Biomimetic Total Synthesis and Paired Omics Identify an Intermolecular Diels-Alder Reaction as the Key Step in Lugdunomycin Biosynthesis

Michiel T. Uiterweerd; Isabel Nuñez Santiago; Ana V. Cunha; Remco W.A. Havenith; Chao Du; Le Zhang; Helga U. van der Heul; Somayah S. Elsayed; Adriaan J. Minnaard; Gilles P. van Wezel

doi:10.1021/jacs.5c01883

Biomimetic Total Synthesis and Paired Omics Identify an Intermolecular Diels-Alder Reaction as the Key Step in Lugdunomycin Biosynthesis

Michiel T. Uiterweerd
, Isabel Nuñez Santiago
, Ana V. Cunha
, Remco W.A. Havenith
, Chao Du
, Le Zhang
, Helga U. van der Heul
, Somayah S. Elsayed
, Adriaan J. Minnaard
, Gilles P. van Wezel

Research output: Contribution to journal › Article › peer-review

Abstract

Microbial natural products are the basis of the majority of clinical drugs, where the discovery of truly novel structural scaffolds to fill the discovery pipelines is a prerequisite. Lugdunomycin is a highly rearranged angucycline polyketide produced by Streptomyces sp. QL37, with an enigmatic biosynthetic pathway. Here we show that lugdunomycin is formed by a rare intermolecular Diels-Alder reaction, with elmonin as a masked diene and iso-maleimycin as a dienophile. Genomics, mutational analysis, and heterologous expression revealed that the biosynthesis of the substrates is encoded by distinct biosynthetic gene clusters (BGCs), whereby elmonin is specified by an angucycline BGC, while the biosynthesis of iso-maleimycin is encoded by a BGC for a β-lactone-like compound. Biomimetic total synthesis of lugdunomycin showed that the Diels-Alder reaction leads to the production of a diastereomer of lugdunomycin as the main product in vitro. The diastereomeric ratio of the in vitro Diels-Alder reaction shifted toward lugdunomycin in the presence of proteinaceous material, suggesting that the in vivo Diels-Alder reaction is templated. Alphafold modeling and experimental data suggest that GarL could potentially function as a Diels-Alder template in lugdunomycin biosynthesis. The requirement of distinct biosynthetic pathways and complex chemical reactions indicates the challenges we face in discovering new chemical space.

Original language	English
Pages (from-to)	13764-13774
Number of pages	11
Journal	Journal of the American Chemical Society
Volume	147
Issue number	16
DOIs	https://doi.org/10.1021/jacs.5c01883
State	Published - Apr 23 2025
Externally published	Yes

Funding

We would like to dedicate this paper to the memory of Helen M. Kieser for her contribution to the field of Streptomyces genetics and her strong commitment to open science. We are grateful to Changsheng Wu, Serina Robinson, Martin Witte, and Hermen Overkleeft for discussions, to Lina Bayona Maldonado for help with metabolomics analysis, to Renze Sneep for HRMS analysis, to Hans van der Velde for X-ray crystallography, and to Johan Kemmink and Pieter van der Meulen for NMR support. The work was supported by the NACTAR program of The Netherlands Organization for Scientific Research (NWO), Grant 16439 to G.P.v.W and A.J.M. and by NWO grant 2022.004 for use of supercomputer facilities. The bioinformatic experiments presented in this work were performed using the computer resources from the Academic Leiden Interdisciplinary Cluster Environment (ALICE) provided by Leiden University.

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Uiterweerd, M. T., Nuñez Santiago, I., Cunha, A. V., Havenith, R. W. A., Du, C., Zhang, L., van der Heul, H. U., Elsayed, S. S., Minnaard, A. J., & van Wezel, G. P. (2025). Biomimetic Total Synthesis and Paired Omics Identify an Intermolecular Diels-Alder Reaction as the Key Step in Lugdunomycin Biosynthesis. Journal of the American Chemical Society, 147(16), 13764-13774. https://doi.org/10.1021/jacs.5c01883

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title = "Biomimetic Total Synthesis and Paired Omics Identify an Intermolecular Diels-Alder Reaction as the Key Step in Lugdunomycin Biosynthesis",

abstract = "Microbial natural products are the basis of the majority of clinical drugs, where the discovery of truly novel structural scaffolds to fill the discovery pipelines is a prerequisite. Lugdunomycin is a highly rearranged angucycline polyketide produced by Streptomyces sp. QL37, with an enigmatic biosynthetic pathway. Here we show that lugdunomycin is formed by a rare intermolecular Diels-Alder reaction, with elmonin as a masked diene and iso-maleimycin as a dienophile. Genomics, mutational analysis, and heterologous expression revealed that the biosynthesis of the substrates is encoded by distinct biosynthetic gene clusters (BGCs), whereby elmonin is specified by an angucycline BGC, while the biosynthesis of iso-maleimycin is encoded by a BGC for a β-lactone-like compound. Biomimetic total synthesis of lugdunomycin showed that the Diels-Alder reaction leads to the production of a diastereomer of lugdunomycin as the main product in vitro. The diastereomeric ratio of the in vitro Diels-Alder reaction shifted toward lugdunomycin in the presence of proteinaceous material, suggesting that the in vivo Diels-Alder reaction is templated. Alphafold modeling and experimental data suggest that GarL could potentially function as a Diels-Alder template in lugdunomycin biosynthesis. The requirement of distinct biosynthetic pathways and complex chemical reactions indicates the challenges we face in discovering new chemical space.",

