Identification of DNA aptazymes for small-molecule sensors

描述(由申请人提供)：这一领域的拟议研究应用寻求利用DNA的结合和催化能力来开发用于小分子传感器的新方法。从药物到制造化学品的许多小分子存在于环境中，并可能影响人类健康。农药是一类对人类构成重大危害的小分子污染物。目前的检测方法依赖于包括质谱仪和液相色谱在内的技术，能够在非常低的浓度下检测到污染物，但需要大量的样品准备，这必须在实验室完成。由于产生小分子污染物抗体所需的努力以及抗体对反应条件的敏感性，开发污染物的ELISA法是具有挑战性的。使用简单的设备在现场检测这些污染物的能力，而不是要求在实验室中运输和处理样本，将允许快速评估潜在的有害情况。适配子是以高亲和力和高选择性结合靶标的能力为特征的核酸序列。脱氧核酶或脱氧核酶是一种催化DNA序列，可以与适体配对形成脱氧核酶。DNA的稳定性使端粒酶对传感器具有吸引力。虽然aptazyme已被开发用于检测金属离子和一些小分子的存在，但我们打算开发新的aptazyme系统来检测农药。DNA传感器的发展依赖于RNA裂解以外的催化活性，这将代表便携式传感器的新方向，将探索DNA催化的新领域，并可以提供关于小分子如何与核酸相互作用的信息。这项提案中讨论的DNA aptazyme的成功识别最终可能为个人提供评估其个人健康风险的新工具，并将很容易应用于其他小分子。该提案的三个具体目标是：(1)鉴定依赖于农药的DNA aptazyme的活性；(2)表征农药特异性的DNA aptazyme；(3)开发DNA aptazyme的信号途径。 公共卫生相关性：这一领域应用的目标是利用DNA的结合和催化能力来开发新的环境污染物小分子传感器。这些传感器的成功开发将为个人提供新的工具来监测他们的个人健康风险，而不需要等待实验室检测结果。

DESCRIPTION (provided by applicant): 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. PUBLIC HEALTH RELEVANCE: 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.