title: DNA methyltransferase activity detection based on fluorescent silver nanocluster hairpin-shaped DNA probe with 5'-C-rich/G-rich-3' tails.
abstract: DNA methylation has received a large amount of attention due to its close relationship to a wide range of biological phenomena, such as gene activation, gene imprinting, and chromatin stability. Herein, we have designed a hairpin-shaped DNA probe with 5'-C-rich/G-rich-3' tails and developed a simple and reliable fluorescence turn-off assay for DNA adenine methylation (Dam) methyltransferase (MTase) detection combining site recognition and the fluorescence enhancement of DNA-templated silver nanoclusters (DNA-AgNCs) by guanine-rich DNA sequences. A designed hairpin probe with 5' CCCTTACCCC and 3' GGGTGGGGTGGGGTGGGG displays a bright red emission after reacting with AgNO3 and NaBH4. In the presence of Dam MTase, the methylation-sensitive restriction endonuclease Dpn I which has the same recognition site with the Dam MTase can split the probe, freeing the G-rich sequence from the C-rich sequence, thus quenching the fluorescence of DNA-AgNCs. Compared to traditional fluorescent-based methods, this strategy is simple and inexpensive. A linear response to concentrations of Dam MTase which range from 1 U/mL to 100 U/mL and a detection limit of 1 U/mL are obtained without any amplification steps. In addition, we also demonstrate the method can be used for evaluation and screening of inhibitors for Dam MTase.
authors: College of Chemistry and Molecular Sciences, Key Laboratory of Biomedical Polymers of Ministry of Education, Wuhan University, Wuhan, Hubei 430072, PR China.
authors: Wenting
authors: W
authors: Liu
authors: College of Chemistry and Molecular Sciences, Key Laboratory of Biomedical Polymers of Ministry of Education, Wuhan University, Wuhan, Hubei 430072, PR China.
authors: Han
authors: H
authors: Lai
authors: College of Chemistry and Molecular Sciences, Key Laboratory of Biomedical Polymers of Ministry of Education, Wuhan University, Wuhan, Hubei 430072, PR China.
authors: Rong
authors: R
authors: Huang
authors: College of Chemistry and Molecular Sciences, Key Laboratory of Biomedical Polymers of Ministry of Education, Wuhan University, Wuhan, Hubei 430072, PR China.
authors: Chuntao
authors: C
authors: Zhao
authors: College of Chemistry and Molecular Sciences, Key Laboratory of Biomedical Polymers of Ministry of Education, Wuhan University, Wuhan, Hubei 430072, PR China.
authors: Yimo
authors: Y
authors: Wang
authors: College of Chemistry and Molecular Sciences, Key Laboratory of Biomedical Polymers of Ministry of Education, Wuhan University, Wuhan, Hubei 430072, PR China. Electronic address: xcweng@whu.edu.cn.
authors: Xiaocheng
authors: X
authors: Weng
authors: College of Chemistry and Molecular Sciences, Key Laboratory of Biomedical Polymers of Ministry of Education, Wuhan University, Wuhan, Hubei 430072, PR China; State Key Laboratory of Natural and Biomimetic Drugs, Peking University, PR China. Electronic address: xzhou@whu.edu.cn.
authors: Xiang
authors: X
authors: Zhou
mesh_terms: D015374:Biosensing Techniques; D004247:DNA / Q000737:chemistry*; D019175:DNA Methylation; D015342:DNA Probes / Q000737:chemistry; D005456:Fluorescent Dyes / Q000737:chemistry; D057230:Limit of Detection; D008780:Methyltransferases / Q000737:chemistry / Q000302:isolation & purification*; D012834:Silver / Q000737:chemistry; D015265:Site-Specific DNA-Methyltransferase (Adenine-Specific)
chemical_list: D015342:DNA Probes; D005456:Fluorescent Dyes; D012834:Silver; D004247:DNA; D008780:Methyltransferases; D015265:Site-Specific DNA-Methyltransferase (Adenine-Specific)
keywords: DNA methyltransferase; Dam MTase; Fluorescence assay; Silver nanoclusters (AgNCs)