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Obtaining Structural Insights on Bacterial Protein Complexes Using Time-Resolved Hydrogen-Deuterium Exchange Mass Spectrometry

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dc.contributor.advisorAudette, Gerald F.
dc.creatorLento, Cristina
dc.date.accessioned2015-12-16T19:30:30Z
dc.date.available2015-12-16T19:30:30Z
dc.date.copyright2015-08-25
dc.date.issued2015-12-16
dc.date.updated2015-12-16T19:30:30Z
dc.degree.disciplineChemistry
dc.degree.levelMaster's
dc.degree.nameMSc - Master of Science
dc.description.abstractPersistent infections by Pseudomonas aeruginosa are initiated by interaction of a type IV pilus (T4P) with receptors on the mucosal cells of susceptible hosts. Here, we examine the structural changes occurring between the monomeric and dimeric states of ΔK122 using time-resolved electrospray ionization hydrogen-deuterium exchange mass spectrometry (TRESI-HDX-MS). Based on levels of deuterium uptake, the N-terminal α-helix and the loop connecting the second and third strands of the anti-parallel β-sheet contribute significantly to pilin dimerization. Conversely, the antiparallel β-sheet and αβ loop region exhibit increased flexibility, while the receptor binding domain retains a rigid conformation in the equilibrium state. Additionally, Escherichia coli are able to adapt to changing environmental conditions and develop antibiotic resistance through a process called F-plasmid conjugation, carried out through a type IV secretion system (T4SS). The F-T4SS protein TraF is of particular interest due to its involvement in pilus assembly to mediate the transfer of DNA. Dynamic analysis of a GST-TraF construct through TRESI-HDX-MS was performed to gain further insights on its structure. These studies have revealed that the C-terminal region predicted to contain the thioredoxin-like domain is quite structured compared to the more solvent accessible N-terminal region predicted to form a protein-protein interaction with companion T4SS protein TraH. Structural analysis of a GST-TraF construct is on-going to further characterize the regions responsible for protein-protein interaction and the elucidation of its three-dimensional structure.
dc.identifier.urihttp://hdl.handle.net/10315/30744
dc.language.isoen
dc.rightsAuthor owns copyright, except where explicitly noted. Please contact the author directly with licensing requests.
dc.subjectChemistry
dc.subjectBiochemistry
dc.subjectBiology
dc.subject.keywordschemistry
dc.subject.keywordsmass spectrometry
dc.subject.keywordsbiochemistry
dc.subject.keywordsprotein dynamics
dc.subject.keywordstype IV pilins
dc.subject.keywordsprotein nanotubes
dc.subject.keywordsbacterial conjugation
dc.subject.keywordstime-resolved electrospray ionization
dc.subject.keywordshydrogen-deuterium exchange
dc.titleObtaining Structural Insights on Bacterial Protein Complexes Using Time-Resolved Hydrogen-Deuterium Exchange Mass Spectrometry
dc.typeElectronic Thesis or Dissertation

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