Electrochemical DNA Sensors Based on MoS2-AuNPs for Polynucleotide Kinase Activity and Inhibition Assay

The determination of T4 polynucleotide kinase (PNK) activity and the screening of PNK inhibitors are critical to disease diagnosis and drug discovery. Numerous electrochemical strategies have been developed for the sensitive measurement of PNK activity and inhibition. However, they often suffer from additional labels and multiple steps of the detection process for the electrochemical readout. Herein, we have demonstrated an electrochemical DNA (E-DNA) sensor for the one-step detection of PNK with "signal-on" readout with no need for additional labels. In our design, the highly switchable double-stranded DNA (dsDNA) probes are immobilized on the gold nanoparticle-decorated molybdenum disulfide nanomaterial (MoS₂-AuNPs), which possesses large surface area and high conductivity for elevating the signal gain in the PNK detection. This signal-on E-DNA sensor integrated with MoS₂-AuNPs exhibits a much higher sensitivity than that without MoS₂-AuNPs, showing a detection limit of 2.18 × 10^-4 U/mL. Furthermore, this assay shows high selectivity, with the ability to discriminate PNK from other enzymes and proteins, and can be utilized to screen inhibitors. The proposed sensor is easy to operate with one-step readout and robust for PNK detection in the biological matrix and shows great potential for point-of-care in clinical diagnostics and drug screening.