Publication Date


Date of Final Oral Examination (Defense)


Type of Culminating Activity


Degree Title

Doctor of Philosophy in Biomolecular Sciences



Major Advisor

Juliette K. Tinker, Ph.D.


Allan R. Albig, Ph.D.


Richard S. Beard Jr., Ph.D.


Daniel Fologea, Ph.D.


Bacterial mono-ADP-ribosyltransferases (ARTs) catalyze the singular transfer of an ADP-ribose moiety from an NAD+ molecule onto a target molecule. ARTs contain an ancient and highly conserved tertiary structure and have a wide variety of intracellular targets and effects. Some, but not all, bacterial ARTs have an AB5-type multimeric structure consisting of an enzymatically active subunit non-covalently situated atop of a non-toxic pentamer. The active, or A, subunit of AB5-type toxins has a catalytic action that contributes to bacterial pathogenicity, and it is sometimes, but not always, an ART. ArtAB is an ART with AB5-type structure from the virulent and highly antibiotic resistant Salmonella Typhimurium DT104. In the studies described here, we tested the hypothesis that the active subunit of ArtAB is structurally and enzymatically homologous to that of the well-characterized AB5-type ART pertussis toxin. ArtAB was purified from E. coli and was used to characterize ArtAB’s cellular effects, predicted structure, and biophysical properties. In addition, a set of single-residue mutants was constructed and purified to probe ArtAB’s active site. AB5-type toxins have long been studied for their immunogenic properties, and some of these bacterial munitions have been harnessed and repurposed as vaccines or vaccine adjuvants to prevent infectious disease. Their receptor-binding pentamer, abbreviated as B5, binds to, and facilitates entry into, host cells. In additional work presented here, we tested the hypothesis that the B5 subunit of cholera toxin (CTB) from Vibrio cholerae could be used to construct a safe and effective mucosal vaccine against Staphylococcus aureus-caused mastitis. We constructed a bovine vaccine by conjugating Staphylococcus aureus antigens to the CTB-based adjuvant platform, and the immunogenicity of the vaccine was characterized in a bovine clinical trial. Finally, clinical isolates of caprine S. aureus were screened for the presence of surface antigens that could be use in a caprine version of the vaccine against mastitis. The work on bacterial AB5-type ARTs presented here contributes to a growing global understanding of the bacterial ART family, lays a foundation for the potential incorporation of ArtAB in a vaccine against Salmonella, and advances the development of bovine and caprine vaccines against S. aureus-caused mastitis.