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Entomopathogenic bacteria use multiple mechanisms for bioactive peptide library design

Abstract

The production of natural product compound libraries has been observed in nature for different organisms such as bacteria, fungi and plants; however, little is known about the mechanisms generating such chemically diverse libraries. Here we report mechanisms leading to the biosynthesis of the chemically diverse rhabdopeptide/xenortide peptides (RXPs). They are exclusively present in entomopathogenic bacteria of the genera Photorhabdus and Xenorhabdus that live in symbiosis with nematodes delivering them to insect prey, which is killed and utilized for nutrition by both nematodes and bacteria. Chemical diversity of the biologically active RXPs results from a combination of iterative and flexible use of monomodular nonribosomal peptide synthetases including substrate promiscuity, enzyme cross-talk and enzyme stoichiometry as shown by in vivo and in vitro experiments. Together, this highlights several of nature's methods for diversification, or evolution, of natural products and sheds light on the biosynthesis of the bioactive RXPs.

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Figure 1: Structures of selected RXP derivatives highlighting the chemical diversity.
Figure 2: RXP production from selected Xenorhabdus and Photorhabdus wild-type strains correlating the NRPS organization with the RXPs produced in these strains.
Figure 3: Selected modifications of the RXP cluster from Xenorhabdus KJ12.1 heterologously expressed in E. coli with the RXP derivatives produced from these modifications.
Figure 4: Dependence of RXP chain length on protein stoichiometry of proteins involved in peptide elongation (Kj12B) and peptide termination (Kj12C) as shown by Western blot, HPLC/MS analysis and in vitro experiments using purified Kj12B and Kj12C.
Figure 5: Examples for natural and artificial crosstalk between different RXP-NRPSs analysed by heterologous expression in E. coli.

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Acknowledgements

This work was supported by an ERC Starting Grant to H.B.B. (grant agreement no. 311477). The authors are grateful to Daniela Reimer for her pioneering work on rhabdopeptide biosynthesis and the identification of the first RXP-BGC. Additionally, M.K. and H.B.B. were supported by the European Community's Seventh Framework Program (FP7/2007–2013) under grant agreement no. 602773.

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X.C. and H.B.B. planned the experiments and wrote the paper, all experiments were performed by X.C. except protein expression, quantification and in vitro experiments performed by S.N., chemical synthesis of RXPs performed by F.W. and bioactivity testing performed by M.K., I.B. and R.F. All authors discussed the results and commented on the manuscript.

Corresponding author

Correspondence to Helge. B. Bode.

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The authors declare no competing financial interests.

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Cai, X., Nowak, S., Wesche, F. et al. Entomopathogenic bacteria use multiple mechanisms for bioactive peptide library design. Nature Chem 9, 379–386 (2017). https://doi.org/10.1038/nchem.2671

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