A Rieske oxygenase/epoxide hydrolase-catalysed reaction cascade creates oxygen heterocycles in mupirocin biosynthesis

Abstract

Oxygen heterocycles, in particular tetrahydropyrans 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 tetrahydropyran 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 tetrahydropyran ring of mupirocin. A Rieske non-heme 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 tetrahydropyran. In the absence of MupZ, a 5membered tetrahydrofuran ring is isolated and model studies are consistent with cyclisation occurring via an epoxide intermediate. High resolution X-ray crystallographic studies, molecular modelling and mutagenesis experiments of MupZ provide insights into tetrahydropyran ring formation proceeding via an anti-Baldwin 6-endo-tet cyclisation.

Mixing and matching genes of marine and terrestrial origin in the biosynthesis of the mupirocin antibiotics

“…3B). 14 TmlW and TmlZ from the thiomarinol gene cluster, show 55.8% and 56.3% identity, respectively, to MupW and MupZ. To investigate if the same mechanism of tetrahydropyran formation operates in both thiomarinol and mupirocin biosynthesis, TmlW was heterologously overexpressed in E. coli which is able to provide the additional redox partners and cofactors necessary to constitute an active cascade.…”

Synthetic and biosynthetic methods for selective cyclisations of 4,5-epoxy alcohols to tetrahydropyrans

“…This journal is © The Royal Society of Chemistry 2022 epoxide formation/epoxide opening cascade starting from a non-activated alkane precursor. 110 Notably, all the other epoxidases reviewed herein use alkene precursors as substrates.…”

“…Using whole-cell biotransformations, Crump, Willis and coworkers showed that the C8-C16 single bond in the acyclic precursor 198 is oxidised by the Rieske non-haem oxygenase MupW to give the corresponding 8,16-epoxide 199 ( presumably through an 8,16-alkene intermediate), which spontaneously cyclises giving a five-membered tetrahydrofuran ring product 201 (Scheme 45). 110 When both MupW and the epoxide hydrolase MupZ are included in the biotransformation system, the hydroxylated THP ring product 200 is then generated. Deletion of the mupZ gene in the mupirocin producing strain abolished production of the THP ring metabolites and the THF ring compounds were produced instead.…”

Enlighten2: molecular dynamics simulations of protein–ligand systems made accessible

“…As a first attempt to meet this need, we initially developed efficient protocols (implemented as shellscripts) and a basic PyMOL plugin (for the legacy Tkinter GUI) as Enlighten (https://github.com/marcvanderkamp/enlighten). These protocols have already been used to analyze enzyme-substrate interactions and dynamics (Rice, et al, 2019;Wang, et al, 2018) as well as for preparation for quantum mechanical / molecular mechanical (QM/MM) simulations of enzyme reactions (Byrne, et al, 2016;Drulyte, et al, 2019). Now, we introduce Enlighten2, which includes a Python package for the protocols, a new userfriendly PyMOL plugin and, notably, a Docker image with all the required software that allows seamless running of simulations on Windows, Mac and Linux operating systems without non-trivial software installation.…”