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Binding of distinct substrate conformations enables hydroxylation of remote sties in thaxtomin D by cytochrome P450 TxtC
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Alkhalaf, Lona M., Barry, Sarah M., Rea, Dean, Gallo, Angelo, Griffiths, Daniel, Lewandowski, Józef R., Fülöp, Vilmos and Challis, Gregory L. (2019) Binding of distinct substrate conformations enables hydroxylation of remote sties in thaxtomin D by cytochrome P450 TxtC. Journal of the American Chemical Society, 141 (1). pp. 216-222. doi:10.1021/jacs.8b08864 ISSN 0002-7863.
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WRAP-binding-distinct-substrate-conformations-hydroxylation-remote-cytochrome-Fulop-2018.pdf - Accepted Version - Requires a PDF viewer. Download (1260Kb) | Preview |
Official URL: https://doi.org/10.1021/jacs.8b08864
Abstract
Cytochromes P450 (CYPs) catalyze various oxidative transformations in drug metabolism, xenobiotic degradation and natural product biosynthesis. Here we report biochemical, structural and theoretical studies of TxtC, an unusual bifunctional CYP involved in the biosynthesis of the EPA-approved herbicide thaxtomin A. TxtC was shown to hydroxylate two remote sites within the Phe residue of its diketopiperazine substrate thaxtomin D. The reactions follow a preferred order, with hydroxylation of the -carbon preceding functionalization of the phenyl group. To illuminate the molecular basis for remote site functionalization, X-ray crystal structures of TxtC in complex with the substrate and monohydroxylated intermediate were determined. Electron density cor-responding to a diatomic molecule (probably dioxygen) was sandwiched between the heme iron atom and Thr237 in the TxtC-inter-mediate structure, providing insight into the mechanism for conversion of the ferrous-dioxygen complex into the reactive ferryl intermediate. The substrate and monohydroxylated intermediate adopted similar conformations in the active site, with the -face of the phenyl group positioned over the heme iron atom. Docking simulations reproduced this observation and identified a second, energetically similar but conformationally-distinct binding mode in which the -hydrogen of the Phe residue is positioned over the heme prosthetic group. Molecular dynamics simulations confirmed that the -hydrogen is sufficiently close to the ferryl oxygen atom to be extracted by it and indicated that the two substrate conformations cannot readily interconvert in the active site. These results indicate that TxtC is able to hydroxylate two spatially remote sites by binding distinct conformations of the substrate and monohy-droxylated intermediate.
| Item Type: | Journal Article | ||||||||||||
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| Subjects: | Q Science > QD Chemistry Q Science > QP Physiology S Agriculture > SB Plant culture |
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| Divisions: | Faculty of Science, Engineering and Medicine > Science > Chemistry Faculty of Science, Engineering and Medicine > Science > Life Sciences (2010- ) |
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| Library of Congress Subject Headings (LCSH): | Herbicides -- Analysis, Cytochromes, Molecular dynamics, Catalysis | ||||||||||||
| Journal or Publication Title: | Journal of the American Chemical Society | ||||||||||||
| Publisher: | American Chemical Society | ||||||||||||
| ISSN: | 0002-7863 | ||||||||||||
| Official Date: | 9 January 2019 | ||||||||||||
| Dates: |
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| Volume: | 141 | ||||||||||||
| Number: | 1 | ||||||||||||
| Page Range: | pp. 216-222 | ||||||||||||
| DOI: | 10.1021/jacs.8b08864 | ||||||||||||
| Status: | Peer Reviewed | ||||||||||||
| Publication Status: | Published | ||||||||||||
| Access rights to Published version: | Restricted or Subscription Access | ||||||||||||
| Date of first compliant deposit: | 11 December 2018 | ||||||||||||
| Date of first compliant Open Access: | 5 December 2019 | ||||||||||||
| RIOXX Funder/Project Grant: |
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