The proposed research in this AREA application seeks to harness the binding and catalytic abilities of DNA to develop new approaches for small-molecule sensors. Many small molecules ranging from pharmaceuticals to manufacturing chemicals are present in the environment and potentially impact human health. Pesticides are one class of small-molecule pollutants that represent a significant hazard to humans. Current testing methods relying on techniques including mass spectrometry and liquid chromatography are able to detect pollutants at very low concentrations, but require extensive sample preparation that must be done in a lab. Developing ELISA assays for pollutants is challenging because of the efforts required to generate antibodies for small- molecule pollutants and the sensitivity of antibodies to reaction conditions. The ability to detect these pollutants on-site wih simple equipment rather than requiring samples be transported and processed in a lab would allow for rapid assessment of potentially harmful situations. Aptamers are nucleic acid sequences characterized by their ability to bind a target with high affinity and selectivity. Deoxyribozymes or DNAzymes are catalytic DNA sequences that can be paired with aptamers to form aptazymes. The stability of DNA makes aptazymes attractive components for sensors. While aptazymes have been developed to detect the presence of metal ions and, in proof-of-principle studies, some small molecules, we intend to develop new aptazyme systems for the detection of pesticides. The development of DNA sensors for pollutants that rely on catalytic activities other than RNA cleavage would represent new directions for portable sensors, would explore new areas of DNA catalysis, and could provide information about how small molecules interact with nucleic acids. The successful identification of the DNA aptazymes discussed in this proposal could eventually provide individuals with new tools for assessing their personal health risks and would be easily applied to other small molecules. The three specific aims of the proposal are to (1) identify DNA aptazymes dependent on pesticides for activity;(2) characterize pesticide-specific DNA aptazymes;and (3) develop signaling approaches for DNA aptazymes.
The goal of this AREA application is to use the binding and catalytic abilities of DNA to develop new small- molecule sensors for environmental pollutants. The successful development of these sensors will provide individuals with new tools for monitoring their personal health risks without needing to wait on lab testing results.