title: A new versatile microarray-based method for high throughput screening of carbohydrate-active enzymes.
abstract: Carbohydrate-active enzymes have multiple biological roles and industrial applications. Advances in genome and transcriptome sequencing together with associated bioinformatics tools have identified vast numbers of putative carbohydrate-degrading and -modifying enzymes including glycoside hydrolases and lytic polysaccharide monooxygenases. However, there is a paucity of methods for rapidly screening the activities of these enzymes. By combining the multiplexing capacity of carbohydrate microarrays with the specificity of molecular probes, we have developed a sensitive, high throughput, and versatile semiquantitative enzyme screening technique that requires low amounts of enzyme and substrate. The method can be used to assess the activities of single enzymes, enzyme mixtures, and crude culture broths against single substrates, substrate mixtures, and biomass samples. Moreover, we show that the technique can be used to analyze both endo-acting and exo-acting glycoside hydrolases, polysaccharide lyases, carbohydrate esterases, and lytic polysaccharide monooxygenases. We demonstrate the potential of the technique by identifying the substrate specificities of purified uncharacterized enzymes and by screening enzyme activities from fungal culture broths.
authors: From the Department of Plant and Environmental Sciences, Faculty of Science, University of Copenhagen, 1871 Frederiksberg, Denmark.
authors: Silvia
authors: S
authors: Vidal-Melgosa
authors: From the Department of Plant and Environmental Sciences, Faculty of Science, University of Copenhagen, 1871 Frederiksberg, Denmark.
authors: Henriette L
authors: HL
authors: Pedersen
authors: From the Department of Plant and Environmental Sciences, Faculty of Science, University of Copenhagen, 1871 Frederiksberg, Denmark.
authors: Julia
authors: J
authors: Schückel
authors: INRA, UMR792 Ingénierie des Systèmes Biologiques et des Procédés, Toulouse F-31400, France, Université de Toulouse, INSA, UPS, INP, LISBP, 135 Avenue de Rangueil, F-31077 Toulouse, France, CNRS, UMR5504, F-31400 Toulouse, France.
authors: Grégory
authors: G
authors: Arnal
authors: INRA, UMR792 Ingénierie des Systèmes Biologiques et des Procédés, Toulouse F-31400, France, Université de Toulouse, INSA, UPS, INP, LISBP, 135 Avenue de Rangueil, F-31077 Toulouse, France, CNRS, UMR5504, F-31400 Toulouse, France.
authors: Claire
authors: C
authors: Dumon
authors: From the Department of Plant and Environmental Sciences, Faculty of Science, University of Copenhagen, 1871 Frederiksberg, Denmark.
authors: Daniel B
authors: DB
authors: Amby
authors: Novozymes A/S, Krogshøjvej 36, 2880 Bagsværd, Denmark, and.
authors: Rune Nygaard
authors: RN
authors: Monrad
authors: From the Department of Plant and Environmental Sciences, Faculty of Science, University of Copenhagen, 1871 Frederiksberg, Denmark, Department of Chemistry, Biotechnology and Food Science, Norwegian University of Life Sciences, 1432 Aas, Norway.
authors: Bjørge
authors: B
authors: Westereng
authors: From the Department of Plant and Environmental Sciences, Faculty of Science, University of Copenhagen, 1871 Frederiksberg, Denmark, willats@plen.ku.dk.
authors: William G T
authors: WG
authors: Willats
mesh_terms: D002241:Carbohydrates / Q000737:chemistry*; D004798:Enzymes / Q000378:metabolism*; D057166:High-Throughput Screening Assays; D020411:Oligonucleotide Array Sequence Analysis
chemical_list: D002241:Carbohydrates; D004798:Enzymes
keywords: CAZy Database; Carbohydrate-active Enzymes (CAZymes); Carbohydrate-binding Module; Enzyme Activity; Glycoside Hydrolase; High Throughput Screening (HTS); Microarray; Monoclonal Antibody; Plant Cell Wall; Polysaccharide Degradation
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