Homogeneous Electrochemiluminescence for Highly Sensitive Determination of Demethylase FTO Based on Target-Regulated DNAzyme Cleavage and Host–Guest Interaction

作者：Xia Yang, Shuai Qiao, Wei Zhao, Sijia Li, Yanxia Qiao, Yang Jiang, Yaqian Zhou, Yan Li · 发表于：Analytical Chemistry · 年份：2023 · DOI：10.1021/acs.analchem.3c01661 · 被引用次数：23 · 研究领域：RNA modifications and cancer、Advanced biosensing and bioanalysis techniques、MicroRNA in disease regulation

Fat mass and obesity-associated protein (FTO) is the first reported N 6 -methyladenosine (m 6 A) RNA demethylase. The dysregulation of FTO demethylation is strongly associated with various human cancers in a m 6 A-dependent manner. Herein, a homogeneous electrochemiluminescence (ECL) method for the determination of FTO was proposed based on the target-regulated DNAzyme cleavage. Moreover, the ECL signal was highly enhanced by host–guest interaction between β-cyclodextrin (β-CD) and tri- n -propylamine (TPrA). The m 6 A caged DNAzyme 17E-Me acted as a padlock, while the FTO served as the corresponding key. As the key, FTO could specifically remove m 6 A modification, restoring the cleavage activity of DNAzyme 17E. With the assistance of the Zn 2+ cofactor, the substrate strand was cleaved at a specific site, and the ECL indicator of Ru(phen) 3 2+ was discharged to produce an ECL signal. On the contrary, 17E-Me was blocked and no cleavage reaction occurred without the key. For the ECL detection, the electrode modification of β-CD@AuNPs concentrated Ru(phen) 3 2+ species through electrostatic adsorption and gathered TPrA molecules through host–guest interaction with β-CD, which resulted in an intense ECL response. The results demonstrated the ECL intensity linearly correlated with the logarithm of the FTO concentration (from 0.0001 to 100 nM) with a low detection limit (30 fM). The IC 50 value for FTO inhibitors rhein and meclofenamic acid were 35.6 μM and 20.3 μM, respectively....