Novel Method for Protein Stability and Delivery through the Formation of Complex Coacervates

Polyelectrolyte complexes and complex coacervation offer a novel means for the delivery of proteins and other bioactive materials. Polypeptides are biocompatible materials whose tunable properties lend themselves as viable vehicles for the delivery of drugs and, in particular, proteins. We also investigated the encapsulation of green fluorescent protein (GFP), bovine serum albumin (BSA), and lysozyme as model protein systems into polypeptide-based complex coacervates. These polypeptides are capable of mimicking the milieu found in cells, particularly cells’ natural characteristics of compartmentalization, crowding and soft interactions. The phase separation that occurs with coacervate formation gives way to compartmentalization and the high concentration of molecules simulates the biomolecules content, which also favors soft interactions between involved species. Additionally, these coacervates are aqueous, just like cells, and have tunable chemistry for furthering their cell-like characteristics. As such, the ability for peptide based coacervate systems to partially mimic this biological environment presents a viable means for potentially allowing proteins to exist in a wide array of environmental conditions without concern for their degradation. To further investigate protein encapsulation, we propose the creation of polypeptide-based block copolymers built from poly-lysine and polystyrene. We will focus on the characterization of polyelectrolyte complex micelle formation between various combinations of polypeptide and polystyrene. Previous work has shown that different ratio of hydrophobic to hydrophilic parts has a large impact on the geometry of micelle formation. By controlling the molecular weight of the block copolymer, the behavior of micelle stability will be investigated, which is very useful in protein encapsulation. It has been shown that the ionic strength and pH of the environment heavily influences coacervate formation and affects the encapsulation efficiency of proteins, thus, various mixtures of polycation/polyanion/protein mixtures were investigated. Trends from these environmental factors will then used to design delivery systems in the future. Key features of these mixtures that were varied were total monomer concentration, the geometry of micelle formation when using complex coacervate micelles, ratio of polycation to polyanion and initial concentration of protein. The stability and geometry of micelle will be determined by light scattering and transmission electron microscopy. Encapsulation efficiencies were determined using a Bradford assay and UV/Vis spectroscopy and stability data were acquired using circular dichroism. Polypeptide-based coacervates are a powerful platform technology to understand the role of the chemical environment on the stability of a protein formulation. Ultimately, we look to harvest these materials to enable the development of highly stable therapeutics and biologics such as refrigeration-free vaccines. This novel system is a major stepping-stone for the effective treatment of therapeutic regiments, providing not only a milieu that mimics the biological environment, but a means for stabilization of bioactive materials.