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Combining De Ley–Doudoroff and methylerythritol phosphate pathways for enhanced isoprene biosynthesis from d-galactose

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Abstract

An engineered Escherichia coli strain was developed for enhanced isoprene production using d-galactose as substrate. Isoprene is a valuable compound that can be biosynthetically produced from pyruvate and glyceraldehyde-3-phosphate (G3P) through the methylerythritol phosphate pathway (MEP). The Leloir and De Ley–Doudoroff (DD) pathways are known existing routes in E. coli that can supply the MEP precursors from d-galactose. The DD pathway was selected as it is capable of supplying equimolar amounts of pyruvate and G3P simultaneously. To exclusively direct d-galactose toward the DD pathway, an E. coli ΔgalK strain with blocked Leloir pathway was used as the host. To obtain a fully functional DD pathway, a dehydrogenase encoding gene (gld) was recruited from Pseudomonas syringae to catalyze d-galactose conversion to d-galactonate. Overexpressions of endogenous genes known as MEP bottlenecks, and a heterologous gene, were conducted to enhance and enable isoprene production, respectively. Growth test confirmed a functional DD pathway concomitant with equimolar generation of pyruvate and G3P, in contrast to the wild-type strain where G3P was limiting. Finally, the engineered strain with combined DD–MEP pathway exhibited the highest isoprene production. This suggests that the equimolar pyruvate and G3P pools resulted in a more efficient carbon flux toward isoprene production. This strategy provides a new platform for developing improved isoprenoid producing strains through the combined DD–MEP pathway.

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Acknowledgments

This work was supported by the Priority Research Centers Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Education, Science, and Technology (2009-0093816).

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Authors and Affiliations

  1. Department of Energy and Biotechnology (DEB), Energy and Environment Fusion Technology Center (E2FTC), Myongji University, 116 Myongji-ro, Cheoin-gu, Yongin-si, Gyeonggi-do, 449-728, Republic of Korea

    Kristine Rose M. Ramos, Kris Niño G. Valdehuesa, Huaiwei Liu, Grace M. Nisola & Wook-Jin Chung

  2. Division of Bioscience and Bioinformatics, Myongji University, 116 Myongji-ro, Cheoin-gu, Yongin-si, Gyeonggi-do, 449-728, Republic of Korea

    Won-Keun Lee

Authors
  1. Kristine Rose M. Ramos
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  2. Kris Niño G. Valdehuesa
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  3. Huaiwei Liu
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  4. Grace M. Nisola
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  5. Won-Keun Lee
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  6. Wook-Jin Chung
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Correspondence to Wook-Jin Chung.

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Ramos, K.R.M., Valdehuesa, K.N.G., Liu, H. et al. Combining De Ley–Doudoroff and methylerythritol phosphate pathways for enhanced isoprene biosynthesis from d-galactose. Bioprocess Biosyst Eng 37, 2505–2513 (2014). https://doi.org/10.1007/s00449-014-1228-z

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  • DOI: https://doi.org/10.1007/s00449-014-1228-z

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