Label-free colorimetric assay for T4 polynucleotide kinase/phosphatase activity and its inhibitors based on G-quadruplex/hemin DNAzyme

Abstract

Here we report a new approach for label-free colorimetric assay of T4 polynucleotide kinase/phosphatase (PNKP) activity based on G-quadruplex/hemin DNAzyme. In the presence of T4 PNKP, the DNA primer with a 3'-phosphate can be dephosphorylated into a 3'-hydroxyl and initiate a primer elongation reaction to open the hairpin probe, and leading to releasing the G-quartets. Under optimal conditions, the proposed method exhibited a considerable performance with a detection limit of 0.01 U/mL. Furthermore, the present assay can be used to study the potential T4 PNKP inhibitor screening, making it promise to be applied in the fields of drug discovery.

Keywords: Colorimetric assay; G-quadruplex/hemin DNAzyme; T4 polynucleotide kinase/phosphatase.