Identification of Novel Bacterial M.SssI DNA Methyltransferase Inhibitors

Abstract

DNA methylation is an important epigenetic regulator of gene expression. Abnormalities in DNA methylation patterns have been associated with various developmental and proliferative diseases, particularly cancer. Targeting DNA methyltransferases (DNMTs) represents a promising strategy for the treatment of such diseases. Current DNMT inhibitors suffer important drawbacks with respect to their efficacy, specificity, and toxicity. In this study, we have set up a robust in vitro bacterial M.SssI DNMT activity assay to systematically screen a collection of 26 240 compounds that were predicted to compete with the S-adenosyl-L-methionine (SAM) substrate of DNMT. This resulted in the identification of a novel set of structurally distinct inhibitors of M.SssI DNMT activity. Although molecular docking studies using an M.SssI homology model suggest that these compounds might compete with SAM binding, mode of activity (MoA) assays are still needed to confirm this hypothesis. Our set of novel M.SssI DNMT inhibitors, once confirmed in an orthogonal DNMT assay, may thus serve as a starting point to identify and characterize suitable lead candidates for further drug optimization.

In Vitro and In Vivo Enzyme Activity Screening via RNA-Based Fluorescent Biosensors for S-Adenosyl-l-homocysteine (SAH)

“…As shown, enzymatic reactions were performed, then a fixed aliquot of the reaction mixture was diluted into the assay reaction containing the RNA biosensor, DFHBI, and buffer. The bacterial DNA MTase M.SssI possesses robust enzymatic activity and shares structural similarities with human DNA MTase 1 (DNMT1), an important epigenetic regulator and drug target. , With the use of any of the fluorescent biosensors, M.SssI activity was detected as an increase in fluorescence only in the presence of both SAM and dsDNA substrate (Figure S11). To determine the generality of the strategy, we also tested this assay on a histone protein MTase.…”

Bacterial Epigenomics: Coming of Age

“…While some selective inhibitors of Dam were previously proposed ( 22 ), there have been no further advances over the past decade. Since camA and dam are part of a much larger list of 145 genes recently reported in a study investigating highly conserved MTases in bacteria ( 14 ), I foresee a renewed interest in the exploitation of such targets for the development of next-generation epigenetic drugs ( 23 – 25 ).…”