skip to main content
DOE PAGES title logo U.S. Department of Energy
Office of Scientific and Technical Information

Title: Structural studies of Neurospora crassa LPMO9D and redox partner CDHIIA using neutron crystallography and small-angle scattering

Abstract

Sensitivity to hydrogen/deuterium and lack of observable radiation damage makes cold neutrons an ideal probe the structural studies of proteins with highly photosensitive groups such as the copper center of lytic polysaccharide monooxygenases (LPMOs) and flavin adenine dinucleotide (FAD) and heme redox cofactors of cellobiose dehydrogenases (CDHs). In this paper, neutron crystallography and small-angle neutron scattering are used to investigate Neurospora crassa LPMO9D (NcLPMO9D) and CDHIIA (NcCDHIIA), respectively. The presence of LPMO greatly enhances the efficiency of commercial glycoside hydrolase cocktails in the depolymerization of cellulose. LPMOs can receive electrons from CDHs to activate molecular dioxygen for the oxidation of cellulose resulting in chain cleavage and disruption of local crystallinity. Using neutron protein crystallography, the hydrogen/deuterium atoms of NcLPMO9D could be located throughout the structure. At the copper active site, the protonation states of the side chains of His1, His84, His157 and Tyr168, and the orientation of water molecules could be determined. Small-angle neutron scattering measurements provided low resolution models of NcCDHIIA with both the dehydrogenase and cytochrome domains in oxidized states that exhibited elongated conformations. Finally, this work demonstrates the suitability of neutron diffraction and scattering for characterizing enzymes critical to oxidative cellulose deconstruction.

Authors:
 [1]; ORCiD logo [1];  [2];  [1]
  1. North Carolina State Univ., Raleigh, NC (United States). Dept. of Molecular and Structural Biochemistry; Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States). Neutron Sciences Directorate
  2. Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States). Neutron Sciences Directorate
Publication Date:
Research Org.:
Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States); North Carolina State Univ., Raleigh, NC (United States)
Sponsoring Org.:
USDOE Office of Science (SC), Biological and Environmental Research (BER); USDOE Office of Science (SC), Basic Energy Sciences (BES); National Science Foundation (NSF); USDA National Inst. of Food and Agriculture (NIFA)
OSTI Identifier:
1424447
Alternate Identifier(s):
OSTI ID: 1576621
Grant/Contract Number:  
AC05-00OR22725; MRI 09229719; IGERT1069091; 1010523
Resource Type:
Accepted Manuscript
Journal Name:
Carbohydrate Research
Additional Journal Information:
Journal Volume: 448; Journal ID: ISSN 0008-6215
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES; polysaccharide monooxygenases; cellobiose dehydrogenases; neutron diffraction; neutron scattering; x-ray induced photoreduction

Citation Formats

Bodenheimer, Annette M., O'Dell, William B., Stanley, Christopher B., and Meilleur, Flora. Structural studies of Neurospora crassa LPMO9D and redox partner CDHIIA using neutron crystallography and small-angle scattering. United States: N. p., 2017. Web. https://doi.org/10.1016/j.carres.2017.03.001.
Bodenheimer, Annette M., O'Dell, William B., Stanley, Christopher B., & Meilleur, Flora. Structural studies of Neurospora crassa LPMO9D and redox partner CDHIIA using neutron crystallography and small-angle scattering. United States. https://doi.org/10.1016/j.carres.2017.03.001
Bodenheimer, Annette M., O'Dell, William B., Stanley, Christopher B., and Meilleur, Flora. Sat . "Structural studies of Neurospora crassa LPMO9D and redox partner CDHIIA using neutron crystallography and small-angle scattering". United States. https://doi.org/10.1016/j.carres.2017.03.001. https://www.osti.gov/servlets/purl/1424447.
@article{osti_1424447,
title = {Structural studies of Neurospora crassa LPMO9D and redox partner CDHIIA using neutron crystallography and small-angle scattering},
author = {Bodenheimer, Annette M. and O'Dell, William B. and Stanley, Christopher B. and Meilleur, Flora},
abstractNote = {Sensitivity to hydrogen/deuterium and lack of observable radiation damage makes cold neutrons an ideal probe the structural studies of proteins with highly photosensitive groups such as the copper center of lytic polysaccharide monooxygenases (LPMOs) and flavin adenine dinucleotide (FAD) and heme redox cofactors of cellobiose dehydrogenases (CDHs). In this paper, neutron crystallography and small-angle neutron scattering are used to investigate Neurospora crassa LPMO9D (NcLPMO9D) and CDHIIA (NcCDHIIA), respectively. The presence of LPMO greatly enhances the efficiency of commercial glycoside hydrolase cocktails in the depolymerization of cellulose. LPMOs can receive electrons from CDHs to activate molecular dioxygen for the oxidation of cellulose resulting in chain cleavage and disruption of local crystallinity. Using neutron protein crystallography, the hydrogen/deuterium atoms of NcLPMO9D could be located throughout the structure. At the copper active site, the protonation states of the side chains of His1, His84, His157 and Tyr168, and the orientation of water molecules could be determined. Small-angle neutron scattering measurements provided low resolution models of NcCDHIIA with both the dehydrogenase and cytochrome domains in oxidized states that exhibited elongated conformations. Finally, this work demonstrates the suitability of neutron diffraction and scattering for characterizing enzymes critical to oxidative cellulose deconstruction.},
doi = {10.1016/j.carres.2017.03.001},
journal = {Carbohydrate Research},
number = ,
volume = 448,
place = {United States},
year = {2017},
month = {3}
}

