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Cutting the Gordian knot: early and complete amino acid sequence confirmation of class II lasso peptides by HCD fragmentation

The Journal of Antibiotics volume 73, pages 772–779 (2020)Cite this article

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Abstract

Lasso peptides are a diverse class of ribosomally synthesized and post-translationally modified peptides (RiPPs). Their proteolytic and thermal stability alongside their growing potential as therapeutics has increased attention to these antimicrobial peptides. With the advent of genome mining, the discovery of RiPPs allows for the accurate prediction of putatively encoded structures, however, MSn experiments only provide partial sequence confirmation, therefore 2D NMR experiments are necessary for characterisation. Multiple MS/MS techniques were applied to two structurally characterized lasso peptides, huascopeptin and leepeptin, and one uncharacterized lasso peptide, citrulassin C, which was not isolable in sufficient quantity for NMR analysis. We have shown that MS2 can be used to elucidate the full amino acid sequences previously predicted with genome mining for this compound class. HCD was able to open the macrocycles and fragment the newly opened linear peptides, confirming the complete amino acid sequences of the characterised lasso peptides. In addition, to determine if this technique could be applied at the earliest stages of the isolation process, we targeted a lasso peptide found by genome mining, citrulassin C, and were able to fully elucidate the amino acid sequence using only MS2 from a semi-crude extract of Streptomyces huasconensis HST28T.

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Acknowledgements

SAJ would like to thank the University of Aberdeen for an Elphinstone Scholarship. CC-A thanks CONICYT PFCHA/DOCTORADO BECAS CHILE/2016 (#21160585) fellowship and CONICYT Basal Centre Grant for the Centre for Biotechnology and Bioengineering, CeBiB (FB0001). JFC also thanks CONICYT for a National PhD Scholarship (#21110356) and a Visiting Student Scholarship.

Author information

Author notes

  1. Scott A. Jarmusch

    Present address: Department of Pharmaceutical Biosciences—Pharmacognosy, Uppsala University, Uppsala, Sweden

  2. Jean Franco Castro

    Present address: Instituto de Investigaciones Agropecuarias (INIA), Chillán, Chile

Authors and Affiliations

  1. Department of Chemistry, Marine Biodiscovery Centre, University of Aberdeen, Old Aberdeen, Aberdeen, AB24 3UE, Scotland, UK

    Scott A. Jarmusch, Jean Franco Castro, Rainer Ebel & Marcel Jaspars

  2. Leibniz-Institut für Analytische Wissenschaften—ISAS—e.V., Dortmund, Germany

    Ingo Feldmann, Bernhard Blank-Landeshammer & Albert Sickmann

  3. Centre for Biotechnology and Bioengineering, University of Chile, Beauchef 851, Santiago, Chile

    Carlos Cortés-Albayay, Jean Franco Castro, Barbara Andrews & Juan A. Asenjo

  4. Department of Molecular Microbiology, John Innes Centre, Norwich, NR4 7UH, UK

    Jean Franco Castro

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Correspondence to Marcel Jaspars.

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This study is dedicated to Professor William Fenical in recognition of his work on establishing marine microbial natural products as a force to be reckoned with.

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Jarmusch, S.A., Feldmann, I., Blank-Landeshammer, B. et al. Cutting the Gordian knot: early and complete amino acid sequence confirmation of class II lasso peptides by HCD fragmentation. J Antibiot 73, 772–779 (2020). https://doi.org/10.1038/s41429-020-00369-z

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