Heat-activated growth of metastable and length-defined DNA fibers expands traditional polymer assembly.

Biopolymers such as nucleic acids and proteins exhibit dynamic backbone folding, wherein site-specific intramolecular interactions determine overall structure. Proteins then hierarchically assemble into supramolecular polymers such as microtubules, that are robust yet dynamic, constantly growing or shortening to adjust to cellular needs. The combination of dynamic, energy-driven folding and growth with structural stiffness and length control is difficult to achieve in synthetic polymer self-assembly. Here we show that highly charged, monodisperse DNA-oligomers assemble via seeded growth into length-controlled supramolecular fibers during heating; when the temperature is lowered, these metastable fibers slowly disassemble. Furthermore, the specific molecular structures of oligomers that promote fiber formation contradict the typical theory of block copolymer self-assembly. Efficient curling and packing of the oligomers - or 'curlamers' - determine morphology, rather than hydrophobic to hydrophilic ratio. Addition of a small molecule stabilises the DNA fibers, enabling temporal control of polymer lifetime and underscoring their potential use in nucleic-acid delivery, stimuli-responsive biomaterials, and soft robotics.

Divergent Polymer Superstructures from Protonated Poly(adenine) DNA and RNA.

Studies have shown that poly(adenine) DNA and RNA strands protonate at a low pH to form self-associating duplexes; however, the nanoscopic morphology of these structures is unclear. Here, we use Transition Electron Microscopy (TEM), Atomic Force Microscopy (AFM), dynamic light scattering (DLS), and fluorescence spectroscopy to show that both ribose identity (DNA or RNA) and assembly conditions (thermal or room-temperature annealing) dictate unique hierarchical structures for poly(adenine) sequences at a low pH. We show that while the thermodynamic product of protonating poly(adenine) DNA is a discrete dimer of two DNA strands, the kinetic product is a supramolecular polymer that branches and aggregates to form micron-diameter superstructures. In contrast, we find that protonated poly(A) RNA polymerizes into micrometer-length, twisted fibers under the same conditions. These divergent hierarchical morphologies highlight the amplification of subtle chemical differences between RNA and DNA into unique nanoscale behaviors. With the use of poly(adenine) strands spanning vaccine technologies, sensing, and dynamic biotechnology, understanding and controlling the underlying assembly pathways of these structures are critical to developing robust, programmable nanotechnologies.

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.

Programming rigidity into size-defined wireframe DNA nanotubes.

Nanotubes built from DNA hold promise for several biological and materials applications, due to their high aspect ratio and encapsulation potential. A particularly appealing goal is to control the size, shape, and dynamic behaviour of DNA nanotubes with minimal design alteration, as nanostructures of varying morphologies and lengths have been shown to exhibit distinct cellular uptake, encapsulation behaviour, and in vivo biodistribution. Herein, we report a systematic investigation, combining experimental and computational design, to modulate the length, flexibility, and longitudinal patterns of wireframe DNA nanotubes. Subtle design changes govern the structure and properties of our nanotubes, which are built from a custom-made, long, and size-defined template strand to which DNA rungs and linkers are attached. Unlike DNA origami, these custom-made strands possess regions with repeating sequences at strategic locations, thereby reducing the number of strands necessary for assembly. Through strand displacement, the nanotubes can be reversibly altered between extended and collapsed morphologies. These design concepts enable fine-tuning of the nanotube stiffness and may pave the way for the development of designer nanotubes for a variety of applications, including the study of cellular internalization, biodistribution, and uptake mechanisms for structures of varied shapes and sizes.

Modulating the Lifetime of DNA Motifs Using Visible Light and Small Molecules.

Here we regulate the formation of dissipative assemblies built from DNA using a merocyanine photoacid that responds to visible light. The operation of our system and the relative distribution of species within it are controlled by irradiation time, initial pH value, and the concentration of a small-molecule binder that inhibits the reaction cycle. This approach is modular, does not require DNA modification, and can be used for several DNA sequences and lengths. Our system design allows for waste-free control of dissipative DNA nanotechnology, toward the generation of nonequilibrium, life-like nanodevices.

DNA Sequence and Length Dictate the Assembly of Nucleic Acid Block Copolymers.

