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Influence of Point Mutations on PR65 Conformational Adaptability: Insights from Nanoaperture Optical Tweezer
Ivet Bahar1, Anupam Banerjee1, Samuel Mathew2
1Stony Brook University.
Research Square
|November 28, 2023
Summary
Protein phosphatase 2A scaffold subunit PR65
Area of Science:
- Biochemistry and structural biology
- Protein dynamics and mechanics
- Single-molecule biophysics
Background:
- PR65 is the HEAT-repeat scaffold of protein phosphatase 2A (PP2A), crucial for its function.
- PR65's spring-like structure and conformational changes regulate substrate binding and catalysis.
- Understanding PR65 mechanics is key to deciphering PP2A regulation.
Approach:
- Utilized *in silico* saturation mutagenesis to identify key 'hinge' residues in PR65.
- Performed molecular simulations to predict the impact of hinge mutations on PR65 conformation.
- Employed nanoaperture optical tweezers to experimentally characterize PR65 motion and conformational states.
Key Points:
- Identified specific PR65 'hinge' residues whose mutations alter conformational adaptability.
- Computational predictions of conformational changes (stabilized open or closed states) were confirmed experimentally.
- Mutations shifting PR65 towards extended conformations showed distinct biophysical properties (higher corner frequencies, lower scattering).
Conclusions:
- Experimental data validated *in silico* predictions regarding PR65 hinge mutations and conformational mechanics.
- Nanoaperture optical tweezers are effective for studying protein dynamics at the single-molecule level.
- Integrating computational and single-molecule experimental approaches provides powerful insights into protein function.
