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首页> 外文期刊>Sensors and Actuators >G-triplex molecular beacon-based fluorescence biosensor for sensitive detection of small molecule-protein interaction via exonuclease Ill-assisted recycling amplification
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G-triplex molecular beacon-based fluorescence biosensor for sensitive detection of small molecule-protein interaction via exonuclease Ill-assisted recycling amplification

机译：基于三重信标的基于分子信标的荧光生物传感器，用于通过核酸外切酶病态辅助循环扩增灵敏检测小分子-蛋白质相互作用

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摘要

Small molecule-protein interaction plays a significant role in disease diagnosis and biomedical research. Most of the reported fluorescence strategies for small molecule-protein interaction still involve the use of costly fluor-ophore-labelled probes. In this work, a label-free fluorescent method for sensitive detection of small molecule-protein interaction is proposed. It is based on G-triplex-based molecular beacon (G3-MB) and exonuclease HI (Exo III)-aided DNA recycling amplification. Biotin-streptavidin interaction is selected as one model for proof-of-concept. The biotin-linked double-strand DNA (dsDNA) is used as the binding probe. In the presence of streptavidin, Exo III hydrolyzes the binding probe to release the trigger DNA, and the released trigger DNA can hybridize G3-MB to trigger the Exo HI cleavage process, accompanied by releasing the trigger DNA and G3 sequences. The trigger DNA can recycle, generating a large number of G3 sequences. The generated G3 sequences bind thioflavin T (ThT) to produce strong fluorescence emission. The interaction between small molecule-protein can be detected by the fluorescence reader. This method exhibits low detection limit (5.6 pg/mL), and owns the merits of the simplicity, high specificity and satisfactory applicability in serum sample.
机译：小分子-蛋白质相互作用在疾病诊断和生物医学研究中起着重要作用。大多数已报道的小分子与蛋白质相互作用的荧光策略仍然涉及使用昂贵的荧光载体标记的探针。在这项工作中，提出了一种用于小分子-蛋白质相互作用的灵敏检测的无标记荧光方法。它基于基于G-triplex的分子信标（G3-MB）和核酸外切酶HI（Exo III）辅助的DNA循环扩增。选择生物素-链霉亲和素相互作用作为概念验证的一种模型。生物素连接的双链DNA（dsDNA）用作结合探针。在链霉亲和素的存在下，Exo III水解结合探针以释放触发DNA，释放的触发DNA可以与G3-MB杂交以触发Exo HI裂解过程，并释放触发DNA和G3序列。触发DNA可以回收，产生大量的G3序列。产生的G3序列与硫黄素T（ThT）结合以产生强荧光发射。小分子-蛋白质之间的相互作用可以通过荧光读取器检测。该方法的检出限低（5.6 pg / mL），具有操作简便，特异性强，在血清样品中令人满意的适用性等优点。

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