Physics
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Item USING SINGLE MOLECULE TECHNIQUES TO DETERMINE THE MECHANISM OF DNA TOPOLOGY SIMPLIFICATION BY TYPE IIA TOPOISOMERASES(2011) Hardin, Ashley Harris; Thirumalai, Devarajan; Neuman, Keir C; Chemical Physics; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)Type IIA topoisomerases are essential, universally conserved proteins that modify DNA topology by passing one segment of duplex DNA (the transfer, or T-segment) through a transient double strand break in a second segment of DNA (the gate, or G-segment) in an ATP-dependent reaction. Type IIA topoisomerases decatenate, unknot, and relax supercoiling in DNA to levels below equilibrium, resulting in global topology simplification. The mechanism underlying non-equilibrium topology simplification remains speculative, though several plausible models have been proposed. This thesis tests two of these, the bend angle and kinetic proofreading models, using single-molecule techniques. The bend angle model postulates that non-equilibrium topology simplification scales with the bend angle imposed on the G-segment DNA by a type IIA topoisomerase. To test this model, we used atomic force microscopy and single molecule Förster resonance energy transfer to measure the extent of bending imposed on DNA by three type IIA topoisomerases that span the range of topology simplification activity. We found that all proteins bent DNA, but the imposed bends are similar and cannot account for the differences among the enzymes. These data do not support the bend angle model and suggest that DNA bending is not the sole determinant of non-equilibrium topology simplification. Based on the assumption that the rates of collision between DNA segments is higher in knotted, linked, and supercoiled DNA than in topologically free or relaxed DNA, the kinetic proofreading model proposes that two successive binding events between a G-segment bound topoisomerase and a putative T-segment are required to initiate strand passage. As a result of the two step process, the overall rate of strand passage should scale with the square of the collision probability of two DNA segments. To test this model, we used magnetic tweezers to manipulate a paramagnetic bead tethered to the surface by two DNA molecules. By rotating the bead, we varied the proximity, and thus collision rate, of the two molecules to determine the relationship between collision probability and rate of strand passage. Our data indicate that the strand passage rate scales linearly with the collision probability, which is inconsistent with the kinetic proofreading model.Item Ferrocene-based molecular electronics and nanomanufacturing of Pd nanowires.(2007-11-27) Wang, Lixin; Sita, Lawrence; Chemical Physics; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)Two test structures were tried out for molecular junction formation and subsequent I-V characteristics measurements. One is formed by insertion of certain dithiol molecules into an alkanethiol self-assembled monolayer (SAM), followed by tethering the free thiol end with gold nanoparticles. The test structure can then be measured with CP-AFM. The matrix SAM, mixed monolayer with inserted dithiol molecules, and final test structure with gold nanoparticles were prepared and characterized by ellipsometry, AFM and STM. However, the CP-AFM measurements were very irreproducible, even on an alkanethiol SAM. This problem was analyzed and believed to come from two possible causes, namely thermal drift and deformation of the metalized tips. The other test structure was from insertion of molecules into nanogaps made by electromigration technique. Two molecules were tested and drastically different properties were observed from junctions with each molecule. For Fc-OPE molecules, near perfect conductance peaks (>0.6G0) were observed in some junctions and analysis indicates that such molecular junction contains only one or two molecules inside the nanogap. The formation of conductance peaks was analyzed with Landauer formula and a simple metal-molecule-metal model. Computational calculation also predicted high conductance through such junctions and the existence of resonant peaks. The junctions with OPE molecules, however, showed poor conductance. Possible causes such as molecular structure and easiness of molecular junction formation were discussed. In the second part of this dissertation, a new method was developed to fabricate Pd nanowires on HOPG surface using a sacrificial Cu film. The morphology and composition of the nanowires were characterized by AFM, SEM and XPS. The formation of such Pd nanowires was explained with a galvanic displacement mechanism and some test experiments were carried out to prove such growth mechanism. It was also found that the size of the Pd nanowires can be directly controlled by the thickness of the Cu film that was initially deposited. However, attempts to make Au, Pt and Ag nanowires with the same method failed, and possible causes were discussed.