Solid surface modification for force spectroscopy measurements on interactions at the single-molecule level

Atomic Force Microscope (AFM) based force spectroscopy has been widely employed in quantitative assessments of interactions at the molecular level. Force spectroscopy is often used to characterize specific interactions between biological molecules. More recently it is becoming a tool of research in chemistry. Applying this approach in chemistry requires modification of the technique. The primary goal of this work is to consider and implement changes to force spectroscopy methodology, improving its ability to characterize interactions between individual molecules in aqueous solution.;To separate possible surface adhesion from the molecular unbinding events under study, molecules are bound by relatively long polymeric tethers. The rupture events are detected at the tip position of approximately twice the tether length (double tether approach). In addition, relatively large areas that polymeric tethers occupy on surfaces limit possible number of interacting molecules to a few due to the typical nanoscale size of AFM tips. Polyethylene glycol (PEG) is a good candidate as a polymer tether because it is inert and non-adhering in water. Many functionalized PEG are commercially available and can be used for grafting to the surfaces of tips and substrates. Different grafting strategies should be adopted according to specific properties of molecules tethered by PEG. The sample preparation approach generally adopted in our experiments was first tested in the measurements of catechol---catechol interactions mediated by Fe(III) (Chapter 2).;Force spectroscopy experiments directly probe intermolecular forces subjected to an external mechanical load and register either conformational transitions or ruptures of intermolecular bonds. Application of a mechanical load lowers the molecular energy barrier in the direction of applied force thus directing the reaction pathway. The rupture forces and the dissociation rates under applied load are measured directly. Extrapolation of these results to zero force is often used for interpretation and subsequent comparison of the extracted kinetic parameters. Common data analysis uses a simplified model of the potential of mean force. However, the data analysis introduced in Chapter 3 does not make typical simplifying assumptions. The results show that the standard data analysis may lead to significant systematic errors in estimating kinetic parameters.;An analytical model for the probability density of simultaneous rupture of two bonds in force spectroscopy experiments is proposed and used in the data analysis in Chapter 4. It is suggested that the observed effects of multiple bonds rupture may be a common feature of force spectroscopy experiments and therefore should be considered to reduce the systematic errors in extracted kinetic parameters.;Research presented here establishes an experimental approach to determine kinetic parameters of non-covalent bonds in solution and provides molecular level insights into hydrophobic interactions. For single C60 fullerene-C 60 fullerene interactions in water, experimentally determined activation barrier width is ∼0.39 nm and activation energy is ∼61 kJ·mol -1. For hexadecane-hexadecane interactions in water, the activation barrier widths varies from 0.42 to 0.7 nm depending on the placement of the attachment point of the polymer tether on the hexadecane molecules, while the corresponding activation energies (-50 kJ·mol-1) doesn't change significantly. (Abstract shortened by UMI.).