Journal Article:

Citation Metrics:
Cited by: 6 works
Citation information provided by
Web of Science

Save / Share:

Works referenced in this record:

Novel enzymes for the degradation of cellulose
journal, January 2012

  • Horn, Svein; Vaaje-Kolstad, Gustav; Westereng, Bjørge
  • Biotechnology for Biofuels, Vol. 5, Issue 1, Article No. 45
  • DOI: 10.1186/1754-6834-5-45

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

Oxygen Activation at the Active Site of a Fungal Lytic Polysaccharide Monooxygenase
journal, December 2016

  • O'Dell, William B.; Agarwal, Pratul K.; Meilleur, Flora
  • Angewandte Chemie International Edition, Vol. 56, Issue 3
  • DOI: 10.1002/anie.201610502

Cellulose Degradation by Polysaccharide Monooxygenases
journal, June 2015


Cellobiose Dehydrogenase: A Versatile Catalyst for Electrochemical Applications
journal, July 2010

  • Ludwig, Roland; Harreither, Wolfgang; Tasca, Federico
  • ChemPhysChem, Vol. 11, Issue 13
  • DOI: 10.1002/cphc.201000216

Structural basis for cellobiose dehydrogenase action during oxidative cellulose degradation
journal, July 2015

  • Tan, Tien-Chye; Kracher, Daniel; Gandini, Rosaria
  • Nature Communications, Vol. 6, Issue 1
  • DOI: 10.1038/ncomms8542

Photoreduction of the active site of the metalloprotein putidaredoxin by synchrotron radiation
journal, August 2007

  • Corbett, Mary C.; Latimer, Matthew J.; Poulos, Thomas L.
  • Acta Crystallographica Section D Biological Crystallography, Vol. 63, Issue 9
  • DOI: 10.1107/S0907444907035160

Towards accurate structural characterization of metal centres in protein crystals: the structures of Ni and Cu T 6 bovine insulin derivatives
journal, December 2013

  • Frankaer, Christian Grundahl; Mossin, Susanne; Ståhl, Kenny
  • Acta Crystallographica Section D Biological Crystallography, Vol. 70, Issue 1
  • DOI: 10.1107/S1399004713029040

Structural and Electronic Snapshots during the Transition from a Cu(II) to Cu(I) Metal Center of a Lytic Polysaccharide Monooxygenase by X-ray Photoreduction
journal, May 2014

  • Gudmundsson, Mikael; Kim, Seonah; Wu, Miao
  • Journal of Biological Chemistry, Vol. 289, Issue 27, p. 18782-18792
  • DOI: 10.1074/jbc.M114.563494

Lytic polysaccharide monooxygenases: a crystallographer's view on a new class of biomass-degrading enzymes
journal, October 2016


Limiting radiation damage for high-brilliance biological solution scattering: practical experience at the EMBL P12 beamline PETRAIII
journal, February 2015

  • Jeffries, Cy M.; Graewert, Melissa A.; Svergun, Dmitri I.
  • Journal of Synchrotron Radiation, Vol. 22, Issue 2
  • DOI: 10.1107/S1600577515000375

Radiation damage to a protein solution, detected by synchrotron X-ray small-angle scattering: dose-related considerations and suppression by cryoprotectants
journal, October 2004

  • Kuwamoto, Shigeo; Akiyama, Shuji; Fujisawa, Tetsuro
  • Journal of Synchrotron Radiation, Vol. 11, Issue 6
  • DOI: 10.1107/S0909049504019272

Domain Motion in Cytochrome P450 Reductase: CONFORMATIONAL EQUILIBRIA REVEALED BY NMR AND SMALL-ANGLE X-RAY SCATTERING
journal, October 2009

  • Ellis, Jacqueline; Gutierrez, Aldo; Barsukov, Igor L.
  • Journal of Biological Chemistry, Vol. 284, Issue 52
  • DOI: 10.1074/jbc.M109.054304

Redox-Linked Domain Movements in the Catalytic Cycle of Cytochrome P450 Reductase
journal, September 2013


Neutron protein crystallography: A complementary tool for locating hydrogens in proteins
journal, July 2016

  • O'Dell, William B.; Bodenheimer, Annette M.; Meilleur, Flora
  • Archives of Biochemistry and Biophysics, Vol. 602
  • DOI: 10.1016/j.abb.2015.11.033

Crystallization of a fungal lytic polysaccharide monooxygenase expressed from glycoengineered Pichia pastoris for X-ray and neutron diffraction
journal, January 2017

  • O'Dell, William B.; Swartz, Paul D.; Weiss, Kevin L.
  • Acta Crystallographica Section F Structural Biology Communications, Vol. 73, Issue 2
  • DOI: 10.1107/S2053230X16020318

