Microplate-Based Detection of Lytic Polysaccharide Monooxygenase Activity by Fluorescence-Labeling of Insoluble Oxidized Products.
Identifieur interne : 000174 ( Main/Corpus ); précédent : 000173; suivant : 000175Microplate-Based Detection of Lytic Polysaccharide Monooxygenase Activity by Fluorescence-Labeling of Insoluble Oxidized Products.
Auteurs : Thu V. Vuong ; Bing Liu ; Mats Sandgren ; Emma R. MasterSource :
- Biomacromolecules [ 1526-4602 ] ; 2017.
English descriptors
- KwdEn :
- Cellulose (chemistry), Chitin (chemistry), Fluorescence (MeSH), Microtechnology (methods), Mixed Function Oxygenases (metabolism), Oxidation-Reduction (MeSH), Phanerochaete (enzymology), Phanerochaete (growth & development), Photoelectron Spectroscopy (MeSH), Polysaccharides (chemistry), Substrate Specificity (MeSH).
- MESH :
- chemical , chemistry : Cellulose, Chitin, Polysaccharides.
- chemical , metabolism : Mixed Function Oxygenases.
- enzymology : Phanerochaete.
- growth & development : Phanerochaete.
- methods : Microtechnology.
- Fluorescence, Oxidation-Reduction, Photoelectron Spectroscopy, Substrate Specificity.
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.
DOI: 10.1021/acs.biomac.6b01790
PubMed: 28125213
Links to Exploration step
pubmed:28125213Le document en format XML
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<author><name sortKey="Vuong, Thu V" sort="Vuong, Thu V" uniqKey="Vuong T" first="Thu V" last="Vuong">Thu V. Vuong</name>
<affiliation><nlm:affiliation>Department of Chemical Engineering and Applied Chemistry, University of Toronto , 200 College Street, Toronto, Ontario M5S 3E5, Canada.</nlm:affiliation>
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<author><name sortKey="Liu, Bing" sort="Liu, Bing" uniqKey="Liu B" first="Bing" last="Liu">Bing Liu</name>
<affiliation><nlm:affiliation>Department of Molecular Sciences, Swedish University of Agricultural Sciences , 750 07 Uppsala, Sweden.</nlm:affiliation>
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<author><name sortKey="Sandgren, Mats" sort="Sandgren, Mats" uniqKey="Sandgren M" first="Mats" last="Sandgren">Mats Sandgren</name>
<affiliation><nlm:affiliation>Department of Molecular Sciences, Swedish University of Agricultural Sciences , 750 07 Uppsala, Sweden.</nlm:affiliation>
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<author><name sortKey="Master, Emma R" sort="Master, Emma R" uniqKey="Master E" first="Emma R" last="Master">Emma R. Master</name>
<affiliation><nlm:affiliation>Department of Chemical Engineering and Applied Chemistry, University of Toronto , 200 College Street, Toronto, Ontario M5S 3E5, Canada.</nlm:affiliation>
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<sourceDesc><biblStruct><analytic><title xml:lang="en">Microplate-Based Detection of Lytic Polysaccharide Monooxygenase Activity by Fluorescence-Labeling of Insoluble Oxidized Products.</title>
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<author><name sortKey="Liu, Bing" sort="Liu, Bing" uniqKey="Liu B" first="Bing" last="Liu">Bing Liu</name>
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<author><name sortKey="Sandgren, Mats" sort="Sandgren, Mats" uniqKey="Sandgren M" first="Mats" last="Sandgren">Mats Sandgren</name>
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<author><name sortKey="Master, Emma R" sort="Master, Emma R" uniqKey="Master E" first="Emma R" last="Master">Emma R. Master</name>
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<series><title level="j">Biomacromolecules</title>
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<profileDesc><textClass><keywords scheme="KwdEn" xml:lang="en"><term>Cellulose (chemistry)</term>
<term>Chitin (chemistry)</term>
<term>Fluorescence (MeSH)</term>
<term>Microtechnology (methods)</term>
<term>Mixed Function Oxygenases (metabolism)</term>
<term>Oxidation-Reduction (MeSH)</term>
<term>Phanerochaete (enzymology)</term>
<term>Phanerochaete (growth & development)</term>
<term>Photoelectron Spectroscopy (MeSH)</term>
<term>Polysaccharides (chemistry)</term>
<term>Substrate Specificity (MeSH)</term>
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<term>Chitin</term>
<term>Polysaccharides</term>
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<keywords scheme="MESH" qualifier="enzymology" xml:lang="en"><term>Phanerochaete</term>
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<keywords scheme="MESH" qualifier="growth & development" xml:lang="en"><term>Phanerochaete</term>
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<keywords scheme="MESH" qualifier="methods" xml:lang="en"><term>Microtechnology</term>
</keywords>
<keywords scheme="MESH" xml:lang="en"><term>Fluorescence</term>
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<term>Photoelectron Spectroscopy</term>
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<front><div type="abstract" xml:lang="en">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.</div>
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<Abstract><AbstractText>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.</AbstractText>
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