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Substrate control in stereoselective lanthionine biosynthesis

Nature Chemistry volume 7, pages 57–64 (2015)Cite this article

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Abstract

Enzymes are typically highly stereoselective catalysts that enforce a reactive conformation on their native substrates. We report here a rare example in which the substrate controls the stereoselectivity of an enzyme-catalysed Michael-type addition during the biosynthesis of lanthipeptides. These natural products contain thioether crosslinks formed by a cysteine attack on dehydrated Ser and Thr residues. We demonstrate that several lanthionine synthetases catalyse highly selective anti-additions in which the substrate (and not the enzyme) determines whether the addition occurs from the re or si face. A single point mutation in the peptide substrate completely inverted the stereochemical outcome of the enzymatic modification. Quantum mechanical calculations reproduced the experimentally observed selectivity and suggest that conformational restraints imposed by the amino-acid sequence on the transition states determine the face selectivity of the Michael-type cyclization.

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Figure 1: Biosynthesis of class II lanthipeptides.
Figure 2: GC-MS analysis of in vivo modified HalA2-CylLS, LtnA2-CylLS and ProcA3.2-CylLS chimeric peptides.
Figure 3: Structural characterization of the HalM2-modified HalA2-T2A peptide.
Figure 4: Secondary structure of the A ring of the CylLS core sequence derived from MD simulations.
Figure 5: Lowest-energy QM TSs for the re- and si-face Michael-type additions that generate the A ring in the CylLS core sequence.

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Acknowledgements

We thank M. C. Martínez-Cuesta for providing the pBAC105 plasmid encoding LtnM2 from Lactococcus lactis IFPL105 and L. A. Furgerson for constructing the pET15b-HalA2-T2A plasmid. We thank F. Sun for assistance with GC-MS analysis. This work was supported by the National Institutes of Health (GM 58822 to W.A.v.d.D. and GM 075962 to K.N.H.). Calculations were performed on the Hoffman2 Cluster at the University of California, Los Angeles, and the Extreme Science and Engineering Discovery Environment, which is supported by the National Science Foundation (OCI-1053575).

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Authors and Affiliations

  1. Department of Chemistry and Howard Hughes Medical Institute, University of Illinois at Urbana-Champaign, Urbana, 61801, Illinois, USA

    Weixin Tang & Wilfred A. van der Donk

  2. Department of Chemistry and Biochemistry, University of California, Los Angeles, Los Angeles, 90095-1569, California, USA

    Gonzalo Jiménez-Osés & K. N. Houk

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  1. Weixin Tang
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Contributions

W.T. and W.A.v.d.D. designed the study. W.T. performed all the experiments. G.J.O. performed all the theoretical studies. W.T., G.J.O., K.N.H. and W.A.v.d.D. analysed the data and wrote the manuscript.

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Correspondence to K. N. Houk or Wilfred A. van der Donk.

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Tang, W., Jiménez-Osés, G., Houk, K. et al. Substrate control in stereoselective lanthionine biosynthesis. Nature Chem 7, 57–64 (2015). https://doi.org/10.1038/nchem.2113

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