Dynamical Regulation of Enzyme Cascade Amplification by a Regenerated DNA Nanotweezer for Ultrasensitive Electrochemical DNA Detection

摘要

Traditional scaffolds such as metal nanoparticles and DNA origami remain a considerable challenge to regulate the enzyme cascade catalytic efficiency dynamically and reversibly on account of their irreversible conformation. To address these issues, a regenerated DNA tweezer was designed to dynamically regulate the interenzyme spacing for high-efficiency enzyme cascade amplification for homogeneous determination of target DNA related to cancer diseases. Initially, the enzyme-functionalized DNA tweezer was maintained at the opened state with a relatively distant interenzyme distance (19–24 nm), leading to a low catalytic efficiency. Benefiting from target induced Mg~(2+)-dependent DNAzyme cleavage recycling, the one input target could be transduced to multiple corresponding methylene blue (MB) labeled DNA (S5), which served not only as the signal probe to provide a detectable electrochemical signal but also fuel to switch the DNA tweezer from the opened to closed state, leading to cascaded enzymes close enough (5–10 nm) for enhancing the catalytic efficiency for sensitive target DNA analysis with a low detection limit down to 30 fM. In the presence of antifuels, the closed DNA tweezer easily switched back to the opened state via a one-step strand displacement, and the obtained DNA tweezer achieved regeneration for subsequently recycling target detection. With the dynamical regulation of interenzyme distance in an “open–close–open” way, the enzyme cascade catalytic efficiency became dynamically controllable, and the DNA tweezer realized simple reutilization over five times, overcoming the drawbacks of inflexible, time-consuming operation and false positive signal induced by traditional scaffolds. More importantly, this method opened a new avenue for employing the arbitrary change of enzyme cascade catalytic efficiency for sensitive detection various biomolecules.