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Computation-guided optimization of split protein systems

View ORCID ProfileTaylor B. Dolberg, View ORCID ProfileAnthony T. Meger, View ORCID ProfileJonathan D. Boucher, View ORCID ProfileWilliam K. Corcoran, View ORCID ProfileElizabeth E. Schauer, View ORCID ProfileAlexis N. Prybutok, View ORCID ProfileSrivatsan Raman, View ORCID ProfileJoshua N. Leonard
doi: https://doi.org/10.1101/863530
Taylor B. Dolberg
1Department of Chemical and Biological Engineering, Northwestern University, Evanston, Illinois 60208, United States
2Center for Synthetic Biology, Northwestern University, Evanston, Illinois 60208, United States
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Anthony T. Meger
3Department of Biochemistry, University of Wisconsin-Madison, Madison, Wisconsin 53706, United States
4Great Lakes Bioenergy Research Center, University of Wisconsin-Madison, Madison, Wisconsin 53706, United States
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Jonathan D. Boucher
2Center for Synthetic Biology, Northwestern University, Evanston, Illinois 60208, United States
5Interdisciplinary Biological Sciences Program, Northwestern University, Evanston, Illinois 60208, United States
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William K. Corcoran
2Center for Synthetic Biology, Northwestern University, Evanston, Illinois 60208, United States
5Interdisciplinary Biological Sciences Program, Northwestern University, Evanston, Illinois 60208, United States
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Elizabeth E. Schauer
1Department of Chemical and Biological Engineering, Northwestern University, Evanston, Illinois 60208, United States
2Center for Synthetic Biology, Northwestern University, Evanston, Illinois 60208, United States
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Alexis N. Prybutok
1Department of Chemical and Biological Engineering, Northwestern University, Evanston, Illinois 60208, United States
2Center for Synthetic Biology, Northwestern University, Evanston, Illinois 60208, United States
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Srivatsan Raman
3Department of Biochemistry, University of Wisconsin-Madison, Madison, Wisconsin 53706, United States
4Great Lakes Bioenergy Research Center, University of Wisconsin-Madison, Madison, Wisconsin 53706, United States
8Department of Bacteriology, University of Wisconsin-Madison, Madison, Wisconsin 53706, United States
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  • For correspondence: sraman4@wisc.edu j-leonard@northwestern.edu
Joshua N. Leonard
1Department of Chemical and Biological Engineering, Northwestern University, Evanston, Illinois 60208, United States
2Center for Synthetic Biology, Northwestern University, Evanston, Illinois 60208, United States
5Interdisciplinary Biological Sciences Program, Northwestern University, Evanston, Illinois 60208, United States
6Chemistry of Life Processes Institute, Northwestern University, Evanston, Illinois 60208, United States
7Member, Robert H. Lurie Comprehensive Cancer Center, Northwestern University, Evanston, Illinois 60208, United States
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  • For correspondence: sraman4@wisc.edu j-leonard@northwestern.edu
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ABSTRACT

Splitting bioactive proteins, such as enzymes or fluorescent reporters, into conditionally reconstituting fragments is a powerful strategy for building tools to study and control biochemical systems. However, split proteins often exhibit a high propensity to reconstitute even in the absence of the conditional trigger, which limits their utility. Current approaches for tuning reconstitution propensity are laborious, context-specific, or often ineffective. Here, we report a computational design-driven strategy that is grounded in fundamental protein biophysics and which guides the experimental evaluation of a focused, sparse set of mutants—which vary in the degree of interfacial destabilization while preserving features such as stability and catalytic activity—to identify an optimal functional window. We validate our method by solving two distinct split protein design challenges, generating both broad insights and new technology platforms. This method will streamline the generation and use of split protein systems for diverse applications.

Footnotes

  • One author name was corrected (Srivatsan Raman) in the bioRxiv metadata associated with this submission. Submitted files are unchanged.

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The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. All rights reserved. No reuse allowed without permission.
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Posted December 05, 2019.
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Computation-guided optimization of split protein systems
Taylor B. Dolberg, Anthony T. Meger, Jonathan D. Boucher, William K. Corcoran, Elizabeth E. Schauer, Alexis N. Prybutok, Srivatsan Raman, Joshua N. Leonard
bioRxiv 863530; doi: https://doi.org/10.1101/863530
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Computation-guided optimization of split protein systems
Taylor B. Dolberg, Anthony T. Meger, Jonathan D. Boucher, William K. Corcoran, Elizabeth E. Schauer, Alexis N. Prybutok, Srivatsan Raman, Joshua N. Leonard
bioRxiv 863530; doi: https://doi.org/10.1101/863530

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