Chaplin, Amanda K., Wilson, Michael T., Hough, Michael A. et al. (5 more authors) (2016) Heterogeneity in the histidine-brace copper coordination sphere in auxiliary activity family 10 (AA10) lytic polysaccharide monooxygenases. Journal of Biological Chemistry. pp. 12838-12850. ISSN 1083-351X
Abstract
Copper-dependent lytic polysaccharide monooxygenases (LPMOs) are enzymes that oxidatively deconstruct polysaccharides. The active site copper in LPMOs is coordinated by a histidine-brace. This utilizes the amino group and side chain of the N-terminal His residue with the side chain of a second His residue to create a T-shaped arrangement of nitrogen ligands. We report a structural, kinetic, and thermodynamic appraisal of copper binding to the histidine-brace in an auxiliary activity family 10 (AA10) LPMO from Streptomyces lividans (SliLPMO10E). Unexpectedly, we discovered the existence of two apo-SliLPMO10E species in solution that can each bind copper at a single site with distinct kinetic and thermodynamic (exothermic and endothermic) properties. The experimental EPR spectrum of copper-bound SliLPMO10E requires the simulation of two different line shapes, implying two different copper-bound species, indicative of three and two nitrogen ligands coordinating the copper. Amino group coordination was probed through the creation of an N-terminal extension variant (SliLPMO10E- Ext). The kinetics and thermodynamics of copper binding to SliLPMO10E-Ext are in accord with copper binding to one of the apo-forms in the wild-type protein, suggesting that amino group coordination is absent in the two-nitrogen coordinate form of SliLPMO10E. Copper binding to SliLPMO10B was also investigated, and again it revealed the presence of two apo-forms with kinetics and stoichiometry of copper binding identical to that of SliLPMO10E. Our findings highlight that heterogeneity exists in the active site copper coordination sphere of LPMOs that may have implications for the mechanism of loading copper in the cell.
Metadata
Item Type: | Article |
---|---|
Authors/Creators: |
|
Copyright, Publisher and Additional Information: | © 2016 by The American Society for Biochemistry and Molecular Biology, Inc. Uploaded in accordance with the publisher’s self-archiving policy. Further copying may not be permitted; contact the publisher for details. |
Dates: |
|
Institution: | The University of York |
Academic Units: | The University of York > Faculty of Sciences (York) > Chemistry (York) |
Depositing User: | Pure (York) |
Date Deposited: | 02 Sep 2016 09:58 |
Last Modified: | 25 Dec 2024 00:14 |
Published Version: | https://doi.org/10.1074/jbc.M116.722447 |
Status: | Published |
Refereed: | Yes |
Identification Number: | 10.1074/jbc.M116.722447 |
Related URLs: | |
Open Archives Initiative ID (OAI ID): | oai:eprints.whiterose.ac.uk:104306 |
Download
Filename: J._Biol._Chem._2016_Chaplin_12838_50.pdf
Description: J. Biol. Chem.-2016-Chaplin-12838-50