Letter | Published:

Optogenetic regulation of engineered cellular metabolism for microbial chemical production

Nature volume 555, pages 683687 (29 March 2018) | Download Citation

Abstract

The optimization of engineered metabolic pathways requires careful control over the levels and timing of metabolic enzyme expression1,2,3,4. Optogenetic tools are ideal for achieving such precise control, as light can be applied and removed instantly without complex media changes. Here we show that light-controlled transcription can be used to enhance the biosynthesis of valuable products in engineered Saccharomyces cerevisiae. We introduce new optogenetic circuits to shift cells from a light-induced growth phase to a darkness-induced production phase, which allows us to control fermentation with only light. Furthermore, optogenetic control of engineered pathways enables a new mode of bioreactor operation using periodic light pulses to tune enzyme expression during the production phase of fermentation to increase yields. Using these advances, we control the mitochondrial isobutanol pathway to produce up to 8.49 ± 0.31 g l−1 of isobutanol and 2.38 ± 0.06 g l−1 of 2-methyl-1-butanol micro-aerobically from glucose. These results make a compelling case for the application of optogenetics to metabolic engineering for the production of valuable products.

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Acknowledgements

We thank K. Gardner and L. Motta-Mena for providing the plasmids and maps for the EL222 system (pVP16–EL222 and pC120-Fluc)12, D. Pincus for plasmid pNH603, J. J. Lee for plasmid pET28a Ldh, C. Taxis for plasmid pDS143, C. Nelson for sharing her qPCR equipment, S. Han for assistance in qPCR experiments and figure presentation, J. Rabinowitz and J. Storey for sharing their 500-ml Sixfors fermentation system for fed-batch fermentation experiments, S. Silverman for technical assistance on this equipment and C. DeCoste and the Princeton Molecular Biology Flow Cytometry Resource Center for assistance in flow cytometry experiments. This work was supported by the Alfred P. Sloan Foundation (to J.L.A.), The Pew Charitable Trusts (to J.L.A.), National Institutes of Health grant DP2EB024247 (to J.E.T.) and an Eric and Wendy Schmidt Transformative Technology Fund grant (to J.L.A. and J.E.T.).

Author information

Affiliations

  1. Department of Chemical and Biological Engineering, Hoyt Laboratory, Princeton University, 25 William Street, Princeton, New Jersey 08544, USA

    • Evan M. Zhao
    • , Yanfei Zhang
    • , Justin Mehl
    • , Helen Park
    • , Makoto A. Lalwani
    •  & José L. Avalos
  2. Department of Molecular Biology, 140 Lewis Thomas Laboratory, Washington Road, Princeton, New Jersey 08544, USA

    • Jared E. Toettcher
  3. The Andlinger Center for Energy and the Environment, Princeton University, 86 Olden Street, Princeton, New Jersey 08544, USA

    • José L. Avalos

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Contributions

E.M.Z., J.E.T. and J.L.A. conceived this project and designed the experiments. E.M.Z., Y.Z. and J.L.A. constructed the strains and plasmids. E.M.Z. and H.P. performed the experiments that are shown in Fig. 1; E.M.Z. and J.M. conducted experiments illustrated in Fig. 2; E.M.Z. performed experiments that are shown in Fig. 3; E.M.Z. performed experiments illustrated in Extended Data Figs 1, 2, 3, 4, 5, 6, 7, 8, 9, 10. Y.Z. performed experiments illustrated in Extended Data Figs 1, 8. M.A.L. performed experiments illustrated in Extended Data Fig. 10. E.M.Z., J.E.T. and J.L.A. analysed the data and wrote the paper.

Competing interests

The authors declare no competing financial interests.

Corresponding authors

Correspondence to Jared E. Toettcher or José L. Avalos.

Reviewer Information Nature thanks L. Larrondo and the other anonymous reviewer(s) for their contribution to the peer review of this work.

Publisher's note: Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Extended data

Supplementary information

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  1. 1.

    Life Sciences Reporting Summary

  2. 2.

    Supplementary Information

    This file contains the Detailed Results, a Supplementary Discussion, Supplementary Gene Sequences, Supplementary Tables 1-7, a Supplementary Note and Supplementary References.

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DOI

https://doi.org/10.1038/nature26141

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