title: Simultaneous detection of kinase and phosphatase activities of polynucleotide kinase using molecular beacon probes.
abstract: Phosphorylation and dephosphorylation of DNA by polynucleotide kinase (PNK) has an important role in DNA damage repair, replication, and recombination. Traditionally, it is assayed by denaturing gel electrophoresis and autoradiography, which are tedious and not sensitive. We report on the development of a sensitive and simple method for PNK assay based on DNA ligation using a molecular beacon. Enzyme activity of PNK is measured down to a limit of 0.002 unit/ml. The method not only provides a universal platform for simultaneous monitoring of kinase and phosphatase activities, but also shows great potential in biological research, drug discovery, and clinical diagnostics.
authors.affiliation: School of Life Sciences, The State Key Labratory of Medical Genetics, Central South University, Changsha 410013, China. Electronic address: macb2012@csu.edu.cn.
authors.forename: Changbei
authors.initials: C
authors.lastname: Ma
authors.forename: Hefei
authors.initials: H
authors.lastname: Fang
authors.forename: Kemin
authors.initials: K
authors.lastname: Wang
authors.forename: Kun
authors.initials: K
authors.lastname: Xia
authors.forename: Hanchun
authors.initials: H
authors.lastname: Chen
authors.forename: Hailun
authors.initials: H
authors.lastname: He
authors.forename: Weimin
authors.initials: W
authors.lastname: Zeng
mesh_terms: D017122:Bacteriophage T4 / Q000201:enzymology*; D004247:DNA / Q000378:metabolism; D057075:Enzyme Assays / Q000379:methods*; D005453:Fluorescence; D015335:Molecular Probes / Q000378:metabolism*; D010744:Phosphoric Monoester Hydrolases / Q000378:metabolism; D010766:Phosphorylation; D011116:Polynucleotide 5'-Hydroxyl-Kinase / Q000378:metabolism*; D013050:Spectrometry, Fluorescence / Q000379:methods
chemical_list: D015335:Molecular Probes; D004247:DNA; D011116:Polynucleotide 5'-Hydroxyl-Kinase; D010744:Phosphoric Monoester Hydrolases
keywords: Kinase and phosphatase activities; Molecular beacon; Polynucleotide kinase