Osmolyte Structural and Thermodynamic Effects Across the Protein Folding Landscape.

Area of Science:

• Biochemistry and Molecular Biophysics
• Computational Chemistry
• Protein Dynamics

Background:

• Understanding protein folding and the role of osmolytes like urea and TMAO is crucial for molecular biology.
• The interplay between cosolvents and protein conformational changes influences protein stability and function.
• Previous studies have explored osmolyte effects, but a comprehensive view across complete folding landscapes was lacking.

Purpose of the Study:

• To characterize the structure and thermodynamics of urea and TMAO preferential interactions across the entire folding landscape of two model proteins: the SRC Homology 3 (SH3) domain and the B domain of protein A (BdpA).
• To investigate how these interactions correlate with changes in surface area during protein denaturation.
• To elucidate the specific contributions of backbone, side chain, and individual residues to solvation mechanisms.

Main Methods:

• Utilized a novel computational pipeline combining coarse-grained and atomistic simulations to model solvation structures across complete folding landscapes.
• Applied minimum-distance distribution functions and Kirkwood-Buff solvation theory for detailed solvation analysis.
• Calculated residue-specific contributions to distribution functions and transfer free energies.

Main Results:

• A high correlation was found between preferential interactions and the surface area of denatured states for both SH3 and BdpA proteins.
• Urea destabilized partially denatured states of BdpA, modulating its unfolding pathway, while SH3 denaturation consistently increased surface area.
• Hydrogen bonds with urea and TMAO weakened upon denaturation, while nonspecific interactions strengthened in unfolded structures; urea-backbone and nonpolar-cosolvent interactions were key solvation mechanisms.
• Cosolvent effects on transfer free energies closely matched experimental data (within 1 kcal mol⁻¹).

Conclusions:

• Preferential interactions with urea and TMAO are strongly linked to the surface area exposed during protein denaturation.
• The unfolding pathways of proteins can be significantly modulated by cosolvents, depending on the protein's specific folding landscape.
• The developed computational methods provide a robust framework for studying solvation in complex molecular systems with large conformational changes.