(181g) Protein Design Criteria for Intracellular Delivery Via Polyelectrolyte Complex Micelles

Protein-polyelectrolyte complexes have shown great potential in protein purification and protein delivery. Simple mixing of charged proteins with oppositely charged block copolymer, containing an ionic and water soluble neutral block, can result in the formation of protein-rich micelles. However, many proteins do not form stable complexes or do not phase separate with oppositely charged polyelectrolytes. To address this challenge, the influence of protein charge density and anisotropy on micelle formation with charged block copolymers was tested by engineering the surface charge of model proteins. The formation of micelles was evaluated as a function of protein charge and charge distribution. Additionally, micelle stability over time and to physiologically relevant pH's and ionic strengths was evaluated by dynamic light scattering and transmission electron microscopy. After identification of protein-polymer formulations that resulted in the formation of stable micelles, the ability of the micelles to be internalized in cells was determined by flow cytometry and confocal microscopy. We have found that there is a balance between increased protein charge and micelle stability and efficient protein delivery. The findings described here have informed the design of protein-rich nanoparticles capable of intracellular protein delivery using non-model proteins, demonstrating the general utility of protein-polyelectrolyte complexes for protein encapsulation and delivery.