Solution thermodynamic approach to analyze protein stability in aqueous solutions

Area of Science:

• Biochemistry
• Physical Chemistry
• Thermodynamics

Background:

• Protein thermal stability is crucial for protein function and is influenced by solution conditions.
• Hydrogen bonds (HB) and water activity (A_w) are key factors affecting protein structure and stability.
• Understanding these factors is essential for protein engineering and biopharmaceutical development.

Purpose of the Study:

• To analyze protein thermal stability using a solution thermodynamic approach.
• To investigate the role of water activity (A_w) in protein stability.
• To establish the relationship between A_w, solution structure, and protein unfolding.

Main Methods:

• Solution thermodynamic analysis of protein thermal stability.
• Precise determination of water activity (A_w) in various solutions.
• Wyman-Tanford analysis to correlate protein unfolding ratio with A_w.

Main Results:

• Small energetic differences in hydrogen bonds (HB) are amplified, shifting the protein folding-unfolding equilibrium.
• A linear regression was observed between protein unfolding ratio and A_w for lysozyme, ribonuclease A, and α-chymotrypsinogen A.
• The free energy difference (ΔΔG) was readily obtained from the linear regression, quantifying stability changes.

Conclusions:

• Protein stability is determined by hydration, solute-binding effects, and solution structure, which influences hydrophobic interactions.
• Water activity (A_w) is a critical determinant of protein stability in aqueous solutions.
• Temperature dependence of HB is interrelated with hydrophobic interactions, further influencing protein stability.