Ultrastable Bimolecular G-Quadruplexes Programmed DNA Nanoassemblies for Reconfigurable Biomimetic DNAzymes

The relatively low stability and polymorphism of bimolecular G-quadruplexes (bi-G4s) are big difficulties that are faced in employing them to guide DNA assembly, as they are usually subject to a transformation into more stable tetramolecular or G-wire structures favored by K⁺ or Mg²⁺. Although bi-G4s benefit by additional duplex handles, a challenge remains in tailoring their intrinsic properties to resolve the above difficulties. Toward this challenge, here we engineer several ultrastable bi-G4s via replacing their nucleotide loops with special mini-hairpins, which consist of a GAA loop and a short GC-paired stem. Such a structural alteration favors the formation of G:C:G:C tetrads in the head-to-head folding topologies of bi-G4s and improves their thermal stability, with an increase in the melting temperature by up to 25 °C. It dramatically reduces their structural conversion into G-wires, verified by atomic force microscopy. These features enable the utilization of two well-chosen bi-G4s to shape a DNA nanotriangle into the desired framework nucleic acid (FNA) architectures such as "bowknot" and "butterfly" that are reversibly switched by the bi-G4s. On this basis, we further build a reconfigurable DNAzyme device to mimic the activation of human telomerase that is modulated by the G4 dimerization. Our designed ultrastable bi-G4s will offer a promising tool for dynamically manipulating intracellular DNA nanoassemblies with endogenous K⁺ and exploring the relationship between dimerization and function in some physiological processes.