DNA aptamers for the separation of metal ions

Aptamers are short oligonucleotides that selectively bind to a target compound with high binding affinity. This ability to selectively bind a target compound is often facilitated by the well-defined secondary structure of the aptamer. Prior interest in aptamers has focused mainly on their interactions with the target molecule against which they were isolated, in either a sensing or pharmaceutical application. Our interest in aptamers lies in the interactions they show with molecules other than the target species. Just as the unique, three-dimensional conformation of the aptamer imparts binding selectivity toward its target, it is our supposition that the conformation will enable the aptamer to show a range of weaker binding interactions with a series of non-target molecules. This range of interactions would indicate that aptamers have potential as a stationary phase for a chemical separation.; This dissertation focuses on the interactions of lanthanide ions with an aptamer selected to bind to thrombin. The aptamer forms a G-quartet conformation that is characterized by two planes of guanine tetrads. In addition to the thrombin-binding ligand, several sequential variations of this aptamer are discussed. The first part of this work utilized spectroscopy techniques such as absorbance circular dichroism, thermal melt analysis, and fluorescence energy transfer measurements to demonstrate that the aptamers showed a range of binding associations among the lanthanide metal ions. Absorbance circular dichroism was used to monitor the changes in the conformation of the aptamer in the presence of different lanthanide ions and compare the extent of interaction among the different ions. Thermal melts were used to compare the differences in interactions among the metals by examining the extent to which the ions affected the temperature at which the aptamer changed from a G-quartet to a non-G-quartet-conformation. Fluorescence energy transfer measurements were made to study possible binding sites heterogeneity. In addition to spectroscopic measurements, lanthanide-shifted proton and phosphorus nuclear magnetic resonance was used to investigate the location of the binding site.; Based upon the results of these spectroscopic studies, a new aptamer was designed to increase differences observed among the lanthanide ions. The aptamer was attached to the surface of a fused-silia capillary and electrochromatography was performed to separate mixtures of lanthanide ions. The effects of several different parameters were investigated, including column length, column inner diameter, and the used of surfactants. These results indicate that aptamers do exhibit potential for the separation of lanthanide metal ions.