Topological Links between Duplex DNA and a Circular DNA Single Strand

DNA nanostructures of building blocks topologically linked at a precise position can be assembled from DNA duplexes and circularized oligonucleotides with the aid of peptide nucleic acids (PNAs). Shown schematically is a linked catenane yielding an earring topological DNA label.     

Bioimaging Based on Nucleic Acid Nanostructures

Nucleic acid nanostructures with structural programmability, spatial addressability and excellent biocompatibility have drawn much attention in various biomedical applications, such as bioimaging, biosensing and drug delivery. In this review, we summarize the recent research progress in the field of bioimaging based on nucleic acid nanostructures with different imaging models, including fluorescent imaging(FI), magnetic resonance imaging(MRI), photoacoustic imaging(PAI) and positron emission tomography/computed tomography(PET/CT) imaging. We also discuss the remaining challenges and further opportunities involved in the bioimaging research based on nucleic acid nanostructures.     

Structural Aspects of Nucleic Acid Analogs and Antisense Oligonucleotides

Hybridization of complementary oligonucleotides is essential to highly valuable research tools in many fields including genetics, molecular biology, and cell biology. For example, an antisense molecule for a particular segment of sense messenger RNA allows gene expression to be selectively turned off, and the polymerase chain reaction requires complementary primers in order to proceed. It is hoped that the antisense approach may lead to therapeutics for treatment of various diseases including cancer. Areas of active research in the antisense field focus on the mechanisms of cellular uptake of antisense molecules and their delivery to specific cell sites, an improved understanding of how these molecules inhibit the production of proteins, as well as the optimization of the chemical stability of antisense molecules and the thermodynamic stability of the duplexes they form with the mRNA targets. The last two issues in particular have prompted chemists to launch an extensive search for oligonucleotide analogs with improved binding properties for hybridization with RNA and higher resistance toward nuclease degradation. During the last years this research has resulted in a flurry of new chemical analogs of DNA and RNA with modifications in the sugar–phosphate backbone as well as in the nucleobase sites. However, to date little effort has been directed toward uncovering the exact origins of the gain or loss in stability when nucleic acid analogs bind to RNA. Although large amounts of thermodynamic data have been collected, the structural perturbations induced by the modifications in hybrid duplexes are only poorly understood. For many modified oligonucleotides the compatibility of protection, coupling, and deprotection chemistry with standard DNA and RNA synthesis protocols makes it now possible to generate modified nucleic acid fragments or mixed oligonucleotides containing modifications at selected sites in quantities suitable for three-dimensional structure investigations. Such studies should reveal the structural origins of the observed changes in affinity and specificity of binding for particular modifications and may guide the development of second-and third-generation antisense molecules. In addition, the availability of a previously unimaginable variety of modified building blocks and the investigation of their structures provides the basis for a deeper understanding of the native DNA and RNA structures. This contribution will summarize the results of X-ray crystallographic structure determinations of modified nucleic acid fragments conducted in our laboratory during the last three years and the insights gained from them.     

AIEgens/Nucleic Acid Nanostructures for Bioanalytical Applications

DNA occupies significant roles in life processes, which include encoding the sequences of proteins and accurately transferring genetic information from generation to generation. Recent discoveries have demonstrated that a variety of biological functions are correlated with DNA′s conformational transitions. The non‐B form has attained great attention among the diverse forms of DNA over the past several years. The main reason for this is that a large number of studies have shown that the non‐B form of DNA is associated with gross deletions, inversions, duplications, translocations as well as simple repeating sequences, which therefore causes human diseases. Consequently, the conformational transition of DNA between the B‐form and the non‐B form is important for biology. Conventional fluorescence probes based on the conformational transitions of DNA usually need a fluorophore and a quencher group, which suffers from the complex design of the structure and tedious synthetic procedures. Moreover, conventional fluorescence probes are subject to the aggregation‐caused quenching (ACQ) effect, which limits their application toward imaging and analyte detection. Fluorogens exhibiting aggregation‐induced emission (AIE) have attracted tremendous attention over the past decade. By taking advantage of this unique behavior, plenty of fluorescent switch‐on probes without the incorporation of fluorescent quenchers/fluorophore pairs have been widely developed as biosensors for imaging a variety of analytes. Herein, the recent progress in bioanalytical applications on the basis of aggregation‐induced emission luminogens (AIEgens)/nucleic acid nanostructures are presented and discussed.     

