(451d) Evaluating the Role of Solid Surfaces in Inducing Conformational Changes of Adsorbed and Desorbed Proteins
Presently, more than 130 biologically-active pharmaceuticals are approved for clinical use by the FDA. The successful formulation of these biologics is highly dependent on maintaining the stability of the drug throughout its lifecycle. The most common route of administration of these protein and peptide-based drugs is intravenous injection. During storage and delivery of solution-based proteins, they interact with various types of surfaces, often for extended periods of time under sub-optimal conditions. Surface-induced perturbation of protein structure can occur, potentially causing modifications in conformation and activity, and resulting in population of non-native, aggregate-prone states. Such behavior may ultimately compromise the clinical safety and efficacy of these high-value biologics. Here, we present a model system for evaluating surface-induced changes in protein secondary structure on both hydrophilic and hydrophobic surfaces. The system allows insight into protein-surface affinity, kinetics of adsorption and real-time monitoring of adsorbed and desorbed protein structure using circular dichroism. Conformational changes of model proteins interacting with silica nanoparticles were monitored, and both the adsorbed and desorbed states of the protein were characterized. The results show significant loss of structure after adsorption of the particles, which is accompanied by a helix to sheet transition. We present a technique for measuring the secondary structure of proteins in the adsorbed state, and results show the adsorbed spectra of various model proteins does not seem to be as dependent on surface coverage as previously thought. Desorbed proteins were also isolated and characterized. Intrinsically stable proteins, such as lysozyme, show structural similarity between desorbed and native protein, indicating refolding upon desorption. Less stable proteins, on the other hand, show desorbed proteins with altered structure compared to the native, indicating that structural loss in the adsorbed state is partially retained after desorption. Finally, structural states of adsorbed protein are reported following the desorption of various populations of proteins. Results indicate that the most highly unfolded states remain tightly bound to the surface following the desorption of more structurally ordered, loosely bound proteins. The kinetics of helix to sheet transition of these various states of adsorbed proteins are shown.