title: Quencher-free hairpin probes for real-time detection of T4 polynucleotide kinase activity.
abstract: Traditional methods of assaying polynucleotide kinase (PNK) activity are discontinuous, time-consuming, and laborious. Here we report a new quencher-free approach to real-time monitoring of PNK activity using a 2-aminopurine probe. When the 2-aminopurine probe was 5'-phosphorylated by PNK, it could be efficiently degraded by lambda exonuclease to release free 2-aminopurine molecules and generate a fluorescence signal. This method not only provides a universal approach to real-time monitoring of PNK activity, but also shows great potential for screening suitable inhibitor drugs for PNK.
authors.affiliation: State Key Laboratory of Medical Genetics & School of Life Sciences, Central South University, Changsha 410013, China; State Key Laboratory of Chemo/Biosensing and Chemometrics, Hunan University, Changsha 410081, China. Electronic address: macb2012@csu.edu.cn.
authors.forename: Changbei
authors.initials: C
authors.lastname: Ma
authors.affiliation: State Key Laboratory of Medical Genetics & School of Life Sciences, Central South University, Changsha 410013, China.
authors.forename: Haisheng
authors.initials: H
authors.lastname: Liu
authors.affiliation: State Key Laboratory of Medical Genetics & School of Life Sciences, Central South University, Changsha 410013, China.
authors.forename: Junyan
authors.initials: J
authors.lastname: Du
authors.affiliation: State Key Laboratory of Medical Genetics & School of Life Sciences, Central South University, Changsha 410013, China.
authors.forename: Hanchun
authors.initials: H
authors.lastname: Chen
authors.affiliation: State Key Laboratory of Medical Genetics & School of Life Sciences, Central South University, Changsha 410013, China.
authors.forename: Hailun
authors.initials: H
authors.lastname: He
authors.affiliation: State Key Laboratory of Medical Genetics & School of Life Sciences, Central South University, Changsha 410013, China.
authors.forename: Shunxin
authors.initials: S
authors.lastname: Jin
authors.affiliation: State Key Laboratory of Chemo/Biosensing and Chemometrics, Hunan University, Changsha 410081, China.
authors.forename: Kemin
authors.initials: K
authors.lastname: Wang
authors.affiliation: State Key Laboratory of Medical Genetics & School of Life Sciences, Central South University, Changsha 410013, China.
authors.forename: Jun
authors.initials: J
authors.lastname: Wang
mesh_terms: D015075:2-Aminopurine / Q000378:metabolism; D017122:Bacteriophage T4 / Q000201:enzymology*; D010582:Bacteriophage lambda / Q000201:enzymology; D005092:Exonucleases / Q000378:metabolism; D010766:Phosphorylation; D011116:Polynucleotide 5'-Hydroxyl-Kinase / Q000032:analysis* / Q000378:metabolism; D013050:Spectrometry, Fluorescence
chemical_list: D015075:2-Aminopurine; D011116:Polynucleotide 5'-Hydroxyl-Kinase; D005092:Exonucleases
keywords: 2-Aminopurine; Inhibitor; Polynucleotide kinase activity; Quencher-free