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Catch and Release: Engineered Allosterically Regulated β-Roll Peptides Enable On/Off Biomolecular Recognition
Beyza Bulutoglu1, Kevin Dooley1, Géza Szilvay1
1Department of Chemical Engineering, Columbia University , New York, New York 10027, United States.
ACS Synthetic Biology
|May 19, 2017
Summary
Researchers developed a novel calcium-activated protein scaffold, the repeat-in-toxin (RTX) domain, for targeted biomolecular recognition. This stimulus-responsive peptide binds lysozyme with high affinity and can be switched on/off, enabling controlled molecular capture and release.
Area of Science:
- Biochemistry
- Molecular Biology
- Protein Engineering
Background:
- Immunoglobulin domains have limitations for biomolecular recognition.
- Repeat-in-toxin (RTX) domains reversibly adopt a β-roll structure upon calcium binding.
Purpose of the Study:
- To engineer RTX peptides for stimulus-responsive biomolecular recognition.
- To develop an alternative scaffold for controlled molecular interactions.
Main Methods:
- Selection of an RTX peptide library using ribosome display against lysozyme.
- Identification and concatenation of high-affinity RTX mutants.
- Immobilization of RTX peptides for affinity chromatography.
Main Results:
- Identified RTX mutants with low micromolar dissociation constants for lysozyme.
- Achieved a 500-fold increase in affinity, resulting in a 65 nM apparent dissociation constant.
- Demonstrated on/off molecular recognition via calcium-dependent binding and release.
Conclusions:
- Designed a novel stimulus-responsive scaffold based on the RTX domain.
- RTX peptides enable environmentally responsive specific molecular recognition and self-assembly.
- This approach offers a new tool for controlled biomolecular interactions.
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