A new versatile microarray-based method for high-throughput screening of carbohydrate-active enzymes

Silvia Vidal Melgosa; Henriette Lodberg Pedersen; Julia Schückel; Grégory Arnal; Claire Dumon; Daniel Buchvaldt Amby; Rune Nygaard Monrad; Bjørge Westereng; William George Tycho Willats

doi:10.1074/jbc.m114.630673

A new versatile microarray-based method for high-throughput screening of carbohydrate-active enzymes

Silvia Vidal Melgosa, Henriette Lodberg Pedersen, Julia Schückel, Grégory Arnal, Claire Dumon, Daniel Buchvaldt Amby, Rune Nygaard Monrad, Bjørge Westereng, William George Tycho Willats

36 Citationer (Scopus)

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.

Originalsprog	Engelsk
Tidsskrift	Journal of Biological Chemistry
Vol/bind	290
Sider (fra-til)	9020-9036
Antal sider	17
ISSN	0021-9258
DOI	https://doi.org/10.1074/jbc.m114.630673
Status	Udgivet - 3 apr. 2015

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10.1074/jbc.m114.630673

http://www.jbc.org/content/290/14/9020.full.pdfLicens: CC BY

Citationsformater

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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.",

author = "{Vidal Melgosa}, Silvia and Pedersen, {Henriette Lodberg} and Julia Sch{\"u}ckel and Gr{\'e}gory Arnal and Claire Dumon and Amby, {Daniel Buchvaldt} and Monrad, {Rune Nygaard} and Bj{\o}rge Westereng and Willats, {William George Tycho}",

note = "Copyright {\textcopyright} 2015, The American Society for Biochemistry and Molecular Biology.",

year = "2015",

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doi = "10.1074/jbc.m114.630673",

language = "English",

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T1 - A new versatile microarray-based method for high-throughput screening of carbohydrate-active enzymes

AU - Vidal Melgosa, Silvia

AU - Pedersen, Henriette Lodberg

AU - Schückel, Julia

AU - Arnal, Grégory

AU - Dumon, Claire

AU - Amby, Daniel Buchvaldt

AU - Monrad, Rune Nygaard

AU - Westereng, Bjørge

AU - Willats, William George Tycho

PY - 2015/4/3

Y1 - 2015/4/3

N2 - 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.

AB - 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.

U2 - 10.1074/jbc.m114.630673

DO - 10.1074/jbc.m114.630673

M3 - Journal article

C2 - 25657012

SN - 0021-9258

VL - 290

SP - 9020

EP - 9036

JO - Journal of Biological Chemistry

JF - Journal of Biological Chemistry

ER -

A new versatile microarray-based method for high-throughput screening of carbohydrate-active enzymes

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