Issue 15, 2022

Molecular beacons with oxidized bases report on substrate specificity of DNA oxoguanine glycosylases

Abstract

DNA glycosylase enzymes recognize and remove structurally distinct modified forms of DNA bases, thereby repairing genomic DNA from chemically induced damage or erasing epigenetic marks. However, these enzymes are often promiscuous, and advanced tools are needed to evaluate and engineer their substrate specificity. Thus, in the present study, we developed a new strategy to rapidly profile the substrate specificity of 8-oxoguanine glycosylases, which cleave biologically relevant oxidized forms of guanine. We monitored the enzymatic excision of fluorophore-labeled oligonucleotides containing synthetic modifications 8-oxoG and FapyG, or G. Using this molecular beacon approach, we identified several hOGG1 mutants with higher specificity for FapyG than 8-oxoG. This approach and the newly synthesized probes will be useful for the characterization of glycosylase substrate specificity and damage excision mechanisms, as well as for evaluating engineered enzymes with altered reactivities.

Graphical abstract: Molecular beacons with oxidized bases report on substrate specificity of DNA oxoguanine glycosylases

Supplementary files

Article information

Article type
Edge Article
Submitted
13 Oct 2021
Accepted
15 Feb 2022
First published
16 Feb 2022
This article is Open Access

All publication charges for this article have been paid for by the Royal Society of Chemistry
Creative Commons BY license

Chem. Sci., 2022,13, 4295-4302

Molecular beacons with oxidized bases report on substrate specificity of DNA oxoguanine glycosylases

J. Sun, N. M. Antczak, H. L. Gahlon and S. J. Sturla, Chem. Sci., 2022, 13, 4295 DOI: 10.1039/D1SC05648D

This article is licensed under a Creative Commons Attribution 3.0 Unported Licence. You can use material from this article in other publications without requesting further permissions from the RSC, provided that the correct acknowledgement is given.

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