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Title: Structure of a Thermobifida fusca lytic polysaccharide monooxygenase and mutagenesis of key residues

Abstract

Auxiliary activity (AA) enzymes are produced by numerous bacterial and fungal species to assist in the degradation of biomass. These enzymes are abundant but have yet to be fully characterized. Here, we report the X-ray structure of Thermobifida fusca AA10A (TfAA10A), investigate mutational characterization of key surface residues near its active site, and explore the importance of the various domains of Thermobifida fusca AA10B (TfAA10B). The structure of TfAA10A is similar to other bacterial LPMOs (lytic polysaccharide monooxygenases), including signs of photo-reduction and a distorted active site, with mixed features showing both type I and II copper coordination. The point mutation experiments of TfAA10A show that Trp82 and Asn83 are needed for binding, but only Trp82 affects activity. The TfAA10B domain truncation mutants reveal that CBM2 is crucial for the binding of substrate, but that the X1 module does not affect binding or activity. In TfAA10A, Trp82 and Asn83 are needed for binding, but only Trp82 affects activity. The TfAA10B domain truncation mutants reveal that CBM2 is crucial for substrate binding, but that the X1 module does not affect binding or activity. The structure of TfAA10A is similar to other bacterial lytic polysaccharide monooxygenases with mixed features showing both typemore » I and II copper coordination. The role of LPMOs and the variability of abundance in genomes are not fully explored. LPMOs likely perform initial attacks into crystalline cellulose to allow larger processive cellulases to bind and attack, but the precise nature of their synergistic behavior remains to be definitively characterized.« less

Authors:
 [1];  [2];  [2];  [2]; ORCiD logo [2];  [1]
  1. Cornell Univ., Ithaca, NY (United States). Department of Molecular Biology and Genetics
  2. National Renewable Energy Lab. (NREL), Golden, CO (United States)
Publication Date:
Research Org.:
National Renewable Energy Lab. (NREL), Golden, CO (United States)
Sponsoring Org.:
USDOE Office of Science (SC), Biological and Environmental Research (BER) (SC-23)
OSTI Identifier:
1415124
Report Number(s):
NREL/JA-2700-70439
Journal ID: ISSN 1754-6834
Grant/Contract Number:  
AC36-08GO28308
Resource Type:
Journal Article: Accepted Manuscript
Journal Name:
Biotechnology for Biofuels
Additional Journal Information:
Journal Volume: 10; Journal Issue: 1; Journal ID: ISSN 1754-6834
Publisher:
BioMed Central
Country of Publication:
United States
Language:
English
Subject:
09 BIOMASS FUELS; Thermobifida fusca; biofuels; biomass degrading enzymes; LPMO; cellulose; oxidative chemistry

Citation Formats

Kruer-Zerhusen, Nathan, Alahuhta, Markus, Lunin, Vladimir V., Himmel, Michael E., Bomble, Yannick J., and Wilson, David B.. Structure of a Thermobifida fusca lytic polysaccharide monooxygenase and mutagenesis of key residues. United States: N. p., 2017. Web. doi:10.1186/s13068-017-0925-7.
Kruer-Zerhusen, Nathan, Alahuhta, Markus, Lunin, Vladimir V., Himmel, Michael E., Bomble, Yannick J., & Wilson, David B.. Structure of a Thermobifida fusca lytic polysaccharide monooxygenase and mutagenesis of key residues. United States. doi:10.1186/s13068-017-0925-7.
Kruer-Zerhusen, Nathan, Alahuhta, Markus, Lunin, Vladimir V., Himmel, Michael E., Bomble, Yannick J., and Wilson, David B.. Thu . "Structure of a Thermobifida fusca lytic polysaccharide monooxygenase and mutagenesis of key residues". United States. doi:10.1186/s13068-017-0925-7. https://www.osti.gov/servlets/purl/1415124.
@article{osti_1415124,
title = {Structure of a Thermobifida fusca lytic polysaccharide monooxygenase and mutagenesis of key residues},
author = {Kruer-Zerhusen, Nathan and Alahuhta, Markus and Lunin, Vladimir V. and Himmel, Michael E. and Bomble, Yannick J. and Wilson, David B.},
abstractNote = {Auxiliary activity (AA) enzymes are produced by numerous bacterial and fungal species to assist in the degradation of biomass. These enzymes are abundant but have yet to be fully characterized. Here, we report the X-ray structure of Thermobifida fusca AA10A (TfAA10A), investigate mutational characterization of key surface residues near its active site, and explore the importance of the various domains of Thermobifida fusca AA10B (TfAA10B). The structure of TfAA10A is similar to other bacterial LPMOs (lytic polysaccharide monooxygenases), including signs of photo-reduction and a distorted active site, with mixed features showing both type I and II copper coordination. The point mutation experiments of TfAA10A show that Trp82 and Asn83 are needed for binding, but only Trp82 affects activity. The TfAA10B domain truncation mutants reveal that CBM2 is crucial for the binding of substrate, but that the X1 module does not affect binding or activity. In TfAA10A, Trp82 and Asn83 are needed for binding, but only Trp82 affects activity. The TfAA10B domain truncation mutants reveal that CBM2 is crucial for substrate binding, but that the X1 module does not affect binding or activity. The structure of TfAA10A is similar to other bacterial lytic polysaccharide monooxygenases with mixed features showing both type I and II copper coordination. The role of LPMOs and the variability of abundance in genomes are not fully explored. LPMOs likely perform initial attacks into crystalline cellulose to allow larger processive cellulases to bind and attack, but the precise nature of their synergistic behavior remains to be definitively characterized.},
doi = {10.1186/s13068-017-0925-7},
journal = {Biotechnology for Biofuels},
number = 1,
volume = 10,
place = {United States},
year = {Thu Nov 30 00:00:00 EST 2017},
month = {Thu Nov 30 00:00:00 EST 2017}
}

