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Reporter-recruiting bifunctional aptasensor for bioluminescent analytical assays

机译:记者 - 招募生物发光分析测定的双功能aptasensor

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We report a novel bioluminescent aptasensor, which consists of 2′-F-RNA aptamer modules joined into a bi-specific aptamer construct. One aptamer module binds the analyte, then after structural rearrangement the second module recruits non-covalently Ca ~(2+) -dependent photoprotein obelin from the solution, thus providing a bioluminescent signal. This concept allows using free protein as a reporter, which brings such advantages as no need for aptamer–protein conjugation, a possibility of thermal re-folding of aptamer component with no harm to a protein, and simpler detection protocol. We developed the new 2′-F-RNA aptamer for obelin, and proposed the strategy for engineering structure-switching bi-modular aptamer constructs which bind the analyte and the obelin in a sequential manner. With the use of hemoglobin as a model analyte, we showed the feasibility of utilizing the aptasensor in a fast and straightforward bioluminescent microplate assay. With a proper design of a secondary structure, this strategy of aptasensor engineering might be further extended to bi-specific aptamer-based bioluminescent sensors for other analytes of interest.
机译:我们报告了一种新的生物发光Aptasensor,其由连接到双特异性适体构建体的2'-F-RNA适体模块组成。一个适体模块结合分析物,然后在结构重新排列之后,第二模块从溶液中重新募集非共价Ca〜(2+)依赖性光蛋白质,从而提供生物发光信号。该概念允许使用游离蛋白作为记者,这为不需要适体 - 蛋白质缀合而产生这种优点,这种优点是对适体组分的热再折叠不损害蛋白质的可能性,以及更简单的检测方案。我们为Obelin开发了新的2'-F-RNA适体,并提出了工程结构切换双模型适体构建体的策略,其以顺序方式结合分析物和obelin。通过使用血红蛋白作为模型分析物,我们展示了利用快速和直接的生物发光微孔板测定中的Aptasensor的可行性。通过适当的次结构设计,这种Aptasentor工程策略可能进一步扩展到基于双特异性适体的生物发光传感器,用于其他目的的其他分析。

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