Fluorescent copper nanoclusters as a nano-dye for DNA methyltransferase activity analysis and inhibitor screening

Anal Biochem. 2018 Oct 15:559:5-10. doi: 10.1016/j.ab.2018.08.011. Epub 2018 Aug 16.

Abstract

Fluorescent copper nanoslusters (CuNCs) as a new class of fluorophores have attracted more and more attention due to their ease of synthesis, excellent optical properties, and low cost. In this study, a novel label-free fluorescent method was developed for the detection of DNA methyltransferases based on template length-dependent of dsDNA-CuNCs. In the absence of DNA adenine methylation methyltransferase (Dam MTase), the dsDNA containing the methylation-responsive sequence could effectively template the formation of fluorescent CuNCs with bright fluorescence. When the dsDNA substrate is methylated by Dam MTase, the methylation-sensitive restriction endonuclease Dpn I cleaves the methylated dsDNA and produces shorter dsDNA product, which fails to template fluorescent CuNCs. So, the Dam MTase activity could be identified by the changes of CuNCs' fluorescence. Based on this method, a linear range of 0.5-10 U/mL was achieved with high sensitivity and selectivity. Moreover, we also demonstrate the proposed method can be applied to evaluation and screening of inhibitors for Dam MTase.

Keywords: Copper nanoslusters; DNA methyltransferase; Fluorescence; Inhibitor screening.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Copper / chemistry*
  • DNA Modification Methylases / analysis*
  • DNA Modification Methylases / antagonists & inhibitors
  • DNA Modification Methylases / metabolism
  • Drug Evaluation, Preclinical / methods*
  • Enzyme Inhibitors / analysis*
  • Enzyme Inhibitors / pharmacology*
  • Fluorescent Dyes / chemistry*
  • Fluorouracil / analysis
  • Fluorouracil / pharmacology
  • Metal Nanoparticles / chemistry*
  • Spectrometry, Fluorescence

Substances

  • Enzyme Inhibitors
  • Fluorescent Dyes
  • Copper
  • DNA Modification Methylases
  • Fluorouracil