Triplex DNA Nanostructures: From Basic Properties to Applications

Triplex nucleic acids have recently attracted interest as part of the rich “toolbox” of structures used to develop DNA‐based nanostructures and materials. This Review addresses the use of DNA triplexes to assemble sensing platforms and molecular switches. Furthermore, the pH‐induced, switchable assembly and dissociation of triplex‐DNA‐bridged nanostructures are presented. Specifically, the aggregation/deaggregation of nanoparticles, the reversible oligomerization of origami tiles and DNA circles, and the use of triplex DNA structures as functional units for the assembly of pH‐responsive systems and materials are described. Examples include semiconductor‐loaded DNA‐stabilized microcapsules, DNA‐functionalized dye‐loaded metal–organic frameworks (MOFs), and the pH‐induced release of the loads. Furthermore, the design of stimuli‐responsive DNA‐based hydrogels undergoing reversible pH‐induced hydrogel‐to‐solution transitions using triplex nucleic acids is introduced, and the use of triplex DNA to assemble shape‐memory hydrogels is discussed. An outlook for possible future applications of triplex nucleic acids is also provided.     

Modular Reconfigurable DNA Origami: From Two-Dimensional to Three-Dimensional Structures

DNA origami enables the manipulation of objects at nanoscale, and demonstrates unprecedented versatility for fabricating both static and dynamic nanostructures. In this work, we introduce a new strategy for transferring modular reconfigurable DNA nanostructures from two-dimensional to three-dimensional. A 2D DNA sheet could be modularized into connected parts (e.g., two, three, and four parts in this work), which can be independently transformed between two conformations with a few DNA “trigger” strands. More interestingly, the transformation of the connected 2D modules can lead to the controlled, resettable structural conversion of a 2D sheet to a 3D architecture, due to the constraints induced by the connections between the 2D modules. This new approach can provide an efficient mean for constructing programmable, higher-order, and complex DNA objects, as well as sophisticated dynamic substrates for various applications.     

Light-Activated Assembly of DNA Origami into Dissipative Fibrils

Hierarchical DNA nanostructures offer programmable functions at scale, but making these structures dynamic, while keeping individual components intact, is challenging. Here we show that the DNA A-motif—protonated, self-complementary poly(adenine) sequences—can propagate DNA origami into one-dimensional, micron-length fibrils. When coupled to a small molecule pH regulator, visible light can activate the hierarchical assembly of our DNA origami into dissipative fibrils. This system is recyclable and does not require DNA modification. By employing a modular and waste-free strategy to assemble and disassemble hierarchical structures built from DNA origami, we offer a facile and accessible route to developing well-defined, dynamic, and large DNA assemblies with temporal control. As a general tool, we envision that coupling the A-motif to cycles of dissipative protonation will allow the transient construction of diverse DNA nanostructures, finding broad applications in dynamic and non-equilibrium nanotechnology.     

Switchable Reconfiguration of an Interlocked DNA Olympiadane Nanostructure

Interlocked DNA rings (catenanes) are interesting reconfigurable nanostructures. The synthesis of catenanes with more than two rings is, however, hampered, owing to low yields of these systems. We report a new method for the synthesis of catenanes with a controlled number of rings in satisfactory yields. Our approach is exemplified by the synthesis of a five‐ring DNA catenane that exists in four different configurations. By the use of nucleic acids as “fuels” and “antifuels”, the cyclic reconfiguration of the system across four states is demonstrated. One of the states, olympiadane, corresponds to the symbol of the Olympic Games. The five‐ring catenane was implemented as a mechanical scaffold for the reconfiguration of Au NPs. The advantages of DNA catenanes over supramolecular catenanes include the possibility of generating highly populated defined states and the feasibility of tethering nanoobjects to the catenanes, which act as a mechanical scaffold to reconfigure the nanoobjects.     

