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Structural Dynamics of Lytic Polysaccharide Monooxygenase during Catalysis
F. Filandr, D. Kavan, D. Kracher, CVFP. Laurent, R. Ludwig, P. Man, P. Halada
Language English Country Switzerland
Document type Journal Article, Research Support, Non-U.S. Gov't
Grant support
16-34818L
Grantová Agentura České Republiky - International
I 2385-N28
Austrian Science Fund - International
W1224
Austrian Science Fund - International
J-4154
Austrian Science Fund - International
LQ1604
Ministry of Education, Youth and Science - International
CZ.1.05/1.1.00/02.0109
European Regional Development Fund - International
LM2015043
Ministry of Education, Youth and Science - International
SVV260427/2019
Univerzita Karlova v Praze - International
E-resources NLK Online Full text
Directory of Open Access Journals from 2011PubMed Central from 2011
Europe PubMed Central from 2011
ProQuest Central from 2011-01-01
Open Access Digital Library from 2011-01-01
Open Access Digital Library from 2011-01-01
Health & Medicine (ProQuest) from 2011-01-01
ROAD: Directory of Open Access Scholarly Resources from 2011
Links
PubMed
32033404
DOI
10.3390/biom10020242
Knihovny.cz E-resources
- MeSH
- Cellulose chemistry MeSH
- Fungal Proteins chemistry MeSH
- Spectrometry, Mass, Electrospray Ionization MeSH
- Mass Spectrometry MeSH
- Catalytic Domain MeSH
- Catalysis MeSH
- Hydrogen-Ion Concentration MeSH
- Protein Conformation MeSH
- Oxygen chemistry MeSH
- Lignin chemistry MeSH
- Copper chemistry MeSH
- Neurospora crassa enzymology MeSH
- Oxidative Stress MeSH
- Oxidoreductases chemistry MeSH
- Mixed Function Oxygenases chemistry MeSH
- Polysaccharides chemistry MeSH
- Reactive Oxygen Species chemistry MeSH
- Substrate Specificity MeSH
- Protein Binding MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
Lytic polysaccharide monooxygenases (LPMOs) are industrially important oxidoreductases employed in lignocellulose saccharification. Using advanced time-resolved mass spectrometric techniques, we elucidated the structural determinants for substrate-mediated stabilization of the fungal LPMO9C from Neurosporacrassa during catalysis. LPMOs require a reduction in the active-site copper for catalytic activity. We show that copper reduction in NcLPMO9C leads to structural rearrangements and compaction around the active site. However, longer exposure to the reducing agent ascorbic acid also initiated an uncoupling reaction of the bound oxygen species, leading to oxidative damage, partial unfolding, and even fragmentation of NcLPMO9C. Interestingly, no changes in the hydrogen/deuterium exchange rate were detected upon incubation of oxidized or reduced LPMO with crystalline cellulose, indicating that the LPMO-substrate interactions are mainly side-chain mediated and neither affect intraprotein hydrogen bonding nor induce significant shielding of the protein surface. On the other hand, we observed a protective effect of the substrate, which slowed down the autooxidative damage induced by the uncoupling reaction. These observations further complement the picture of structural changes during LPMO catalysis.
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- $a Lytic polysaccharide monooxygenases (LPMOs) are industrially important oxidoreductases employed in lignocellulose saccharification. Using advanced time-resolved mass spectrometric techniques, we elucidated the structural determinants for substrate-mediated stabilization of the fungal LPMO9C from Neurosporacrassa during catalysis. LPMOs require a reduction in the active-site copper for catalytic activity. We show that copper reduction in NcLPMO9C leads to structural rearrangements and compaction around the active site. However, longer exposure to the reducing agent ascorbic acid also initiated an uncoupling reaction of the bound oxygen species, leading to oxidative damage, partial unfolding, and even fragmentation of NcLPMO9C. Interestingly, no changes in the hydrogen/deuterium exchange rate were detected upon incubation of oxidized or reduced LPMO with crystalline cellulose, indicating that the LPMO-substrate interactions are mainly side-chain mediated and neither affect intraprotein hydrogen bonding nor induce significant shielding of the protein surface. On the other hand, we observed a protective effect of the substrate, which slowed down the autooxidative damage induced by the uncoupling reaction. These observations further complement the picture of structural changes during LPMO catalysis.
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