Madhu Nair Muraleedharan Maria Dimarogona Evangelos Topakas Ulrika Rova Mats Sandgren Paul Christakopoulos |
| author_sort | Anthi Karnaouri |
| collection | DOAJ |
| description | Abstract Background Filamentous fungi are among the most powerful cellulolytic organisms in terrestrial ecosystems. To perform the degradation of lignocellulosic substrates, these microorganisms employ both hydrolytic and oxidative mechanisms that involve the secretion and synergism of a wide variety of enzymes. Interactions between these enzymes occur on the level of saccharification, i.e., the release of neutral and oxidized products, but sometimes also reflected in the substrate liquefaction. Although the synergism regarding the yield of neutral sugars has been extensively studied, further studies should focus on the oxidized sugars, as well as the effect of enzyme combinations on the viscosity properties of the substrates. Results In the present study, the heterologous expression of an endoglucanase (EG) and its combined activity together with a lytic polysaccharide monooxygenase (LPMO), both from the thermophilic fungus Myceliophthora thermophila, are described. The EG gene, belonging to the glycoside hydrolase family 5, was functionally expressed in the methylotrophic yeast Pichia pastoris. The produced MtEG5A (75 kDa) featured remarkable thermal stability and showed high specific activity on microcrystalline cellulose compared to CMC, which is indicative of its processivity properties. The enzyme was capable of releasing high amounts of cellobiose from wheat straw, birch, and spruce biomass. Addition of MtLPMO9 together with MtEG5A showed enhanced enzymatic hydrolysis yields against regenerated amorphous cellulose (PASC) by improving the release not only of the neutral but also of the oxidized sugars. Assessment of activity of MtEG5A on the reduction of viscosity of PASC and pretreated wheat straw using dynamic viscosity measurements revealed that the enzyme is able to perform liquefaction of the model substrate and the natural lignocellulosic material, while when added together with MtLPMO9, no further synergistic effect was observed. Conclusions The endoglucanase MtEG5A from the thermophilic fungus M. thermophila exhibited excellent properties that render it a suitable candidate for use in biotechnological applications. Its strong synergism with LPMO was reflected in sugars release, but not in substrate viscosity reduction. Based on the level of oxidative sugar formation, this is the first indication of synergy between LPMO and EG reported. |
| first_indexed | 2024-12-12T08:20:29Z |
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| institution | Directory Open Access Journal |
| issn | 1754-6834 |
| language | English |
| last_indexed | 2024-12-12T08:20:29Z |
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| series | Biotechnology for Biofuels |
| spelling | doaj.art-f30eaa93fb274f1b93fc2189fa23ab4c2022-12-22T00:31:25ZengBMCBiotechnology for Biofuels1754-68342017-05-0110111710.1186/s13068-017-0813-1Recombinant expression of thermostable processive MtEG5 endoglucanase and its synergism with MtLPMO from Myceliophthora thermophila during the hydrolysis of lignocellulosic substratesAnthi Karnaouri0Madhu Nair Muraleedharan1Maria Dimarogona2Evangelos Topakas3Ulrika Rova4Mats Sandgren5Paul Christakopoulos6Biochemical Process Engineering, Chemical Engineering, Department of Civil, Environmental and Natural Resources Engineering, Luleå University of TechnologyBiochemical Process Engineering, Chemical Engineering, Department of Civil, Environmental and Natural Resources Engineering, Luleå University of TechnologyBiotechnology Laboratory, Department of Synthesis and Development of Industrial Processes, School of Chemical Engineering, National Technical University of AthensBiochemical Process Engineering, Chemical Engineering, Department of Civil, Environmental and Natural Resources Engineering, Luleå University of TechnologyBiochemical Process Engineering, Chemical Engineering, Department of Civil, Environmental and Natural Resources Engineering, Luleå University of TechnologyDepartment of Chemistry and Biotechnology, Swedish University of Agricultural SciencesBiochemical Process Engineering, Chemical Engineering, Department of Civil, Environmental and Natural Resources Engineering, Luleå University of TechnologyAbstract