Tang Zhiwen, Mallikaratchy Prabodhika, Yang Ronghua, Kim Youngmi, Zhu Zhi, Wang Hui, Tan Weihong
Center for Research at the Bio/Nano Interface, Department of Chemistry, Shands Cancer Center and UF Genetics Institute, University of Florida, Gainesville, Florida 32611-7200, USA.
J Am Chem Soc. 2008 Aug 27;130(34):11268-9. doi: 10.1021/ja804119s. Epub 2008 Aug 5.
A novel aptamer-based molecular probe design employing intramolecular signal transduction is demonstrated. The probe is composed of three elements: an aptamer, a short, partially cDNA sequence, and a PEG linker conjugating the aptamer with the short DNA strand. We have termed this aptamer probe an "aptamer switch probe", or ASP. The ASP design utilizes both a fluorophore and a quencher which are respectively modified at the two termini of the ASP. In the absence of the target molecule, the short DNA will hybridize with the aptamer, keeping the fluorophore and quencher in close proximity, thus switching off the fluorescence. However, when the ASP meets its target, the binding between the aptamer and the target molecule will disturb the intramolecular DNA hybridization, move the quencher away from the fluorophore, and, in effect, switch on the fluorescence. Both ATP and human alpha-thrombin aptamers were engineered to demonstrate this design, and both showed that fluorescence enhancement could be quantitatively mediated by the addition of various amounts of target molecules. Both of these ASPs presented excellent selectivity and prompt response toward their targets. With intrinsic advantages resulting from its intramolecular signal transduction architecture, the ASP design holds promising potential for future applications, such as biochip and in situ imaging, which require reusability, excellent stability, prompt response, and high sensitivity.
展示了一种基于新型适体的分子探针设计，该设计采用分子内信号转导。该探针由三个元件组成：一个适体、一个短的部分cDNA序列，以及一个将适体与短DNA链连接的聚乙二醇接头。我们将这种适体探针称为“适体开关探针”，即ASP。ASP设计利用了分别修饰在ASP两端的一个荧光团和一个猝灭剂。在没有目标分子的情况下，短DNA会与适体杂交，使荧光团和猝灭剂保持紧密接近，从而关闭荧光。然而，当ASP遇到其目标时，适体与目标分子之间的结合会干扰分子内DNA杂交，使猝灭剂远离荧光团，实际上开启荧光。ATP和人α-凝血酶适体都经过改造以证明这种设计，并且都表明荧光增强可以通过添加不同量的目标分子进行定量介导。这两种ASP对其目标都表现出优异的选择性和快速响应。由于其分子内信号转导结构具有内在优势，ASP设计在未来应用中具有广阔的潜力，如生物芯片和原位成像，这些应用需要可重复使用性、出色的稳定性、快速响应和高灵敏度。
