ENZYME-FREE UBIQUITIN LIGATION. FROM NATIVE CHEMICAL LIGATION AND SPIN LABELING TO DIMERIZATION BY INTER-UBIQUITIN MIMICKING LINKAGE AND PH-DEPENDENT CONFORMATIONAL SWITCH

The delicate balance between the production and disposal of proteins is vital for the changes required in the cell to respond to given stimulus. Ubiquitination is a protein modification with a range of signaling outcomes when ubiquitin is attached to a protein through a highly ordered enzymatic cascade process. Understanding ubiquitination is a growing field and nowadays the application of chemical reactions allows the isolation of quantitative materials for structural studies. Therefore, in this dissertation it is described some of these suitable chemical methodologies to produce an isopeptide bond toward the polymerization of ubiquitin bypassing the enzymatic control with the purpose of showing if these chemical modifications have a direct impact on the structure of ubiquitin. First, the possibility of incorporating non-natural lysine analogs known as mercaptolysines into the polypeptide chain of Ubiquitin was explored when they were attached to ubiquitin by native chemical ligation at its C terminus. The sulfhydryl group was used for the attachment of a paramagnetic label to map the surface of ubiquitin. Second, the condensation catalyzed by silver nitrate was used for the dimer assembly. In particular, the main focus was on examining whether orthogonal protection and deprotection of each monomer have an impact on the reaction yield, since the synthetic strategy has been previously attempted successfully. Third, the formation of ubiquitin dimers was approached by building an inter-ubiquitin linkage mimicking the isopeptide bond with two approaches, the classic disulfide exchange as well as the thiol-ene click reaction by thermal initiation in aqueous conditions. After assembling the dimeric units, they were studied by Nuclear Magnetic Resonance, in order to establish a conformational state profile which depends on the pH conditions. The latter is a very important concept since some ligands have a preferred affinity when the protein-protein hydrophobic patches are in close proximity.

(inter)FACE: A Study of Black Families Advocating for their Children’s Education

Black students are consistently overrepresented in categories of academic underachievement. Parent engagement has long been touted as an effective strategy for improving the educational outcomes of Black children. However, most parent engagement research reflects deficit based perspectives frame Black parents as problems that must be fixed or mitigated before they can positively contribute to their children’s education. Consequently, parent engagement research and frameworks ignore the perspectives of Black parents and the assets they use to participate effectively in parent engagement. In this case study, I draw on individual and focus group interview data, documents, and observations, to examine how fifteen Black families, collectively known as FACE: 1) define and participate in parental engagement, 2) experience barriers to and opportunities for engagement, and 3) experience benefits of engagement for their children and their own personal development. Guided by Black Feminist and Critical Race Theories, I show how Black families in this study used a myriad of engagement strategies to improve their children’s educational experiences which were invisible to schools and how they used school-sanctioned engagement activities to meet their own objectives. Ultimately, I argue that school-centered parent engagement frameworks and models are ineffective for empowering Black families and accounting for the essential ways that these families contribute to the well-being of their children. Based on my findings, I discuss implications for theory, practice and policy, and research, and make recommendations for a more family-centered approach to parent engagement.