Lytic polysaccharide monooxygenases (LPMOs) have emerged as key proteins for depolymerization of cellulose. These copper-containing enzymes oxidize C-1 and/or C-4 bonds in cellulose, promoting increased hydrolysis of the oxidized cellulose chains. The LPMO from Thermoascus aurantiacus, a thermophilic ascomycete fungus, has been extensively studied and has served as a model LPMO. A method was developed to purify the LPMO from culture filtrates of T. aurantiacus along with its native cellobiohydrolase and endoglucanase. The activity of the purified LPMO was measured with a colorimetric assay that established the Topt of the native LPMO at 60 °C. Purification of the components of the T. aurantiacus cellulase mixture also enabled quantification of the amounts of cellobiohydrolase, endoglucanase and LPMO present in the T. aurantiacus culture filtrate, establishing that the LPMO was the most abundant protein in the culture supernatants. The importance of the LPMO to activity of the mixture was demonstrated by saccharifications with Avicel and acid-pretreated corn stover.
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The authors are grateful for funding support from the DOE Joint BioEnergy Institute (https://www.jbei.org) supported by the U.S. Department of Energy, Office of Science, Office of Biological and Environmental Research, through contract DE-AC02-05CH11231 between Lawrence Berkeley National Laboratory and the U.S. Department of Energy. Novozymes is acknowledged for the generous gift of Cellic CTec2. The U.S. Government retains and the publisher, by accepting the article for publication, acknowledges that the U.S. Government retains a nonexclusive, paid up, irrevocable, worldwide license to publish or reproduce the published form of this work, or allow others to do so, for U.S. Government purposes. The authors thank Melvin Tucker of the National Renewable Energy Laboratory for providing acid-pretreated corn stover. We thank Andy DiGiovanni of the Joint BioEnergy Institute for assistance with protein purification.
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Fritsche, S., Hopson, C., Gorman, J. et al. Purification and characterization of a native lytic polysaccharide monooxygenase from Thermoascus aurantiacus. Biotechnol Lett (2020). https://doi.org/10.1007/s10529-020-02942-w
- Lytic polysaccharide monooxygenase
- Biomass deconstruction