Department of Chemical and Biomolecular Engineering (BK 21 + program), KAIST, Daehak-ro 291, Yuseong-gu, Daejeon 34141, Republic of Korea.
Analyst. 2018 Apr 30;143(9):2023-2028. doi: 10.1039/c8an00099a.
We herein describe a simple and sensitive strategy to detect a small molecule-protein interaction based on terminal protection-mediated exponential strand displacement amplification (eSDA). In principle, the small molecule linked to a DNA probe protects the DNA probe against the exonuclease I-catalyzed degradation after its binding to the corresponding target protein. The protected DNA probe then serves as a template to promote eSDA. Consequently, a large number of duplexes are produced, which leads to a high fluorescence from a double-stranded DNA specific fluorescent dye, SYBR Green I. As a model system to prove this sensing strategy, the interaction between biotin and streptavidin (SA), which is known to be the strongest among the non-covalent biological interactions, was selected and its analytical performance was thoroughly investigated. As a result, SA was sensitively detected with the limit of detection of 16 pM. In addition, the practical applicability of this method was successfully demonstrated by reliably determining the SA in human serum.
我们在此描述了一种基于末端保护介导的指数链置换扩增（eSDA）检测小分子-蛋白质相互作用的简单而灵敏的策略。从原理上讲，与 DNA 探针连接的小分子在与相应靶蛋白结合后，可保护 DNA 探针免受外切酶 I 催化的降解。然后，受保护的 DNA 探针充当模板以促进 eSDA。结果，产生了大量的双链体，从而导致双链体特异性荧光染料 SYBR Green I 产生高荧光。作为证明这种传感策略的模型系统，选择了生物素和链霉亲和素（SA）之间的相互作用，众所周知，这是所有非共价生物相互作用中最强的相互作用之一，并对其分析性能进行了深入研究。结果，SA 的检测灵敏度达到 16 pM。此外，通过可靠地测定人血清中的 SA，成功地证明了该方法的实际适用性。
