Denatured state aggregation parameters derived from concentration dependence of protein stability

Area of Science:

• Biochemistry
• Protein Science
• Biophysical Chemistry

Background:

• Protein aggregation significantly impacts the long-term stability of protein formulations.
• Denatured or partially denatured proteins are prone to aggregation due to exposed hydrophobic cores.
• Protein aggregation shifts equilibrium, increasing denatured and aggregated protein populations.

Purpose of the Study:

• To present a new approach for simultaneously determining protein structural stability and the extent of denaturation and aggregation.
• To enable early recognition and quantification of denatured and aggregated proteins.
• To facilitate the selection of optimal solvent conditions and protein engineering for enhanced stability.

Main Methods:

• Analysis of the concentration dependence of the Gibbs energy (ΔG) of protein stability.
• Simultaneous evaluation of three key parameters related to protein conformation and aggregation.

Main Results:

• The method allows for the simultaneous determination of:
• The population of denatured protein.
• The population of aggregated protein.
• The fraction of denatured protein that has aggregated.

Conclusions:

• The developed approach provides a comprehensive assessment of protein stability and aggregation.
• This method is beneficial for identifying optimal conditions and engineering proteins with improved stability profiles.
• Early quantification of denaturation and aggregation is crucial for protein formulation and development.