Magnetic bead-liposome hybrids enable sensitive and portable detection of DNA methyltransferase activity using personal glucose meter.
DNA methyltransferase (MTase) plays a critical role in maintaining genome methylation patterns, which has a close relationship to cancer and bacterial diseases. This encouraged the need to develop highly sensitive, simple, and robust assays for DNA MTase detection and inhibitor screening. Herein, a simple, sensitive, and specific DNA MTase activity assay was developed based on magnetic beads-liposome hybrids combined with personal glucose meter (PGM) for quantitative detection of DNA MTase and inhibitor screening. First, a magnetic beads-liposome hybrid probe is designed by the hybridization of p1DNA-functionalized magnetic bead with p2DNA-functionalized glucoamylase-encapsulated liposome (GEL). It integrates target recognition, magnetic separation and signal amplification within one multifunctional design. Then, in the presence of Dam MTase, the hybrids probe was methylated, and cleaved by methylation-sensitive restriction endonuclease Dpn I, making liposome separated from magnetic bead by magnetic separation. Finally, the separated liposome was decomposed, liberating the encapsulated glucoamylase to catalyze the hydrolysis of the signal substrate amylose with multiple turnovers, producing a large amount of glucose for quantitative readout by the PGM. In the proposed assay, the magnetic beads-liposome hybrids offered excellent sensitivity due to primary amplification via releasing numerous glucoamylase from a liposome followed by a secondary enzymatic amplification. The use of portable quantitative device PGM bypasses the requirement of complicated instruments and sophisticated operations, making the method simple and feasible for on-site detection. Moreover, the proposed assay was successfully applied in complex biological matrix and screen suitable inhibitor drugs for DAM for disease(s) treatment. The results reveal that the approach provides a simple, sensitive, and robust platform for DNA MTases detection and screening potential drugs in medical research and early clinical diagnostics.
Department of Chemistry, Liaocheng University, Liaocheng, Shandong 252059, China.
Youna
Y
Zhang
Department of Chemistry, Liaocheng University, Liaocheng, Shandong 252059, China. Electronic address: xueqingwang1983@163.com.
Qingwang
Q
Xue
Key Laboratory of Food Nutrition and Safety, Ministry of Education of China, Tianjin University of Science and Technology, Tianjin 300457, China. Electronic address: liujifeng111@gmail.com.
Jifeng
J
Liu
Department of Chemistry, Liaocheng University, Liaocheng, Shandong 252059, China. Electronic address: hswang@lcu.edu.cn.
Huaisheng
H
Wang
D000688:Amylose / Q000378:metabolism; D015374:Biosensing Techniques / Q000379:methods*; D015190:Blood Glucose Self-Monitoring / Q000379:methods*; D004353:Drug Evaluation, Preclinical / Q000379:methods; D004800:Enzymes, Immobilized / Q000378:metabolism; D005087:Glucan 1,4-alpha-Glucosidase / Q000378:metabolism; D005947:Glucose / Q000032:analysis / Q000378:metabolism; D006801:Humans; D008081:Liposomes / Q000737:chemistry*; D059346:Magnets / Q000737:chemistry*; D015265:Site-Specific DNA-Methyltransferase (Adenine-Specific) / Q000037:antagonists & inhibitors / Q000097:blood* / Q000378:metabolism*
D004800:Enzymes, Immobilized; D008081:Liposomes; D000688:Amylose; C058693:Dam methyltransferase; D015265:Site-Specific DNA-Methyltransferase (Adenine-Specific); D005087:Glucan 1,4-alpha-Glucosidase; D005947:Glucose
DNA methyltransferase; Enzymatic amplification; Magnetic beads-liposome hybrids; PGM