Abstract
Oxygen heterocycles—in particular, tetrahydropyrans (THPs) and tetrahydrofurans—are common structural features of many biologically active polyketide natural products. Mupirocin is a clinically important antibiotic isolated from Pseudomonas fluorescens and is assembled on a THP ring, which is essential for bioactivity. However, the biosynthesis of this moiety has remained elusive. Here, we show an oxidative enzyme-catalysed cascade that generates the THP ring of mupirocin. Rieske non-haem oxygenase (MupW)-catalysed selective oxidation of the C8–C16 single bond in a complex acyclic precursor is combined with an epoxide hydrolase (MupZ) to catalyse the subsequent regioselective ring formation to give the hydroxylated THP. In the absence of MupZ, a five-membered tetrahydrofuran ring is isolated, and model studies are consistent with cyclization occurring via an epoxide intermediate. High-resolution X-ray crystallographic studies, molecular modelling and mutagenesis experiments of MupZ provide insights into THP ring formation proceeding via an anti-Baldwin 6-endo-tet cyclization.
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Data availability
The data that support the plots within this paper and other findings of this study are available from the corresponding authors upon reasonable request. X-ray crystallographic data are available in the EMBL-EBI PDB under accession number 6FXD.
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Acknowledgements
We are grateful to the BBSRC and EPSRC for funding through the Bristol Centre for Synthetic Biology (BB/L01386X/1), and the BBSRC for funding through BB/M012107/1 and BB/R007853/1, as well as a David Phillips Fellowship (BB/M026280/1) to M.W.v.d.K. We thank M. Malaysia for a scholarship to N.A.B., and H. P. Schweizer for kindly providing plasmids for gene inactivation and complementation.
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L.W. and C.L.W. designed the experiments, analysed the data and, together with M.P.C. and T.J.S., drafted the manuscript. L.W. conducted the heterologous expression, protein purification, biotransformations, isolation and characterization of metabolites, and site-directed mutagenesis. A.P. and C.W. assisted with protein crystallization and structure determination. M.R.C. assisted with cloning and protein purification. N.A.B. conducted substrate synthesis and model cyclization. M.W.v.d.K. performed the molecular modelling. P.R.R. and M.P.C. led the protein structural studies and contributed to writing the manuscript.
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Wang, L., Parnell, A., Williams, C. et al. A Rieske oxygenase/epoxide hydrolase-catalysed reaction cascade creates oxygen heterocycles in mupirocin biosynthesis. Nat Catal 1, 968–976 (2018). https://doi.org/10.1038/s41929-018-0183-5
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DOI: https://doi.org/10.1038/s41929-018-0183-5