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.

Ocean acidification in the subpolar North Atlantic: rates and mechanisms controlling pH changes

Repeated hydrographic sections provide critically needed data on and understanding of changes in basin-wide ocean CO2 chemistry over multi-decadal timescales. Here, high-quality measurements collected at twelve cruises carried out along the same track between 1991 and 2015 have been used to determine long-term changes in ocean CO2 chemistry and ocean acidification in the Irminger and Iceland basins of the North Atlantic Ocean. Trends were determined for each of the main water masses present and are discussed in the context of the basin-wide circulation. The pH has decreased in all water masses of the Irminger and Iceland basins over the past 25 years with the greatest changes in surface and intermediate waters (between −0.0010 ± 0.0001 and −0.0018 ± 0.0001 pH units yr−1). In order to disentangle the drivers of the pH changes, we decomposed the trends into their principal drivers: changes in temperature, salinity, total alkalinity (AT) and total dissolved inorganic carbon (both its natural and anthropogenic components). The increase in anthropogenic CO2 (Cant) was identified as the main agent of the pH decline, partially offset by AT increases. The acidification of intermediate waters caused by Cant uptake has been reinforced by the aging of the water masses over the period of our analysis. The pH decrease of the deep overflow waters in the Irminger basin was similar to that observed in the upper ocean and was mainly linked to the Cant increase, thus reflecting the recent contact of these deep waters with the atmosphere.

Surface enrichment in gold/silver alloys: effects of temperature studied by atom probe tomography

Bimetallic alloys are increasingly used in heterogeneous catalysis. This interest is explained by the emergence of new features that are absent in the parent single metals. Synergistic effects between the two combined elements create a more efficient catalyst. One of the most challenging aspect of multicomponent materials in catalysis is the ability to fine-tune the catalytic properties of an alloy by controlling the nature and composition of the surface [1]. For example, the gold/silver alloy combines a high activity and a large selectivity for a broad range of oxidation reaction.It is well established that the surface composition of alloys may deviate from that of the bulk phase. Surface enrichment has also important consequences in some applications of heterogeneous catalysis. In some cases, the thermal and chemical treatments can lead to opposite trends regarding the nature of the metal prone to surface enrichment. Using atom probe tomography we aim to link the physicochemical conditions the composition of the very first atomic layers of bimetallic catalysts and eventually to fine-tune the catalytic features of the latter.

Surface Activity and Bulk Defect Chemistry of Solid Oxide Fuel Cell Cathodes

In the first half of this thesis, a new robotic instrument called a scanning impedance probe is presented that can acquire electrochemical impedance spectra in automated fashion from hundreds of thin film microelectrodes with systematically varied properties. Results from this instrument are presented for three catalyst compositions that are commonly considered for use in state-of-the-art solid oxide fuel cell cathodes. For (La_0.8Sr_0.2)_0.95Mn_O3+δ (LSM), the impedance spectra are well fit by a through-the-film reaction pathway. Transport rates are extracted, and the surface activity towards oxygen reduction is found to be correlated with the number of exposed grain boundary sites, suggesting that grain boundaries are more surface-active than grains. For La_0.5Sr_0.5Co_O3-δ (LSC), the surface activity degrades ~50x initially and then stabilizes at a comparable activity to that of previously measured Ba_0.5Sr_0.5Co_0.8Fe_0.2O3-δ films. For Sr_0.06Nb_0.06Bi_1.87O₃ (SNB), an example of a doped bismuth oxide, the activity of the metal-SNB boundary is measured.

In the second half of this thesis, SrCo_0.9Nb_0.1O_3-δ is selected as a case study of perovskites containing Sr and Co, which are the most active oxygen reduction catalysts known. Several bulk properties are measured, and synchrotron data are presented that provide strong evidence of substantial cobalt-oxygen covalency at high temperatures. This covalent bonding may be the underlying source of the high surface activity.

Voltammetric Investigation of Xanthate Chemisorption on a Chalcopyrite Surface

Cyclic Voltammetry experiments have been conducted on copper, iron, and chalcopyrite (CuFeS2) and compared to mass-balanced EH-pH Diagrams. Potassium ethyl xanthate (KEX) was added to solution and additional voltammetry experiments were performed to determine the surface chemistry reactions of flotation collector in solution with these minerals. The ultimate goal of this research was to investigate the possibility of xanthate chemisorption onto the chalcopyrite mineral surface. Results of the copper mineral testing confirm previous literature studies and corroborate published isotherm data. Results of the iron mineral testing showed changes in surface reactions with the addition of potassium ethyl xanthate to solution, however, these results were not attributed to the chemisorption of xanthate. Results of the chalcopyrite mineral testing indicate that the surface of the mineral oxidizes to chalcocite (Cu2S). In the presence of ethyl xanthate, small currents were observed and attributed to chemisorption of the potassium ethyl xanthate at the chalcocite surface, suggesting that the mineral's hydrophobicity is induced by more than dixanthogen. This phenomenon was found to be pH-dependent under a range of alkaline conditions (i.e., pH 7-12) at narrow potentials (i.e., 0 to -200mV).

Effect of surface conditions on flow of a micropolar fluid driven by a porous stretching sheet

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Some surface profiles of a strip after plane-strain indentation by rigid bodies with serrated surfaces

This paper is concerned with the surface profiles of a strip after rigid bodies with serrated (saw-teeth) surfaces indent the strip and are subsequently removed. Plane-strain conditions are assumed. This has application in roughness transfer of final metal forming process. The effects of the semi-angle of the teeth, the depth of indentation and the friction on the contact surface on the profile are considered.