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ORCID

http://orcid.org/0000-0003-3216-1792

Date of Award

Spring 5-15-2020

Author's School

Graduate School of Arts and Sciences

Author's Department

Biology & Biomedical Sciences (Computational & Molecular Biophysics)

Degree Name

Doctor of Philosophy (PhD)

Degree Type

Dissertation

Abstract

Antibiotic-resistant bacterial infections are a serious and immediate threat to global public health. In the United States alone, over 2 million individuals develop antibiotic-resistant infections annually, resulting in 23,000 deaths and $20 billion in excess health care costs. Virulence factors that allow bacteria to invade and persist within the host are promising targets for novel antimicrobial agents that could be used to curb the spread of antibiotic resistance. Development of therapeutics that can selectively eliminate pathogenic bacteria while sparing the beneficial host microbiota requires a detailed molecular understanding of critical virulence factors that facilitate interactions between pathogens and their environments. Two such factors are Chaperone-Usher Pathway (CUP) pili, which mediate bacterial adhesion, and curli, which constitute the major extracellular matrix component of many biofilms. In this dissertation, I have elucidated the structure of two pilus adhesins thought to mediate colonization of the gastrointestinal tract and determined the mechanism of CsgE assembly factor function in curli biogenesis. CUP pili comprise hundreds of repeating structural subunits capped by an adhesin whose N-terminal lectin domain binds host receptors with stereochemical specificity, thus facilitating the colonization of specific host niches by Gram-negative bacteria. Using X-ray crystallography, I determined that the UclD adhesin on the F17-like pilus contains a transverse binding groove that binds O-glycans in the colonic crypts. Conversely, I found that the YehD adhesin contains a unique helix-loop-helix motif that can pivot open to reveal a deep hydrophobic pocket. Additional structural and genetic studies of the Yeh pilus rod reveal a tightly-wound right-handed helix that has maintained its biomechanical response to external force through purifying evolutionary selection. I have also utilized X-ray crystallography to elucidate the structure-activity relationship of high-affinity compounds that inhibit the binding of two additional pilus adhesins, FimH and FmlH, to the urinary tract during acute and chronic urinary tract infection. Finally, I have performed genetic and biochemical studies to elucidate the role of CsgE in bacterial biofilm formation. During curli biogenesis, I found that CsgE uses a series of specific and nonspecific electrostatic interactions to bind the major amyloid subunit CsgA and deliver it to the CsgG outer membrane pore, thus preventing premature aggregation and cellular toxicity. In aggregate, these findings have advanced our molecular understanding of bacterial adhesion during pathogenesis and illuminated novel pathways for the treatment of antibiotic-resistant infections.

Language

English (en)

Chair and Committee

Scott J. Hultgren

Committee Members

Michael Caparon, Matthew Chapman, Daved Fremont, Carl Frieden,

Available for download on Wednesday, May 15, 2120

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