Date of Award

Spring 2012

Degree Type

Thesis

Degree Name

Doctor of Philosophy (PhD)

Department

Biochemistry

Supervisor

Brian K Coombes

Language

English

Abstract

Bacteria employ virulence mechanisms to promote fitness that are generally detrimental to a host organism. The Gram-negative pathogen Salmonella enterica utilizes type three secretion systems (T3SS) to inject proteins termed effectors into the host cell cytoplasm where normal cellular function is modified. The coordinated T3SS assembly, and delivery of effectors to the cytoplasmic face of the T3SS is aided by virulence chaperones. The interaction of effector-chaperone complex with the T3SS occurs via an ATPase protein, where the complex is dissociated and the effector is unfolded, presumably for passage through the T3SS. The virulence chaperone network associated with the Salmonella pathogenicity island two (SPI-2) encoded T3SS has not been fully characterized. Additionally, the T3SS ATPase protein encoded within SPI-2, SsaN, has yet to be examined for functional motifs or a precise role in effector secretion. The contents of this thesis describe the characterization of two novel virulence chaperones, SrcA and SscA, and the T3SS ATPase SsaN. SrcA is a virulence chaperone for the effector substrates SseL and PipB2, and adopts the characteristic horseshoe-like structure common amongst effector chaperones. SscA is a chaperone for the translocon component SseC of the T3SS structure, and both proteins impact the regulation of SPI-2 promoters. The structure of SsaN resembles other T3SS ATPases, although different conformations exist between the structures, potentially highlighting regions with T3SS function. Additionally, an N-terminal domain was found to be dispensable for membrane localization, and residues within the predicted hexamer model impact effector secretion. These results identify novel virulence chaperones essential for T3SS function, and characterize the T3SS ATPase protein encoded within SPI-2. These findings greatly expand our knowledge of the virulence mechanisms utilized by S. enterica.

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