Protein-induced metamorphosis of unilamellar lipid vesicles to multilamellar hybrid vesicles

Protein encapsulation into nanocarriers has been extensively studied to improve the efficacy and stability of therapeutic proteins. However, the chemical modification of proteins or new synthetic carrier materials are essential to achieve a high encapsulation efficiency and structural stability of proteins, which hinders their clinical applications. New strategies to physically incorporate proteins into nanocarriers feasible for clinical uses are required to overcome the current limitation. Here we report the spontaneous protein-induced reorganization of ‘pre-formed’ unilamellar lipid vesicles to efficiently incorporate proteins within multilamellar protein-lipid hybrid vesicles without chemical modification. Epidermal growth factor (EGF) binds to the surface of cationic unilamellar lipid vesicles and induces layer-by-layer self-assembly of the vesicles. The protein is spontaneously entrapped in the interstitial layers of a multilamellar structure with extremely high loading efficiency, ~99%, through polyionic interactions as predicted by molecular dynamics simulation. The loaded protein exhibits much higher structural, chemical, and biological stability compared to free protein. The method is also successfully applied to several other proteins. This work provides a promising method for the highly efficient encapsulation of therapeutic proteins into multilamellar lipid vesicles without the use of specialized instruments, high energy, coupling agents, or organic solvents. Multilamellar Protein-Lipid Vesicles (MPLVs) fabricated in a mild environment with low pressure and no organic solvent at room temperature were devised for linker-free and highly-efficient protein encapsulation. MPLVs have a high loading efficiency and long-term dispersion stability. The morphological changes, such as folding and engulfing steps, of unilamellar lipid vesicles by protein-induced lipid molecular asymmetry evolve into a multilamellar structure. [Display omitted] •Proteins can induce the folding and engulfing of unilamellar lipid vesicles.•Proteins can be efficiently intercalated into multilamellar lipid structures.•Multilamellar protein-lipid vesicles (MPLVs) are prepared by simple mixing.•The protein loading yield and efficiency are ~33% and ~99%, respectively.•MPLVs exhibit long-term structural, chemical, and biological integrities of proteins.