author = "Uiterweerd, \{Michiel T.\} and \{Nu{\~n}ez Santiago\}, Isabel and Cunha, \{Ana V.\} and Havenith, \{Remco W.A.\} and Chao Du and Le Zhang and \{van der Heul\}, \{Helga U.\} and Elsayed, \{Somayah S.\} and Minnaard, \{Adriaan J.\} and \{van Wezel\}, \{Gilles P.\}",

note = "Publisher Copyright: {\textcopyright} 2025 The Authors. Published by American Chemical Society.",

year = "2025",

month = apr,

day = "23",

doi = "10.1021/jacs.5c01883",

language = "English",

volume = "147",

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Uiterweerd, MT, Nuñez Santiago, I, Cunha, AV, Havenith, RWA, Du, C, Zhang, L, van der Heul, HU, Elsayed, SS, Minnaard, AJ & van Wezel, GP 2025, 'Biomimetic Total Synthesis and Paired Omics Identify an Intermolecular Diels-Alder Reaction as the Key Step in Lugdunomycin Biosynthesis', Journal of the American Chemical Society, vol. 147, no. 16, pp. 13764-13774. https://doi.org/10.1021/jacs.5c01883

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AU - Uiterweerd, Michiel T.

AU - Nuñez Santiago, Isabel

AU - Cunha, Ana V.

AU - Havenith, Remco W.A.

AU - Du, Chao

AU - Zhang, Le

AU - van der Heul, Helga U.

AU - Elsayed, Somayah S.

AU - Minnaard, Adriaan J.

AU - van Wezel, Gilles P.

PY - 2025/4/23

Y1 - 2025/4/23

N2 - Microbial natural products are the basis of the majority of clinical drugs, where the discovery of truly novel structural scaffolds to fill the discovery pipelines is a prerequisite. Lugdunomycin is a highly rearranged angucycline polyketide produced by Streptomyces sp. QL37, with an enigmatic biosynthetic pathway. Here we show that lugdunomycin is formed by a rare intermolecular Diels-Alder reaction, with elmonin as a masked diene and iso-maleimycin as a dienophile. Genomics, mutational analysis, and heterologous expression revealed that the biosynthesis of the substrates is encoded by distinct biosynthetic gene clusters (BGCs), whereby elmonin is specified by an angucycline BGC, while the biosynthesis of iso-maleimycin is encoded by a BGC for a β-lactone-like compound. Biomimetic total synthesis of lugdunomycin showed that the Diels-Alder reaction leads to the production of a diastereomer of lugdunomycin as the main product in vitro. The diastereomeric ratio of the in vitro Diels-Alder reaction shifted toward lugdunomycin in the presence of proteinaceous material, suggesting that the in vivo Diels-Alder reaction is templated. Alphafold modeling and experimental data suggest that GarL could potentially function as a Diels-Alder template in lugdunomycin biosynthesis. The requirement of distinct biosynthetic pathways and complex chemical reactions indicates the challenges we face in discovering new chemical space.

AB - Microbial natural products are the basis of the majority of clinical drugs, where the discovery of truly novel structural scaffolds to fill the discovery pipelines is a prerequisite. Lugdunomycin is a highly rearranged angucycline polyketide produced by Streptomyces sp. QL37, with an enigmatic biosynthetic pathway. Here we show that lugdunomycin is formed by a rare intermolecular Diels-Alder reaction, with elmonin as a masked diene and iso-maleimycin as a dienophile. Genomics, mutational analysis, and heterologous expression revealed that the biosynthesis of the substrates is encoded by distinct biosynthetic gene clusters (BGCs), whereby elmonin is specified by an angucycline BGC, while the biosynthesis of iso-maleimycin is encoded by a BGC for a β-lactone-like compound. Biomimetic total synthesis of lugdunomycin showed that the Diels-Alder reaction leads to the production of a diastereomer of lugdunomycin as the main product in vitro. The diastereomeric ratio of the in vitro Diels-Alder reaction shifted toward lugdunomycin in the presence of proteinaceous material, suggesting that the in vivo Diels-Alder reaction is templated. Alphafold modeling and experimental data suggest that GarL could potentially function as a Diels-Alder template in lugdunomycin biosynthesis. The requirement of distinct biosynthetic pathways and complex chemical reactions indicates the challenges we face in discovering new chemical space.

UR - https://www.scopus.com/pages/publications/105003246156

U2 - 10.1021/jacs.5c01883

DO - 10.1021/jacs.5c01883

M3 - Article

C2 - 40215358

AN - SCOPUS:105003246156

SN - 0002-7863

VL - 147

SP - 13764

EP - 13774

JO - Journal of the American Chemical Society

JF - Journal of the American Chemical Society

IS - 16

ER -

Biomimetic Total Synthesis and Paired Omics Identify an Intermolecular Diels-Alder Reaction as the Key Step in Lugdunomycin Biosynthesis

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