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Improved production of adipate with Escherichia coli by reversal of β-oxidation

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Abstract

The linear C6 dicarboxylic acid adipic acid is an important bulk chemical in the petrochemical industry as precursor of the polymer nylon-6,6-polyamide. In recent years, efforts were made towards the biotechnological production of adipate from renewable carbon sources using microbial cells. One strategy is to produce adipate via a reversed β-oxidation pathway. Hitherto, the adipate titers were very low due to limiting enzyme activities for this pathway. In most cases, the CoA intermediates are non-natural substrates for the tested enzymes and were therefore barely converted. We here tested heterologous enzymes in Escherichia coli to overcome these limitations and to improve the production of adipate via a reverse β-oxidation pathway. We tested in vitro selected enzymes for the efficient reduction of the enoyl-CoA and in the final reaction for the thioester cleavage. The genes encoding the enzymes which showed in vitro the highest activity were then used to construct an expression plasmid for a synthetic adipate pathway. Expression of paaJ, paaH, paaF, dcaA, and tesB in E. coli BL21(DE3) resulted in the production of up to 36 mg/L of adipate after 30 h of cultivation. Beside the activities of the pathway enzymes, the availability of metabolic precursors may limit the synthesis of adipate, providing another key target for further strain engineering towards high-yield production of adipate with E. coli.

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Authors and Affiliations

  1. IBG-1: Biotechnology, Institute of Bio- and Geosciences, Forschungszentrum Jülich GmbH, 52425, Jülich, Germany

    Nicolai Kallscheuer, Jochem Gätgens, Michael Bott & Tino Polen

  2. Institute of Bioorganic Chemistry, Heinrich-Heine-Universität, Düsseldorf, 52425, Jülich, Germany

    Marvin Lübcke & Jörg Pietruszka

Authors
  1. Nicolai Kallscheuer
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  2. Jochem Gätgens
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  3. Marvin Lübcke
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  4. Jörg Pietruszka
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  5. Michael Bott
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  6. Tino Polen
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Correspondence to Nicolai Kallscheuer or Tino Polen.

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Kallscheuer, N., Gätgens, J., Lübcke, M. et al. Improved production of adipate with Escherichia coli by reversal of β-oxidation. Appl Microbiol Biotechnol 101, 2371–2382 (2017). https://doi.org/10.1007/s00253-016-8033-3

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  • DOI: https://doi.org/10.1007/s00253-016-8033-3

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