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Lasso peptides from proteobacteria: Genome mining employing heterologous expression and mass spectrometry

Authors

  • Julian D. Hegemann,

  • Marcel Zimmermann,

  • Shaozhou Zhu,

  • Dennis Klug,

  • Mohamed A. Marahiel

    Corresponding author
    1. Department of Chemistry, Philipps-University Marburg, Hans-Meerwein-Strasse 4 and LOEWE-Center for Synthetic Microbiology, Marburg, Germany
    • Correspondence to: Mohamed A. Marahiel, Department of Chemistry, Philipps-University Marburg, Hans-Meerwein-Strasse 4 and LOEWE-Center for Synthetic Microbiology, Marburg D-35032, Germany; e-mail: marahiel@staff.uni-marburg.de

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  • This article was originally published online as an accepted preprint. The “Published Online” date corresponds to the preprint version. You can request a copy of the preprint by emailing the Biopolymers editorial office at biopolymers@wiley.com

ABSTRACT

Lasso peptides are natural products with a unique three dimensional structure resembling a lariat knot. They are from ribosomal origin and are post-translationally modified by two enzymes (B and C), one of which shares little similarity to enzymes outside of lasso peptide biosynthetic gene clusters and as such is a useful target for genome mining. In this study, we demonstrate a B protein–centric genome mining approach through which we were able to identify 102 putative lasso peptide biosynthetic gene clusters from a total of 87 different proteobacterial strains. Ten of these clusters were cloned into the pET41a expression vector, optimized through incorporation of a ribosomal binding site and heterologously expressed in Escherichia coli BL21(DE3). All 12 predicted lasso peptides (namely burhizin, caulonodin I, caulonodin II, caulonodin III, rhodanodin, rubrivinodin, sphingonodin I, sphingonodin II, syanodin I, sphingopyxin I, sphingopyxin II, and zucinodin) were detected by high-resolution Fourier transform mass spectrometry and their proposed primary structure was confirmed through tandem mass spectrometry. High yields (ranging from 0.4 to 5.2 mg/L) were observable for eight of these compounds, while thermostability assays revealed five new representatives of heat labile lasso peptides. © 2013 Wiley Periodicals, Inc. Biopolymers (Pept Sci) 100: 527–542, 2013.

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