Comparative evaluation of single-site mutations in the West Nile virus envelope protein using a mouse model

Abstract

For many years, West Nile virus (WNV) has been a leading cause of viral encephalitis across the globe. However, there are no human vaccines licensed for use and only four veterinary vaccines available. This thesis aims to investigate the similarities across WNV and other flaviviruses with the goal of developing a common attenuation mechanism. This mechanism could aid in the development of a vaccine candidate that could be used for WNV as well as other viruses in the Flaviviridae family. Viruses in the Flavivirus genus share significant homology among genomes, including around 40% homology in the envelope (E) protein. The E protein of WNV functions as a class II fusion protein, facilitating viral entry and acting as the main target of neutralizing antibodies. This makes this protein an ideal target for vaccine development technology. The E protein contains 500 amino acid residues and consists of three domains: EDI, EDII, and EDIII. Between two of these domains, EDI and EDII, lies the hinge region. This region appears to be necessary for the pH-dependent conformational changes necessary for endosomal fusion. In this study, eight residues located in the hinge region were investigated: E-A54, E-I130, E-I135, E-I196, E-Y201, E-A269, E-V272, and E-L281. These highly conserved hydrophobic residues were selected to analyze single-site mutations for the development of a flavivirus common attenuation mechanism for candidate live-attenuated vaccine creation. In this thesis, it was determined that one selected single-site mutation, E-Y201P, results in a fully attenuated, immunogenic phenotype, while the mutation E-A54S results in a partially attenuated, immunogenic phenotype. With both mutation sites being highly conserved among flaviviruses, this study lays the foundation for further studies to examine alternate mutations, combinations of these mutations or other attenuating mutations, and/or candidate live-attenuated vaccine development using this attenuation mechanism.