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Formation of a Copper(II)–Tyrosyl Complex at the Active Site of Lytic Polysaccharide Monooxygenases Following Oxidation by H2O2

Paradisi, Alessandro; Johnston, Esther M.; Tovborg, Morten; Nicoll, Callum R.; Ciano, Luisa; Dowle, Adam; McMaster, Jonathan; Hancock, Y.; Davies, Gideon J.; Walton, Paul H.

Formation of a Copper(II)–Tyrosyl Complex at the Active Site of Lytic Polysaccharide Monooxygenases Following Oxidation by H2O2 Thumbnail


Authors

Alessandro Paradisi

Esther M. Johnston

Morten Tovborg

Callum R. Nicoll

Adam Dowle

Y. Hancock

Gideon J. Davies

Paul H. Walton



Abstract

Hydrogen peroxide is a cosubstrate for the oxidative cleavage of saccharidic substrates by copper-containing lytic polysaccharide monooxygenases (LPMOs). The rate of reaction of LPMOs with hydrogen peroxide is high, but it is accompanied by rapid inactivation of the enzymes, presumably through protein oxidation. Herein, we use UV− vis, CD, XAS, EPR, VT/VH-MCD, and resonance Raman spectroscopies, augmented with mass spectrometry and DFT calculations, to show that the product of reaction of an AA9 LPMO with H2O2 at higher pHs is a singlet Cu(II)−tyrosyl radical species, which is inactive for the oxidation of saccharidic substrates. The Cu(II)−tyrosyl radical center entails the formation of significant Cu(II)−(●OTyr) overlap, which in turn requires that the plane of the d(x2−y2) SOMO of the Cu(II) is orientated toward the tyrosyl radical. We propose from the Marcus cross-relation that the active site tyrosine is part of a “hole-hopping” charge-transfer mechanism formed of a pathway of conserved tyrosine and tryptophan residues, which can protect the protein active site from inactivation during uncoupled turnover.

Citation

Paradisi, A., Johnston, E. M., Tovborg, M., Nicoll, C. R., Ciano, L., Dowle, A., McMaster, J., Hancock, Y., Davies, G. J., & Walton, P. H. (2019). Formation of a Copper(II)–Tyrosyl Complex at the Active Site of Lytic Polysaccharide Monooxygenases Following Oxidation by H2O2. Journal of the American Chemical Society, 141(46), 18585-18599. https://doi.org/10.1021/jacs.9b09833

Journal Article Type Article
Acceptance Date Nov 1, 2019
Online Publication Date Nov 1, 2019
Publication Date Nov 20, 2019
Deposit Date Nov 18, 2019
Publicly Available Date Nov 25, 2019
Journal Journal of the American Chemical Society
Print ISSN 0002-7863
Electronic ISSN 1520-5126
Publisher American Chemical Society
Peer Reviewed Peer Reviewed
Volume 141
Issue 46
Pages 18585-18599
DOI https://doi.org/10.1021/jacs.9b09833
Keywords Colloid and surface chemistry; Biochemistry; General chemistry; Catalysis
Public URL https://nottingham-repository.worktribe.com/output/3322229
Publisher URL https://pubs.acs.org/doi/abs/10.1021/jacs.9b09833
Contract Date Nov 18, 2019