Biochemical and thermodynamic effects of Hofmeister salts on β-trypsin proteoform

Protein stability is profoundly influenced by the ionic environment, which impacts both structural conformation and catalytic function. In this study, we examined the effects of various Hofmeister salts on the biochemical, structural, and thermodynamic properties of the β-trypsin proteoform. We employed enzymatic assays, fluorescence spectroscopy, and differential scanning calorimetry (DSC) to evaluate how specific cations and anions modulate the enzyme's behavior. Our results show that low concentrations of CaCl₂ stabilize the native protein structure and enhance enzymatic activity, while higher concentrations induce destabilization, likely due to chaotropic interactions. In addition, it was possible to observe the thermodynamic panorama of the interaction of the ions with the solvation layers and with the proteins. These findings offer a comprehensive perspective on ion-protein interactions and have practical implications for the rational design of enzyme formulations in biotechnological applications.