Protein vesicles incorporating functional, globular proteins have enormous potential in a number of bio-applications such as drug delivery systems, micro bioreactors, and mimicking artificial cells. We have designed globule-zipper-polypeptide protein complexes (mCherry-ZE/ZR-ELP), consisting of a model fluorescent globular protein (mCherry), high affinity leucine zipper pair (ZE/ZR), and thermo-responsive elastin-like polypeptide (ELP). These complexes self-assemble into vesicles under warming. In order to propose rational guidelines to engineer the protein vesicles, we have traced the phase transition of the recombinant fusion proteins as a function of concentration and temperature using characterization methods including dynamic light scattering, transmission electron microscopy and small angle neutron scattering. The fusion protein complexes at a constant concentration undergo the phase transition from soluble monomers, through dynamic coacervates, to stable hollow vesicles as operating temperature (To) increases over transition temperature (Tt). Since the Tt is decreased with increasing protein concentration, diverse self-assembled structures from spherical particles to vesicles form with the increased average sizes as protein concentration increases at the same To. Also, the protein complexes can self-assemble into either single-layered or bi-layered vesicles according to the hydrophobic conformation of ELP domain. The phase diagram of mCherry-ZE/ZR-ELP complexes established in this study reveals the effect of difference in the To and Tt on the self-assembled protein vesicle size and structures, which provide a simple design rule to engineer the functional, globular domain-containing protein vesicles with desired size and structure for target applications. Functional demonstration of the design rule was applied to ScFvF5-ZE/ZR-ELP to create nanoscale vesicles that target HER2+ breast cancer cells.

Self-assembled protein vesicles made from recombinant fusion proteins

253rd National Meeting (2017)

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