BRIGE: Fluorescence Based Single Molecule Force Spectroscopy with DNA Nanotechnology

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This Broadening Participation Research Initiation Grant in Engineering (BRIGE) provides funding for the development of a nanoscale device for fluorescence-based high throughput single molecule force spectroscopy. The device will be assembled using the recently developed nanotechnology DNA origami. DNA origami enables the construction of nanoscale objects with unprecedented geometric complexity via programmed molecular self-assembly. The device will comprise a stiff framework of bundles of double-stranded DNA, attachment points for two biomolecules, a flexible polymer force probe, and fluorescent molecules to act as a readout of the interaction between the two biomolecules of interest. The force probe will facilitate binding between the biomolecules, and subsequently apply a known force acting to rupture the interaction. The bond lifetime will be monitored using a fluorescence readout, which is amenable to high throughput data collection. Interaction lifetimes will be measured as a function of force to determine kinetic parameters that govern the molecular interaction. Initial proof-of-principle experiments will probe DNA and RNA base-pairing interactions. Ultimately, the device will be implemented to study protein-DNA, protein-RNA, and protein-protein systems.Single molecule force spectroscopy studies have provided critical insight into the interactions that stabilize biomolecular machinery and regulate the function of cells. However, the widespread application of force spectroscopy has been hindered by the need for cumbersome and expensive equipment and low throughput data acquisition. If successful, this research will enable economical and high-throughput force spectroscopy studies of biomolecular machinery. The ultimate goal is to develop a device that can be packaged and distributed for widespread application on basic laboratory fluorescence microscopes. This award will also support the establishment of a biomolecular machinery short course to be offered to high school age students at summer engineering camps and the development of a research seminar course aimed to recruit underrepresented engineering students to pursue graduate level research.