Reconfigurable Plasmonic Nanostructures Controlled by DNA Origami

Precise surface functionalization and reconfigurable capability of nanomaterials are essential to construct complex nanostructures with specific functions.Here we show tire assembly of a reconfigurable plasmonic nanostructure,which executes both conformational and plasmonic changes in response to DNA strands.In this work,different sized gold nanoparticles(AuNPs)were arranged site-specifically on the surface of a DNA origami clamp nanostructure.The opening and closing of the DNA origami clamp could be precisely controlled by a series of strand emplacement reactions.Therefore,the patterns of these AuNPs could be switched between two different configura-tions.The observed plasmon band shift indicates the change of the plasmonic interactions among the assembled AuNPs.Our study achieves the construction of reconfigurable nanomaterials with tunable plasmonic interactions,and will enrich the toolbox of DNA-based functional nanomachinery.     

Block Copolymer Micellization as a Protection Strategy for DNA Origami

DNA nanotechnology enables the synthesis of nanometer‐sized objects that can be site‐specifically functionalized with a large variety of materials. For these reasons, DNA‐based devices such as DNA origami are being considered for applications in molecular biology and nanomedicine. However, many DNA structures need a higher ionic strength than that of common cell culture buffers or bodily fluids to maintain their integrity and can be degraded quickly by nucleases. To overcome these deficiencies, we coated several different DNA origami structures with a cationic poly(ethylene glycol)–polylysine block copolymer, which electrostatically covered the DNA nanostructures to form DNA origami polyplex micelles (DOPMs). This straightforward, cost‐effective, and robust route to protect DNA‐based structures could therefore enable applications in biology and nanomedicine where unprotected DNA origami would be degraded.     

Gold‐Nanoparticle‐Mediated Jigsaw‐Puzzle‐like Assembly of Supersized Plasmonic DNA Origami

DNA origami has rapidly emerged as a powerful and programmable method to construct functional nanostructures. However, the size limitation of approximately 100 nm in classic DNA origami hampers its plasmonic applications. Herein, we report a jigsaw‐puzzle‐like assembly strategy mediated by gold nanoparticles (AuNPs) to break the size limitation of DNA origami. We demonstrated that oligonucleotide‐functionalized AuNPs function as universal joint units for the one‐pot assembly of parent DNA origami of triangular shape to form sub‐microscale super‐origami nanostructures. AuNPs anchored at predefined positions of the super‐origami exhibited strong interparticle plasmonic coupling. This AuNP‐mediated strategy offers new opportunities to drive macroscopic self‐assembly and to fabricate well‐defined nanophotonic materials and devices.     

Construction of Smart Stimuli-Responsive DNA Nanostructures for Biomedical Applications

DNA nanostructures have recently attracted increasing interest in biological and biomedical applications by virtue of their unique properties, such as structural programmability, multi-functionality, stimuli-responsive behaviors, and excellent biocompatibility. In particular, the intelligent responsiveness of smart DNA nanostructures to specific stimuli has facilitated their extensive development in the field of high-performance biosensing and controllable drug delivery. This minireview begins with different self-assembly strategies for the construction of various DNA nanostructures, followed by the introduction of a variety of stimuli-responsive functional DNA nanostructures for assembling metastable soft materials and for facilitating amplified biosensing. The recent achievements of smart DNA nanostructures for controllable drug delivery are highlighted. Finally, the current challenges and possible developments of this promising research are discussed in the fields of intelligent nanomedicine.     

Patterning Porous Networks through Self-Assembly of Programmed Biomacromolecules

Two-dimensional (2D) porous networks are of great interest for the fabrication of complex organized functional materials for potential applications in nanotechnologies and nanoelectronics. This review aims at providing an overview of bottom-up approaches towards the engineering of 2D porous networks by using biomacromolecules, with a particular focus on nucleic acids and proteins. The first part illustrates how the advancements in DNA nanotechnology allowed for the attainment of complex ordered porous two-dimensional DNA nanostructures, thanks to a biomimetic approach based on DNA molecules self-assembly through specific hydrogen-bond base pairing. The second part focuses the attention on how polypeptides and proteins structural properties could be used to engineer organized networks templating the formation of multifunctional materials. The structural organization of all examples is discussed as revealed by scanning probe microscopy or transmission electron microscopy imaging techniques.     

