Three-Dimensional Nucleic-Acid-Based Constitutional Dynamic Networks: Enhancing Diversity through Complexity of the Systems

Inspired by nature, where gene regulatory networks consisting of intercommunicating constituents, each composed of three or more components, play a central role in the development of living systems, we use the information encoded in the base sequences of nucleic acids to construct three-dimensional constitutional dynamic networks (3D CDNs) consisting of eight three-component constituents (A_iB_jC_k). Upon subjecting the parent 3D CDN I to four auxiliary nucleic acid triggers (T₁, T₂, T₃, or T₄), the adaptive reconfiguration of CDN I into four different CDNs (II, III, IV, or V) is demonstrated, and by applying two consecutive triggers or counter triggers, the adaptive reversible hierarchical control over the compositions of new CDN systems (VI, VII, VIII, or IX) is demonstrated. The labeling of the constituents with nine different Mg²⁺-ion-dependent DNAzyme reporter units and the incorporation of a fluorescent dye/anticocaine aptamer complex into the structure of one of the constituents enable the quantitative evaluation of the contents of the constituents in the different CDNs. The quantification of the compositions of the CDNs is based on the activities of the DNAzymes conjugated to the constituents, the fluorescence signals upon the cocaine-induced separation of the dye/aptamer complex, appropriate calibration curves, and the set of equations. These assessments are further supported by quantitative electrophoretic experiments of the respective CDNs.