Protein corona formation and RNA packaging in barley stripe mosaic virus-like particles

• Christina Rodriguez, Applied Molecular Biology & Biotechnology, University of Delaware

• Kevin Solomon, Department of Chemical and Biomolecular Engineering, University of Delaware

The efficacy of nanoparticle-based vaccines primarily depends on the targeted delivery of antigens to immune cells and stimulating immune responses. Precise control over surface composition and cargo encapsulation is essential for adjuvanticity and antigen delivery.  Rod-shaped plant viruses, such as barley stripe mosaic viruses (BSMV), are hierarchically assembled protein nanorods that allow control over these properties.  Each nanorod consists of several hundred to thousands of copies of the capsid protein that self-assemble on an RNA template, enabling high-density surface display and RNA encapsidation. We leverage a bacterial platform to produce non-infectious BSMV VLPs, in which the viral genome is replaced by a user-defined RNA template for additional control over surface properties. This flexible engineering platform allows the insertion of amino acids at the solvent-exposed C-terminus. Light scattering and electron microscopy confirms the assembly of mutated coat proteins into rods. Through the introduction of specific residues, we achieve control over the surface charge of BSMV VLPs, as confirmed by zeta potential measurements. Furthermore, we confirm the presence of a protein corona formed by the adsorption of serum proteins on the surface of BSMV VLPs via gel electrophoresis. We also identify an RNA packaging signal necessary for the self-assembly of coat proteins into rods through plasmid cloning and electron microscopy.  Future work will include characterizing the effect of surface properties on protein corona composition and utilizing the identified RNA packaging signal for in vitro assembly of nanorods.  These advances will set the stage for the development of BSMV VLPs as a platform for vaccine delivery.