title: Sensitive Electrochemical Detection of Human Methyltransferase Based on a Dual Signal Amplification Strategy Coupling Gold Nanoparticle-DNA Complexes with Ru(III) Redox Recycling.
abstract: Effective detection of DNA methyltransferase (DNMT) activity is significant for cancer research. Herein, we developed a sensitive electroanalytical method to detect human DNA (cytosine-5)-methyltransferase 1 (DNMT1) from crude lysates of cancer cells. In this assay, capture DNA having a preferred DNMT1 methylation site was immobilized on a gold electrode and then hybridized with gold nanoparticle (Au NP)-DNA complexes. The modified electrodes were equilibrated with the lysate and then incubated with methylation-sensitive restriction enzyme. If the lysate was negative for DNMT1 activity, the Au NP-DNA complexes would be cut by the restriction enzyme and released from the electrode. Conversely, restriction enzyme cleavage would be blocked by the fully methylated duplexes, and the Au NP-DNA complexes would remain on the electrode. Electroactive Ru(NH3)63+was used as the signal reporter, because of its electrostatic attraction to DNA, resulting in an electrochemical signal. Since the electrochemical signal reflects the amount of Ru(III) redox and the amount of Ru(III) redox is correlated with the activity of DNMT1, the activity of DNMT1 is proportional to the electrochemical signal. The signal could be amplified by the numerous DNAs on the Au NPs and further amplified by Ru(III) redox recycling. With this method, a detection limit down to 0.3 U/mL for pure DNMT1 and 8 MCF-7 cells was achieved. DNMT1 activities of different cell lines were also successfully evaluated.
authors.affiliation: Jiangsu Key Laboratory of New Power Batteries, Jiangsu Collaborative Innovation Center of Biomedical Functional Materials, Jiangsu Key Laboratory of Biomedical Materials, National and Local Joint Engineering Research Center of Biomedical Functional Materials, College of Chemistry and Materials Science, Nanjing Normal University , Nanjing, Jiangsu 210097, P. R. China.
authors.forename: Hui
authors.initials: H
authors.lastname: Zhang
authors.affiliation: Jiangsu Key Laboratory of New Power Batteries, Jiangsu Collaborative Innovation Center of Biomedical Functional Materials, Jiangsu Key Laboratory of Biomedical Materials, National and Local Joint Engineering Research Center of Biomedical Functional Materials, College of Chemistry and Materials Science, Nanjing Normal University , Nanjing, Jiangsu 210097, P. R. China.
authors.forename: Huilei
authors.initials: H
authors.lastname: Dong
authors.affiliation: Jiangsu Key Laboratory of New Power Batteries, Jiangsu Collaborative Innovation Center of Biomedical Functional Materials, Jiangsu Key Laboratory of Biomedical Materials, National and Local Joint Engineering Research Center of Biomedical Functional Materials, College of Chemistry and Materials Science, Nanjing Normal University , Nanjing, Jiangsu 210097, P. R. China.
authors.forename: Guoqing
authors.initials: G
authors.lastname: Yang
authors.affiliation: Jiangsu Key Laboratory of New Power Batteries, Jiangsu Collaborative Innovation Center of Biomedical Functional Materials, Jiangsu Key Laboratory of Biomedical Materials, National and Local Joint Engineering Research Center of Biomedical Functional Materials, College of Chemistry and Materials Science, Nanjing Normal University , Nanjing, Jiangsu 210097, P. R. China.
authors.forename: Hongfei
authors.initials: H
authors.lastname: Chen
authors.affiliation: Jiangsu Key Laboratory of New Power Batteries, Jiangsu Collaborative Innovation Center of Biomedical Functional Materials, Jiangsu Key Laboratory of Biomedical Materials, National and Local Joint Engineering Research Center of Biomedical Functional Materials, College of Chemistry and Materials Science, Nanjing Normal University , Nanjing, Jiangsu 210097, P. R. China.
authors.forename: Chenxin
authors.initials: C
authors.lastname: Cai
mesh_terms: D015374:Biosensing Techniques; D002478:Cells, Cultured; D004247:DNA / Q000737:chemistry*; D000076123:DNA (Cytosine-5-)-Methyltransferase 1 / Q000032:analysis* / Q000378:metabolism*; D055664:Electrochemical Techniques; D006046:Gold / Q000737:chemistry*; D006801:Humans; D053768:Metal Nanoparticles / Q000737:chemistry*; D046529:Microscopy, Electron, Transmission; D009693:Nucleic Acid Hybridization; D010084:Oxidation-Reduction; D017975:Ruthenium Compounds / Q000737:chemistry*
chemical_list: D017975:Ruthenium Compounds; D006046:Gold; D004247:DNA; D000076123:DNA (Cytosine-5-)-Methyltransferase 1; C000619380:DNMT1 protein, human