Volume changes of proteins adsorbed on silica particles

Abstract

Proteins often stay at interfaces, where their conformation and biological activity can be altered. In this study, we investigate the underlying changes in the folding stability and molecular volume of proteins using staphylococcal nuclease (SNase) as the model protein and colloidal silica particles as the model adsorbent. The folding stability and molecular volume have been determined by high-pressure fluorescence experiments in the range of 1–2500 bar utilizing the intrinsic Trp fluorescence of SNase. At pH = 7.0 and 25 °C, SNase, dissolved in bulk solution, is characterized by a volume change of unfolding of −73 mL mol⁻¹. This value is drastically reduced, when SNase is adsorbed on silica particles. Here, volume changes in the range of 41 to 32 mL mol⁻¹ can be measured at silica particles differing in their surface charge density. In addition, the standard Gibbs energy change and the pressure of unfolding are strongly reduced in the adsorbed state of SNase indicating a surface-induced destabilization of the protein native structure. The effect of the pH-value has been studied as well. Whereas a pH-change in the range of 7–10 has no significant effect on the pressure-driven unfolding of dissolved SNase, strong pH-dependence is observed for the adsorbed SNase. As the pH is approaching the isoelectric point of SNase, the conformational stability of the adsorbed protein is lowered drastically. The results of this study reveal a novel view on conformational changes upon protein adsorption. It is suggested that the molecular volume of SNase is lowered by a partial filling of the protein void volume corresponding to about two water molecules.