Self-assembly of Micrometer-long DNA Nanoribbons with Four Oligonucleotides

DNA nanostructures have found widespread applications in areas including nanoelectronics and biomedicine. However, traditional DNA origami needs a long single‐stranded virus DNA and hundreds of short DNA strands, which make this method complicated and money‐consuming. Here, we present a protocol for the assembly of DNA nanoribbons with only four oligonucleotides. DNA nanoribbons with different dimensions were successfully assembled with a 96‐base scafford strand and three short staples. These biotinylated nanoribbons could also be decorated with streptavidins. This approach suggests that there exist great design spaces for the creation of simple nucleic acid nanostructures which could facilitate their application in plasmonic or drug delivery.     

Engineering Flexible Metal-Phenolic Networks with Guest Responsiveness via Intermolecular Interactions

Flexible metal-organic materials are of growing interest owing to their ability to undergo reversible structural transformations under external stimuli. Here, we report flexible metal-phenolic networks (MPNs) featuring stimuli-responsive behavior to diverse solute guests. The competitive coordination of metal ions to phenolic ligands of multiple coordination sites and solute guests (e.g., glucose) primarily determines the responsive behavior of the MPNs, as revealed experimentally and computationally. Glucose molecules can be embedded into the dynamic MPNs upon mixing, leading to the reconfiguration of the metal-organic networks and thus changes in their physicochemical properties for targeting applications. This study expands the library of stimuli-responsive flexible metal-organic materials and the understanding of intermolecular interactions between metal-organic materials and solute guests, which is essential for the rational design of responsive materials for various applications.     

Reversible Reconfiguration of DNA Origami Nanochambers Monitored by Single‐Molecule FRET

Today, DNA nanotechnology is one of the methods of choice to achieve spatiotemporal control of matter at the nanoscale. By combining the peculiar spatial addressability of DNA origami structures with the switchable mechanical movement of small DNA motifs, we constructed reconfigurable DNA nanochambers as dynamic compartmentalization systems. The reversible extension and contraction of the inner cavity of the structures was used to control the distance‐dependent energy transfer between two preloaded fluorophores. Interestingly, single‐molecule FRET studies revealed that the kinetics of the process are strongly affected by the choice of the switchable motifs and/or actuator sequences, thus offering a valid method for fine‐tuning the dynamic properties of large DNA nanostructures. We envisage that the proposed DNA nanochambers may function as model structures for artificial biomimetic compartments and transport systems.     

Surface‐Assisted Large‐Scale Ordering of DNA Origami Tiles

The arrangement of DNA‐based nanostructures into extended higher order assemblies is an important step towards their utilization as functional molecular materials. We herein demonstrate that by electrostatically controlling the adhesion and mobility of DNA origami structures on mica surfaces by the simple addition of monovalent cations, large ordered 2D arrays of origami tiles can be generated. The lattices can be formed either by close‐packing of symmetric, non‐interacting DNA origami structures, or by utilizing blunt‐end stacking interactions between the origami units. The resulting crystalline lattices can be readily utilized as templates for the ordered arrangement of proteins.     

Designed Intercalators for Modification of DNA Origami Surface Properties

The modification of the backbone properties of DNA origami nanostructures through noncovalent interactions with designed intercalators, based on acridine derivatized with side chains containing esterified fatty acids or oligo(ethylene glycol) residues is reported. Spectroscopic analyses indicate that these intercalators bind to DNA origami structures. Atomic force microscopy studies reveal that intercalator binding does not affect the structural intactness but leads to altered surface properties of the highly negatively charged nanostructures, as demonstrated by their interaction with solid mica or graphite supports. Moreover, the noncovalent interaction between the intercalators and the origami structures leads to alteration in cellular uptake, as shown by confocal microscopy studies using two different eukaryotic cell lines. Hence, the intercalator approach offers a potential means for tailoring the surface properties of DNA nanostructures.     

Information Coding in a Reconfigurable DNA Origami Domino Array

DNA nanostructures with programmable nanoscale patterns has been achieved in the past decades, and molecular information coding (MIC) on those designed nanostructures has gained increasing attention for information security. However, achieving steganography and cryptography synchronously on DNA nanostructures remains a challenge. Herein, we demonstrated MIC in a reconfigurable DNA origami domino array (DODA), which can reconfigure intrinsic patterns but keep the DODA outline the same for steganography. When a set of keys (DNA strands) are added, the cryptographic data can be translated into visible patterns within DODA. More complex cryptography with the ASCII code within a programmable 6×6 lattice is demonstrated to demosntrate the versatility of MIC in the DODA. Furthermore, an anti‐counterfeiting approach based on conformational transformation‐mediated toehold strand displacement reaction is designed to protect MIC from decoding and falsification.     

