Protein Structure Dynamics In Gram Negative Bacterial Secretion Systems

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dc.contributor.advisorAudette, Gerald F.
dc.contributor.authorNicholas Joseph Bragagnolo
dc.date.accessioned2025-04-10T10:54:09Z
dc.date.available2025-04-10T10:54:09Z
dc.date.copyright2024-12-02
dc.date.issued2025-04-10
dc.date.updated2025-04-10T10:54:08Z
dc.degree.disciplineChemistry
dc.degree.levelDoctoral
dc.degree.namePhD - Doctor of Philosophy
dc.description.abstractBacteria use a variety of secretion systems that enhance their virulence. The conjugative type IV secretion system attributes transmission of mobile DNA elements in bacteria, whereas type IV pili are akin to the type II secretion system and are responsible for host adhesion and twitching motility. Understanding the structure and function of components in these systems has the potential to elucidate the mechanisms of bacterial commensalism and antagonism, and would aid in the development of inhibitors to bacterial pathogenesis by disrupting the tools virulent species use for stable, long-term infections. The entry exclusion protein of the F-like type IV secretion system, TraG, consists of a membrane-bound N-terminal domain and a periplasmic C-terminal domain, denoted TraG*. TraG* is essential in preventing redundant DNA transfer through interaction with a cognate TraS in the inner membrane of the recipient cell to prevent conjugation when the recipient cell carries the same plasmid. Using a multitude of biophysical methods including thermofluor, circular dichroism, collision induced unfolding mass spectrometry, and small angle X-ray scattering, the structural dynamics of TraG* was investigated. This allowed for the characterisation of the 50 N-terminal residues, the 77 C-terminal residues, and the TraS-interacting region of the protein as highly dynamic, providing evidence for the mechanism of the long-distance interactions made by TraG. This dissertation also describes the 1.3 Å crystal structure of the N-terminally truncated Type IV pilin of Pseudomonas aeruginosa from strain P1 (ΔP1), the first structure of its phylogenetically linked group (group Ia) to be reported. The structure was solved from X-ray diffraction data collected 20 years ago with a molecular replacement search model generated using AlphaFold. Comparisons of the ΔP1 structure to other Type IV pilins using ProSMART indicated that there are cases of higher structural homology between different phylogenetic groups of P. aeruginosa than there are between pilins from the same phylogenetic group, indicating that the classification of pilins based on structural homology may be necessary in developing anti-virulence drugs and vaccines.
dc.identifier.urihttps://hdl.handle.net/10315/42846
dc.languageen
dc.rightsAuthor owns copyright, except where explicitly noted. Please contact the author directly with licensing requests.
dc.subject.keywordsStructural biology
dc.subject.keywordsBacterial secretion systems
dc.subject.keywordsX-ray crystallography
dc.subject.keywordsSmall angle X-ray scattering
dc.subject.keywordsBacterial pathogens
dc.subject.keywordsCollision induced unfolding mass spectrometry
dc.subject.keywordsMulti-angle light scattering
dc.subject.keywordsCircular dichroism
dc.subject.keywordsAlphaFold
dc.subject.keywordsAntibiotic resistance
dc.subject.keywordsBionanotechnology
dc.subject.keywordsProtein structure determination
dc.titleProtein Structure Dynamics In Gram Negative Bacterial Secretion Systems
dc.typeElectronic Thesis or Dissertation

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