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Structural insight into antibiotic fosfomycin biosynthesis by a mononuclear iron enzyme

Nature volume 437, pages 838844 (06 October 2005) | Download Citation

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Abstract

The biosynthetic pathway of the clinically important antibiotic fosfomycin uses enzymes that catalyse reactions without precedent in biology. Among these is hydroxypropylphosphonic acid epoxidase, which represents a new subfamily of non-haem mononuclear iron enzymes. Here we present six X-ray structures of this enzyme: the apoenzyme at 2.0 Å resolution; a native Fe(ii)-bound form at 2.4 Å resolution; a tris(hydroxymethyl)aminomethane–Co(ii)-enzyme complex structure at 1.8 Å resolution; a substrate–Co(ii)-enzyme complex structure at 2.5 Å resolution; and two substrate–Fe(ii)-enzyme complexes at 2.1 and 2.3 Å resolution. These structural data lead us to suggest how this enzyme is able to recognize and respond to its substrate with a conformational change that protects the radical-based intermediates formed during catalysis. Comparisons with other family members suggest why substrate binding is able to prime iron for dioxygen binding in the absence of α-ketoglutarate (a co-substrate required by many mononuclear iron enzymes), and how the unique epoxidation reaction of hydroxypropylphosphonic acid epoxidase may occur.

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Acknowledgements

This research is supported in part by the National Institutes of Health (C.L.D. and H-W.L.), the National Institute of Environmental Health Sciences (L.J.H.), the Searle Scholars Program (C.L.D.), Alfred P. Sloan Foundation (C.L.D.), and a Lester Wolfe Predoctoral Fellowship (L.J.H.). Synchrotron facilities are funded by the NIH National Center of Research Resources (Advanced Photon Source NE-CAT 8BM) and the US Department of Energy, Division of Materials Sciences and Division of Chemical Sciences (National Synchrotron Light Source, Brookhaven National Laboratory).

Author information

Affiliations

  1. Department of Chemistry, Massachusetts Institute of Technology, Cambridge, Massachusetts 02139, USA

    • Luke J. Higgins
    • , Pinghua Liu
    •  & Catherine L. Drennan
  2. Department of Chemistry and Biochemistry, University of Texas at Austin, Austin, Texas 78712, USA

    • Feng Yan
    • , Pinghua Liu
    •  & Hung-wen Liu

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Competing interests

Atomic coordinates for the following structures have been deposited in the Protein Data Bank: apo-HppE (1ZZ6), Fe(ii)-HppE (1ZZ9), form-1 S-HPP–Fe(ii)-HppE (1ZZ7), form-2 S-HPP–Fe(ii)-HppE (1ZZ8), Tris–Co(ii)-HppE (1ZZC) and S-HPP–Co(ii)-HppE (1ZZB). Reprints and permissions information is available at npg.nature.com/reprintsandpermissions. The authors declare no competing financial interests.

Corresponding author

Correspondence to Catherine L. Drennan.

Supplementary information

Word documents

  1. 1.

    Supplementary Methods

    Site-directed mutagenesis, protein purification, assays, crystallization and data collection.

  2. 2.

    Supplementary Figure S1

    Stereoviews of HppE active sites displayed with 2Fo-Fc maps contoured from 1.0-1.5ω .

  3. 3.

    Supplementary Figure S2

    Structural insights into catalysis (stereoviews).

  4. 4.

    Supplementary Figure S3

    Stereoviews of S-HPP-Co(II)-HppE active sites displayed with 2Fo-Fc maps contoured from 1.0-1.5ω.

  5. 5.

    Supplementary Tables

    Supplementary Table S1 details data and refinement statistics for Co and Apo HppE structures. Supplementary Table S2 details data and refinement statistics for Fe HppE structures

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DOI

https://doi.org/10.1038/nature03924

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