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Dissecting the Determinants of Domain Insertion Tolerance and Allostery in Proteins

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Mathony, Jan, Aschenbrenner, Sabine, Becker, Philipp, Niopek, Dominik·2023·bioRxiv (Cold Spring Harbor Laboratory)
DOI 10.1101/2023.04.11.536407

Preprint — published as 10.1002/advs.202303496 · Paper page

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Abstract

Domain insertion engineering is a promising approach to recombine the functions of evolutionarily unrelated proteins. Insertion of light-switchable receptor domains into a selected effector protein, for instance, can yield allosteric effectors with light-dependent activity. However, the parameters that determine domain insertion tolerance are poorly understood. Here, we used an unbiased screen to systematically assess the domain insertion permissibility of several evolutionary unrelated proteins. Training machine learning models on the resulting data allowed us to dissect features informative for domain insertion tolerance and revealed sequence conservation statistics as the strongest indicators of suitable insertion sites. Finally, extending our experimental pipeline towards the identification of switchable hybrids resulted in opto-chemogenetic derivatives of the transcription factor AraC that function as single-protein Boolean logic gates. Our study reveals determinants of domain insertion tolerance and facilitates the engineering of switchable proteins with unique mechanistic properties.

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Metadata

Authors
Mathony, J., Aschenbrenner, S., Becker, P., & Niopek, D.
Affiliations
  • Technische Universität Darmstadt (DE)
  • Heidelberg University (DE)
Corresponding author
  • Mathony, Jan
  • Niopek, Dominik
ORCID
Venue
bioRxiv (Cold Spring Harbor Laboratory)
Publisher
openRxiv
Published
2023-04-11
DOI
10.1101/2023.04.11.536407
OpenAlex
W4365148007
Cited by
0
References
2
Type
posted content
Language
en
Open access
Licence
cc-by
Locations
  • bioRxiv (Cold Spring Harbor Laboratory) (acceptedVersion, cc-by)
  • Technical University of Denmark, DTU Orbit (Technical University of Denmark, DTU) (submittedVersion, cc-by)
  • TUbilio (Technical University of Darmstadt) (submittedVersion, cc-by)
  • Technischen Universität Darmstadt (submittedVersion, cc-by)
  • Technischen Universität Darmstadt (cc-by)
Funders
  • European Commission (101041570)
  • Deutsche Forschungsgemeinschaft (453202693)
  • Aventis Foundation
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Protein Structure and Dynamics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Biochemical and Structural Characterization
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Gene Regulatory Network Analysis
Keywords
protein allostery, allosteric effectors, sequence conservation, machine learning prediction, optogenetics, chemogenetics, Boolean logic gates, protein engineering

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CoChat. (2026). Citation check: Dissecting the Determinants of Domain Insertion Tolerance and Allostery in Proteins. https://app.cochat.ai/papers/10.1101/2023.04.11.536407
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Mathony, J., Aschenbrenner, S., Becker, P., & Niopek, D. (2023). Dissecting the Determinants of Domain Insertion Tolerance and Allostery in Proteins. bioRxiv (Cold Spring Harbor Laboratory). https://doi.org/10.1101/2023.04.11.536407