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Evolutionary Imprint of Catalytic Domains in Fungal PKS–NRPS Hybrids

Authors

  • Daniela Boettger,

    1. Department of Biomolecular Chemistry, Leibniz Institute for Natural Product Research and Infection Biology—, Hans Knöll Institute, Beutenbergstrasse 11a, 07743 Jena (Germany)
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  • Holger Bergmann,

    1. Department of Systems Biology/Bioinformatics, Leibniz Institute for Natural Product Research and Infection Biology—, Hans Knöll Institute, Beutenbergstrasse 11a, 07743 Jena (Germany)
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  • Barbara Kuehn,

    1. Department of Biomolecular Chemistry, Leibniz Institute for Natural Product Research and Infection Biology—, Hans Knöll Institute, Beutenbergstrasse 11a, 07743 Jena (Germany)
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  • Dr. Ekaterina Shelest,

    Corresponding author
    1. Department of Systems Biology/Bioinformatics, Leibniz Institute for Natural Product Research and Infection Biology—, Hans Knöll Institute, Beutenbergstrasse 11a, 07743 Jena (Germany)
    • Department of Systems Biology/Bioinformatics, Leibniz Institute for Natural Product Research and Infection Biology—, Hans Knöll Institute, Beutenbergstrasse 11a, 07743 Jena (Germany)
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  • Prof. Dr. Christian Hertweck

    Corresponding author
    1. Department of Biomolecular Chemistry, Leibniz Institute for Natural Product Research and Infection Biology—, Hans Knöll Institute, Beutenbergstrasse 11a, 07743 Jena (Germany)
    2. Chair for Natural Product Chemistry, Friedrich Schiller University, Jena (Germany)
    • Department of Biomolecular Chemistry, Leibniz Institute for Natural Product Research and Infection Biology—, Hans Knöll Institute, Beutenbergstrasse 11a, 07743 Jena (Germany)
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Abstract

Fungal hybrid enzymes consisting of a polyketide synthase (PKS) and a nonribosomal peptide synthetase (NRPS) module are involved in the biosynthesis of a vast array of ecologically and medicinally relevant natural products. Whereas a dozen gene clusters could be assigned to the requisite PKS–NRPS pathways, the programming of the multifunctional enzymes is still enigmatic. Through engineering and heterologously expressing a chimera of PKS (lovastatin synthase, LovB) and NRPS (cytochalasin synthase, CheA) in Aspergillus terreus, we noted the potential incompatibility of a fungal highly reducing PKS (hrPKS) with the NRPS component of fungal PKS–NRPS hybrids. To rationalize the unexpected outcome of the gene fusion experiments, we conducted extensive bioinformatic analyses of fungal PKS–NRPS hybrids and LovB-type PKS. From motif studies and the function of the engineered chimeras, a noncanonical function of C-terminal condensation (C) domains in truncated PKS–NRPS homologues was inferred. More importantly, sequence alignments and phylogenetic trees revealed an evolutionary imprint of the PKS–NRPS domains, which reflect the evolutionary history of the entire megasynthase. Furthermore, a detailed investigation of C and adenylation (A) domains provides support for a scenario in which not only the A domain but also the C domain participates in amino acid selection. These findings shed new light on the complex code of this emerging class of multifunctional enzymes and will greatly facilitate future combinatorial biosynthesis and pathway engineering approaches towards natural product analogues.

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