A bioluminescence assay for DNA methyltransferase activity based on methylation-resistant cleavage.

DNA methyltransferase (MTase) is a kind of important regulatory factor in various biological processes. Current methods to investigate DNA MTase activity are still limited in the sensitivity and/or generality. Therefore, developing methods with high sensitivity and improved generality is needed. Here, we develop a new bioluminescence strategy based on methylation-resistant cleavage and protein expression in vitro to detect DNA MTase activity. In the strategy, Dam MTase was used as a model enzyme and MboI as the methylation-resistant endonuclease, and luciferase reporter DNA (LR-DNA) was used as their action target. Because the completely methylated LR-DNA could be expressed as detectable luciferase, Dam MTase activity was quantified by measuring the luminescence intensity of the expressed luciferase. The assay provides a very low detection limit (0.08 U/ml) as well as a wide linear range (0.2-100 U/ml). Besides, the analysis mode has improved generality and could be extended to the detection of other DNA MTases and the corresponding inhibitor screening. Copyright Â