Background Filamentous fungi are among the most powerful cellulolytic organisms in terrestrial ecosystems. To perform the degradation of lignocellulosic substrates, these microorganisms employ both hydrolytic and oxidative mechanisms that involve the secretion and synergism of a wide variety of enzymes. Interactions between these enzymes occur on the level of saccharification, i.e., the release of neutral and oxidized products, but sometimes also reflected in the substrate liquefaction. Although the synergism regarding the yield of neutral sugars has been extensively studied, further studies should focus on the oxidized sugars, as well as the effect of enzyme combinations on the viscosity properties of the substrates. Results In the present study, the heterologous expression of an endoglucanase (EG) and its combined activity together with a lytic polysaccharide monooxygenase (LPMO), both from the thermophilic fungus Myceliophthora thermophila, are described. The EG gene, belonging to the glycoside hydrolase family 5, was functionally expressed in the methylotrophic yeast Pichia pastoris. The produced MtEG5A (75 kDa) featured remarkable thermal stability and showed high specific activity on microcrystalline cellulose compared to CMC, which is indicative of its processivity properties. The enzyme was capable of releasing high amounts of cellobiose from wheat straw, birch, and spruce biomass. Addition of MtLPMO9 together with MtEG5A showed enhanced enzymatic hydrolysis yields against regenerated amorphous cellulose (PASC) by improving the release not only of the neutral but also of the oxidized sugars. Assessment of activity of MtEG5A on the reduction of viscosity of PASC and pretreated wheat straw using dynamic viscosity measurements revealed that the enzyme is able to perform liquefaction of the model substrate and the natural lignocellulosic material, while when added together with MtLPMO9, no further synergistic effect was observed. Conclusions The endoglucanase MtEG5A from the thermophilic fungus M. thermophila exhibited excellent properties that render it a suitable candidate for use in biotechnological applications. Its strong synergism with LPMO was reflected in sugars release, but not in substrate viscosity reduction. Based on the level of oxidative sugar formation, this is the first indication of synergy between LPMO and EG reported.http://link.springer.com/article/10.1186/s13068-017-0813-1Myceliophthora thermophilaEndoglucanasePichia pastorisProcessivityLPMOViscosity |
| spellingShingle | Anthi Karnaouri Madhu Nair Muraleedharan Maria Dimarogona Evangelos Topakas Ulrika Rova Mats Sandgren Paul Christakopoulos Recombinant expression of thermostable processive MtEG5 endoglucanase and its synergism with MtLPMO from Myceliophthora thermophila during the hydrolysis of lignocellulosic substrates Biotechnology for Biofuels Myceliophthora thermophila Endoglucanase Pichia pastoris Processivity LPMO Viscosity |
| title | Recombinant expression of thermostable processive MtEG5 endoglucanase and its synergism with MtLPMO from Myceliophthora thermophila during the hydrolysis of lignocellulosic substrates |
| title_full | Recombinant expression of thermostable processive MtEG5 endoglucanase and its synergism with MtLPMO from Myceliophthora thermophila during the hydrolysis of lignocellulosic substrates |
| title_fullStr | Recombinant expression of thermostable processive MtEG5 endoglucanase and its synergism with MtLPMO from Myceliophthora thermophila during the hydrolysis of lignocellulosic substrates |
| title_full_unstemmed | Recombinant expression of thermostable processive MtEG5 endoglucanase and its synergism with MtLPMO from Myceliophthora thermophila during the hydrolysis of lignocellulosic substrates |
| title_short | Recombinant expression of thermostable processive MtEG5 endoglucanase and its synergism with MtLPMO from Myceliophthora thermophila during the hydrolysis of lignocellulosic substrates |
| title_sort | recombinant expression of thermostable processive mteg5 endoglucanase and its synergism with mtlpmo from myceliophthora thermophila during the hydrolysis of lignocellulosic substrates |
| topic | Myceliophthora thermophila Endoglucanase Pichia pastoris Processivity LPMO Viscosity |
| url | http://link.springer.com/article/10.1186/s13068-017-0813-1 |
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