Assembling a plug-and-play production line for combinatorial biosynthesis of aromatic polyketides in Escherichia coliCitation formats

  • External authors:
  • Matthew Cummings
  • Anna Peters
  • George Whitehead
  • Binuraj Menon

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Assembling a plug-and-play production line for combinatorial biosynthesis of aromatic polyketides in Escherichia coli. / Cummings, Matthew; Peters, Anna; Whitehead, George; Menon, Binuraj; Micklefield, Jason; Webb, Simon; Takano, Eriko.

In: PL o S Biology, 2019.

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@article{c9e60122a8bb496299545962da9eeb67,
title = "Assembling a plug-and-play production line for combinatorial biosynthesis of aromatic polyketides in Escherichia coli",
abstract = "Polyketides are a class of specialised metabolites synthesised by both eukaryotes and prokaryotes. These chemically and structurally diverse molecules are heavily used in the clinic and include frontline antimicrobial and anticancer drugs such as erythromycin and doxorubicin. To replenish the clinicians{\textquoteright} diminishing arsenal of bioactive molecules, a promising strategy aims at transferring polyketide biosynthetic pathways from their native producers into the biotechnologically desirable host Escherichia coli. This approach has been successful for type I modular polyketide synthases; however, despite more than three decades of research, the large and important group of type II polyketide synthases has until now been elusive in E. coli. Here we report on a versatile polyketide biosynthesis pipeline, based on identification of E. coli-compatible type II polyketide synthases. We successfully express 5 ketosynthase (KS) and chain length factor pairs, e.g., from Photorhabdus luminescens TT01, Streptomyces resistomycificus, Streptoccocus sp. GMD2s, Pseudoalteromonas luteoviolacea, and Ktedonobacter racemifer as soluble heterodimeric recombinant proteins in E. coli for the first time. We define the anthraquinone minimal polyketide synthase components, and utilise this biosynthetic system to synthesise anthraquinones, dianthrones and benzoisochromanequinones (BIQs). Furthermore, we demonstrate the tolerance and promiscuity of the anthraquinone heterologous biosynthetic pathway in E. coli to act as genetically applicable plug-and-play scaffold, showing it to function successfully when combined with enzymes from phylogenetically distant species, endophytic fungi and plants, which resulted in two new-to-nature compounds, neomedicamycin and neochaetomycin. This work enables plug-and-play combinatorial biosynthesis of aromatic polyketides using bacterial type II polyketide synthases in E. coli, providing full access to its many advantages in terms of easy and fast genetic manipulation, accessibility for highthroughput robotics, and convenient biotechnological scale-up. Using the synthetic and systems biology toolbox, this plug-and-play biosynthetic platform can serve as an engine for the production of new and diversified bioactive polyketides in an automated, rapid and versatile fashion.",
keywords = "Refactoring, plug-and-play, combinatorial biosynthesis, polyketide synthase, aromatic polyketides, natural products",
author = "Matthew Cummings and Anna Peters and George Whitehead and Binuraj Menon and Jason Micklefield and Simon Webb and Eriko Takano",
year = "2019",
doi = "10.1371/journal.pbio.3000347",
language = "English",
journal = "PL o S Biology",
issn = "1545-7885",
publisher = "Public Library of Science",

}

RIS

TY - JOUR

T1 - Assembling a plug-and-play production line for combinatorial biosynthesis of aromatic polyketides in Escherichia coli

