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PHAB toxins: a unique family of predatory sea anemone toxins evolving via intra-gene concerted evolution defines a new peptide fold

Abstract

Sea anemone venoms have long been recognized as a rich source of peptides with interesting pharmacological and structural properties, but they still contain many uncharacterized bioactive compounds. Here we report the discovery, three-dimensional structure, activity, tissue localization, and putative function of a novel sea anemone peptide toxin that constitutes a new, sixth type of voltage-gated potassium channel (KV) toxin from sea anemones. Comprised of just 17 residues, κ-actitoxin-Ate1a (Ate1a) is the shortest sea anemone toxin reported to date, and it adopts a novel three-dimensional structure that we have named the Proline-Hinged Asymmetric β-hairpin (PHAB) fold. Mass spectrometry imaging and bioassays suggest that Ate1a serves a primarily predatory function by immobilising prey, and we show this is achieved through inhibition of Shaker-type KV channels. Ate1a is encoded as a multi-domain precursor protein that yields multiple identical mature peptides, which likely evolved by multiple domain duplication events in an actinioidean ancestor. Despite this ancient evolutionary history, the PHAB-encoding gene family exhibits remarkable sequence conservation in the mature peptide domains. We demonstrate that this conservation is likely due to intra-gene concerted evolution, which has to our knowledge not previously been reported for toxin genes. We propose that the concerted evolution of toxin domains provides a hitherto unrecognised way to circumvent the effects of the costly evolutionary arms race considered to drive toxin gene evolution by ensuring efficient secretion of ecologically important predatory toxins.

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Acknowledgements

This work was supported by the Brazilian Government (Science Without Borders PhD scholarship to BM), Australian Research Council (DECRA Fellowship DE160101142 to EABU, Future Fellowship FT150100398 to STH, ARC Linkage Grant LP140100832 to BRH and GFK), and National Health & Medical Research Council (Principal Research Fellowship APP1044414 to GFK). We thank Jason Cockington and Gillian Lawrence for maintenance of sea anemones, and Dr. Alun Jones for help with mass spectrometry experiments. Antimicrobial screens were performed by CO-ADD, funded by the Wellcome Trust (UK) and The University of Queensland (Australia). We thank Dr. Lachlan Rash for assistance with ASIC clone acquisition; Prof. John Wood for ASIC1a, ASIC2a, and ASIC3 clones; Prof. Stefan Gründer for the ASIC1b clone; Prof. Alan Goldin for the NaV1.6 clone; and Prof. Frank Bosmans for the NaV1.7 clone.

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Correspondence to Glenn F. King or Eivind A. B. Undheim.

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Madio, B., Peigneur, S., Chin, Y.K.Y. et al. PHAB toxins: a unique family of predatory sea anemone toxins evolving via intra-gene concerted evolution defines a new peptide fold. Cell. Mol. Life Sci. 75, 4511–4524 (2018) doi:10.1007/s00018-018-2897-6

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Keywords

  • Neurotoxin
  • Ion channel
  • Mass spectrometry imaging
  • 3D structure
  • Concerted evolution
  • Extreme resolution mass spectrometry imaging
  • On-tissue reduction alkylation