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Polyketide Synthase-Mediated O-Methyloxime Formation in the Biosynthesis of the Oximidine Anticancer Agents |
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| Authors: | Eveline Vriens Dr Dries De Ruysscher Dr Angus N M Weir Dr Sofie Dekimpe Dr Gert Steurs Ahmed Shemy Leentje Persoons Dr Ana Rita Santos Dr Christopher Williams Prof Dirk Daelemans Prof Matthew P Crump Prof Arnout Voet Prof Wim De Borggraeve Prof Eveline Lescrinier Prof Joleen Masschelein |
| Institution: | 1. Laboratory for Biomolecular Discovery and Engineering, Department of Biology, KU Leuven, 3001 Heverlee, Belgium
VIB-KU Leuven Center for Microbiology, 3001 Heverlee, Belgium These authors contributed equally to this work.;2. Laboratory for Biomolecular Discovery and Engineering, Department of Biology, KU Leuven, 3001 Heverlee, Belgium VIB-KU Leuven Center for Microbiology, 3001 Heverlee, Belgium;3. Department of Chemistry, KU Leuven, 3001 Heverlee, Belgium;4. Laboratory for Biomolecular Modelling and Design, Department of Chemistry, KU Leuven, 3001 Heverlee, Belgium;5. Laboratory of Virology and Chemotherapy, Rega Institute for Medical Research, KU Leuven, 3000 Leuven, Belgium;6. VIB Discovery Sciences, 3001 Heverlee, Belgium;7. School of Chemistry, University of Bristol, Bristol, BS8 1TS UK;8. Sustainable Chemistry for Metals and Molecules, Department of Chemistry, KU Leuven, 3001 Heverlee, Belgium;9. Laboratory for Medicinal Chemistry, Rega Institute for Medical Research, KU Leuven, 3000 Leuven, Belgium;10. Laboratory for Biomolecular Discovery and Engineering, Department of Biology, KU Leuven, 3001 Heverlee, Belgium |
| Abstract: | Bacterial trans-acyltransferase polyketide synthases (trans-AT PKSs) are modular megaenzymes that employ unusual catalytic domains to assemble diverse bioactive natural products. One such PKS is responsible for the biosynthesis of the oximidine anticancer agents, oxime-substituted benzolactone enamides that inhibit vacuolar H+-ATPases. Here, we describe the identification of the oximidine gene cluster in Pseudomonas baetica and the characterization of four novel oximidine variants, including a structurally simpler intermediate that retains potent anticancer activity. Using a combination of in vivo, in vitro and computational approaches, we experimentally elucidate the oximidine biosynthetic pathway and reveal an unprecedented mechanism for O-methyloxime formation. We show that this process involves a specialized monooxygenase and methyltransferase domain and provide insight into their activity, mechanism and specificity. Our findings expand the catalytic capabilities of trans-AT PKSs and identify potential strategies for the production of novel oximidine analogues. |
| Keywords: | Bacterial Natural Products Biosynthesis Enzymology Genome Mining Polyketides |