Synthetically Diverse Triazabutadienes and Their Applications Towards Biological Systems
AuthorShepard, Abigail J.
Keywordsaryl diazonium ion
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PublisherThe University of Arizona.
RightsCopyright © is held by the author. Digital access to this material is made possible by the University Libraries, University of Arizona. Further transmission, reproduction, presentation (such as public display or performance) of protected items is prohibited except with permission of the author.
AbstractSmall molecule tools for interrogating biological systems are important for studying the inner workings of the cell, from protein-protein interactions to understanding protein fluctuations and dynamics in different disease states. Aryl diazonium ions have been found to be a useful tool for studying proteins, capable of labeling tyrosine residues on a proteins surface. To prevent this electrophile from labeling everything in sight affording nonspecific reactions, a protecting group can be invoked. Once such example is the triazabutadiene, a nitrogen rich structural motif capable of falling apart upon protonation or in the presence of light. Previous studies have focused on characterizing deprotection kinetics at a physiological pH, and aryl diazonium ion adducts have been characterized on single proteins using a fluorophore conjugate. To further the understanding of the triazabutadiene system for both synthetic and biological applications, more complex probes must be designed, synthesized, and characterized to move towards in cellulo experiments. A triazabutadiene was designed bearing an alkyne handle on the sacrificial imidazole ring, which can be functionalized using copper-click chemistry to add on cell targeting moieties such as vitamins for active transport or small molecule signals for a specific organelle of interest. The development of such probes adds layers of synthetic complexity to the system as an acid and base sensitive functional group have been installed. Difficulties associated with methyl ester deprotection led to the explorative synthesis of less stable alkyl and silyl functional groups. While informative on general reactivity, all efforts were futile, either due to azide instability, triazabutadiene instability, or unsuccessful ester deprotection. Synthetic efforts for triazabutadiene functionalization have traditionally stayed away from the use of heat, due to the possibility of thermal intramolecular rearrangements, and metal catalysts, due to the chelating potential of the nitrogen rich scaffold. Previous findings have shown the triazabutadiene scaffold is compatible with copper-click chemistry conditions. These findings inspired the work of optimizing a Suzuki-coupling method for facile attachment of aryl rings to the triazabutadiene scaffold. Although there is a lack of support from literature, it was determined the scaffold is compatible with palladium-ferrocene complexes and can withstand temperatures of 95 °C for up to one hour. The biphenyl triazabutadienes, a novel synthetic class of compounds, are capable of deprotecting at physiological pH and labeling proteins, as determined using native mass spectrometry. An interesting nitrobenzoxadiazole triazabutadiene probe was discovered which displays a turn-on fluorescence profile upon protein labeling, with the intact triazabutadiene being non-fluorogenic. The aryl diazonium ion released upon deprotection was synthesized by an alternate route and tested on proteins to confirm these findings. A triazabutadiene containing a mitochondria targeting small molecule, a triphenylphosphonium ion, and a bromobenzene ring for mass spectrometry identification was synthesized and evaluated on proteins. The bromobenzene azo-adduct was found to label proteins, as confirmed using native mass spectrometry, and the adduct also remained intact during tandem mass spectrometry/mass spectrometry allowing for easy identification of labeled protein residues. The triazabutadiene was found to fall apart at pH 7 with the help of ultraviolet light, giving support for moving this system into a cellular setting for protein identification. In conclusion, this dissertation highlights the synthetic explorations of functionalizing triazabutadienes using novel synthetic plans, and the development of more complex probes with unexpected characteristics which will certainly find a use in the field of biological chemistry.
Degree ProgramGraduate College