UC Berkeley

In this dissertation, I describe the development of bioorthogonal chemical tools toward the study of bacterial pathogens during infection. Bioorthogonal chemistry has enabled the visualization of biomolecules not amenable to fluorescent protein fusion. By feeding an organism an unnatural metabolite bearing a bioorthogonal chemical reporter, the reporter is incorporated into the cellular biopolymer of interest and can be detected by bioorthogonal reaction with an imaging agent. To adapt this approach towards visualizing pathogenic bacteria in an infection setting, my dissertation research focused on two main areas. First, I developed a family of fluorogenic azide probes activated by bioorthogonal click reactions with alkynes. These probes offer significant advantages in imaging studies where it is challenging to wash away unreacted probe, such as within live cells or in live organisms, the settings in which bacterial pathogens reside. Secondly, I developed a family of unnatural D-alanine analogues bearing cyclooctynes, which are incorporated into nascent peptidoglycan in a wide variety of bacterial species and undergo bioorthogonal reactions with azides. In conjunction with the fluorogenic azide probes I developed, these analogues enabled the visualization of cell wall synthesis in an intracellular bacterial pathogen during infection.