FRET-based method for direct, real-time measurement of DNA methyltransferase activity

Research output: Contribution to journalArticlepeer-review

Colleges, School and Institutes

External organisations

  • Southern University of Science and Technology
  • Université Clermont Auvergne

Abstract

DNA methyltransferase activity is associated with a host of diseases, including cancers, where global hypomethylation of the genome, as well as marked changes in local DNA methylation patterns, can be both diagnostic and prognostic for the disease. Despite this, we currently lack a method for directly measuring the activity of the DNA methyltransferases, which would support the development of DNA methyltransferase-targeted therapies. Here, we demonstrate an assay for the direct measurement of methyltransferase activity, in real time. We employ a fluorescent methyltransferase cofactor analogue, which when bound by the enzyme to a labeled target DNA sequence results in fluorescence resonance energy transfer (FRET) between the donor dye (DNA) and the acceptor dye (cofactor). We demonstrate that the method can be used to monitor the activity of DNA MTases in real time and can be applied to screen inhibitors of the DNA methyltransferases. We show this in both bulk phase and single molecule imaging experiments, highlighting the potential application of the assay in screening and biophysical studies of methyltransferase function.

Bibliographic note

Funding Information: This work was supported by the Engineering and Physical Sciences Research Council (EPSRC, grant number EP/N020901/1). Copyright: © 2020 American Chemical Society

Details

Original languageEnglish
Pages (from-to)192-198
Number of pages7
JournalBioconjugate Chemistry
Volume32
Issue number1
Early online date11 Dec 2020
Publication statusPublished - 20 Jan 2021