The recently engineered reversal of the β-oxidation cycle has been proposed as a potential platform for the efficient synthesis of longer chain (C ≥ 4) fuels and chemicals. Here, we demonstrate the utility of this platform for the synthesis of medium-chain length (C6–C10) products through the manipulation of key components of the pathway. Deletion of endogenous thioesterases provided a clean background in which the expression of various thiolase and termination components, along with required core enzymes, resulted in the ability to alter the chain length distribution and functionality of target products. This approach enabled the synthesis of medium-chain length carboxylic acids and primary alcohols from glycerol, a low-value feedstock. The use of BktB as the thiolase component with thioesterase TesA’ as the termination enzyme enabled the synthesis of about 1.3 g/L C6–C10 saturated carboxylic acids. Tailoring of product formation to primary alcohol synthesis was achieved with the use of various acyl-CoA reductases. The combination of AtoB and FadA as the thiolase components with the alcohol-forming acyl-CoA reductase Maqu2507 from M. aquaeolei resulted in the synthesis of nearly 0.3 g/L C6–C10 alcohols. These results further demonstrate the versatile nature of a β-oxidation reversal, and highlight several key aspects and control points that can be further manipulated to fine-tune the synthesis of various fuels and chemicals.
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This work was supported by grants from the U.S. National Science Foundation (CBET-1134541, CBET-1067565).
Special Issue: Metabolic Engineering.
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Kim, S., Clomburg, J.M. & Gonzalez, R. Synthesis of medium-chain length (C6–C10) fuels and chemicals via β-oxidation reversal in Escherichia coli . J Ind Microbiol Biotechnol 42, 465–475 (2015). https://doi.org/10.1007/s10295-015-1589-6
- β-Oxidation reversal
- Metabolic engineering
- Synthetic biology
- Fuels and chemicals
- Medium-chain acids
- Medium-chain alcohols