Intraprotein electron transfer processes are crucial to the maintenance of cellular pathways that enable life as we know. Well-studied and well-characterized proteins such as cytochrome c and azurin have long been used to explore these processes. The pathways present in these proteins and others, while varying in length can include covalent bonds, hydrogen bonds, and through-space jumps. Of interest are interactions in yeast cytochrome c that are similar to hydrogen bonds. We aimed to explore a pathway where the hydroxyl of Tyr67 interacts with the Met80 sulfur. To probe the importance of the interaction in the context of electron transfer, Tyr67 was replaced with different fluorotyrosines of varying pKas. We then evaluated a second pathway containing two hydrogen bonds of which were removed by mutating to different amino acids. The elimination of these hydrogen bonds did not influence the rate of intramolecular electron transfer so an alternative pathway was examined. From this pathway, a substitution to Met64 to increase the length of a through-space jump decreased the electron transfer rate by a factor of two suggesting the initial pathway is non-operative. Lastly, the protein, azurin, was used as a model to investigate the properties of unnatural fluorotyrosines. The Trp48 amino acid was replaced with various fluorotyrosines in order to develop a system where the unnatural amino acid properties can be probed in relation to electron transfer processes.
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Thesis advisor: J., Warren, Jeffrey
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