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Direct nitration and azidation of aliphatic carbons by an iron-dependent halogenase

Nature Chemical Biology volume 10, pages 209–215 (2014)Cite this article

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Abstract

Iron-dependent halogenases employ cis-halo-Fe(IV)-oxo (haloferryl) complexes to functionalize unactivated aliphatic carbon centers, a capability elusive to synthetic chemists. Halogenation requires (i) coordination of a halide anion (Cl− or Br−) to the enzyme's Fe(II) cofactor, (ii) coupled activation of O2 and decarboxylation of α-ketoglutarate to generate the haloferryl intermediate, (iii) abstraction of hydrogen (H•) from the substrate by the ferryl and (iv) transfer of the cis halogen as Cl• or Br• to the substrate radical. This enzymatic solution to an unsolved chemical challenge is potentially generalizable to installation of other functional groups, provided that the corresponding anions can support the four requisite steps. We show here that the wild-type halogenase SyrB2 can indeed direct aliphatic nitration and azidation reactions by the same chemical logic. The discovery and enhancement by mutagenesis of these previously unknown reaction types suggest unrecognized or untapped versatility in ferryl-mediated enzymatic C-H bond activation.

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Figure 1: Related mechanistic logic of the Fe/αKG halogenases and hydroxylases.
Figure 2: LC/MS analysis demonstrating the new C-N coupling reactions catalyzed by SyrB2.
Figure 3: Binding of chloride, azide and nitrite to the SyrB2–Fe(II)–αKG complex monitored by perturbations to the absorption spectrum from the Fe(II)→αKG MLCT transition.
Figure 4: Triggering of O2 addition and ferryl formation in SyrB2 by binding of the anion.
Figure 5: Enhanced efficiency of C-N coupling in the A118G variant of SyrB2.

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Acknowledgements

This work was supported by the National Institutes of Health (GM-69657 to C.K. and J.M.B. Jr.) and the National Science Foundation (MCB-642058 and CHE-724084 to C.K. and J.M.B. Jr.). We thank C. Drennan and M. Dey at the Massachusetts Institute of Technology for the kind gift of the plasmid to express SyrB2A118G.

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  1. Megan L Matthews & Linde A Miles

    Present address: Present addresses: Department of Chemical Physiology, The Scripps Research Institute, La Jolla, California, USA (M.L.M.); Department of Medicine, Memorial Sloan-Kettering Cancer Center, New York, New York, USA (L.A.M.).,

Authors and Affiliations

  1. Department of Chemistry, The Pennsylvania State University, University Park, Pennsylvania, USA

    Megan L Matthews, Wei-chen Chang, Andrew P Layne, Carsten Krebs & J Martin Bollinger Jr

  2. Department of Biochemistry and Molecular Biology, The Pennsylvania State University, University Park, Pennsylvania, USA

    Linde A Miles, Carsten Krebs & J Martin Bollinger Jr

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Contributions

M.L.M. prepared reagents, designed and carried out experiments, analyzed data and composed and edited the manuscript. W-c.C. prepared reagents, designed and carried out experiments, analyzed data and edited the manuscript. A.P.L. prepared reagents, carried out experiments and analyzed data. L.A.M. prepared reagents, carried out experiments and analyzed data. C.K. designed experiments, analyzed data and edited the manuscript. J.M.B. Jr. designed experiments and composed and edited the manuscript.

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Correspondence to Megan L Matthews or J Martin Bollinger Jr.

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Matthews, M., Chang, Wc., Layne, A. et al. Direct nitration and azidation of aliphatic carbons by an iron-dependent halogenase. Nat Chem Biol 10, 209–215 (2014). https://doi.org/10.1038/nchembio.1438

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