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.

Glutamine supplementation and renal health

Gemstone Team Juiced

Between Nation and State: Albanian Associations from Ottoman Origins to a Communist Party, 1880 - 1945

This dissertation addresses the broader antecedents of the Communist Party of Albania (CPA) as one of a number of associations whose experience was central to Albanian political history. This long experience dates back to the informal national associations formed in the Ottoman Empire of the late nineteenth century. The dissertation examines the role of these associations which, pursuing language rights and political representation through imperial state reforms, set a pattern that struggled to connect nation and state, rather than asserting the territorial demands for a nation-state familiar across the region. Starting out in the Ottoman Empire, but then maturing in the Albanian diaspora in Romania, Bulgaria, Egypt and the United States, this dissertation shows politically significant processes of longer-term adaptation that created informal associations as institutional structures able to channel collective action. It then traces the reframing of these patterns through their destruction in the Balkan Wars and the First World War to the emergence of communist associations in the interwar period and beyond. This dissertation is a sustained study that traces long-term Ottoman imperial political legacies in the Albanian successor state. The story of the associations, based on hitherto unexamined archival documents, shows that the Albanians possessed a far greater capacity for political mobilization that previously acknowledged by historians. Moreover, the dissertation successfully challenges the conventional wisdom that portrays the Albanians as irreparably divided along sectarian and regional faultlines. It finds that Albanian national activism was civic in character rather than ethnic as elsewhere in the Balkans. The Albanians fought to remain within a multinational framework because this afforded them political security, social advancement and potential economic growth. In the late Ottoman period, this political objective was manifested in the acceptance of the supranational imperial order whereas during the Second World War, in the aspiration to become members of the Comintern internationalist movement. Another important find, is the newly-discovered evidence concerning the founding of the CPA and its wartime conduct as an organization created and led by the Albanians themselves, albeit with Yugoslav ideological assistance under the transnational umbrella of the Comintern.