Surface-Functionalization of PP7 Virus-Like Particles with Spytag for Bioconjugation Applications.

Abstract

Virus-like particles (VLPs) are protein-based nanoscale assemblies derived from structural proteins of viruses. They are non-infectious scaffolds and are highly immunogenic; thus, they widely used as scaffolds to display and enhance the immunogenicity of less immunogenic foreign antigens. Different approaches, including genetic insertions, can be used to display foreign antigens on VLPs; however, some of these approaches have limitations, i.e., inability of viral coat proteins to tolerate a foreign insertion. In this study, we assessed the tolerability of bacteriophage Qβ and PP7 coat proteins to insertions of Spytag003 or Spytag peptides; we assessed whether the insertions of these peptides to the coat proteins will affect the ability of the recombinant coat proteins to assemble into VLPs; chimeric VLPs displaying these peptides can be used to conjugate antigens for vaccine studies. While the insertion of Spytag003 peptide at either the N or C termini of the coat protein of Qβ and at the C terminus of the coat protein of PP7 did not affect the expression of the recombinant proteins, the expressed proteins were not soluble. In contrast, insertion of the shorter Spytag at either the N terminus or AB-loop of PP7 gave rise to soluble proteins that assembled into VLPs. PP7-Spytag VLPs were successfully conjugated with SpyCatcher003 and a fungal antigen. Immunization studies revealed that Spycatcher003 protein conjugated on PP7-SpyTag VLPs elicited significantly higher anti-Spycatcher003 antibody titers compared to unconjugated Spycatcher003 (p = 0.0286). Together, these findings establish proof of concept that PP7-Spytag VLPs should be explored as a platform to conjugate foreign proteins.