title: A methylation-blocked cascade amplification strategy for label-free colorimetric detection of DNA methyltransferase activity.
abstract: DNA methyltransferase (MTase), catalyzing DNA methylation in both eukaryotes and prokaryotes, is closely related with cancer and bacterial diseases. Although there are various methods focusing on DNA MTase detection, most of them share common defects such as complicated setup, laborious operation and requirement of expensive analytical instruments. In this work, a simple strategy based on methylation-blocked cascade amplification is developed for label-free colorimetric assay of MTase activity. When DNA adenine methylation (Dam) MTase is introduced, the hairpin probe is methylated. This blocks the amplified generation of G-riched DNAzyme by nicking endonuclease and DNA polymerase, and inhibits the DNAzyme-catalyzed colorimetric reaction. Contrarily, an effective colorimetric reaction is initiated and high color signal is clearly observed by the naked eye in the absence of Dam MTase. A satisfying sensitivity and high selectivity are readily achieved within a short assay time of 77 min, which are superior to those of some existing approaches. Additionally, the application of the sensing system in human serum is successfully verified with good recovery and reproducibility, indicating great potential for the practicality in high concentrations of interfering species. By using several anticancer and antimicrobial drugs as model, the inhibition of Dam MTase is well investigated. Therefore, the proposed method is not only promising and convenient in visualized analysis of MTase, but also useful for further application in fundamental biological research, early clinical diagnosis and drug discovery.
authors: Key Laboratory of Biomedical Information Engineering of Education Ministry, School of Life Science and Technology, Xi'an Jiaotong University, Xi'an, Shaanxi 710049, PR China; Division of Physical Biology, and Bioimaging Center, Shanghai Synchrotron Radiation Facility, CAS Key Laboraotory of Interfacial Physics and Technology, Shanghai Institute of Applied Physics, Chinese Academy of Sciences, Shanghai 201800, PR China. Electronic address: yxzhao@mail.xjtu.edu.cn.
authors: Yongxi
authors: Y
authors: Zhao
authors: Key Laboratory of Biomedical Information Engineering of Education Ministry, School of Life Science and Technology, Xi'an Jiaotong University, Xi'an, Shaanxi 710049, PR China.
authors: Feng
authors: F
authors: Chen
authors: Key Laboratory of Biomedical Information Engineering of Education Ministry, School of Life Science and Technology, Xi'an Jiaotong University, Xi'an, Shaanxi 710049, PR China.
authors: Manli
authors: M
authors: Lin
authors: Division of Physical Biology, and Bioimaging Center, Shanghai Synchrotron Radiation Facility, CAS Key Laboraotory of Interfacial Physics and Technology, Shanghai Institute of Applied Physics, Chinese Academy of Sciences, Shanghai 201800, PR China. Electronic address: fchh@sinap.ac.cn.
authors: Chunhai
authors: C
authors: Fan
mesh_terms: D015374:Biosensing Techniques / Q000379:methods*; D003124:Colorimetry / Q000379:methods; D019175:DNA Methylation; D015342:DNA Probes / Q000378:metabolism; D021881:DNA, Catalytic / Q000378:metabolism; D057075:Enzyme Assays / Q000379:methods*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D015265:Site-Specific DNA-Methyltransferase (Adenine-Specific) / Q000037:antagonists & inhibitors / Q000097:blood* / Q000378:metabolism*
chemical_list: D015342:DNA Probes; D021881:DNA, Catalytic; D004791:Enzyme Inhibitors; D015265:Site-Specific DNA-Methyltransferase (Adenine-Specific)
keywords: DNA methyltransferase; DNAzyme; Inhibitor; Label-free colorimetric detection; Methylation-blocked cascade amplification