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FimH as a scaffold for regulated molecular recognition
Shivani Gupta Ludwig1, Casey L Kiyohara1, Laura A Carlucci1
1Department of Bioengineering, University of Washington, 3720 15th Ave NE. Foege N430P, Box 355061, Seattle, USA.
Journal of Biological Engineering
|January 13, 2021
Summary
Researchers engineered the bacterial FimH protein to create regulated molecular recognition. This scaffold maintains conformation-dependent binding, enabling tunable affinity for new targets like Penta-His antibody through allosteric and parasteric mechanisms.
Area of Science:
- Biotechnology
- Protein Engineering
- Molecular Recognition
Background:
- Antibodies are the standard for molecular recognition but lack convenient regulation.
- Alternative protein scaffolds offer opportunities for novel functionalities, including regulated binding.
- The bacterial adhesin FimH exhibits conformation-dependent binding to mannose, regulated by allosteric and parasteric mechanisms.
Purpose of the Study:
- To investigate FimH as a scaffold for regulated molecular recognition.
- To determine if FimH's conformational regulation is retained after altering its binding site.
- To explore the utility of FimH's native regulatory mechanisms for new targets.
Main Methods:
- Reengineering the FimH binding site to recognize non-mannosylated targets (nickel, Penta-His antibody).
- Assessing changes in binding affinity (KD) between different FimH conformations.
- Utilizing allosteric and parasteric regulation with native ligands and antibodies to modulate binding.
Main Results:
- Engineered FimH variants maintained conformational regulation, showing up to a 7-fold difference in KD for new targets.
- Both allosteric and parasteric mechanisms successfully regulated binding to Penta-His antibody.
- Allosteric regulation by mab21 reduced affinity 7-fold, inducing 98% target detachment.
Conclusions:
- FimH serves as a versatile scaffold for developing conformationally regulated binding proteins.
- The deeply studied, conformation-dependent binding of FimH can be adapted for multiple regulatory strategies.
- This approach offers potential for advanced biotechnology applications requiring tunable molecular recognition.
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