(141i) Thermodynamic Signature of Binding at the Nano-Bio Interface

Following the advances in design and characterization of engineered nanoparticles, interactions of biopolymers with these particles have been one research topic of high interest to both industry and academia. At the nanoparticle-biopolymer interface, dynamic interactions are complicated but have to be explored in order to implement nanotechnology for the benefit of living organisms including humans. In the current study, a model system composed of monodisperse silica nanoparticles (negatively charged) and the globular protein lysozyme (positively charged) was investigated via isothermal titration calorimetry (ITC), static-dynamic-electrophoretic light scattering and circular di-chroism (CD). A new approach and model was developed to quantitatively assess the binding of the protein on the nano-particles and an adsorption mechanism that accounts for both enthalpic and entrophic contributions were proposed for binding. The novel model is supported by a number of experimental evidences performed using the same model system and under the same conditions (e.g. temperature, ionic strength, the concentration ratios, mixing time, equilibration time) for each technique. To compare the new model with the Langmuir model, traditional adsorption isotherms were obtained with depletion method. Calculated theoretical surface coverage and adsorbed protein thickness was compared with the Transmission Electron Microscopy Images to investigate the extent of the protein layer formed on nanoparticles. Even though each experimental method has its own limitations and theoretical modeling is only feasible after a few assumptions, thermal footprints of protein titration into nanoparticle suspensions indicate two types of interactions. The first type of interaction has higher binding affinity and driven by a higher enthalpic contribution compared to the second type. The protein bound with high affinity looses 45% of its helical structure according to deconvolution of CD signal whereas entropically driven second type interaction does not cause a significant change in protein secondary structure. Zeta potential of silica- protein complex aggregates reaches to equilibrium (+10 mV ) at the concentration ratio that refers to the equilibrium of first type of interaction ( Silica/Protein= 15 ) and multi angle dynamic light scattering  does not indicate further grow of aggregates after the first equilibrium. Based on all these findings, it is proposed that lysozyme adsorption on nano-silica is a result of protein-nanoparticle and protein-protein interactions that are driven by both enthalpic and entopic contributions depending on the extent of adsorption which can not be modeled by monolayer Langmuir model.  As more studies conducted with different model systems, under different conditions; mechanism of interactions at the nano-bio interface will be understood better. Finally a better fundamental understanding will lead more applications that use or include proteins and nanoparticles in food, pharmaceutical, cosmetic and biomedical fields.