The self-assembly of block copolymers is often rationalized by structure and microphase separation; pathways that diverge from this parameter space may provide new mechanisms of polymer assembly. Here, we show that the sequence and length of single-stranded DNA directly influence the self-assembly of sequence-defined DNA block copolymers. While increasing the length of DNA led to predictable changes in self-assembly, changing only the sequence of DNA produced three distinct structures: spherical micelles (spherical nucleic acids, SNAs) from flexible poly(thymine) DNA, fibers from semirigid mixed-sequence DNA, and networked superstructures from rigid poly(adenine) DNA. The secondary structure of poly(adenine) DNA strands drives a temperature-dependent polymerization and assembly mechanism: copolymers stored in an SNA reservoir form fibers after thermal activation, which then aggregate upon cooling to form interwoven networks. DNA is often used as a programming code that aids in nanostructure addressability and function. Here, we show that the inherent physical and chemical properties of single-stranded DNA sequences also make them an ideal material to direct self-assembled morphologies and select for new methods of supramolecular polymerization.

Incorporation of a Phosphino(pyridine) Subcomponent Enables the Formation of Cages with Homobimetallic and Heterobimetallic Vertices.

Biological systems employ multimetallic assemblies to achieve a range of functions. Here we demonstrate the preparation of metal-organic cages that contain either homobimetallic or heterobimetallic vertices. These vertices are constructed using 2-formyl-6-diphenylphosphinopyridine, which forms ligands that readily bridge between a pair of metal centers, thus enforcing the formation of bimetallic coordination motifs. Two pseudo-octahedral homometallic MI12L4 cages (MI = CuI or AgI) were prepared, with a head-to-head configuration of their vertices confirmed by X-ray crystallography and multinuclear NMR for AgI. The phosphino-pyridine subcomponent also enabled the formation of a class of octanuclear CdII4CuI4L4 tetrahedral cages, representing an initial example of self-assembled cages containing well-defined heterobimetallic vertices.

A dissipative pathway for the structural evolution of DNA fibres.

Biochemical networks interconnect, grow and evolve to express new properties as different chemical pathways are selected during a continuous cycle of energy consumption and transformation. In contrast, synthetic systems that push away from equilibrium usually return to the same self-assembled state, often generating waste that limits system recyclability and prevents the formation of adaptable networks. Here we show that annealing by slow proton dissipation selects for otherwise inaccessible morphologies of fibres built from DNA and cyanuric acid. Using single-molecule fluorescence microscopy, we observe that proton dissipation influences the growth mechanism of supramolecular polymerization, healing gaps within fibres and converting highly branched, interwoven networks into nanocable superstructures. Just as the growth kinetics of natural fibres determine their structural attributes to modulate function, our system of photoacid-enabled depolymerization and repolymerization selects for healed materials to yield organized, robust fibres. Our method provides a chemical route for error-checking, distinct from thermal annealing, that improves the morphologies and properties of supramolecular materials using out-of-equilibrium systems.

Asymmetric patterning drives the folding of a tripodal DNA nanotweezer.

DNA tweezers have emerged as powerful devices for a wide range of biochemical and sensing applications; however, most DNA tweezers consist of single units activated by DNA recognition, limiting their range of motion and ability to respond to complex stimuli. Herein, we present an extended, tripodal DNA nanotweezer with a small molecule junction. Simultaneous, asymmetric elongation of our molecular core is achieved using polymerase chain reaction (PCR) to produce length- and sequence-specific DNA arms with repeating DNA regions. When rigidified, our DNA tweezer can be addressed with streptavidin-binding ligands. Full control over the number, separation, and location of these ligands enables site-specific streptavidin recognition; all three arms of the DNA nanotweezer wrap around multiple streptavidin units simultaneously. Our approach combines the simplicity of DNA tile arrays with the size regime normally provided by DNA origami, offering an integrated platform for the use of branched DNA scaffolds as structural building blocks, protein sensors, and dynamic, stimuli-responsive materials.

A poly(thymine)-melamine duplex for the assembly of DNA nanomaterials.

The diversity of DNA duplex structures is limited by a binary pair of hydrogen-bonded motifs. Here we show that poly(thymine) self-associates into antiparallel, right-handed duplexes in the presence of melamine, a small molecule that presents a triplicate set of the hydrogen-bonding face of adenine. X-ray crystallography shows that in the complex two poly(thymine) strands wrap around a helical column of melamine, which hydrogen bonds to thymine residues on two of its three faces. The mechanical strength of the thymine-melamine-thymine triplet surpasses that of adenine-thymine base pairs, which enables a sensitive detection of melamine at 3 pM. The poly(thymine)-melamine duplex is orthogonal to native DNA base pairing and can undergo strand displacement without the need for overhangs. Its incorporation into two-dimensional grids and hybrid DNA-small-molecule polymers highlights the poly(thymine)-melamine duplex as an additional tool for DNA nanotechnology.

Single-molecule methods in structural DNA nanotechnology.

