Type IV pili (T4P) are long, fibrous surface appendages involved in attachment, motility, biofilm formation and DNA uptake that are expressed by bacteria and archaea. They are an important virulence factor for a number of bacteria, including Pseudomonas aeruginosa, an opportunistic pathogen that is a common cause of nosocomial infections. T4P are composed mainly of monomers of the major pilin subunit, PilA, although several low abundance proteins called minor pilins are also present. These surface-exposed proteins are potential vaccine candidates, although a more complete understanding of their diversity and function is required for the rational development of a pilus-based vaccine. There are five distinct groups of P. aeruginosa major pilins, which vary based on their sequence and their associated accessory proteins, and two distinct sets of minor pilins, although the roles of the latter in pilus biology are poorly understood. This study focuses on the structural characterization of major and minor pilins and functional implications for pilus assembly and disassembly dynamics. The structural analysis of major pilins from groups III and V revealed specific differences in pilin structure that may affect subunit interactions within the pilus fibre and interactions with their specific accessory proteins and minor pilins. The minor pilins PilVWX were shown to form a putative subcomplex with the adhesin and anti-retraction protein PilY1, which is proposed to prime pilus assembly and thus traffic PilY1 to the bacterial surface. High resolution X-ray crystal structures of the minor pilins FimU and PilE were solved and functional characterization suggested that FimU and PilE are necessary for efficient pilus assembly to stably connect the priming subcomplex to the major pilin subunits. Together, this work has increased our understanding of pilin diversity and defined a concrete role for the minor pilins in pilus assembly. / Thesis / Doctor of Philosophy (PhD) / Pseudomonas aeruginosa is a bacterium that can take advantage of a weakened immune system to cause lethal infections. The first step of infection involves attachment to the host using long sticky fibres called type IV pili. Each fibre is composed primarily of a single protein, the major pilin, but also contains low abundance proteins called minor pilins. Without these proteins, the bacteria can’t attach and cause infections, making pilins excellent vaccine candidates. This study focused on the characterization of major and minor pilins to understand the diversity of these proteins and how these differences might affect pilus assembly. We show that the molecular structure of the major pilin differs between strains although the core architecture is the same, and that the minor pilins are required for initiation of pilus assembly. This work furthers our understanding of the structures and functions of pilin proteins, and provides information helpful for the development of vaccines.
| Identifer | oai:union.ndltd.org:mcmaster.ca/oai:macsphere.mcmaster.ca:11375/17277 |
| Date | 08 May 2015 |
| Creators | Nguyen, Ylan |
| Contributors | Burrows, Lori, Biochemistry and Biomedical Sciences |
| Source Sets | McMaster University |
| Language | English |
| Detected Language | English |
| Type | Thesis |
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