UC Davis

Sterols are part of an important class of molecules that modulate cellular signaling, recognition, and structure which make sterols an ideal probe to understand life processes. Dehydroepiandrosterone (DHEA) and sitosterol were used to synthesize novel chemical probes bearing an azide or alkyne as well as optimize reactions of known bioorthogonal sterols in Chapter 1. Routes to sterols with both bioorthogonal handles allows for a more comprehensive understanding of whether the type of or position of a bioorthogonal group on the sterol changes the metabolic outcomes when fed to an organism. The x-ray crystal structures of alkyne tagged DHEA and sitosterol analogs, as well as intermediates synthesized in effort to produce the bioorthogonal sterols, were collected and conformational analyses in Chapter 2 showed highly similar structures to that of endogenous sterols such as DHEA, cholesterol, and sitosterol. From the development of sterol probes arose the optimized synthesis of a fluorescent alkyne and invention of a novel fluorescent azide (Chapter 3). Routes to covalently attach the sterol and fluorophore by ‘click chemistry’ were developed for temperature-sensitive samples as well as for high atom economy with catalytic reagents. The culmination of this work was in the application of synthesized bioorthogonal sterols, fluorophores, and ‘click chemistry’ methods in Helicobacter pylori (Chapter 4). Target-based screening for the production of steryl glycoside metabolite analogs of bioorthogonal sterols by UPLC-fluorescence and characterization by UPLC-MS/MS was accomplished, showing the potential for this platform to be used in other living systems.