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An in vivo gene amplification system for high level expression in Saccharomyces cerevisiae

Author

Listed:
  • Bingyin Peng

    (The University of Queensland
    Commonwealth Scientific and Industrial Research Organisation (CSIRO)
    Queensland University of Technology
    Queensland University of Technology)

  • Lygie Esquirol

    (The University of Queensland
    Griffith University)

  • Zeyu Lu

    (The University of Queensland
    Queensland University of Technology
    Queensland University of Technology)

  • Qianyi Shen

    (The University of Queensland
    Queensland University of Technology
    Queensland University of Technology)

  • Li Chen Cheah

    (The University of Queensland
    Queensland University of Technology)

  • Christopher B. Howard

    (The University of Queensland)

  • Colin Scott

    (Commonwealth Scientific and Industrial Research Organisation (CSIRO)
    Black Mountain Science and Innovation Park)

  • Matt Trau

    (The University of Queensland
    The University of Queensland)

  • Geoff Dumsday

    (CSIRO Manufacturing)

  • Claudia E. Vickers

    (Commonwealth Scientific and Industrial Research Organisation (CSIRO)
    Queensland University of Technology
    Queensland University of Technology
    Griffith University)

Abstract

Bottlenecks in metabolic pathways due to insufficient gene expression levels remain a significant problem for industrial bioproduction using microbial cell factories. Increasing gene dosage can overcome these bottlenecks, but current approaches suffer from numerous drawbacks. Here, we describe HapAmp, a method that uses haploinsufficiency as evolutionary force to drive in vivo gene amplification. HapAmp enables efficient, titratable, and stable integration of heterologous gene copies, delivering up to 47 copies onto the yeast genome. The method is exemplified in metabolic engineering to significantly improve production of the sesquiterpene nerolidol, the monoterpene limonene, and the tetraterpene lycopene. Limonene titre is improved by 20-fold in a single engineering step, delivering ∼1 g L−1 in the flask cultivation. We also show a significant increase in heterologous protein production in yeast. HapAmp is an efficient approach to unlock metabolic bottlenecks rapidly for development of microbial cell factories.

Suggested Citation

  • Bingyin Peng & Lygie Esquirol & Zeyu Lu & Qianyi Shen & Li Chen Cheah & Christopher B. Howard & Colin Scott & Matt Trau & Geoff Dumsday & Claudia E. Vickers, 2022. "An in vivo gene amplification system for high level expression in Saccharomyces cerevisiae," Nature Communications, Nature, vol. 13(1), pages 1-12, December.
    • Handle: RePEc:nat:natcom:v:13:y:2022:i:1:d:10.1038_s41467-022-30529-8
    DOI: 10.1038/s41467-022-30529-8
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    References listed on IDEAS

    as
    1. Codruta Ignea & Morten H. Raadam & Mohammed S. Motawia & Antonios M. Makris & Claudia E. Vickers & Sotirios C. Kampranis, 2019. "Orthogonal monoterpenoid biosynthesis in yeast constructed on an isomeric substrate," Nature Communications, Nature, vol. 10(1), pages 1-15, December.
    2. Adam L. Meadows & Kristy M. Hawkins & Yoseph Tsegaye & Eugene Antipov & Youngnyun Kim & Lauren Raetz & Robert H. Dahl & Anna Tai & Tina Mahatdejkul-Meadows & Lan Xu & Lishan Zhao & Madhukar S. Dasika , 2016. "Rewriting yeast central carbon metabolism for industrial isoprenoid production," Nature, Nature, vol. 537(7622), pages 694-697, September.
    3. Zeyu Lu & Bingyin Peng & Birgitta E. Ebert & Geoff Dumsday & Claudia E. Vickers, 2021. "Auxin-mediated protein depletion for metabolic engineering in terpene-producing yeast," Nature Communications, Nature, vol. 12(1), pages 1-13, December.
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