DNAzyme and rGO based fluorescence assay for Fpg activity analysis, drug screening, and bacterial imaging.
DNAzyme and rGO based fluorescence assay for Fpg activity analysis, drug screening, and bacterial imaging.
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DOI：
10.1016/j.talanta.2020.121158
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发表时间：
2020-10
期刊：
影响因子：
6.1
通讯作者：
Y. Qiu;Wenya Dang;Jialong Fan;Ting Zhou;Bin Li;Yang Liu;Yan Qin;C. Tong;M. Daniyal
中科院分区：
文献类型：
--
作者：
Y. Qiu;Wenya Dang;Jialong Fan;Ting Zhou;Bin Li;Yang Liu;Yan Qin;C. Tong;M. Daniyal
由于甲酰胺嘧啶DNA糖基化酶（formamidopyrimidine DNA glycosylase，Fpg）在生理过程和DNA氧化损伤相关疾病中的重要作用，建立灵敏的体外和体内检测Fpg活性的方法以阐明其在这些事件中的具体作用是非常必要的。利用DNA酶辅助级联信号放大和还原氧化石墨烯（rGO）超高荧光猝灭效率的优点，建立了一种灵敏、简便、可靠的荧光检测方法。该检测体系由DNAzyme、rGO和荧光探针组成，检测范围为0 ~ 80 U/mL，检测限为0.66 U/mL。利用该方法对11个天然化合物进行了筛选，鉴定出7个化合物为Fpg的激活剂。更重要的是，该方法首次用于通过荧光成像监测活大肠杆菌中Fpg的活性。成像结果直观地显示了天然化合物人参皂苷Re对大肠杆菌Fpg的动态激活作用。上述结果表明，基于DNAzyme和rGO的荧光检测方法为Fpg的体外定量检测和活菌的实时监测提供了一种有效的策略，在生物学研究和Fpg靶向药物筛选方面具有很大的应用潜力。
Due to the significant role of formamidopyrimidine DNA glycosylase (Fpg) in physiological processes and DNA oxidative damage-related diseases, it is essential to establish sensitive methods for monitoring the Fpg activityin vitroandin vivoso as to illustrate its concrete role in these events. In this work, a sensitive, simple and reliable fluorescence assay was developed by taking the advantages of DNAzyme assisted cascade signal amplification and ultra-high fluorescence quenching efficiency of reduced graphene oxide (rGO). This detection system consisted of DNAzyme, rGO and fluorescence probe allows the activity of Fpg to be detected in a linear range from 0 to 80 U/mL with a detection limit of 0.66 U/mL. With the help of this method, 11 natural compounds were screened, and 7 compounds were identified as activators of Fpg. More importantly, the developed assay was used to monitor the activity of Fpg through fluorescence imaging in livingEscherichia colifor the first time. The imaging results visually demonstrated the dynamic activation effect of natural compound Ginsenoside Re on the Fpg ofEscherichia coli. In summary, these results indicated that this DNAzyme and rGO based fluorescence assay provides a potent strategy for Fpg quantitative assayin vitroand real-time monitoring in living bacteria, which holds great potential for applying on biological study and Fpg-targeted drug screening.