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Evolutionary engineering of methylotrophic E. coli enables fast growth on methanol

Author

Listed:
  • Liang-Yu Nieh

    (Academia Sinica
    National Taiwan University)

  • Frederic Y.-H. Chen

    (Academia Sinica)

  • Hsin-Wei Jung

    (Academia Sinica)

  • Kuan-Yu Su

    (Academia Sinica)

  • Chao-Yin Tsuei

    (Academia Sinica)

  • Chun-Ting Lin

    (Academia Sinica)

  • Yue-Qi Lee

    (Academia Sinica)

  • James C. Liao

    (Academia Sinica)

Abstract

As methanol can be derived from either CO2 or methane, methanol economy can play an important role in combating climate change. In this scenario, rapid utilization of methanol by an industrial microorganism is the first and crucial step for efficient utilization of the C1 feedstock chemical. Here, we report the development of a methylotrophic E. coli strain with a doubling time of 3.5 hours under optimal conditions, comparable or faster than native model methylotrophs Methylorubrum extorquens AM1 (Td~4hr) and Bacillus methanolicus at 37°C (Td~5hr). To accomplish this, we develop a bacterial artificial chromosome (BAC) with dynamic copy number variation (CNV) to facilitate overcoming the formaldehyde-induced DNA-protein cross-linking (DPC) problem in the evolution process. We track the genome variations of 75 cultures along the evolution process by next-generation sequencing, and identified the features of the fast-growing strain. After stabilization, the final strain (SM8) grows to 20 g/L of cell mass within 77 hrs in a bioreactor. This study illustrates the potential of dynamic CNV as an evolution tool and synthetic methylotrophs as a platform for sustainable biotechnological applications.

Suggested Citation

  • Liang-Yu Nieh & Frederic Y.-H. Chen & Hsin-Wei Jung & Kuan-Yu Su & Chao-Yin Tsuei & Chun-Ting Lin & Yue-Qi Lee & James C. Liao, 2024. "Evolutionary engineering of methylotrophic E. coli enables fast growth on methanol," Nature Communications, Nature, vol. 15(1), pages 1-16, December.
    • Handle: RePEc:nat:natcom:v:15:y:2024:i:1:d:10.1038_s41467-024-53206-4
    DOI: 10.1038/s41467-024-53206-4
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    References listed on IDEAS

    as
    1. Fabian Meyer & Philipp Keller & Johannes Hartl & Olivier G. Gröninger & Patrick Kiefer & Julia A. Vorholt, 2018. "Methanol-essential growth of Escherichia coli," Nature Communications, Nature, vol. 9(1), pages 1-10, December.
    2. Philipp Keller & Michael A. Reiter & Patrick Kiefer & Thomas Gassler & Lucas Hemmerle & Philipp Christen & Elad Noor & Julia A. Vorholt, 2022. "Generation of an Escherichia coli strain growing on methanol via the ribulose monophosphate cycle," Nature Communications, Nature, vol. 13(1), pages 1-13, December.
    3. Benjamin M. Woolston & Jason R. King & Michael Reiter & Bob Van Hove & Gregory Stephanopoulos, 2018. "Improving formaldehyde consumption drives methanol assimilation in engineered E. coli," Nature Communications, Nature, vol. 9(1), pages 1-12, December.
    4. Erez Dekel & Uri Alon, 2005. "Optimality and evolutionary tuning of the expression level of a protein," Nature, Nature, vol. 436(7050), pages 588-592, July.
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