Webb, J. orcid.org/0000-0002-2825-7671, Springthorpe, V., Rossoni, L. et al. (11 more authors) (2019) Systems analyses reveal the resilience of Escherichia coli physiology during accumulation and export of the nonnative organic acid citramalate. mSystems, 4 (4). e00187-19. ISSN 2379-5077
Abstract
Productivity of bacterial cell factories is frequently compromised by stresses imposed by recombinant protein synthesis and carbon-to-product conversion, but little is known about these bioprocesses at a systems level. Production of the unnatural metabolite citramalate in Escherichia coli requires the expression of a single gene coding for citramalate synthase. Multiomic analyses of a fermentation producing 25 g liter−1 citramalate were undertaken to uncover the reasons for its productivity. Metabolite, transcript, protein, and lipid profiles of high-cell-density, fed-batch fermentations of E. coli expressing either citramalate synthase or an inactivated enzyme were similar. Both fermentations showed downregulation of flagellar genes and upregulation of chaperones IbpA and IbpB, indicating that these responses were due to recombinant protein synthesis and not citramalate production. Citramalate production did not perturb metabolite pools, except for an increased intracellular pyruvate pool. Gene expression changes in response to citramalate were limited; none of the general stress response regulons were activated. Modeling of transcription factor activities suggested that citramalate invoked a GadW-mediated acid response, and changes in GadY and RprA regulatory small RNA (sRNA) expression supported this. Although changes in membrane lipid composition were observed, none were unique to citramalate production. This systems analysis of the citramalate fermentation shows that E. coli has capacity to readily adjust to the redirection of resources toward recombinant protein and citramalate production, suggesting that it is an excellent chassis choice for manufacturing organic acids.
IMPORTANCE: Citramalate is an attractive biotechnology target because it is a precursor of methylmethacrylate, which is used to manufacture Perspex and other high-value products. Engineered E. coli strains are able to produce high titers of citramalate, despite having to express a foreign enzyme and tolerate the presence of a nonnative biochemical. A systems analysis of the citramalate fermentation was undertaken to uncover the reasons underpinning its productivity. This showed that E. coli readily adjusts to the redirection of metabolic resources toward recombinant protein and citramalate production and suggests that E. coli is an excellent chassis for manufacturing similar small, polar, foreign molecules.
Metadata
Item Type: | Article |
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Authors/Creators: |
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Copyright, Publisher and Additional Information: | Copyright © 2019 The Authors. This is an open-access article distributed under the terms of the Creative Commons Attribution 4.0 International license (https://creativecommons.org/licenses/by/4.0/). |
Keywords: | Escherichia coli; bioproduction of chemicals; citramalate; citramalic acid; fed-batch fermentation; lipidomics; metabolomics; proteomics; transcriptomics |
Dates: |
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Institution: | The University of Sheffield |
Academic Units: | The University of Sheffield > Faculty of Science (Sheffield) > School of Biosciences (Sheffield) > Department of Animal and Plant Sciences (Sheffield) The University of Sheffield > Faculty of Science (Sheffield) > School of Biosciences (Sheffield) > Department of Molecular Biology and Biotechnology (Sheffield) |
Depositing User: | Symplectic Sheffield |
Date Deposited: | 18 Jul 2019 11:48 |
Last Modified: | 18 Jul 2019 11:55 |
Status: | Published |
Publisher: | American Society for Microbiology |
Refereed: | Yes |
Identification Number: | 10.1128/msystems.00187-19 |
Related URLs: | |
Open Archives Initiative ID (OAI ID): | oai:eprints.whiterose.ac.uk:148726 |