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Assembling a plug-and-play production line for combinatorial biosynthesis of aromatic polyketides in Escherichia coli.
PLoS Biol 2019; 17(7):e3000347PB

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' 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 (PKSs); however, despite more than 3 decades of research, the large and important group of type II PKSs 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 PKSs. We successfully express 5 ketosynthase (KS) and chain length factor (CLF) 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 PKS 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 2 new-to-nature compounds, neomedicamycin and neochaetomycin. This work enables plug-and-play combinatorial biosynthesis of aromatic polyketides using bacterial type II PKSs in E. coli, providing full access to its many advantages in terms of easy and fast genetic manipulation, accessibility for high-throughput 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.

Authors+Show Affiliations

Manchester Synthetic Biology Research Centre SYNBIOCHEM, Manchester Institute of Biotechnology, School of Chemistry, The University of Manchester, Manchester, United Kingdom.Manchester Synthetic Biology Research Centre SYNBIOCHEM, Manchester Institute of Biotechnology, School of Chemistry, The University of Manchester, Manchester, United Kingdom.Manchester Synthetic Biology Research Centre SYNBIOCHEM, Manchester Institute of Biotechnology, School of Chemistry, The University of Manchester, Manchester, United Kingdom.Manchester Synthetic Biology Research Centre SYNBIOCHEM, Manchester Institute of Biotechnology, School of Chemistry, The University of Manchester, Manchester, United Kingdom. Warwick Integrative Synthetic Biology Centre, WISB, School of Life Sciences, The University of Warwick, Coventry, United Kingdom.Manchester Synthetic Biology Research Centre SYNBIOCHEM, Manchester Institute of Biotechnology, School of Chemistry, The University of Manchester, Manchester, United Kingdom.Manchester Synthetic Biology Research Centre SYNBIOCHEM, Manchester Institute of Biotechnology, School of Chemistry, The University of Manchester, Manchester, United Kingdom.Manchester Synthetic Biology Research Centre SYNBIOCHEM, Manchester Institute of Biotechnology, School of Chemistry, The University of Manchester, Manchester, United Kingdom.

Pub Type(s)

Journal Article

Language

eng

PubMed ID

31318855

Citation

Cummings, Matthew, et al. "Assembling a Plug-and-play Production Line for Combinatorial Biosynthesis of Aromatic Polyketides in Escherichia Coli." PLoS Biology, vol. 17, no. 7, 2019, pp. e3000347.
Cummings M, Peters AD, Whitehead GFS, et al. Assembling a plug-and-play production line for combinatorial biosynthesis of aromatic polyketides in Escherichia coli. PLoS Biol. 2019;17(7):e3000347.
Cummings, M., Peters, A. D., Whitehead, G. F. S., Menon, B. R. K., Micklefield, J., Webb, S. J., & Takano, E. (2019). Assembling a plug-and-play production line for combinatorial biosynthesis of aromatic polyketides in Escherichia coli. PLoS Biology, 17(7), pp. e3000347. doi:10.1371/journal.pbio.3000347.
Cummings M, et al. Assembling a Plug-and-play Production Line for Combinatorial Biosynthesis of Aromatic Polyketides in Escherichia Coli. PLoS Biol. 2019;17(7):e3000347. PubMed PMID: 31318855.
* Article titles in AMA citation format should be in sentence-case
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 D, AU - Whitehead,George F S, AU - Menon,Binuraj R K, AU - Micklefield,Jason, AU - Webb,Simon J, AU - Takano,Eriko, Y1 - 2019/07/18/ PY - 2019/02/04/received PY - 2019/06/14/accepted PY - 2019/7/19/entrez PY - 2019/7/19/pubmed PY - 2019/7/19/medline SP - e3000347 EP - e3000347 JF - PLoS biology JO - PLoS Biol. VL - 17 IS - 7 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 (PKSs); however, despite more than 3 decades of research, the large and important group of type II PKSs 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 PKSs. We successfully express 5 ketosynthase (KS) and chain length factor (CLF) 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 PKS 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 2 new-to-nature compounds, neomedicamycin and neochaetomycin. This work enables plug-and-play combinatorial biosynthesis of aromatic polyketides using bacterial type II PKSs in E. coli, providing full access to its many advantages in terms of easy and fast genetic manipulation, accessibility for high-throughput 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. SN - 1545-7885 UR - https://www.unboundmedicine.com/medline/citation/31318855/Assembling_a_plug-and-play_production_line_for_combinatorial_biosynthesis_of_aromatic_polyketides_in_Escherichia_coli L2 - http://dx.plos.org/10.1371/journal.pbio.3000347 DB - PRIME DP - Unbound Medicine ER -