Between Dec 19, 2024 and Jan 2, 2025, datasets can be submitted to DRUM but will not be processed until after the break. Staff will not be available to answer email during this period, and will not be able to provide DOIs until after Jan 2. If you are in need of a DOI during this period, consider Dryad or OpenICPSR. Submission responses to the UDC may also be delayed during this time.

Browsing by Author "Dunleavy, Katie"

Now showing 1 - 2 of 2
  • Thumbnail Image
    Item
    Hydration Environment Characterizations of the Folding of IA3, an Intrinsically Disordered Protein (2020-10-02)
    (2020) Dunleavy, Katie; University of Minnesota Duluth. Department of Chemistry and Biochemistry
    The intrinsically disordered protein (IDP), IA3, found in Saccharomyces cerevisiae has been previously shown to adopt ?-helical secondary structure through the N-terminus when bound to yeast proteinase A (YPRA). The helical structure of IA3 can be stabilized in the absence of YPRA, by using the secondary structure stabilizer, 2,2,2 – trifluoroethanol (TFE). To characterize the TFE induced disordered to ordered states of this IDP, site directed spin labeling (SDSL) and electron paramagnetic resonance spectroscopy (EPR) were used. Cysteine scanning throughout IA3 results in varied degrees of helicity dictated by labeling position, most sensitive within the N-termini. These SDSL – EPR studies, combined with structural characterization using circular dichroism spectroscopy have identified labeled variants that weaken, mimic, or strengthen the TFE induced ordered states of IA3, or overall helical propensity. IA3 labeled variants with differing degrees of helical transitions have further been utilized to assess local hydration dynamics surrounding the disordered and ordered state of this IDP in the absence and presence of TFE, respectively. Overhauser dynamic nuclear polarization (ODNP), a combined EPR and nuclear magnetic resonance spectroscopy technique was used to probe hydration dynamics (i.e. water diffusivity) at the surface of these IA3 variants. Results from ODNP studies suggest significant fluctuations in hydration behavior dependent on both the presence of TFE and the varying degrees of structure found within the N- and Ctermini of IA3.
  • Thumbnail Image
    Item
    Induced changes in protein secondary structure due to internal sequence modifications and ligand binding
    (2015-09) Dunleavy, Katie
    Secondary structure of protein sequences is dependent on both internal and external interactions of amino acid residues and ligand-binding partners. Internal residue features can leave a protein in an ordered, folded state, or in a disordered, unfolded state. Structural characteristics can be further influenced by protein-ligand binding interactions with a lipid membrane surface. Structural features can be altered upon membrane binding, causing disordered proteins to become more ordered in structure. The stabilizing influence of methionine (Met) oxidation in an aromatic-Met hydrogen bonding interaction, within a small, 15-residue peptide was studied using Differential Scanning Calorimetry (DSC) and Circular Dichroism (CD) Spectroscopy to observe changes in structural strength. By observing the ordered to disordered transition of this peptide, changes in enthalpy and transition temperature were determined. This added aromatic-oxidized Met interaction causes a stronger and more stable ordered peptide structure due to internal residue interaction. Multiple intrinsically disordered proteins were studied upon binding to a membrane surface to determine the influence that physiological membrane surfaces and curvature have on appropriate conformer formation of synaptic vesicle (SV) binding proteins. Various C2 domains of synaptotagmin I (Syt I) and α-Synuclein (αS) were studied using DSC, CD and Carboxyfluorescein (CF) release assays. The proper folding of these proteins is important for their necessary function, and misfolding or sequence mutation can significantly alter their functionality within neuronal environments. These studies are vital to enhance understanding of the dependence of internal residue and membrane binding interactions on structural properties of proteins, as these specific interactions are not limited to individual systems.