Poly-proline-based chiral stationary phases are relatively new stationary phases and have shown to be competitive to other commercially available chiral stationary phases for high performance liquid chromatography (HPLC). The conformational studies, solvation properties and enantio-selective mechanism of this chiral stationary phase are the main focus of this thesis. Semi-flexible models are developed based on an extensive series of ab initio calculations for proline selectors from di- to hexa-proline and a series of six chiral analytes. Then molecular dynamics simulations are performed to study the solvation, conformational preferences at the interface, and the selectivity. The solvation and conformational preferences of poly-proline selectors at the interface are examined in a normal phase n-hexane/-2propanol and a reverse phase water/methanol solvent. We noticed a significant difference between conformational preferences of poly-proline chains in these solvents indicating the effect of solvent polarity and hydrogen bonding on the relative stabilities of poly-proline conformers. Solvent partitioning occurs at the interface and this creates a polarity gradient between the stationary phase and the bulk that encourages analyte docking at the interface. Hydrogen bonding to the poly-proline selectors is shown to be a function of solvent composition and poly-proline conformation at the interface. The selectivity of the poly-proline chains was studied by molecular dynamics simulations of chiral analytes docking into the interface. The selectivity factors for a set of enantiomers were predicted successfully. Enantio-resolution has been shown to mostly happen with hydrogen bonding to poly-proline carbonyl oxygens located close to the interface. Steric interactions and conformational flexibility of the analytes are also contributing factors for enantio-resolution. / Thesis (Ph.D, Chemistry) -- Queen's University, 2013-01-28 14:31:53.316

The group 2 late embryogenesis abundant (LEA) proteins, also known as the dehydrins, are intrinsically disordered proteins that are expressed in plants experiencing extreme environmental conditions such as drought or low temperature. In this work, we study the potential roles that dehydrins may have in stabilizing membranes and actin microfilaments during cold stress. We have cloned and expressed in E. coli two dehydrins from Thellungiella salsuginea, denoted TsDHN-1 (acidic) and TsDHN-2 (basic). These proteins were expressed as SUMO-fusion proteins for in vitro phosphorylation by casein kinase II (CKII), and for structural analysis by CD and Fourier transform infrared (FTIR) spectroscopy. We show using transmission-FTIR spectroscopy that ordered secondary structure is induced and stabilized in these proteins by association with large unilamellar vesicles emulating the lipid compositions of plant plasma and organellar membranes. The increase in secondary structure by membrane association is further facilitated by the presence of Zn2+. Lipid composition and temperature have synergistic effects on the secondary structure. Our single molecule force spectroscopy studies also suggest tertiary folding of both TsDHN-1 and TsDHN-2 induced by association with lipids. From Langmuir-Blodgett monolayer compression studies, and from topographic studies using atomic force microscopy at variable temperature, we conclude that TsDHN-1 stabilizes the membrane at lower temperatures. Finally, we show that the conformations of TsDHN-1 and TsDHN-2 are affected by pH, interactions with cations and membranes, and phosphorylation. Actin assembly by these dehydrins was assessed by sedimentation assays, and viewed by transmission electron and atomic force microscopy. Phosphorylation enabled both dehydrins to polymerize actin filaments, a phenomenon that may occur in the cytosols of plant cells undergoing environmental stress. These results support the hypothesis that dehydrins stabilize plant organellar membranes and/or the cytoskeleton in conditions of stress, and further that phosphorylation may be an important feature of this stabilization. / NSERC