Dynamic control of pathway expression with riboregulated switchable feedback promoters

Glasscock, C. J.; Lazar, J. T.; Biggs, B. W.; Arnold, J. H.; Kang, M.-K.; Tullman-Ercek, D.; Tyo, K. E. J.; Lucks, J. B.

doi:10.1101/529180

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bioRxiv · 10.1101/529180

Dynamic control of pathway expression with riboregulated switchable feedback promoters

Glasscock, C. J.·Lazar, J. T.·Biggs, B. W.·Arnold, J. H.·Kang, M.-K.·Tullman-Ercek, D.·Tyo, K. E. J.·Lucks, J. B.

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Abstract

Dynamic pathway regulation has emerged as a promising strategy in metabolic engineering for improved system productivity and yield, and continues to grow in sophistication. Bacterial stress-response promoters allow dynamic gene regulation using the hosts natural transcriptional networks, but lack the flexibility to control the expression timing and overall magnitude of pathway genes. Here, we report a strategy that uses RNA transcriptional regulators to introduce another layer of control over the output of natural stress-response promoters. This new class of gene expression cassette, called a riboregulated switchable feedback promoter (rSFP), can be modularly activated using a variety of mechanisms, from manual induction to quorum sensing. We develop and apply rSFPs to regulate a toxic cytochrome P450 enzyme in the context of a Taxol precursor biosynthesis pathway and show this leads to 2.4x fold higher titers than from the best reported strain. We envision that rSFPs will become a valuable tool for flexible and dynamic control of gene expression in metabolic engineering, protein and biologic production, and many other applications.

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Glasscock, C. J., Lazar, J. T., Biggs, B. W., Arnold, J. H., Kang, M.-K., Tullman-Ercek, D., Tyo, K. E. J., Lucks, J. B.. 2019-01-23. Dynamic control of pathway expression with riboregulated switchable feedback promoters. https://doi.org/10.1101/529180

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