Post-translational control of genetic circuits using Potyvirus proteases
Name
Fernandez-Rodriguez-2016-Post-translational c.pdf
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2.48 MB
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Adobe PDF
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Author(s)
Fernandez-Rodriguez, Jesus
Voigt, Christopher A.
Date Issued
June 2016
Journal
Nucleic Acids Research
Publisher
Oxford University Press
Citation
Fernandez-Rodriguez, Jesus, and Christopher A. Voigt. "Post-translational control of genetic circuits using Potyvirus proteases." Nucleic Acids Research (June 13, 2016), pp.1-10.
Version
Final published version
Abstract
Genetic engineering projects often require control over when a protein is degraded. To this end, we use a fusion between a degron and an inactivating peptide that can be added to the N-terminus of a protein. When the corresponding protease is expressed, it cleaves the peptide and the protein is degraded. Three protease:cleavage site pairs from Potyvirus are shown to be orthogonal and active in exposing degrons, releasing inhibitory domains and cleaving polyproteins. This toolbox is applied to the design of genetic circuits as a means to control regulator activity and degradation. First, we demonstrate that a gate can be constructed by constitutively expressing an inactivated repressor and having an input promoter drive the expression of the protease. It is also shown that the proteolytic release of an inhibitory domain can improve the dynamic range of a transcriptional gate (200-fold repression). Next, we design polyproteins containing multiple repressors and show that their cleavage can be used to control multiple outputs. Finally, we demonstrate that the dynamic range of an output can be improved (8-fold to 190-fold) with the addition of a protease-cleaved degron. Thus, controllable proteolysis offers a powerful tool for modulating and expanding the function of synthetic gene circuits.
MIT Department
MIT Synthetic Biology Center
Massachusetts Institute of Technology. Department of Biological Engineering
Terms of Use
Creative Commons Attribution Non-Commercial License
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DOI of Published Version
https://doi.org/10.1093/nar/gkw537
