Ortmayer, M., Hardy, F.J., Quesne, M.G. orcid.org/0000-0001-5130-1266 et al. (8 more authors) (2021) A Noncanonical Tryptophan Analogue Reveals an Active Site Hydrogen Bond Controlling Ferryl Reactivity in a Heme Peroxidase. JACS Au, 1 (7). pp. 913-918. ISSN 2691-3704
Abstract
Nature employs high-energy metal-oxo intermediates embedded within enzyme active sites to perform challenging oxidative transformations with remarkable selectivity. Understanding how different local metal-oxo coordination environments control intermediate reactivity and catalytic function is a long-standing objective. However, conducting structure-activity relationships directly in active sites has proven challenging due to the limited range of amino acid substitutions achievable within the constraints of the genetic code. Here, we use an expanded genetic code to examine the impact of hydrogen bonding interactions on ferryl heme structure and reactivity, by replacing the N-H group of the active site Trp51 of cytochrome c peroxidase by an S atom. Removal of a single hydrogen bond stabilizes the porphyrin π-cation radical state of CcP W191F compound I. In contrast, this modification leads to more basic and reactive neutral ferryl heme states, as found in CcP W191F compound II and the wild-type ferryl heme-Trp191 radical pair of compound I. This increased reactivity manifests in a >60-fold activity increase toward phenolic substrates but remarkably has negligible effects on oxidation of the biological redox partner cytc. Our data highlight how Trp51 tunes the lifetimes of key ferryl intermediates and works in synergy with the redox active Trp191 and a well-defined substrate binding site to regulate catalytic function. More broadly, this work shows how noncanonical substitutions can advance our understanding of active site features governing metal-oxo structure and reactivity.
Metadata
Item Type: | Article |
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Authors/Creators: |
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Copyright, Publisher and Additional Information: | © 2021 The Authors. This is an open access article under the terms of the Creative Commons Attribution License (CC-BY 4.0), which permits unrestricted use, distribution and reproduction in any medium, provided the original work is properly cited. |
Keywords: | heme enzyme; metal-oxo reactivity; hydrogen bonding; proton-coupled electron transfer; genetic code expansion; tryptophan analogue; cytochrome c peroxidase |
Dates: |
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Institution: | The University of Leeds |
Academic Units: | The University of Leeds > Faculty of Engineering & Physical Sciences (Leeds) > School of Chemistry (Leeds) |
Depositing User: | Symplectic Publications |
Date Deposited: | 13 Sep 2024 14:40 |
Last Modified: | 13 Sep 2024 14:40 |
Status: | Published |
Publisher: | American Chemical Society |
Identification Number: | 10.1021/jacsau.1c00145 |
Related URLs: | |
Sustainable Development Goals: | |
Open Archives Initiative ID (OAI ID): | oai:eprints.whiterose.ac.uk:217178 |