Structure and Assembly of a Thermophilic Bacteriophage

Abstract

Bacteriophages (phages) are ubiquitously abundant bacterial viruses and the most numerous biological entities on Earth. In order to fully understand the phage life cycle, we must understand how phage particles self-assemble. Further, an understanding of phage structure and assembly permits the application of phages to human health. Phage assembly is largely governed by two parallel pathways: capsid assembly and tail assembly. The capsid houses the genome, while the tail is the channel through which the genome travels to infect its host, making both components essential for successful infections. In this dissertation, I investigate the assembly pathways of the hyperthermophilic phage P74-26 as a model for understanding long-tailed phages (~85% of all phages). Primarily, I combine cryoEM structures of the P74-26 tail tube with an in vitro system for studying assembly kinetics to propose the first molecular model for the assembly of long-tailed phage tail tubes. Additionally, the tail is attached to a Tail Tip Complex (TTC) which recognizes the surface of the host. I present a cryo-EM structure of the P74-26 TTC, identifying the protein components for the first time. Finally, I explore the self-assembly of the capsid protein with the potential for establishing an in vitro system for studying capsid assembly and present proof-of-concept studies for using the particles as functional nanoparticle therapeutics. Together, this work explores principles of phage assembly and thermostability in tailed bacteriophages and how we can take advantage of these principles for future development of therapeutic delivery tools and nanoparticle applications.