Solution Structure of a Locked Nucleic Acid Modified Quadruplex: Introducing the V4 Folding Topology

Sharp curves : The structure of a locked nucleic acid modified telomeric sequence from Oxytricha nova displays a remarkable folding topology, distinct from the native O. nova quadruplex. Each guanine stretch folds back in a V‐shaped turn that puts the first and fourth guanines in the same tetrad, looping over a tetrad with a sharp turn in the DNA backbone, showing how subtle interplay between sequence and conformation defines the folding topology.

     

Highly Stable Duplex Formation by Artificial Nucleic Acids Acyclic Threoninol Nucleic Acid (aTNA) and Serinol Nucleic Acid (SNA) with Acyclic Scaffolds

The stabilities of duplexes formed by strands of novel artificial nucleic acids composed of acyclic threoninol nucleic acid (aTNA) and serinol nucleic acid (SNA) building blocks were compared with duplexes formed by the acyclic glycol nucleic acid (GNA), peptide nucleic acid (PNA), and native DNA and RNA. All acyclic nucleic acid homoduplexes examined in this study had significantly higher thermal stability than DNA and RNA duplexes. Melting temperatures of homoduplexes were in the order of aTNA>PNA≈GNA≥SNA?RNA>DNA. Thermodynamic analyses revealed that high stabilities of duplexes formed by aTNA and SNA were due to large enthalpy changes upon formation of duplexes compared with DNA and RNA duplexes. The higher stability of the aTNA homoduplex than the SNA duplex was attributed to the less flexible backbone due to the methyl group of D ‐threoninol on aTNA, which induced clockwise winding. Unlike aTNA, the more flexible SNA was able to cross‐hybridize with RNA and DNA. Similarly, the SNA/PNA heteroduplex was more stable than the aTNA/PNA duplex. A 15‐mer SNA/RNA was more stable than an RNA/DNA duplex of the same sequence.     

Mismatches Improve the Performance of Strand‐Displacement Nucleic Acid Circuits

Catalytic hairpin assembly (CHA) has previously proven useful as a transduction and amplification method for nucleic acid detection. However, the two hairpin substrates in a CHA circuit can potentially react non‐specifically even in the absence of a single‐stranded catalyst, and this non‐specific background degrades the signal‐to‐noise ratio. The introduction of mismatched base pairs that impede uncatalyzed strand exchange reactions led to a significant decrease of the background signal, while only partially damping the signal in the presence of a catalyst. Various types and lengths of mismatches were assayed by fluorimetry, and in many instances, our MismatCHA designs yielded 100‐fold increased signal‐to‐background ratios compared to a ratio of 4:1 with the perfectly matched substrates. These observations could be of general utility for the design of non‐enzymatic nucleic acid circuits.     

The Unexpected Base-Pairing Behavior of Cyanuric Acid in RNA and Ribose versus Cyanuric Acid Induced Helicene Assembly of Nucleic Acids: Implications for the Pre-RNA Paradigm

The cyanuric acid (CA) heterocycle forms supramolecular structures with adenine nucleobases/nucleosides and oligonucleotides, leading to speculation that they can act as forerunners to RNA. Herein, the assembly behavior of RNA containing CA and CA–ribose nucleoside was studied. Contrary to previous reports, CA in RNA and the CA-ribonucleoside resulted in destabilization of supramolecular assemblies, which led to a reevaluation of the CA–adenine hexameric rosette structure. An unprecedented noncovalent supramolecular helicene structure is proposed to account for the striking difference in behavior, which has implications for novel paradigms for reorganizing the structures of nucleic acids, the synthesis of long helicenes, and pre-RNA world paradigms. The results caution against extrapolating the self-assembly behavior of individual heterocycles from the level of monomers to oligomers because the base-paring properties of (non-)canonical nucleobases are impacted by the type of oligomeric backbone to which they are attached.     

