Design, Assembly, and Simulation of a Triangular DNA Origami Box: A Wireframe Nanostructure Approach

Abstract DNA nanotechnology enables the programmable construction of complex nanoscale architectures with high precision. In this study, we present the design, synthesis, and computational validation of a three-dimensional rectangular DNA box constructed from triangular wireframe subunits. Using a modular edge-based strategy, triangular motifs were assembled into a closed nanocage. The structure was designed using lattice-based routing and converted into a coarse-grained molecular model for simulation. Molecular dynamics simulations confirm structural stability and mechanical integrity. This work demonstrates a scalable approach for constructing rigid, low-material DNA nanostructures with potential applications in nanoreactors, drug delivery, and biomolecular engineering. 1. Introduction DNA origami has revolutionized nanoscale fabrication by enabling the folding of long single-stranded DNA into predefined shapes using short staple strands. While early designs focused on solid, brick-like architectures, recent advances have shifted toward wireframe structures composed of polygonal units. Triangular motifs provide enhanced rigidity and reduced material consumption, making them ideal building blocks for three-dimensional nanocages. In this study, we design a rectangular DNA box constructed from triangular subunits. The structure is assembled via programmable sticky-end cohesion and validated through coarse-grained molecular dynamics simulations. 2. Materials and Methods 2.1 Structural Design The DNA box was constructed using triangular panels, each composed of three double-helical edges. Each edge consisted of a 21 base pair duplex with 4-nucleotide sticky ends to facilitate inter-edge connectivity. The structure comprises: 12 triangular faces 36 edges Modular connectivity via complementary overhangs 2.2 Sequence Design Edge sequences were designed to: Minimize secondary structure formation Maintain uniform GC content (~45–55%) Avoid repetitive motifs Each edge consisted of two complementary strands forming a stable duplex core, with single-stranded overhangs for assembly. 2.3 caDNAno Modeling The structure was approximated in a honeycomb lattice framework. Scaffold routing was implemented to follow triangular paths, with crossovers placed at periodic intervals to maintain structural integrity. The design was exported as a JSON file and converted into a simulation-ready format. 3. Computational Modeling 3.1 oxDNA Simulation Setup The structure was simulated using a coarse-grained molecular dynamics framework. The topology file defined strand connectivity, while the configuration file specified initial nucleotide positions. Simulation parameters: Temperature: 300 K Salt concentration: 0.5 M Time step: 0.005 simulation units Total steps: 5 × 10⁶ 3.2 Energy Minimization Initial configurations were relaxed to remove steric clashes. Hydrogen bonding and stacking interactions stabilized the structure during equilibration. 4. Results 4.1 Structural Assembly The triangular subunits successfully assembled into a closed rectangular box. Sticky-end cohesion ensured correct edge pairing, while crossover placement stabilized the geometry. 4.2 Simulation Stability Molecular dynamics simulations showed: Stable edge lengths Minimal structural deformation Preservation of triangular geometry Root mean square deviation (RMSD) analysis indicated convergence after equilibration, confirming structural robustness. 4.3 Mechanical Properties The triangular architecture demonstrated: Increased rigidity compared to planar designs Resistance to bending and collapse Efficient stress distribution across edges 5. Discussion The use of triangular subunits provides significant advantages over traditional DNA origami designs. The wireframe approach reduces the number of անհրաժեշտ nucleotides while maintaining structural integrity. This makes the design particularly suitable for applications requiring lightweight yet मजबूत nanostructures. The integration of computational modeling ensures that only structurally viable designs are synthesized experimentally, reducing trial-and-error. 6. Applications 6.1 Nanoreactors The DNA box can encapsulate enzymes, enabling confined biochemical reactions. 6.2 Drug Delivery Programmable opening mechanisms can allow targeted release of therapeutic molecules. 6.3 Metal Nanoparticle Synthesis Functionalization with thiol-modified strands enables selective binding and nucleation of metal nanoparticles. 7. Conclusion We present a complete pipeline for the design, assembly, and validation of a triangular DNA box. The combination of modular sequence design, lattice-based modeling, and molecular simulation provides a robust framework for constructing advanced DNA nanostructures. This approach is scalable and adaptable for a wide range of nanobiotechnological applications. 8. Figures Figure 1: Structural Design of Triangular DNA Box Schematic showing rectangular box composed of triangular panels Dimensions and edge connectivity highlighted Figure 2: Scaffold and Staple Organization Routing of scaffold strand Binding positions of staple strands Crossover locations Figure 3: Folding Pathway Mixing of scaffold and staples Thermal annealing profile Final folded structure Figure 4: Simulation Results Initial vs equilibrated structure RMSD plot over time Energy stabilization curve 9. References