Abstract

Natural product discovery efforts have focused primarily on microbial biosynthetic gene clusters (BGCs) containing large multimodular polyketide synthases and nonribosomal peptide synthetases; however, sequencing of fungal genomes has revealed a vast number of BGCs containing smaller NRPS-like genes of unknown biosynthetic function. Using comparative metabolomics, we show that a BGC in the human pathogen Aspergillus fumigatus named fsq, which contains an NRPS-like gene lacking a condensation domain, produces several new isoquinoline alkaloids known as the fumisoquins. These compounds derive from carbon-carbon bond formation between two amino acid–derived moieties followed by a sequence that is directly analogous to isoquinoline alkaloid biosynthesis in plants. Fumisoquin biosynthesis requires the N-methyltransferase FsqC and the FAD-dependent oxidase FsqB, which represent functional analogs of coclaurine N-methyltransferase and berberine bridge enzyme in plants. Our results show that BGCs containing incomplete NRPS modules may reveal new biosynthetic paradigms and suggest that plant-like isoquinoline biosynthesis occurs in diverse fungi.

  • Compound

    (1S,3S,11R,11aR)-3-amino-1,7,8,11-tetrahydroxy-1,2,3,6,11,11a-hexahydro-4H-pyrido[1,2-b]isoquinolin-4-one

  • Compound

    (1S,3S,11R,11aR)-3-amino-1,7,11-trihydroxy-4-oxo-1,3,4,6,11,11a-hexahydro-2H-pyrido[1,2-b]isoquinolin-8-yl hydrogen sulfate

  • Compound

    (2S,4S)-4,8-dihydroxy-7,10-dioxo-1,2,3,4,7,10-hexahydrobenzo[c][1,5]naphthyridine-2-carboxylic acid

  • Compound

    8-hydroxybenzo[c][1,5]naphthyridine-4,7,10(1H)-trione

  • Compound

    benzo[c][1,5]naphthyridine-7,8,10-triol

  • Compound

    (S)-3,10-dimethoxy-5,8,13,13a-tetrahydro-6H-isoquinolino[3,2-a]isoquinoline-2,9-diol

  • Compound

    5-benzyl-1H-pyrrole-2-carboxylic acid

  • Compound

    5-(4-hydroxybenzyl)-1H-pyrrole-2-carboxylic acid

  • Compound

    3-(3,4-dihydroxyphenyl)-2-(methylamino)propanoic acid

  • Compound

    7,8-dihydroxy-1,2,3,4-tetrahydroisoquinoline-3-carboxylic acid

  • Compound

    6,7-dihydroxy-1,2,3,4-tetrahydroisoquinoline-3-carboxylic acid

  • Compound

    methyl-L-tyrosine

  • Compound

    5,5-dimethylcyclohexane-1,3-dione

  • Compound

    ((4,4-dimethyl-2,6-dioxocyclohexyl)methyl)-L-tyrosine

  • Compound

    2-((7,8-dioxo-7,8-dihydroisoquinolin-5-yl)amino)benzoic acid

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Acknowledgements

This research was funded in part by NIH R01GM112739-01 (to N.P.K. and F.C.S.), NIH CBI training grant T32GM008500 (J.A.B.) and an NSF Graduate Research Fellowship under grant DGE-1256259 (to J.E.S.). A.A.B. and D.H. were supported by the Deutsche Forschungsgemeinschaft (CRC ChemBioSys 1127). The authors would like to thank D. Kiemle for his kind assistance in operating the Bruker Avance III HD 800 MHz NMR spectrometer.

Author information

Author notes

    • Joshua A Baccile
    •  & Joseph E Spraker

    These authors contributed equally to this work.

Affiliations

  1. Boyce Thompson Institute and Department of Chemistry and Chemical Biology, Cornell University, Ithaca, New York, USA.

    • Joshua A Baccile
    • , Henry H Le
    • , Christian Gomez
    •  & Frank C Schroeder
  2. Department of Plant Pathology, University of Wisconsin–Madison, Madison, Wisconsin, USA.

    • Joseph E Spraker
  3. Department of Pharmaceutical Microbiology at the Hans-Knöll-Institute, Friedrich Schiller University, Jena, Germany.

    • Eileen Brandenburger
    •  & Dirk Hoffmeister
  4. Institute for Microbiology, Friedrich Schiller University, Jena, Germany.

    • Jin Woo Bok
    • , Juliane Macheleidt
    •  & Axel A Brakhage
  5. Molecular and Applied Microbiology, Leibniz Institute for Natural Product Research and Infection Biology (HKI), Jena, Germany.

    • Juliane Macheleidt
    •  & Axel A Brakhage
  6. Department of Bacteriology, University of Wisconsin–Madison, Madison, Wisconsin, USA.

    • Nancy P Keller
  7. Department of Medical Microbiology and Immunology, University of Wisconsin–Madison, Madison, Wisconsin, USA.

    • Nancy P Keller

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Contributions

J.A.B. and F.C.S. characterized and identified metabolites and biosynthetic pathways. J.E.S. and J.W.B. created all of the A. fumigatus mutants used. J.A.B. and C.G. performed stable-isotope labeling experiments. H.H.L., E.B. and D.H. contributed biochemical assays. J.M. and A.A.B. contributed to the identification of new metabolites. J.A.B., J.E.S., D.H., N.P.K. and F.C.S. wrote the manuscript.

Competing interests

The authors declare no competing financial interests.

Corresponding authors

Correspondence to Nancy P Keller or Frank C Schroeder.

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    Supplementary Results, Supplementary Tables 1–5, Supplementary Figures 1–17 and Supplementary Note.

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DOI

https://doi.org/10.1038/nchembio.2061

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