title: Microplate-Based Detection of Lytic Polysaccharide Monooxygenase Activity by Fluorescence-Labeling of Insoluble Oxidized Products.
abstract: Most existing methods for screening the activity of lytic polysaccharide mono-oxygenases (LPMOs) on polysaccharides are based on the detection of soluble oxidized sugars. This approach might underestimate the total performance of LPMOs since oxidation events that do not lead to oligosaccharide release are not detected. Using PcLPMO9D as a model enzyme, a microplate-based method has been developed to detect C1-oxidizing LPMO activity by covalently linking a water-soluble fluorophore to oxidized positions within the cellulose fiber. This fluorescence method was validated using X-ray photoelectron spectroscopy and then combined with high-performance anion-exchange chromatography to track total PcLPMO9D activity.
authors.affiliation: Department of Chemical Engineering and Applied Chemistry, University of Toronto , 200 College Street, Toronto, Ontario M5S 3E5, Canada.
authors.forename: Thu V
authors.identifier: 0000-0002-4753-6975
authors.initials: TV
authors.lastname: Vuong
authors.affiliation: Department of Molecular Sciences, Swedish University of Agricultural Sciences , 750 07 Uppsala, Sweden.
authors.forename: Bing
authors.initials: B
authors.lastname: Liu
authors.affiliation: Department of Molecular Sciences, Swedish University of Agricultural Sciences , 750 07 Uppsala, Sweden.
authors.forename: Mats
authors.initials: M
authors.lastname: Sandgren
authors.affiliation: Department of Chemical Engineering and Applied Chemistry, University of Toronto , 200 College Street, Toronto, Ontario M5S 3E5, Canada.
authors.forename: Emma R
authors.initials: ER
authors.lastname: Master
mesh_terms: D002482:Cellulose / Q000737:chemistry; D002686:Chitin / Q000737:chemistry; D005453:Fluorescence; D055616:Microtechnology / Q000379:methods*; D006899:Mixed Function Oxygenases / Q000378:metabolism*; D010084:Oxidation-Reduction; D020075:Phanerochaete / Q000201:enzymology* / Q000254:growth & development; D056951:Photoelectron Spectroscopy; D011134:Polysaccharides / Q000737:chemistry*; D013379:Substrate Specificity
chemical_list: D011134:Polysaccharides; D002686:Chitin; D002482:Cellulose; D006899:Mixed Function Oxygenases