Purification and characterization of a native lytic polysaccharide monooxygenase from Thermoascus aurantiacus
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ORIGINAL RESEARCH PAPER
Purification and characterization of a native lytic polysaccharide monooxygenase from Thermoascus aurantiacus Susanne Fritsche . Cynthia Hopson . Jennifer Gorman . Raphael Gabriel . Steven W. Singer
Received: 5 May 2020 / Accepted: 11 June 2020 Ó Springer Nature B.V. 2020
Abstract 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
S. Fritsche C. Hopson J. Gorman R. Gabriel (&) S. W. Singer (&) Lawrence Berkeley National Laboratory, Joint BioEnergy Institute, Emeryville, CA 94608, USA e-mail: [email protected]
R. Gabriel Institut fu¨r Genetik, Technische Universita¨t Braunschweig, Spielmannstr. 7, 38106 Braunschweig, Germany
S. W. Singer e-mail: [email protected] S. Fritsche C. Hopson J. Gorman R. Gabriel S. W. Singer Lawrence Berkeley National Laboratory, Biological Systems and Engineering Division, Berkeley, CA 94720, USA S. Fritsche University of Natural Resources and Life Sciences Vienna, Muthgasse 18, 1190 Vienna, Austria C. Hopson Department of Chemical Engineering and Materials, Faculty of Chemistry, Complutense University of Madrid, Avda. Complutense s/n, 28040 Madrid, Spain
123
Biotechnol Lett
demonstrated by saccharifications with Avicel and acid-pretreated corn stover. Keywords Lytic polysaccharide monooxygenase Cellulose Biomass deconstruction
Introduction The conversion of plant biomass to biofuels and biochemicals is a critical component in implementing the bioeconomy (Scarlat et al 2015). This conversion comprises chemical pretreatment of lignocellulose followed by enzymatic depolymerization of the polysaccharides to sugars for microbial conversion (Blanch et al 2011). Cellulose is the most abundant polysaccharide in plant biomass, and cellulases produced by filamentous fungi have been harnessed for cellulose depolymerization on an industrial scale (Kuhad et al 2016). Fungi produce a variety of hydrolytic enzymes (cellobiohydrolases, endoglucanases and beta-glucosidases) that hydrolyze glycosidic bonds in the cellulose chain and in cellobiose, the hydrolytic p
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