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.