On the catalytic mechanisms of lytic polysaccharide monooxygenases
journal, April 2016


The Copper Active Site of CBM33 Polysaccharide Oxygenases
journal, April 2013

  • Hemsworth, Glyn R.; Taylor, Edward J.; Kim, Robbert Q.
  • Journal of the American Chemical Society, Vol. 135, Issue 16
  • DOI: 10.1021/ja402106e

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

The molecular basis of polysaccharide cleavage by lytic polysaccharide monooxygenases
journal, February 2016

  • Frandsen, Kristian E. H.; Simmons, Thomas J.; Dupree, Paul
  • Nature Chemical Biology, Vol. 12, Issue 4
  • DOI: 10.1038/nchembio.2029

Insights into the oxidative degradation of cellulose by a copper metalloenzyme that exploits biomass components
journal, August 2011

  • Quinlan, R. J.; Sweeney, M. D.; Lo Leggio, L.
  • Proceedings of the National Academy of Sciences, Vol. 108, Issue 37, p. 15079-15084
  • DOI: 10.1073/pnas.1105776108

FoXS, FoXSDock and MultiFoXS: Single-state and multi-state structural modeling of proteins and their complexes based on SAXS profiles
journal, May 2016

  • Schneidman-Duhovny, Dina; Hammel, Michal; Tainer, John A.
  • Nucleic Acids Research, Vol. 44, Issue W1
  • DOI: 10.1093/nar/gkw389

Accurate SAXS Profile Computation and its Assessment by Contrast Variation Experiments
journal, August 2013

  • Schneidman-Duhovny, Dina; Hammel, Michal; Tainer, John A.
  • Biophysical Journal, Vol. 105, Issue 4
  • DOI: 10.1016/j.bpj.2013.07.020

SASSIE: A program to study intrinsically disordered biological molecules and macromolecular ensembles using experimental scattering restraints
journal, February 2012

  • Curtis, Joseph E.; Raghunandan, Sindhu; Nanda, Hirsh
  • Computer Physics Communications, Vol. 183, Issue 2
  • DOI: 10.1016/j.cpc.2011.09.010

Enhancement of enzymatic activity and catalytic current of cellobiose dehydrogenase by calcium ions
journal, April 2012


Small-angle scattering and 3D structure interpretation
journal, October 2016


The IMAGINE instrument: first neutron protein structure and new capabilities for neutron macromolecular crystallography
journal, September 2013

  • Meilleur, Flora; Munshi, Parthapratim; Robertson, Lee
  • Acta Crystallographica Section D Biological Crystallography, Vol. 69, Issue 10
  • DOI: 10.1107/S0907444913019604

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

Characterization of the Two Neurospora crassa Cellobiose Dehydrogenases and Their Connection to Oxidative Cellulose Degradation
journal, June 2012

  • Sygmund, Christoph; Kracher, Daniel; Scheiblbrandner, Stefan
  • Applied and Environmental Microbiology, Vol. 78, Issue 17
  • DOI: 10.1128/AEM.01503-12

The extended Q -range small-angle neutron scattering diffractometer at the SNS
journal, July 2010


Mantid—Data analysis and visualization package for neutron scattering and μ SR experiments
journal, November 2014

  • Arnold, O.; Bilheux, J. C.; Borreguero, J. M.
  • Nuclear Instruments and Methods in Physics Research Section A: Accelerators, Spectrometers, Detectors and Associated Equipment, Vol. 764
  • DOI: 10.1016/j.nima.2014.07.029

Determination of the regularization parameter in indirect-transform methods using perceptual criteria
journal, August 1992


PRIMUS: a Windows PC-based system for small-angle scattering data analysis
journal, September 2003

  • Konarev, Petr V.; Volkov, Vladimir V.; Sokolova, Anna V.
  • Journal of Applied Crystallography, Vol. 36, Issue 5, p. 1277-1282
  • DOI: 10.1107/S0021889803012779

Glycan reader: Automated sugar identification and simulation preparation for carbohydrates and glycoproteins
journal, August 2011

  • Jo, Sunhwan; Song, Kevin C.; Desaire, Heather
  • Journal of Computational Chemistry, Vol. 32, Issue 14
  • DOI: 10.1002/jcc.21886

    Works referencing / citing this record:

    Neutron scattering in the biological sciences: progress and prospects
    journal, December 2018

    • Ashkar, Rana; Bilheux, Hassina Z.; Bordallo, Heliosa
    • Acta Crystallographica Section D Structural Biology, Vol. 74, Issue 12
    • DOI: 10.1107/s2059798318017503

    Direct Electron Transfer of Enzymes Facilitated by Cytochromes
    journal, December 2018


    Distinct Substrate Specificities and Electron-Donating Systems of Fungal Lytic Polysaccharide Monooxygenases
    journal, May 2018

    • Frommhagen, Matthias; Westphal, Adrie H.; van Berkel, Willem J. H.
    • Frontiers in Microbiology, Vol. 9
    • DOI: 10.3389/fmicb.2018.01080

    Direct Electron Transfer of Enzymes Facilitated by Cytochromes
    journal, January 2019