Nucleic Acid Based Logical Systems

Researchers increasingly visualize a significant role for artificial biochemical logical systems in biological engineering, much like digital logic circuits in electrical engineering. Those logical systems could be utilized as a type of servomechanism to control nanodevices in vitro, monitor chemical reactions in situ, or regulate gene expression in vivo. Nucleic acids (NA), as carriers of genetic information with well‐regulated and predictable structures, are promising materials for the design and engineering of biochemical circuits. A number of logical devices based on nucleic acids (NA) have been designed to handle various processes for technological or biotechnological purposes. This article focuses on the most recent and important developments in NA‐based logical devices and their evolution from in vitro, through cellular, even towards in vivo biological applications.     

Covalent DNA–Streptavidin Conjugates as Building Blocks for Novel Biometallic Nanostructures

Supramolecular aggregates of DNA, RNA, streptavidin, immunoglobulin, and nanocrystalline metal clusters can be generated by self-assembly on the basis of oligonucleotide hybridization (shown schematically). Following selective immunosorption on surface-immobilized antigen, the biometallic hybrid is detectable by electron microscopy.     

Supramolecular Materials Comprising Nucleic Acid Biopolymers

We have generated a supramolecular self-assembling film by exchanging the counter-ions of the phosphate moieties in nucleic acid with those of cationic amphiphiles as didodecyldimethylammonium bromide (or DDAB). SAXS and WAXS data for all film samples showed similar harmonic peaks suggesting a lamellar multilayer structure with layers of nucleic acids being separated by lipid bilayers of DDAB. AFM height images also showed that double stranded nucleic acid film can form the step or plateau type of structure and shorter nucleic acid film showed shorter step feature. Moreover, the length and the molecular structure of DNA and RNA can be used to manipulate the mechanical properties of these self-assembled films.     

Assembly of DNA/Fullerene Hybrid Materials

Ordering of cationic fullerene derivatives on a DNA template by phosphate/fullerene complexation provides a rapid route to organic materials several hundred nanometers in length (shown on the right). Hydrophobic interactions between the fullerene units lead to DNA supercoiling that can be relieved by adding a nonionic surfactant. The products are easily imaged by transmission electron microscopy without the need for heavy-metal staining.     

Transient DNA-Based Nanostructures Controlled by Redox Inputs

Synthetic DNA has emerged as a powerful self-assembled material for the engineering of nanoscale supramolecular devices and materials. Recently dissipative self-assembly of DNA-based supramolecular structures has emerged as a novel approach providing access to a new class of kinetically controlled DNA materials with unprecedented life-like properties. So far, dissipative control has been achieved using DNA-recognizing enzymes as energy dissipating units. Although highly efficient, enzymes pose limits in terms of long-term stability and inhibition of enzyme activity by waste products. Herein, we provide the first example of kinetically controlled DNA nanostructures in which energy dissipation is achieved through a non-enzymatic chemical reaction. More specifically, inspired by redox signalling, we employ redox cycles of disulfide-bond formation/breakage to kinetically control the assembly and disassembly of tubular DNA nanostructures in a highly controllable and reversible fashion.     

Structural Classification and General Principles for the Design of Spherical Molecular Hosts

Cryptands, carcerands, polyoxometalates, and molecular capsules are cagelike hosts that complex guests through encapsulation. Following the discovery of a nanometer scale supramolecular shell-like spheroid, these and other shell-like hosts were structurally classified. Their frameworks may be catalogued according to principles of solid geometry. This has led to the identification of hosts that have not yet been synthesized or discovered (such as the cuboctahedron shown; X=O, S) and should lead to the design of additional container assemblies.     

Tuning Dihydrogen Bonds: Enhanced Solid-State Reactivity in a Dihydrogen-Bonded System with Exceptionally Short H⋅⋅⋅H Distances

Topochemical assembly of a covalent material can be achieved with the complex LiBH₄⋅TEA (TEA=triethanolamine; section of structure shown), a dihydrogen-bonded system which has very short H⋅⋅⋅H contacts and high solid-state reactivity due to acidity enhancement in the OH groups by Li⁺ ion complexation.     