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Works referenced in this record:

Stimulation of Lignocellulosic Biomass Hydrolysis by Proteins of Glycoside Hydrolase Family 61 Structure and Function of a Large, Enigmatic Family
journal, April 2010

  • Harris, Paul; Welner, Ditte; McFarland, K.
  • Biochemistry, Vol. 49, Issue 15, p. 3305-3316
  • DOI: 10.1021/bi100009p

An Oxidative Enzyme Boosting the Enzymatic Conversion of Recalcitrant Polysaccharides
journal, October 2010

  • Vaaje-Kolstad, Gustav; Westereng, Bjørge; Horn, Svein J.
  • Science, Vol. 330, Issue 6001, p. 219-222
  • DOI: 10.1126/science.1192231

The Fibronectin Type 3-Like Repeat from the Clostridium thermocellum Cellobiohydrolase CbhA Promotes Hydrolysis of Cellulose by Modifying Its Surface
journal, September 2002


Kinetic Studies of Thermobifida fusca Cel9A Active Site Mutant Enzymes
journal, August 2004

  • Zhou, Weilin; Irwin, Diana C.; Escovar-Kousen, Jose
  • Biochemistry, Vol. 43, Issue 30, p. 9655-9663
  • DOI: 10.1021/bi049394n

Regulation and characterization of Thermobifida fusca carbohydrate-binding module proteins E7 and E8
journal, January 2008

  • Moser, Felix; Irwin, Diana; Chen, Shaolin
  • Biotechnology and Bioengineering, Vol. 100, Issue 6, p. 1066-1077
  • DOI: 10.1002/bit.21856

Structural Basis for Substrate Targeting and Catalysis by Fungal Polysaccharide Monooxygenases
journal, June 2012

  • Li, Xin; Beeson, William T.; Phillips, Christopher M.
  • Structure, Vol. 20, Issue 6, p. 1051-1061
  • DOI: 10.1016/j.str.2012.04.002

Microbial Cellulose Utilization: Fundamentals and Biotechnology
journal, September 2002

  • Lynd, L. R.; Weimer, P. J.; van Zyl, W. H.
  • Microbiology and Molecular Biology Reviews, Vol. 66, Issue 3, p. 506-577
  • DOI: 10.1128/MMBR.66.3.506-577.2002

PHENIX: a comprehensive Python-based system for macromolecular structure solution
journal, January 2010

  • Adams, Paul D.; Afonine, Pavel V.; Bunkóczi, Gábor
  • Acta Crystallographica Section D Biological Crystallography, Vol. 66, Issue 2, p. 213-221
  • DOI: 10.1107/S0907444909052925

Cellobiose Dehydrogenase and a Copper-Dependent Polysaccharide Monooxygenase Potentiate Cellulose Degradation by Neurospora crassa
journal, December 2011

  • Phillips, Christopher M.; Beeson, William T.; Cate, Jamie H.
  • ACS Chemical Biology, Vol. 6, Issue 12, p. 1399-1406
  • DOI: 10.1021/cb200351y

Recent insights into copper-containing lytic polysaccharide mono-oxygenases
journal, October 2013

  • Hemsworth, Glyn R.; Davies, Gideon J.; Walton, Paul H.
  • Current Opinion in Structural Biology, Vol. 23, Issue 5, p. 660-668
  • DOI: 10.1016/j.sbi.2013.05.006

Expansion of the enzymatic repertoire of the CAZy database to integrate auxiliary redox enzymes
journal, January 2013

  • Levasseur, Anthony; Drula, Elodie; Lombard, Vincent
  • Biotechnology for Biofuels, Vol. 6, Issue 1, Article No. 41
  • DOI: 10.1186/1754-6834-6-41