Renewable DNA tetrahedron Interface enabling ultrasensitive detection of copper via synergetic enhancement of click chemistry and DNAzyme catalysis.

Copper is an essential trace element that plays critical roles in numerous physiological processes, including mitochondrial respiration, antioxidant defense, and neurotransmitter biosynthesis. However, an imbalance in copper homeostasis can lead to severe health disorders. Copper deficiency is linked to diseases such as anemia and neutropenia, while copper overload is associated with neurodegenerative diseases like Wilson's disease and Alzheimer's disease. The sensing performance of copper-mediated click chemistry is hindered by poor nucleic acid ligation efficiency and the difficulty in removing ligation products. To overcome these issues, we developed a renewable framework nucleic acid sensing interface for the ultrasensitive detection of copper ions, leveraging the synergistic effects of click chemistry and DNAzyme catalysis. For the first time, we found that the tetrahedron DNA nanostructure (TDN) could enhance the G-quadruplex/Hemin complex catalysis activity in a noncovalent assembled fashion. This enhancement benefits from the negative charge microenvironment of the TDN skeleton, which improve the binding affinity of DNAzyme toward the positive charge substrate. Accompanying with increasing the local concentration of azide group modified strands, and improving accessibility of that, a high ligation efficiency of split G-rich sequence was implemented on the TDN scaffold via click-chemistry. This synergetic enhancement of click chemistry and DNAzyme catalysis enables ultrasensitive detection of copper ions, and the limit of detection was 28.1 pM, which is 200 times lower than that without TDN manner. More importantly, the click chemistry product can be removed by breaking Hoogsteen hydrogen bond in an alkaline condition, enabling a renewable sensing interface. Furthermore, this approach was also succeed applied for the alkaline phosphatase activity analysis. This work further extended the functional of TDN and provide a reference for the construction of TDN-based multifunctional sensing interface.