Simple and Fast DNA Based Sensor System for Screening of Small-Molecule Compounds Targeting Eukaryotic Topoisomerase 1
Ikusi/ Ireki
Data
2021-08-13Egilea
Petersen, Kamilla Vandsø
Selas Lanseros, Asier
Hymøller, Kirstine Mejlstrup
Mizielinski, Karol
Thorsager, Maria
Stougaard, Magnus
Pérez-Pertejo, Yolanda
Reguera, Rosa M.
Balaña-Fouce, Rafael
Knudsen, Birgitta Ruth
Tesauro, Cinzia
Pharmaceutics 13(8) : (2021) // Article ID 1255
Laburpena
Background: Eukaryotic topoisomerase 1 is a potential target of anti-parasitic and anti-cancer drugs. Parasites require topoisomerase 1 activity for survival and, consequently, compounds that inhibit topoisomerase 1 activity may be of interest. All effective topoisomerase 1 drugs with anti-cancer activity act by inhibiting the ligation reaction of the enzyme. Screening for topoisomerase 1 targeting drugs, therefore, should involve the possibility of dissecting which step of topoisomerase 1 activity is affected. Methods: Here we present a novel DNA-based assay that allows for screening of the effect of small-molecule compounds targeting the binding/cleavage or the ligation steps of topoisomerase 1 catalysis. This novel assay is based on the detection of a rolling circle amplification product generated from a DNA circle resulting from topoisomerase 1 activity. Results: We show that the binding/cleavage and ligation reactions of topoisomerase 1 can be investigated separately in the presented assay termed REEAD (C|L) and demonstrate that the assay can be used to investigate, which of the individual steps of topoisomerase 1 catalysis are affected by small-molecule compounds. The assay is gel-free and the results can be detected by a simple colorimetric readout method using silver-on-gold precipitation rendering large equipment unnecessary. Conclusion: REEAD (C|L) allows for easy and quantitative investigations of topoisomerase 1 targeting compounds and can be performed in non-specialized laboratories.
Collections
Bestelakorik adierazi ezean, itemaren baimena horrela deskribatzen da:2021 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).