Chain Compaction and Synergistic Destabilization of Globular Proteins by Mixture of Denaturants

Abstract

Characterization of unfolded states is essential to understand the mechanism of protein folding. This study examines the unfolded states of RNase A and α‐LA in the mixture of ′′structurally‐similar′′ chemical denaturants, arginine (Arg) and guanidinium chloride (Gdm). Thermal‐ and chemical‐unfolding experiments show that the mixture of denaturants synergistically destabilizes the proteins. Volumetric analysis shows that partial molar volume (V°) and adiabatic compressibility (Ks) of the proteins decreases with increasing [Gdm]. This suggests that the proteins might be gradually losing their intra‐molecular interactions resulting in decreased internal cavity. The addition of Arg to the Gdm‐unfolded states decreases both V° and Ks values indicating compaction of the protein chains. This could be attributed to reduction in the hydration of surface‐exposed residues of the proteins upon direct interaction with Arg and increased non‐native contacts. The results provide a direct experimental evidence for the compaction of unfolded protein chain upon addition of another denaturant.

Cryo vs Thermo: Duality of Ethylene Glycol on the Stability of Proteins

“…Therefore, we propose that the preferential interaction of EG stabilizes the proteins against cold-induced denaturation. However, at higher temperatures, the preferential binding of EG might destabilize the proteins, as commonly observed in the case of chemical denaturants such as urea and arginine. ,, The preferential binding of EG around the hydrophobic residues could decrease the cost of hydrophobic exposure of the residues at higher temperatures, thus destabilizing the native conformations. These observations indicate that the effect of preferential binding of cosolvents on protein stability might change with temperature, particularly at sub-zero temperature conditions, and with the nature of interactions that they have with the proteins.…”