| This research is focused on studying the mechanical property of biopolymers such as polysaccharides, proteins and DNA at a single molecule level using Atomic Force Microscopy (AFM)-based force-spectroscopy techniques. In addition to its basic nanoscience goals, this research aims at: developing new analytical methods for identifying polysaccharides by force spectroscopy that can benefit carbohydrate science; developing new methods for detecting DNA damage at a single-molecule level that potentially can benefit human health; and developing new biophysical methodologies for characterizing the mechanical properties of tandem ankyrin repeats, which can help to elucidate the gating mechanism of ion channels.; The primary experimental method of this research is AFM-based single-molecule force-spectroscopy that allows mechanical manipulations on single biopolymers including accurate measurements of their elasticity, force-induced conformational transition, and unfolding and folding reaction. AFM measurements of polysaccharide elasticity are supplemented by Steered Molecular Dynamics (SMD) calculations, which reveal the details of their behavior under tension, at an atomic level.; 1,6-linked polysaccharides. We observed a new force-induced conformational transition in the beta 1,6-linked D-glucose polysaccharide, pustulan, which involves the forced rotation of the exo-cyclic C5-C 6 group to a high-energy position. This transition produces an unusual linear elasticity of pustulan. We also observed a new force-induced conformational transition in the alpha 1,6-linked D-glucose polysaccharide, dextran, which involves a compound transition of the glucopyranose ring from a chair to a boat-like conformation with a simultaneous rotation of the C 5-C6 bond to a high-energy position.; Diagnosis of UV damage of individual DNA. We found that the characteristic plateau in the force spectrogram of the irradiated DNA is shortened in a dose-dependent manner as compared to native DNA. We also observed sequence-dependent damage saturation on the elasticity of synthetic DNA polydA·polydT and polydG·polydC, which indicates the steady-state of the yield of UV-induced dimers.; Nanospring behavior of ankyrin repeats. We found that ankyrin repeats reveal the fully reversible hookean linear elasticity, which makes them an ideal candidate for controlling mechanotransduction. In addition, we obtained the first direct measurement of the magnitude of refolding forces of a protein domain, indicating that the unfolded polypeptide chain refolds quickly, generating the folding forces. |
