A screening approach for assessing lytic polysaccharide monooxygenase activity in fungal strains
Abstract Background Efforts to develop efficient lignocellulose-degrading enzymatic preparations have led to the relatively recent discovery of a new class of novel cellulase boosters, termed lytic polysaccharide monoxygenases (LPMOs). These enzymes are copper-dependent metalloenzymes that initiate...
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BMC
2019-07-01
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| Series: | Biotechnology for Biofuels |
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| Online Access: | http://link.springer.com/article/10.1186/s13068-019-1526-4 |
| _version_ | 1828340610956787712 |
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| author | Pooja Dixit Biswajit Basu Munish Puri Deepak Kumar Tuli Anshu Shankar Mathur Colin James Barrow |
| author_facet | Pooja Dixit Biswajit Basu Munish Puri Deepak Kumar Tuli Anshu Shankar Mathur Colin James Barrow |
| author_sort | Pooja Dixit |
| collection | DOAJ |
| description | Abstract Background Efforts to develop efficient lignocellulose-degrading enzymatic preparations have led to the relatively recent discovery of a new class of novel cellulase boosters, termed lytic polysaccharide monoxygenases (LPMOs). These enzymes are copper-dependent metalloenzymes that initiate the biomass deconstruction process and subsequently work together with cellulases, hemicellulases, and other accessory enzymes to enhance their hydrolytic action. Given their wide distribution and diversity, screening and isolation of potent LPMOs from natural fungal diversity may provide an important avenue for increasing the efficiency of cellulases and thereby decreasing cellulosic ethanol production costs. However, methods for quick screening and detection are still not widely available. In this article, a simple and sensitive method is described by combining nonhydrolytic activity enhancement followed by LC–MS-based quantitation of LPMOs. Results In this study, a screening approach has been developed for the detection of nonhydrolytic cellulase-enhancing enzymes in crude fungal supernatants. With the application of a saturating benchmark cocktail of Celluclast 1.5L, fungal isolates were selected which had the capability of hydrolyzing pretreated rice straw by their synergistic enzyme fractions. Subsequently, these fungal extracts along with an LPMO-enriched commercial enzyme were investigated for their ability to produce Type I LPMO activity. An LC–MS-based methodology was applied to quantitate gluconic acid in enzymatic hydrolysates as an indirect measurement of Type I LPMO activity. Conclusion The present study describes an LC–MS-based separation method to detect and quantitate LPMO activity in a commercial enzyme. This method was also applied to screen fungal extracts. The developed screening strategy has enabled detection of LPMO activity in two industrially important Penicillium strains. |
| first_indexed | 2024-04-13T22:58:39Z |
| format | Article |
| id | doaj.art-69eb39f9d64e44dbbb2ffbc799b005b5 |
| institution | Directory Open Access Journal |
| issn | 1754-6834 |
| language | English |
| last_indexed | 2024-04-13T22:58:39Z |
| publishDate | 2019-07-01 |
| publisher | BMC |
| record_format | Article |
| series | Biotechnology for Biofuels |
| spelling | doaj.art-69eb39f9d64e44dbbb2ffbc799b005b52022-12-22T02:25:55ZengBMCBiotechnology for Biofuels1754-68342019-07-0112111610.1186/s13068-019-1526-4A screening approach for assessing lytic polysaccharide monooxygenase activity in fungal strainsPooja Dixit0Biswajit Basu1Munish Puri2Deepak Kumar Tuli3Anshu Shankar Mathur4Colin James Barrow5DBT-IOC Centre for Bioenergy Research, R&D Centre, Indian Oil Corporation LimitedDBT-IOC Centre for Bioenergy Research, R&D Centre, Indian Oil Corporation LimitedSchool of Life and Environmental Sciences, Deakin UniversityDBT-IOC Centre for Bioenergy Research, R&D Centre, Indian Oil Corporation LimitedDBT-IOC Centre for Bioenergy Research, R&D Centre, Indian Oil Corporation LimitedSchool of Life and Environmental Sciences, Deakin UniversityAbstract Background Efforts to develop efficient lignocellulose-degrading enzymatic preparations have led to the relatively recent discovery of a new class of novel cellulase boosters, termed lytic polysaccharide monoxygenases (LPMOs). These enzymes are copper-dependent metalloenzymes that initiate the biomass deconstruction process and subsequently work together with cellulases, hemicellulases, and other accessory enzymes to enhance their hydrolytic action. Given their wide distribution and diversity, screening and isolation of potent LPMOs from natural fungal diversity may provide an important avenue for increasing the efficiency of cellulases and thereby decreasing cellulosic ethanol production costs. However, methods for quick screening and detection are still not widely available. In this article, a simple and sensitive method is described by combining nonhydrolytic activity enhancement followed by LC–MS-based quantitation of LPMOs. Results In this study, a screening approach has been developed for the detection of nonhydrolytic cellulase-enhancing enzymes in crude fungal supernatants. With the application of a saturating benchmark cocktail of Celluclast 1.5L, fungal isolates were selected which had the capability of hydrolyzing pretreated rice straw by their synergistic enzyme fractions. Subsequently, these fungal extracts along with an LPMO-enriched commercial enzyme were investigated for their ability to produce Type I LPMO activity. An LC–MS-based methodology was applied to quantitate gluconic acid in enzymatic hydrolysates as an indirect measurement of Type I LPMO activity. Conclusion The present study describes an LC–MS-based separation method to detect and quantitate LPMO activity in a commercial enzyme. This method was also applied to screen fungal extracts. The developed screening strategy has enabled detection of LPMO activity in two industrially important Penicillium strains.http://link.springer.com/article/10.1186/s13068-019-1526-4AA9 LPMOsPenicillium sp. LPMOsSynergy in cellulasesCelluloseHemicelluloseAcid-treated rice straw |
| spellingShingle | Pooja Dixit Biswajit Basu Munish Puri Deepak Kumar Tuli Anshu Shankar Mathur Colin James Barrow A screening approach for assessing lytic polysaccharide monooxygenase activity in fungal strains Biotechnology for Biofuels AA9 LPMOs Penicillium sp. LPMOs Synergy in cellulases Cellulose Hemicellulose Acid-treated rice straw |
| title | A screening approach for assessing lytic polysaccharide monooxygenase activity in fungal strains |
| title_full | A screening approach for assessing lytic polysaccharide monooxygenase activity in fungal strains |
| title_fullStr | A screening approach for assessing lytic polysaccharide monooxygenase activity in fungal strains |
| title_full_unstemmed | A screening approach for assessing lytic polysaccharide monooxygenase activity in fungal strains |
| title_short | A screening approach for assessing lytic polysaccharide monooxygenase activity in fungal strains |
| title_sort | screening approach for assessing lytic polysaccharide monooxygenase activity in fungal strains |
| topic | AA9 LPMOs Penicillium sp. LPMOs Synergy in cellulases Cellulose Hemicellulose Acid-treated rice straw |
| url | http://link.springer.com/article/10.1186/s13068-019-1526-4 |
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