A Chemo-Mechanically Coupled DNA Origami Clamp Capable of Generating Robust Compression Forces.

DNA nanostructures have been utilized to study biological mechanical processes and construct artificial nanosystems. Many application scenarios necessitate nanodevices able to robustly generate large single molecular forces. However, most existing dynamic DNA nanostructures are triggered by probabilistic hybridization reactions between spatially separated DNA strands, which only non-deterministically generate relatively small compression forces (≈0.4 piconewtons (pN)). Here, an intercalator-triggered dynamic DNA origami nanostructure is developed, where large amounts of local binding reactions between intercalators and the nanostructure collectively lead to the robust generation of relatively large compression forces (≈11.2 pN). Biomolecular loads with different stiffnesses, 3, 4, and 6-helix DNA bundles are efficiently bent by the compression forces. This work provides a robust and powerful force-generation tool for building highly chemo-mechanically coupled molecular machines in synthetic nanosystems.

Diverse Chiral Nanotubes Assembled from Identical DNA Strands.

DNA nanotubes with controllable geometries hold a wide range of interdisciplinary applications. When preparing DNA nanotubes of varying widths or distinct chirality, existing methods require repeatedly designing and synthesizing specific DNA sequences, which can be costly and laborious. Here, we proposed an intercalator-assisted DNA tile assembly method which enables the production of DNA nanotubes of diverse widths and chirality using identical DNA strands. Through adjusting the concentration of intercalators during assembly, the twisting direction and extent of DNA tiles could be modulated, leading to the formation of DNA nanotubes featuring controllable widths and chirality. Moreover, through introducing additional intercalators and secondary annealing, right-handed nanotubes could be reconfigured into distinct left-handed nanotubes. We expect that this method could be universally applied to modulating the self-assembly pathways of various DNA tiles and other chiral materials, advancing the landscape of DNA tile assembly.

Multimode adaptive logic gates based on temperature-responsive DNA strand displacement.

Living organisms switch their intrinsic biological states to survive environmental turbulence, in which temperature changes are prevalent in nature. Most artificial temperature-responsive DNA nanosystems work as switch modules that transit between "ON-OFF" states, making it difficult to construct nanosystems with diverse functions. In this study, we present a general strategy to build multimode nanosystems based on a temperature-responsive DNA strand displacement reaction. The temperature-responsive DNA strand displacement was controlled by tuning the sequence of the substrate hairpin strands and the invading strands. The nanosystems were demonstrated as logic gates that performed a set of Boolean logical functions at specific temperatures. In addition, an adaptive logic gate was fabricated that could exhibit different logic functions when placed in different temperatures. Specifically, upon the same input strands, the logic gate worked as an XOR gate at 10 °C, an OR gate at 35 °C, an AND gate at 46 °C, and was reset at 55 °C. The design and fabrication of the multifunctional nanosystems would help construct advanced temperature-responsive systems that may be used for temperature-controlled multi-stage drug delivery and thermally-controlled multi-step assembly of nanostructures.

Presketched DNA Origami Canvas for Polymerase-Driven DNA Kirigami.

Reconfigurable DNA origami provides a versatile tool to manipulate the conformation of matter on the nanometer scale. Typically, the DNA kirigami method enables the transformation of an origami structure from an initial shape to another predesigned shape by reconfiguring the staple strands. In a regular origami structure, since the perfectly matched and densely packed DNA duplexes block the removal of staple strands, the construction of finely trimmed "sub-origami" structures by the DNA kirigami method has remained challenging. Herein, we proposed a strategy to construct the presketched DNA origami canvas, where the offcut area in the canvas was sketched by loosely fixed staple strands with single-base insertion, to enhance the fineness of polymerase-driven DNA kirigami. We successfully trimmed presketched two-dimensional rectangular canvas, three-dimensional Möbius strip, and genie bottle canvases into complex letter patterns, supercoiled rings, and nanorods, respectively. Finally, we demonstrated a size-controlled DNA kirigami system: a presketched 6HB origami was trimmed into a set of shorter nanowires with predefined lengths, which quantitatively characterized the fineness of the improved DNA kirigami. The presketched origami design was a general method that applied to both 2D and 3D DNA origami structures in square and honeycomb lattices. Loosening DNA origami structures by introducing single-base insertions provides a practical approach to constructing dynamic components when designing DNA nanomachines. Furthermore, the delicate trimming of the DNA origami canvas driven by polymerase may inspire strategies for graphical information encryption and storage.

Regulating the Polymerization of DNA Structures via Allosteric Control of Monomers.

Regulation of self-assembly is crucial in constructing structural biomaterials, such as tunable DNA nanostructures. Traditional tuning of self-assembled DNA nanostructures was mainly conducted by introducing external stimuli after the assembly process. Here, we explored the allosteric assembly of DNA structures via introducing external stimuli during the assembly process to produce structurally heterogeneous polymerization products. We demonstrated that ethidium bromide (EB), a DNA intercalator, could increase the left-handed out-of-plane chirality of curved DNA structures. Then, EB and double strands were introduced as competing stimuli to transform monomers into allosteric conformations, leading to three different polymerization products. The steric trap between different polymerization products promoted the polymerized structures to keep their geometric properties, like chirality, under varying intensity of external stimuli. Our strategy harnesses allosteric effects for assembly of DNA-based materials and is expected to expand the design space for advanced control in synthetic materials.

