Two-State Folding Energy Determination Based on Transition Points in Nonequilibrium Single-Molecule Experiments

Area of Science:

• Biophysics
• Single-molecule biophysics
• Statistical mechanics

Background:

• Many biomolecules like proteins and nucleic acids exhibit two-state unfolding-refolding transitions under mechanical stress.
• Single-molecule techniques are crucial for observing these transitions at the molecular level.

Purpose of the Study:

• To develop a simplified analytical method for determining molecular folding energy using the Jarzynski equality (JE).
• To enable accurate folding energy calculations from minimal experimental data.

Main Methods:

• Application of the Jarzynski equality (JE) to analytically express folding energy.
• Utilizing experimentally measured transition points (ξ*) under force (F(t)) and position (R(t)) constraints.
• Demonstrating the method's applicability with magnetic tweezers experiments.

Main Results:

• The folding energy can be determined from a single measured data point, significantly simplifying previous methods.
• The force constraint (F(t)) method yields handle-independent results.
• Successful measurement of folding energies for DNA hairpin, DNA G-quadruplex, and titin I27 domain.

Conclusions:

• The developed JE-based approach offers a more efficient and accessible method for quantifying molecular folding energies.
• This technique has broad applicability for studying the mechanical properties of various biomolecules.