Spectroscopic sensing and quantification of AP-endonucleases using fluorescence-enhancement by cis–trans isomerization of cyanine dyes
| Autor: | Jungmin Yoo, Gwangrog Lee, Sanghoon Oh, JunHo Cho, Jae Won Han, Donghun Lee, Daeho Park |
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| Rok vydání: | 2021 |
| Předmět: |
0303 health sciences
Nuclease biology DNA repair General Chemical Engineering General Chemistry Fluorescence Cis trans isomerization 03 medical and health sciences chemistry.chemical_compound 0302 clinical medicine chemistry 030220 oncology & carcinogenesis biology.protein Biophysics AP site Cyanine Biosensor DNA 030304 developmental biology |
| Zdroj: | RSC Advances. 11:11380-11386 |
| ISSN: | 2046-2069 |
| DOI: | 10.1039/d0ra08051a |
| Popis: | Apurinic/apyrimidinic (AP) endonucleases are vital DNA repair enzymes, and proposed to be a prognostic biomarker for various types of cancer in humans. Numerous DNA sensors have been developed to evaluate the extent of nuclease activity but their DNA termini are not protected against other nucleases, hampering accurate quantification. Here we developed a new fluorescence enhancement (FE)-based method as an enzyme-specific DNA biosensor with nuclease-protection by three functional units (an AP-site, Cy3 and termini that are protected from exonucleolytic cleavage). A robust FE signal arises from the fluorescent cis–trans isomerization of a cyanine dye (e.g., Cy3) upon the enzyme-triggered structural change from double-stranded (ds)DNA to single-stranded (ss)DNA that carries Cy3. The FE-based assay reveals a linear dependency on sub-nanomolar concentrations as low as 10−11 M for the target enzyme and can be also utilized as a sensitive readout of other nuclease activities. |
| Databáze: | OpenAIRE |
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