Spatiotemporal Control over Polynucleotide Brush Growth on DNA Origami Nanostructures

DNA nanotechnology provides an approach to create precise, tunable, and biocompatible nanostructures for biomedical applications. However, the stability of these structures is severely compromised in biological milieu due to their fast degradation by nucleases. Recently, we showed how enzymatic polymerization could be harnessed to grow polynucleotide brushes of tunable length and location on the surface of DNA origami nanostructures, which greatly enhances their nuclease stability. Here, we report on strategies that allow for both spatial and temporal control over polymerization through activatable initiation, cleavage, and regeneration of polynucleotide brushes using restriction enzymes. The ability to site-specifically decorate DNA origami nanostructures with polynucleotide brushes in a spatiotemporally controlled way provides access to “smart” functionalized DNA architectures with potential applications in drug delivery and supramolecular assembly.     

Framework Nucleic Acids for Cell Imaging and Therapy

Over the past decade,structural DNA nanotechnology has been well developed to be a promising and powerful technique to generate various nanostructures with programmability,spatial organization and biocompatibi-lity.With the advent of computer-aided tools,framework nucleic acids have been employed in a series of biomedical applications,ranging from biosensing,bioimaging,diagnosis,to therapeutics.In this review,we summarized recent advances in the construction of precisely assembled DNA nanostructures,and DNA-engineered biomimetics.We also outlined the challenges and opportunities for the translational applications of framework nucleic acids.     

Fabrication of Defined Polydopamine Nanostructures by DNA Origami‐Templated Polymerization

A versatile, bottom‐up approach allows the controlled fabrication of polydopamine (PD) nanostructures on DNA origami. PD is a biosynthetic polymer that has been investigated as an adhesive and promising surface coating material. However, the control of dopamine polymerization is challenged by the multistage‐mediated reaction mechanism and diverse chemical structures in PD. DNA origami decorated with multiple horseradish peroxidase‐mimicking DNAzyme motifs was used to control the shape and size of PD formation with nanometer resolution. These fabricated PD nanostructures can serve as “supramolecular glue” for controlling DNA origami conformations. Facile liberation of the PD nanostructures from the DNA origami templates has been achieved in acidic medium. This presented DNA origami‐controlled polymerization of a highly crosslinked polymer provides a unique access towards anisotropic PD architectures with distinct shapes that were retained even in the absence of the DNA origami template.     

DNA origami: the art of folding DNA

The advent of DNA origami technology greatly simplified the design and construction of nanometer-sized DNA objects. The self-assembly of a DNA-origami structure is a straightforward process in which a long single-stranded scaffold (often from the phage M13mp18) is folded into basically any desired shape with the help of a multitude of short helper strands. This approach enables the ready generation of objects with an addressable surface area of a few thousand nm(2) and with a single "pixel" resolution of about 6 nm. The process is rapid, puts low demands on experimental conditions, and delivers target products in high yields. These features make DNA origami the method of choice in structural DNA nanotechnology when two- and three-dimensional objects are desired. This Minireview summarizes recent advances in the design of DNA origami nanostructures, which open the door to numerous exciting applications.     

Enhancing Biocompatible Stability of DNA Nanostructures Using Dendritic Oligonucleotides and Brick Motifs

The use of DNA‐based nanomaterials in biomedical applications is continuing to grow, yet more emphasis is being put on the need for guaranteed structural stability of DNA nanostructures in physiological conditions. Various methods have been developed to stabilize DNA origami against low concentrations of divalent cations and the presence of nucleases. However, existing strategies typically require the complete encapsulation of nanostructures, which makes accessing the encased DNA strands difficult, or chemical modification, such as covalent crosslinking of DNA strands. We present a stabilization method involving the synthesis of DNA brick nanostructures with dendritic oligonucleotides attached to the outer surface. We find that nanostructures assembled from DNA brick motifs remain stable against denaturation without any chemical modifications. Furthermore, densely coating the outer surface of DNA brick nanostructures with dendritic oligonucleotides prevents nuclease digestion.