Highly Disordered Amyloid-β Monomer Probed by Single-Molecule FRET and MD Simulation

Area of Science:

• Biophysics
• Protein Dynamics
• Neurodegenerative Diseases

Background:

• Amyloid-beta (Aβ) protein monomers are generally considered disordered.
• Controversy exists regarding transient or residual conformations that may drive oligomerization and fibril formation.
• Understanding Aβ conformation is crucial for Alzheimer's disease research.

Purpose of the Study:

• To investigate the conformational ensemble and dynamics of Aβ40 and Aβ42.
• To determine if long-lived conformational states exist in Aβ monomers.
• To provide molecular-level insights into Aβ dynamics using experimental and computational methods.

Main Methods:

• Single-molecule Förster resonance energy transfer (FRET) spectroscopy with site-specific labeling.
• Unnatural amino acid incorporation for precise dye placement.
• Nanosecond fluorescence correlation spectroscopy (nsFCS) for fluctuation timescale measurement.
• Molecular dynamics (MD) simulations with an optimized force field for disordered proteins.

Main Results:

• FRET efficiency distributions showed a single peak for both Aβ40 and Aβ42, suggesting a single apparent state.
• 2D FRET analysis revealed a broad distribution of rapidly interconverting conformations.
• Nanosecond fluorescence correlation spectroscopy measured fluctuation timescales of approximately 35 ns.
• MD simulations corroborated experimental findings, showing random polymer chain configurations with minimal secondary structure.

Conclusions:

• Both Aβ40 and Aβ42 populate an ensemble of rapidly reconfiguring unfolded states.
• No long-lived conformational states were distinguished from the disordered ensemble.
• The findings suggest Aβ monomers exist as dynamic, rapidly fluctuating structures rather than stable intermediates.