Single molecules can now be visualised with unprecedented precision. As the resolution of single-molecule experiments improves, so too does the breadth, quantity and quality of information that can be extracted using these methodologies. In the field of DNA nanotechnology, we use programmable interactions between nucleic acids to generate complex, multidimensional structures. We can use single-molecule techniques - ranging from electron and fluorescence microscopies to electrical and force spectroscopies - to report on the structure, morphology, robustness, sample heterogeneity and other properties of these DNA nanoconstructs. In this Tutorial Review, we will detail how complementarity between static and dynamic single-molecule techniques can provide a unified image of DNA nanoarchitectures. The single-molecule methods that we discuss provide unprecedented insight into chemical and structural behaviour, yielding not just an average outcome but reporting on the distribution of values, ultimately showing how bulk properties arise from the collective behaviour of individual structures. As the fields of both DNA nanotechnology and single-molecule characterisation intertwine, a feedback loop is generated between disciplines, providing new opportunities for the development and operation of DNA-based materials as sensors, delivery vehicles, machinery and structural scaffolds.

Narcissistic, Integrative, and Kinetic Self-Sorting within a System of Coordination Cages.

Many useful principles of self-assembly have been elucidated through studies of systems where multiple components combine to create a single structure. More complex systems, where multiple product structures self-assemble in parallel from a shared set of precursors, are also of great interest, as biological systems exhibit this behavior. The greater complexity of such systems leads to an increased likelihood that discrete species will not be formed, however. Here we show how the kinetics of self-assembly govern the formation of multiple metal-organic architectures from a mixture of five building blocks, preventing the formation of a discrete structure of intermediate size. By varying ligand symmetry, denticity, and orientation, we explore how five distinct polyhedra-a tetrahedron, an octahedron, a cube, a cuboctahedron, and a triangular prism-assemble in concert around CoII template ions. The underlying rules dictating the organization of assemblies into specific shapes are deciphered, explaining the formation of only three discrete entities when five could form in principle.

Oxidation triggers guest dissociation during reorganization of an FeII 4L6 twisted parallelogram.

A three-dimensional FeII 4L6 parallelogram was prepared from ferrocene-containing ditopic ligands. The steric preference of the bulky ferrocene cores towards meridional vertex coordination brought about this new structure type, in which the ferrocene units adopt three distinct conformations. The structure possesses two distinct, bowl-like cavities that host anionic guests. Oxidation of the ferrocene FeII to ferrocenium FeIII causes rotation of the ferrocene hinges, converting the structure to an FeII 1L1 + species with release of anionic guests, even though the average charge per iron increases in a way that would ordinarily increase guest binding strength. The degrees of freedom exhibited by these new structures - derived from the different configurations of the three ligands surrounding a meridional FeII center and the rotation of ferrocene cores - thus underpin their ability to reconfigure and eject guests upon oxidation.

Transition-Metal-Functionalized DNA Double-Crossover Tiles: Enhanced Stability and Chirality Transfer to Metal Centers.

The double crossover junction (DX) is a fundamental building block for generating complex and varied structures from DNA. However, its implementation in functional devices is limited to the inherent properties of DNA itself. Here, we developed design strategies to generate the first metal-DX DNA tiles (DXM ) by site-specifically functionalizing the tile crossovers with tetrahedral binding pockets that coordinate CuI . These DX junctions bind two CuI ions independently at distinct sites, display greater thermal stability than native DX tiles upon metalation, and melt in a cooperative fashion. In addition, the right-handed helical chirality of DNA is transferred to the metal centers. Our tiles display high metal ion selectivity, such that CuII is spontaneously reduced to CuI in situ. By modifying our design over three generations of tiles, we elucidated the thermodynamic and geometric requirements for the successful assembly of DXM tiles, which have direct applicability in developing robust, stable DNA-based materials with electroactive, photoactive, and catalytic properties.

Conformational Control in Main Group Phosphazane Anion Receptors and Transporters.

Anion binding by receptor molecules is a central field of modern chemistry which impacts areas of catalysis as well as biological and materials chemistry. As binding often requires high chemical stability under aerobic and aqueous conditions for practical applications, carbon-based anion receptors have dominated this field, with main group element analogues receiving far less attention. The recent observation that the air- and moisture-stable amino-cyclophosph(V)azanes of the type [RN(E)P(µ-NR)]2 (E = O, S, Se) can exhibit halide binding that is competitive with topologically related organic receptors (such as squaramides and thioureas) has motivated us here to explore how the binding properties of phosphazane receptors can be enhanced further. Coordination of transition metals by the two P,N metal coordination sites of the phosph(III)azane dimer [(2-py)NHP(µ-NtBu)]2 not only activates the receptor for anion binding (by fixing the optimum exo-exo conformation and polarizing the endocyclic N-H substituents) but also stabilizes the P2N2 ring to hydrolysis and oxidation. We show how the binding properties of these receptors can be modulated by the coordinated metal fragments and that they can bind chloride 1 to 2 orders of magnitude stronger than the related squaramides and thioureas. These features can be utilized in anion transport through phospholipid bilayers under aqueous conditions for which transport can be improved by 1 order of magnitude compared to the previous best phosphazane and thiourea transporters. This study demonstrates how careful design of inorganic systems can result in potent supramolecular functionality, beyond that observed for organic counterparts.

