Structure and Function of the α-Hydroxylation Bimodule of the Mupirocin Polyketide Synthase

Ashley J Winter, R Nisha Khanizeman, Abigail M C Barker-Mountford, Andrew J Devine, Luoyi Wang, Zhongshu Song, Jonathan A Davies, Paul R Race, Christopher Williams, Thomas J Simpson, Christine L Willis*, Matthew P Crump*

*Corresponding author for this work

Research output: Contribution to journalArticle (Academic Journal)peer-review

2 Citations (Scopus)


Mupirocin is a clinically important antibiotic produced by a trans-AT Type I polyketide synthase (PKS) in Pseudomonas fluorescens. The major bioactive metabolite, pseudomonic acid A (PA-A), is assembled on a tetrasubstituted tetrahydropyran (THP) core incorporating a 6-hydroxy group proposed to be introduced by α-hydroxylation of the thioester of the acyl carrier protein (ACP) bound polyketide chain. Herein, we describe an in vitro approach combining purified enzyme components, chemical synthesis, isotopic labelling, mass spectrometry and NMR in conjunction with in vivo studies leading to the first characterisation of the α-hydroxylation bimodule of the mupirocin biosynthetic pathway. These studies reveal the precise timing of hydroxylation by MupA, substrate specificity and the ACP dependency of the enzyme components that comprise this α-hydroxylation bimodule. Furthermore, using purified enzyme, it is shown that the MmpA KS 0 shows relaxed substrate specificity, suggesting precise spatiotemporal control of in trans MupA recruitment in the context of the PKS. Finally, the detection of multiple intermodular MupA/ACP interactions suggests these bimodules may integrate MupA into their assembly.

Original languageEnglish
Article numbere202312514
JournalAngewandte Chemie - International Edition
Issue number47
Early online date17 Oct 2023
Publication statusPublished - 20 Nov 2023

Bibliographical note

© 2023 The Authors. Angewandte Chemie International Edition published by Wiley-VCH GmbH.


  • Mupirocin
  • Polyketide Synthases/metabolism
  • Hydroxylation
  • Anti-Bacterial Agents/chemistry


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