Rational Design of Phosphorylation-Responsive Coiled Coil-Peptide Assemblies.
Harry F Thompson1,2, Joseph L Beesley1,2, Hannah D Langlands1,2
1School of Biochemistry, University of Bristol, University Walk, Bristol, BS8 1TD, U.K.
ACS Synthetic Biology
|March 29, 2023
Summary
Engineered de novo proteins can reversibly switch states. Site-specific phosphorylation triggers disassembly, while dephosphorylation causes reassembly, advancing synthetic biology and biomaterials.
Area of Science:
- Protein engineering
- Synthetic biology
- Biomaterials
Background:
- Advancements in protein design and synthetic biology require stimuli-responsive peptides and proteins.
- De novo protein design offers a platform for creating novel molecular systems.
Purpose of the Study:
- To engineer de novo protein systems that reversibly disassemble and reassemble in response to phosphorylation and dephosphorylation.
- To utilize hyperthermostable coiled-coil heterotetramers as a base for controlled protein switching.
Main Methods:
- Designed de novo antiparallel and parallel coiled-coil heterotetramers (A2B2 systems).
- Incorporated protein kinase A phosphorylation sites (R-R-X-S) to disrupt coiled-coil interfaces.
- Utilized protein kinase A for phosphorylation-induced disassembly and Lambda Protein Phosphatase for dephosphorylation-induced reassembly.
Main Results:
- Unphosphorylated peptides assembled as designed and unfolded reversibly upon heating.
- Kinase addition induced disassembly of assembled states with half-lives of ≤5 min.
- Reversal of phosphorylation by Lambda Protein Phosphatase resulted in tetramer reassembly.
Conclusions:
- Demonstrated successful engineering of de novo coiled-coil systems with phosphorylation-dependent switching.
- Developed a mechanistic model for these responsive protein components.
- Highlighted potential applications in synthetic biology, biomaterials, and biotechnology.
Keywords:
coiled coilinducible conformational switchphosphorylationprotein designrational peptide designsynthetic biologyMore Related Videos
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