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A ketosynthase homolog uses malonyl units to form esters in cervimycin biosynthesis

Nature Chemical Biology volume 8, pages 154–161 (2012)Cite this article

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Abstract

Ketosynthases produce the carbon backbones of a vast number of biologically active polyketides by catalyzing Claisen condensations of activated acyl and malonyl building blocks. Here we report that a ketosynthase homolog from Streptomyces tendae, CerJ, unexpectedly forms malonyl esters during the biosynthesis of cervimycin, a glycoside antibiotic against methicillin-resistant Staphylococcus aureus (MRSA). Deletion of cerJ yielded a substantially more active cervimycin variant lacking the malonyl side chain, and in vitro biotransformations revealed that CerJ is capable of transferring malonyl, methylmalonyl and dimethylmalonyl units onto the glycoside. According to phylogenetic analyses and elucidation of the crystal structure, CerJ is functionally and structurally positioned between the ketosynthase catalyzing Claisen condensations and acyl-ACP shuttles, and it features a noncanonical catalytic triad. Site-directed mutagenesis and structures of CerJ in complex with substrates not only allowed us to establish a model for the reaction mechanism but also provided insights into the evolution of this important subclass of the thiolase superfamily.

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Figure 1: Cervimycin structures and results of knocking out CerJ.
Figure 2: Catalysis of CerJ in comparison to typical KS III.
Figure 3: Phylogenetic analysis of CerJ and homologs.
Figure 4: 3D structures of CerJ.
Figure 5: Model for the ping-pong bi-bi mechanism of CerJ.

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Acknowledgements

We thank the German Federal Ministry of Education and Research (GenoMik) for financial support and the Swiss light source beamline X06DA for offering beamtime.

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Authors and Affiliations

  1. Leibniz Institute for Natural Product Research and Infection Biology, Jena, Germany

    Tom Bretschneider, Michelle Unger, Kirstin Scherlach & Christian Hertweck

  2. Interfaculty Institute of Biochemistry, Eberhard Karls University Tübingen, Tübingen, Germany

    Georg Zocher & Thilo Stehle

  3. Department of Pediatrics, Vanderbilt University School of Medicine, Nashville, Tennessee, USA

    Thilo Stehle

  4. Chair for Natural Product Chemistry, Friedrich Schiller University, Jena, Germany

    Christian Hertweck

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  1. Tom Bretschneider
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Contributions

T.B. and M.U. performed biochemical and genetic experiments as well as bioinformatic analysis. K.S. performed chemical analysis. G.Z. and T.S. performed crystallographic studies. G.Z. and C.H. designed research. G.Z., T.B. and C.H. wrote the manuscript.

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Correspondence to Georg Zocher or Christian Hertweck.

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Bretschneider, T., Zocher, G., Unger, M. et al. A ketosynthase homolog uses malonyl units to form esters in cervimycin biosynthesis. Nat Chem Biol 8, 154–161 (2012). https://doi.org/10.1038/nchembio.746

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