Institute of Organic Chemistry II, Friedrich-Alexander-Universität Erlangen-Nürnberg , Henkestr. 42, 91054 Erlangen, Germany.
Bioconjug Chem. 2013 Sep 18;24(9):1533-42. doi: 10.1021/bc400152n. Epub 2013 Sep 3.
Photochemical, nucleic acid-induced reactions, which are controlled by nontoxic red light, are well-suited for detection of nucleic acids in live cells, since they do not require any additives and can be spatially and temporally regulated. We have recently described the first reaction of this type, in which a phenylselenyl derivative of thymidine (5'-PhSeT-ODNa) is cleaved in the presence of singlet oxygen (Fülöp, A., Peng, X., Greenberg, M. M., Mokhir, A. (2010) A nucleic acid directed, red light-induced chemical reaction. Chem. Commun. 46, 5659-5661). The latter reagent is produced upon exposure of a photosensitizer 3'-PS-ODNb (PS = Indium(III)-pyropheophorbide-a-chloride: InPPa) to >630 nm light. In 2012 we reported on a fluorogenic version of this reaction (Dutta, S., Flottmann, B., Heilemann, M., Mokhir, A. (2012) Hybridization and reaction-based, fluorogenic nucleic acid probes. Chem. Commun. 47, 9664-9666), which is potentially applicable for the detection of nucleic acids in cells. Unfortunately, its yield does not exceed 25% and no catalytic turnover could be observed in the presence of substrate excess. This problem occurs due to the efficient, competing oxidation of the substrate containing an electron rich carbon-carbon double bonds (SCH═CHS) in the presence of singlet oxygen with formation of a noncleavable product (SCH═CHSO). Herein we describe a related, but substantially improved photochemical, catalytic transformation of a fluorogenic, organic substrate, which consists of 9,10-dialkoxyanthracene linked to fluorescein, with formation of a bright fluorescent dye. In highly dilute solution this reaction occurs only in the presence of a nucleic acid template. We developed three types of such a reaction and demonstrated that they are high yielding and generate over 7.7 catalytic turnovers, are sensitive to single mismatches in nucleic acid targets, and can be applied for determination of both the amount of nucleic acids and potentially their localization.
光化学反应、核酸诱导反应，这些反应受非毒性的红光控制，非常适合用于检测活细胞中的核酸，因为它们不需要任何添加剂，并且可以进行空间和时间上的调控。我们最近描述了这种类型的第一个反应，其中胸腺嘧啶的苯硒基衍生物（5'-PhSeT-ODNa）在单线态氧的存在下被切割（Fülöp，A.，Peng，X.，Greenberg，M. M.，Mokhir，A.（2010）核酸指导的，红光诱导的化学反应。Chem. Commun. 46, 5659-5661）。该试剂是在光敏剂 3'-PS-ODNb（PS = 铟（III）-叶啉-a-氯化物：InPPa）暴露于 >630nm 光时产生的。2012 年，我们报道了这种反应的荧光版本（Dutta，S.，Flottmann，B.，Heilemann，M.，Mokhir，A.（2012）杂交和基于反应的，荧光核酸探针。Chem. Commun. 47, 9664-9666），它可能适用于细胞中核酸的检测。不幸的是，其产率不超过 25%，并且在底物过量的情况下没有观察到催化周转。这个问题是由于在单线态氧存在下，含有富电子碳-碳双键（SCH=CHS）的底物的有效竞争氧化而引起的，生成了不可切割的产物（SCH=CHSO）。在此，我们描述了一种相关的、但大大改进的光化学、催化转化，该转化涉及将 9,10-二烷氧基蒽连接到荧光素上的荧光有机底物，形成明亮的荧光染料。在高度稀释的溶液中，只有在存在核酸模板的情况下，才会发生这种反应。我们开发了三种这种类型的反应，并证明它们产率高，生成超过 7.7 个催化循环，对核酸靶标中的单个错配敏感，并且可用于测定核酸的量和潜在的定位。
