Rodgers, Jeffrey Michael

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  • Publication
    Development, Characterization, And Application Of Biological Spectroscopic Probes
    (2018-01-01) Rodgers, Jeffrey Michael
    Protein and nucleic acid molecules exhibit a range of motions that occur on various time scales and distances including large conformational changes involved in the formation of folded structures and local rearrangements where intramolecular and/or intermolecular interactions confer biological function. Because of their unique advantages, spectroscopic techniques based on measurement of molecular vibrations and fluorescence emissions have played a key role in studying the structure-dynamics-function relationship of biological molecules. However, intrinsic spectroscopic signals of this kind often lack the desired localization to interrogate a particular region or interaction of interest. Recently, small spectroscopic probe molecules which are sensitive to their local environment have become increasingly popular to investigate biological processes in a site-specific manner. Furthermore, relatively minor alterations to such molecules—including isotopic substitution and the position or identity of functional groups—can have a profound effect on their spectroscopic properties and, by extension, their usefulness. In this thesis, we investigate a number of biologically-relevant infrared probes and their variants in order to further expand their utility; we also implement fluorescence spectroscopic techniques to investigate a putative inhibitor of amyloid aggregation. First, using a set of four isotopomers of the unnatural amino acid p-cyano-phenylalanine, we show that a small change in the reduced mass of the nitrile oscillator has significant effects on both the infrared absorption frequency and the vibrational relaxation process. This characterization improves their value as very similar yet non-degenerate vibrational labels which can be introduced at multiple locations simultaneously while maintaining site-specificity and allowing for energy transfer or coupling experiments. Second, we harness the phenomenon of Fermi resonance in the infrared probe molecules 4-cyanoindole and cyclopentanone as a convenient indicator of hydrogen-bonding status of the nitrile and carbonyl functional groups. Furthermore, we investigate the dependence of the NO stretching vibrations of N-methyl-5-nitroindole on solvent polarity; this model molecule exhibits potential as an infrared probe of hydration in nucleic acids when implemented as an unnatural but universal base. Finally, we apply several spectroscopic techniques including fluorescence correlation spectroscopy to study a common neurological amino acid and its ability to prevent and disrupt the aggregation of amyloid-beta.