适体
化学
DNA
指数富集配体系统进化
荧光各向异性
范德瓦尔斯力
检出限
组合化学
色谱法
分子
核糖核酸
生物化学
分子生物学
生物
有机化学
基因
膜
作者
Cunzheng Zhang,Li Wang,Zhui Tu,Xing Sun,Qinghua He,Zhao-Jing Lei,Chongxin Xu,Yuan Liu,Xiao Zhang,Jingyi Yang,Youlong Xu
标识
DOI:10.1016/j.bios.2013.12.020
摘要
An approach is developed to detect the organophosphorus pesticides via competitive binding to a recombinant broad-specificity DNA aptamer with a molecular beacon (MB), the binding of the MB to the aptamer results in the activation of a fluorescent signal, which can be measured for pesticide quantification. Aptamers selected via the Systematic Evolution of Ligands by Exponential Enrichment (SELEX) were structurally modified and truncated to narrow down the binding region of the target, which indicated that loops of the aptamer contributed different functions for different chemical recognition. Thereafter, a variant fused by two different minimum functional structures, was clarified with broad specificity and increased affinity. Further molecular docking and molecular dynamics simulations was conducted to understand the molecular interaction between DNA structure and chemicals. 3D modeling revealed a hot spot area formed by 3 binding sites, forces including hydrogen bonds and van der Waals interactions appear to play a significant role in enabling and stabilizing the binding of chemicals. Finally, an engineered aptamer based approach for the detection of organophosphorus pesticides was successfully applied in a test using a real sample, the limit of quantification (LOQ) for phorate, profenofos, isocarbophos, and omethoate reached 19.2, 13.4, 17.2, and 23.4 nM (0.005 mg L(-1)), respectively.
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