Induced Mutagenesis in West Nile Virus Non-Structural Protein 3 With a Discussion of the Mutation's Effects on Viral Properties

Authors

Caden Connealy, University of Nebraska-LincolnFollow

First Advisor

Nicole Sexton

Second Advisor

Karin van Dijk

Date of this Version

Spring 3-27-2025

Document Type

Thesis

Citation

Connealy, C.P. 2025. Induced Mutagenesis in West Nile Virus Non-Structural Protein 3 With a Discussion of the Mutation's Effects on Viral Properties. Undergraduate Honors Thesis. University of Nebraska - Lincoln.

Comments

Copyright Caden Connealy 2025.

Abstract

West Nile Virus (WNV), a member of the Flaviviridae family and introduced to the Western hemisphere in 1999, presents a unique and emerging threat to human health. Infection in humans is often mild but can occasionally result in severe neurological impairment and even death. West Nile Virus non-structural protein 3 (NS3) plays an important role in the process of viral genome amplification. More specifically, the C-terminal domain of the protein folds into a helicase enzyme, which is responsible for the “unwinding” of double-stranded RNA intermediates and RNA secondary structure on the WNV genome. This is a vitally important step in the process of viral genome amplification. Certain regions within the NS3 domain are essential for helicase activity. Notably, the amino acid at position 249, when subject to mutation, produces viruses with phenotypes that correspond to irregular helicase activity. This thesis discusses the process by which I produced an infectious clone of West Nile Virus (WNV) with a site-saturation mutation at amino acid position 249. I will discuss the rationale for choosing this amino acid position with references to the literature and structural data. Furthermore, I will present preliminary data that reinforces the assumption that this mutant virus sample presents a variety of helicase phenotypes. The research presented here is meant to highlight the importance of the amino acid 249 in the regulation of NS3 helicase activity, which could be used to inform the development of anti-viral treatments.