Sensitive determination of formamidopyrimidine DNA glucosylase based on phosphate group-modulated multi-enzyme catalysis and fluorescent copper nanoclusters
Li, JY (Li, Junyao)[ 1,2 ] ; Zhang, MY (Zhang, Mengyang)[ 1,2 ] ; Wang, HS (Wang, Huaisheng)[ 3 ] ; Wu, J (Wu, Jie)[ 1,2 ] ; Zheng, RX (Zheng, Ruixue)[ 1,2 ] ; Zhang, JH (Zhang, Jiahui)[ 1,2 ] ; Li, Y (Li, Yan)[ 1,2 ] ; Wang, ZY (Wang, Zhaoyin)[ 1,2 ]*(王兆寅); Dai, ZH (Dai, Zhihui)[ 1,2 ]*(戴志晖)
[ 1 ] Nanjing Normal Univ, Jiangsu Collaborat Innovat Ctr Biomed Funct Mat, Sch Chem & Mat Sci, Nanjing 210023, Peoples R China
[ 2 ] Nanjing Normal Univ, Jiangsu Key Lab Biofunct Mat, Sch Chem & Mat Sci, Nanjing 210023, Peoples R China
[ 3 ] Liaocheng Univ, Dept Chem, Liaocheng 252059, Shandong, Peoples R China
ANALYST,202008,145(15),5174-5179
In this work, a method for quantifying the activity of formamidopyrimidine DNA glucosylase (Fpg) was designed based on phosphate group (P)-modulated multi-enzyme catalysis and fluorescent copper nanoclusters (CuNCs). By eliminating 8-oxoguanine from double-stranded DNA, Fpg generates a nick with P at both 3 ' and 5 ' termini. Subsequently, part of the DNA is digested by 5 ' P-activated lambda exonuclease (lambda Exo), and the generated 3 ' P disables exonuclease I (Exo I), resulting in the generation of single-stranded DNA containing poly(thymine) (poly(T)). Using poly(T) as templates, CuNCs were prepared to emit intense fluorescence as the readout of this method. However, in the absence of Fpg, the originally modified 5 ' P triggers the digestion of lambda Exo. In this case, fluorescence emission is not obtained because CuNCs cannot be formed without DNA templates. Therefore, the catalysis of lambda Exo and Exo I can be tuned by 5 ' P and 3 ' P, which can be further used to determine the activity of Fpg. The fluorescent Fpg biosensor works in a "signal-on" manner with the feature of "zero" background noise, and thus shows desirable analytical features and good performance. Besides, Fpg in serum samples and cell lysate could be accurately detected with the biosensor, indicating the great value of the proposed system in practical and clinical analysis.
文章链接:
https://pubs.rsc.org/en/content/articlelanding/2020/AN/D0AN00928H#!divAbstract
版权与免责声明:本网页的内容由收集互联网上公开发布的信息整理获得。目的在于传递信息及分享,并不意味着赞同其观点或证实其真实性,也不构成其他建议。仅提供交流平台,不为其版权负责。如涉及侵权,请联系我们及时修改或删除。邮箱:sales@allpeptide.com