(320g) An Electrokinetic Probe of DNA Binding Interactions Via Resonant Entropic Trapping
- Conference: AIChE Annual Meeting
- Year: 2013
- Proceeding: 2013 AIChE Annual Meeting
- Group: 2013 Annual Meeting of the American Electrophoresis Society (AES)
- Time: Tuesday, November 5, 2013 - 2:42pm-3:04pm
A host of biological processes are orchestrated by complexation between DNA and binding agents (small molecules, proteins, adducts), and a fundamental understanding of these interactions is critical to unlocking their envisioned functionality. Nanopore-based methods have recently demonstrated potential to screen binding events and infer their kinetics, but conformational changes induced in the DNA complex upon binding must be quantitatively mapped in order to ascertain their biological function. These structural details can currently be elucidated by single molecule force measurements employing optical or magnetic tweezers and atomic force microscopy (AFM), but the corresponding experimental platforms are challenging to parallelize and analysis is inherently restricted to large sized biomolecules that can be readily imaged (i.e., λ-phage DNA or longer, contour lengths ~ μm).
Here we show how these limitations can be uniquely overcome by exploiting resonant entropic trapping (RET) imposed when DNA molecules experience confinement within a nanoscale pore network of comparable size to their equilibrium random coil conformation. This approach enables fundamental structural parameters associated with binding and complexation (contour and persistence lengths), including adduct formation, to be finely resolved in DNA much smaller than the size limits of conventional measurement techniques. In this way, we are able to gather structural information associated with DNA-drug complexation by performing a single microchip electrophoresis experiment (~20 min run time). RET can be accessed within the inherently nanoporous architecture of conventional polymeric hydrogels, eliminating the need to construct artificial nanofabricated topologies and making it possible for untethered DNA of arbitrary length to be rapidly analyzed in a convenient highly scalable format.