Structure and Size Control of Globular Protein Vesicles for Potential Biomedical Applications

Many diseases and illnesses curable with medication require accurate transportation and release of therapeutic proteins and drugs within the body. Globular protein vesicles (GPVs), made by self-assembly of recombinant proteins, are a potential platform that can be used for such biomedical applications or other target applications. GPVs are composed of two different proteins, a hydrophilic globular protein fused with acidic leucine zipper and a basic leucine zipper fused with thermally induced inverse phase transition biopolymer, called elastin-like polypeptide (ELP). These protein building blocks self-assemble to form a capsule-like vesicle membrane structure upon warming to potentially carry therapeutic cargo. During my REU program at the University of Florida (UF), I investigated the changes in GPV structure and stability while modifying various parameters that impact protein assemblies, such as salt concentration and incorporation of photo-crosslinkable nonstandard amino acid. We aimed at making GPVs smaller than 200 nm in diameter with a narrow polydispersity index not only for potential drug delivery applications, but also to enable membrane structure characterization using cryo transmission electron microscopy (cryo-TEM). As a result, we were able to achieve stable GPVs in physiological conditions by photocrosslinking of the fusion proteins with para-azidophenylalanine (pAzF) groups present in the ELP domains. We analyzed the size and morphology of GPVs by dynamic light scattering, epi-fluorescent microscopy, and cryo-TEM. This work would give basic inspiration and strategies to control GPV size and structure for target applications.