Disassembly of DNA origami dimers controlled by programmable polymerase primers.

Dynamic regulation of DNA origami nanostructures is important for the fabrication of intelligent DNA nanodevices. Toehold-mediated strand displacement is a common regulation strategy, which utilizes trigger strands to assemble and disassemble nanostructures. Such trigger strands are required to be completely complementary to the corresponding substrate strands, which strictly demands orthogonality and accuracy of the sequence design. Herein, we present a disassembly strategy of DNA origami dimers based on polymerase-triggered strand displacement, where the polymerase primers, as the trigger strands, were only partially complementary to the toehold region of the substrate strands. To demonstrate the programmability of trigger strands, we utilized primers with different sequence combination patterns to disassemble DNA origami dimers. The statistical summary of AFM images and fluorescence curves proved the feasibility of the new strategy. The utilization of polymerase-triggered strand displacement on the disassembly of DNA origami structures enriches the toolbox for the dynamic regulation of DNA nanostructures.

Cyclic transitions of DNA origami dimers driven by thermal cycling.

It is widely observed that life activities are regulated through conformational transitions of biological macromolecules, which inspires the construction of environmental responsive nanomachines in recent years. Here we present a thermal responsive DNA origami dimers system, whose conformations can be cyclically switched by thermal cycling. In our strategy, origami dimers are assembled at high temperatures and disassembled at low temperatures, which is different from the conventional strategy of breaking nanostructures using high temperatures. The advantage of this strategy is that the dimers system can be repeatedly operated without significant performance degradation, compared to traditional strategies such as conformational transitions via i-motif and G-quadruplexes, whose performance degrades with sample dilution due to repeated addition of trigger solutions. The cyclic conformational transitions of the dimers system are verified by fluorescence curves and AFM images. This research offered a new way to construct cyclic transformational nanodevices, such as reusable nanomedicine delivery systems or nanorobots with long service lifetimes.

Tuning curved DNA origami structures through mechanical design and chemical adducts.

The bending and twisting of DNA origami structures are important features for controlling the physical properties of DNA nanodevices. It has not been fully explored yet how to finely tune the bending and twisting of curved DNA structures. Traditional tuning of the curved DNA structures was limited to controlling the in-plane-bending angle through varying the numbers of base pairs of deletions and insertions. Here, we developed two tuning strategies of curved DNA origami structures fromin silicoandin vitroaspects.In silico, the out-of-plane bending and twisting angles of curved structures were introduced, and were tuned through varying the patterns of base pair deletions and insertions.In vitro, a chemical adduct (ethidium bromide) was applied to dynamically tune a curved spiral. The 3D structural conformations, like chirality, of the curved DNA structures were finely tuned through these two strategies. The simulation and TEM results demonstrated that the patterns of base pair insertions and deletions and chemical adducts could effectively tune the bending and twisting of curved DNA origami structures. These strategies expand the programmable accuracy of curved DNA origami structures and have potential in building efficient dynamic functional nanodevices.

DNA Kirigami Driven by Polymerase-Triggered Strand Displacement.

The precursors of functional biomolecules in living cells are synthesized in a bottom-up manner and subsequently activated by modification into a delicate structure with near-atomic precision. DNA origami technology provides a promising way to mimic the synthesis of precursors, although mimicking the modification process is a challenge. Herein, a DNA paper-cutting (DNA kirigami) method to trim origami into designer nanostructures is proposed, where the modification is implemented by a polymerase-triggered DNA strand displacement reaction. Six geometric shapes are created by cutting rectangular DNA origami. Gel electrophoresis and atomic force microscopy results demonstrate the feasibility and capability of the DNA paper-cutting method. The proposed DNA paper-cutting strategy can enrich the toolbox for dynamically transforming DNA origami and has potential applications in biomimetics. .

Programmable allosteric DNA regulations for molecular networks and nanomachines.

Structure-based molecular regulations have been widely adopted to modulate protein networks in cells and recently developed to control allosteric DNA operations in vitro. However, current examples of programmable allosteric signal transmission through integrated DNA networks are stringently constrained by specific design requirements. Developing a new, more general, and programmable scheme for establishing allosteric DNA networks remains challenging. Here, we developed a general strategy for programmable allosteric DNA regulations that can be finely tuned by varying the dimensions, positions, and number of conformational signals. By programming the allosteric signals, we realized fan-out/fan-in DNA gates and multiple-layer DNA cascading networks, as well as expanding the approach to long-range allosteric signal transmission through tunable DNA origami nanomachines ~100 nm in size. This strategy will enable programmable and complex allosteric DNA networks and nanodevices for nanoengineering, chemical, and biomedical applications displaying sense-compute-actuate molecular functionalities.