Daisy Chain Rotaxanes Made from Interlocked DNA Nanostructures

We report the stepwise assembly of supramolecular daisy chain rotaxanes (DCR) made of double‐stranded DNA: Small dsDNA macrocycles bearing an axle assemble into a pseudo‐DCR precursor that was connected to rigid DNA stoppers to form DCR with the macrocycles hybridized to the axles. In presence of release oligodeoxynucleotides (rODNs), the macrocycles are released from their respective hybridization sites on the axles, leading to stable mechanically interlocked DCRs. Besides the expected threaded DCRs, certain amounts of externally hybridized structures were observed, which dissociate into dumbbell structures in presence of rODNs. We show that the genuine DCRs have significantly higher degrees of freedom in their movement along the thread axle than the hybridized DCR precursors. Interlocking of DNA in DCRs might serve as a versatile principle for constructing functional DNA nanostructures where the movement of the subunits is restricted within precisely confined tolerance ranges.     

Transient DNA‐Based Nanostructures Controlled by Redox Inputs

Synthetic DNA has emerged as a powerful self‐assembled material for the engineering of nanoscale supramolecular devices and materials. Recently dissipative self‐assembly of DNA‐based supramolecular structures has emerged as a novel approach providing access to a new class of kinetically controlled DNA materials with unprecedented life‐like properties. So far, dissipative control has been achieved using DNA‐recognizing enzymes as energy dissipating units. Although highly efficient, enzymes pose limits in terms of long‐term stability and inhibition of enzyme activity by waste products. Herein, we provide the first example of kinetically controlled DNA nanostructures in which energy dissipation is achieved through a non‐enzymatic chemical reaction. More specifically, inspired by redox signalling, we employ redox cycles of disulfide‐bond formation/breakage to kinetically control the assembly and disassembly of tubular DNA nanostructures in a highly controllable and reversible fashion.     

Topological Single-stranded DNA Encoding and Programmable Assembly of Molecular Nanostructures for NIR-II Cancer Theranostics

Molecular nanotechnology promises to offer privileged access to developing NIR-II materials with precise structural and functional manipulation for transformable theranostic applications. However, the lack of an affordable, yet general, method makes this goal currently inaccessible. By virtue of the intriguing nucleic acid chemistry, here we present an artificial base-directed topological single-strand DNA encoding design that enables one-step synthesis of valence-controlled NIR-II molecular nanostructures and spatial assembly of these nanostructures to modulate their behaviors in living systems. As proof-of-concept studies, we construct ultrasmall Ag₂S quantum dots and pH-responsive, size-tunable CuS assemblies for in vivo NIR-II fluorescence imaging and deep tumor photothermal therapy. This work paves a new way for creating functionally diversified architectures and broadens the scope of DNA-encoded material engineering.     

Programming Non-Nucleic Acid Molecules into Computational Nucleic Acid Systems

Nucleic acid (NA) computation has been widely developed in the past years to solve kinds of logic and mathematic issues in both information technologies and biomedical analysis. However, the difficulty to integrate non-NA molecules limits its power as a universal platform for molecular computation. Here, we report a versatile prototype of hybridized computation integrated with both nucleic acids and non-NA molecules. Employing the conformationally controlled ligand converters, we demonstrate that non-NA molecules, including both small molecules and proteins, can be computed as nucleic acid strands to construct the circuitry with increased complexity and scalability, and can be even programmed to solve arithmetical calculations within the computational nucleic acid system. This study opens a new door for molecular computation in which all-NA circuits can be expanded with integration of various ligands, and meanwhile, ligands can be precisely programmed by the nuclei acid computation.     

Computer‐Aided Production of Scaffolded DNA Nanostructures from Flat Sheet Meshes

The use of DNA as a nanoscale construction material has been a rapidly developing field since the 1980s, in particular since the introduction of scaffolded DNA origami in 2006. Although software is available for DNA origami design, the user is generally limited to architectures where finding the scaffold path through the object is trivial. Herein, we demonstrate the automated conversion of arbitrary two‐dimensional sheets in the form of digital meshes into scaffolded DNA nanostructures. We investigate the properties of DNA meshes based on three different internal frameworks in standard folding buffer and physiological salt buffers. We then employ the triangulated internal framework and produce four 2D structures with complex outlines and internal features. We demonstrate that this highly automated technique is capable of producing complex DNA nanostructures that fold with high yield to their programmed configurations, covering around 70 % more surface area than classic origami flat sheets.