AU - Cummings, Matthew

AU - Peters, Anna

AU - Whitehead, George

AU - Menon, Binuraj

AU - Micklefield, Jason

AU - Webb, Simon

AU - Takano, Eriko

PY - 2019

Y1 - 2019

N2 - Polyketides are a class of specialised metabolites synthesised by both eukaryotes and prokaryotes. These chemically and structurally diverse molecules are heavily used in the clinic and include frontline antimicrobial and anticancer drugs such as erythromycin and doxorubicin. To replenish the clinicians’ diminishing arsenal of bioactive molecules, a promising strategy aims at transferring polyketide biosynthetic pathways from their native producers into the biotechnologically desirable host Escherichia coli. This approach has been successful for type I modular polyketide synthases; however, despite more than three decades of research, the large and important group of type II polyketide synthases has until now been elusive in E. coli. Here we report on a versatile polyketide biosynthesis pipeline, based on identification of E. coli-compatible type II polyketide synthases. We successfully express 5 ketosynthase (KS) and chain length factor pairs, e.g., from Photorhabdus luminescens TT01, Streptomyces resistomycificus, Streptoccocus sp. GMD2s, Pseudoalteromonas luteoviolacea, and Ktedonobacter racemifer as soluble heterodimeric recombinant proteins in E. coli for the first time. We define the anthraquinone minimal polyketide synthase components, and utilise this biosynthetic system to synthesise anthraquinones, dianthrones and benzoisochromanequinones (BIQs). Furthermore, we demonstrate the tolerance and promiscuity of the anthraquinone heterologous biosynthetic pathway in E. coli to act as genetically applicable plug-and-play scaffold, showing it to function successfully when combined with enzymes from phylogenetically distant species, endophytic fungi and plants, which resulted in two new-to-nature compounds, neomedicamycin and neochaetomycin. This work enables plug-and-play combinatorial biosynthesis of aromatic polyketides using bacterial type II polyketide synthases in E. coli, providing full access to its many advantages in terms of easy and fast genetic manipulation, accessibility for highthroughput robotics, and convenient biotechnological scale-up. Using the synthetic and systems biology toolbox, this plug-and-play biosynthetic platform can serve as an engine for the production of new and diversified bioactive polyketides in an automated, rapid and versatile fashion.

AB - Polyketides are a class of specialised metabolites synthesised by both eukaryotes and prokaryotes. These chemically and structurally diverse molecules are heavily used in the clinic and include frontline antimicrobial and anticancer drugs such as erythromycin and doxorubicin. To replenish the clinicians’ diminishing arsenal of bioactive molecules, a promising strategy aims at transferring polyketide biosynthetic pathways from their native producers into the biotechnologically desirable host Escherichia coli. This approach has been successful for type I modular polyketide synthases; however, despite more than three decades of research, the large and important group of type II polyketide synthases has until now been elusive in E. coli. Here we report on a versatile polyketide biosynthesis pipeline, based on identification of E. coli-compatible type II polyketide synthases. We successfully express 5 ketosynthase (KS) and chain length factor pairs, e.g., from Photorhabdus luminescens TT01, Streptomyces resistomycificus, Streptoccocus sp. GMD2s, Pseudoalteromonas luteoviolacea, and Ktedonobacter racemifer as soluble heterodimeric recombinant proteins in E. coli for the first time. We define the anthraquinone minimal polyketide synthase components, and utilise this biosynthetic system to synthesise anthraquinones, dianthrones and benzoisochromanequinones (BIQs). Furthermore, we demonstrate the tolerance and promiscuity of the anthraquinone heterologous biosynthetic pathway in E. coli to act as genetically applicable plug-and-play scaffold, showing it to function successfully when combined with enzymes from phylogenetically distant species, endophytic fungi and plants, which resulted in two new-to-nature compounds, neomedicamycin and neochaetomycin. This work enables plug-and-play combinatorial biosynthesis of aromatic polyketides using bacterial type II polyketide synthases in E. coli, providing full access to its many advantages in terms of easy and fast genetic manipulation, accessibility for highthroughput robotics, and convenient biotechnological scale-up. Using the synthetic and systems biology toolbox, this plug-and-play biosynthetic platform can serve as an engine for the production of new and diversified bioactive polyketides in an automated, rapid and versatile fashion.

KW - Refactoring

KW - plug-and-play

KW - combinatorial biosynthesis

KW - polyketide synthase

KW - aromatic polyketides

KW - natural products

U2 - 10.1371/journal.pbio.3000347

DO - 10.1371/journal.pbio.3000347

M3 - Article

JO - PL o S Biology

JF - PL o S Biology

SN - 1545-7885

ER -