Protein structure and bioactivity upon adsorption and desorption from nanosilicate sustained release delivery devices.

Sustained local delivery of biologics via hydrogel carriers is a promising approach to enhance protein safety and efficacy. The addition of nanoparticles to polymeric hydrogels has been shown to further improve the retention and delivery kinetics of biologics. Specifically, nanoparticles with high surface area, such as nanosilicates, have shown potential for complexing with biologics to enable highly tunable release profiles. Here, LAPONITE® XLG nanosilicate (NS) was examined due to its platelet-like structure with negatively charged faces and positively charged edges. Our previous results have shown NS to greatly slow the release of model proteins from poly(ethylene glycol) (PEG) hydrogels due to NS-protein complexation. This work aims to determine the structure and stability of several NS-protein complexes, as well as protein activity and structure upon complexation. Binding affinity assays revealed a strong correlation between affinity and protein charge, with positively charged proteins being more attracted to NS. Proteins were shown to unfold in the presence of NS in solution, leading to a partial loss in bioactivity. However, this unfolding was determined to be temporary, as proteins released from PEG-NS hydrogels recovered secondary structure and bioactivity. Binding to NS also provided some protection against protein denaturant guanidine thiocyanate. Through understanding the interactions between proteins and NS, this study paves the way for the application of these NS-protein complexes as tunable, sustained-release delivery devices.