title: One-pot synthesis of GO/AgNPs/luminol composites with electrochemiluminescence activity for sensitive detection of DNA methyltransferase activity.
abstract: DNA methyltransferases catalyze the transfer of a methyl group from S-adenosylmethionine to the target adenine or cytosine, eventually inducing the DNA methylation in both prokaryotes and eukaryotes. Herein, we developed a novel electrochemiluminescence biosensor to quantify DNA adenine methylation (Dam) methyltransferase (MTase) employing signal amplification of GO/AgNPs/luminol composites to enhance the assay sensitivity. The method was developed by designing a capture probe DNA, which was immobilized on gold electrode surface, to hybridize with azide complementary DNA to form the azide-terminated dsDNA. Then, alkynyl functionalized GO/AgNPs/luminol composites as the signal probe were immobilized to azide-terminated dsDNA modified electrode via click chemistry, resulting in a high electrochemiluminescence (ECL) signal. Once the DNA hybrid was methylated (under catalysis of Dam MTase) and further cleaved by Dpn I endonuclease (a site-specific endonuclease recognizing the duplex symmetrical sequence of 5'-G-Am-T-C-3'), GO/AgNPs/luminol composites release from the electrode surface to the solution, leading to significant reduction of the ECL signal. The change of the ECL intensity is related to the methylation status and MTase activity, which forms the basis of MTase activity assay and site-specific methylation determination. This novel strategy can be further used as a universal method for other transferase determination by designing various transferase-specific DNA sequences. In addition, this method can be used for the screening of antimicrobial drugs and has a great potential to be further applied in early clinical diagnosis.
authors.affiliation: Department of Chemistry, Nanchang University, Nanchang 330031, PR China.
authors.forename: Hui-Fang
authors.initials: HF
authors.lastname: Zhao
authors.affiliation: Department of Chemistry, Nanchang University, Nanchang 330031, PR China.
authors.forename: Ru-Ping
authors.initials: RP
authors.lastname: Liang
authors.affiliation: Department of Chemistry, Nanchang University, Nanchang 330031, PR China.
authors.forename: Jing-Wu
authors.initials: JW
authors.lastname: Wang
authors.affiliation: Department of Chemistry, Nanchang University, Nanchang 330031, PR China. Electronic address: jdqiu@ncu.edu.cn.
authors.forename: Jian-Ding
authors.initials: JD
authors.lastname: Qiu
mesh_terms: D015374:Biosensing Techniques / Q000295:instrumentation; D003217:Conductometry / Q000295:instrumentation*; D004789:Enzyme Activation; D004867:Equipment Design; D019544:Equipment Failure Analysis; D006108:Graphite / Q000737:chemistry; D008163:Luminescent Measurements / Q000295:instrumentation*; D008165:Luminol / Q000737:chemistry*; D053768:Metal Nanoparticles / Q000737:chemistry* / Q000648:ultrastructure; D053761:Nanocomposites / Q000737:chemistry / Q000648:ultrastructure; D010087:Oxides / Q000737:chemistry; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D012834:Silver / Q000737:chemistry*; D015265:Site-Specific DNA-Methyltransferase (Adenine-Specific) / Q000032:analysis* / Q000737:chemistry
chemical_list: D010087:Oxides; D012834:Silver; D008165:Luminol; D006108:Graphite; C058693:Dam methyltransferase; D015265:Site-Specific DNA-Methyltransferase (Adenine-Specific)
keywords: Biosensor; Click chemistry; Dam MTase; ECL; GO/AgNPs/luminol