The study of protein–protein interactions (PPIs) plays an important role in the understanding of biological systems; however, the established methods for PPI analysis often involve cumbersome sample preparation, multiple detecting steps, and costly instruments. Here we report a versatile and sensitive electrochemical method based on PPI-induced distinctive migration behavior of DNA deoxyribozyme (DNAzyme) on an electrode surface. In this method, the cleavage activity of DNAzyme toward the substrate DNA modified on the electrode surface is inversely correlated with the hydrodynamic diameter of the macromolecule attached to it. By making full use of this principle in an inexpensive electrochemical format that is named the dynamic sandwich-type electrochemical assay (dSTEA), we can probe into the presence of large macromolecules in a single-step procedure. Moreover, we can not only detect sub-picomolar protein interaction events but also analyze the assembly of kinase in the whole cell extract. This novel signaling mechanism proposed in this work may broaden the applicability of DNAzyme-based electrochemical assays and it may also have great potential for applications in other interfacial sensor developments.

中文翻译：

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用于蛋白质定量和蛋白质-蛋白质相互作用的动态夹心型电化学分析

蛋白质间相互作用（PPI）的研究在理解生物系统中起着重要作用。但是，用于PPI分析的既定方法通常涉及繁琐的样品制备，多个检测步骤以及昂贵的仪器。在这里，我们报告了一种基于PPI诱导的电极表面DNA脱氧核酶（DNAzyme）的独特迁移行为的通用且灵敏的电化学方法。在这种方法中，DNAzyme对在电极表面修饰的底物DNA的切割活性与附着在其上的大分子的流体动力学直径成反比。通过以便宜的电化学形式充分利用这一原理，即动态夹心型电化学分析（dSTEA），我们可以通过一步操作来探查大分子的存在。而且，我们不仅可以检测亚皮摩尔蛋白相互作用事件，还可以分析全细胞提取物中激酶的组装。在这项工作中提出的这种新颖的信号传导机制可能会拓宽基于DNAzyme的电化学检测的适用性，并且在其他界面传感器的开发中也可能具有巨大的潜力。