Remote control of charge transport and chiral induction along a DNA-metallohelicate.

Herein we present a new strategy to achieve chiral induction and redox switching along the backbone of metallohelicate architectures, wherein a DNA duplex directs the handedness and charge transport properties of a metal-organic assembly more than 60 bonds away (a distance of >10 nm). The quantitative and site-specific binding of copper(i) ions to DNA-templated coordination sites imparts enhanced thermodynamic stability to the assembly, while the DNA duplex transfers its natural right-handed helicity to the proximal and distal metal centers of the helicates. When copper(ii) ions are employed instead of copper(i) ions, spontaneous DNA-mediated reduction occurs, which we propose is followed by a slower change in coordination environment (from pentacoordinate CuII to tetrahedral CuI) to generate copper(i) helicates. We demonstrate that the reduction of the adjacent and distal bis-phenanthroline sites is dependent on their proximity to DNA guanine bases (which act as the electron source). The kinetics of helical charge transport can thus be tuned based on guanine-CuII separation, resulting in a sequence- and distance-dependent redox switch that transfers electronic information from DNA to multiple linearly-arranged metal centers.

Guest Binding via N-H⋠⋠⋠π Bonding and Kinetic Entrapment by an Inorganic Macrocycle.

Modern supramolecular chemistry is overwhelmingly based on non-covalent interactions involving organic architectures. However, the question of what happens when you depart from this area to the supramolecular chemistry of structures based on non-carbon frameworks remains largely unanswered, and is an area that potentially provides new directions in molecular activation, host-guest chemistry, and biomimetic chemistry. In this work, we explore the unusual host-guest chemistry of the pentameric macrocycle [{P(µ-Nt Bu}2 NH]5 with a range of anionic and neutral guests. The polar coordination site of this host promotes new modes of guest encapsulation via hydrogen bonding with the π systems of the unsaturated C≡C and C≡N bonds of acetylenes and nitriles as well as with the PCO- anion. Halide guests can be kinetically locked within the structure by oxidation of the phosphorus periphery by oxidation to PV . Our study underscores the future promise of p-block macrocyclic chemistry.

Multisite Binding of Drugs and Natural Products in an Entropically Favorable, Heteroleptic Receptor.

The cavities of artificial receptors are defined by how their components fit together. The encapsulation of specific molecules can thus be engineered by considering geometric principles; however, intermolecular interactions and steric fit scale with receptor size, such that the ability to bind multiple guests from a specific class of compounds remains a current challenge. By employing metal-organic self-assembly, we have prepared a triangular prism from two different ligands that is capable of binding more than 20 different natural products, drugs, and steroid derivatives within its prolate cavity. Encapsulation inflates the host, enhancing its ability to bind other guests in peripheral pockets and thus enabling our system to bind combinations of different drug and natural product cargoes in different locations simultaneously. This new mode of entropically favorable self-assembly thus enables central encapsulation to amplify guest-binding events around the periphery of an artificial receptor.

Hydrogen-Bond-Assisted Symmetry Breaking in a Network of Chiral Metal-Organic Assemblies.

Herein we elucidate the interplay of chiral, chelate, solvent, and hydrogen-bonding information in the self-assembly of a series of new three-dimensional metal-organic architectures. Enantiopure ligands, each containing H-bond donors and acceptors, form different structures, depending on the ratio in which they are combined: enantiopure components form M4L4 assemblies, whereas racemic mixtures form M3L3 stacks. Chiral amplification within M3L3 enantiomers was observed when a 2:1 ratio of R and S subcomponent enantiomers was employed. Simply switching the solvent (from MeCN to MeOH) or chelating unit (from bidentate to tridentate) increased the diversity of structures that can be generated from these building blocks, leading to the selective formation of novel M2L2 and M3L2 assemblies. The addition of achiral ligand building blocks resulted in the formation of further structures: When an achiral subcomponent was combined with its R and S chiral congeners, a three-layer heteroleptic architecture was generated, with the achiral unit sitting at the top of the stack. When combined with the S enantiomer only, however, the achiral unit assembled in the center of the structure, thus demonstrating the selective placement of achiral units within chiral systems. Further sorting experiments revealed that combining R and S stereocenters within a single ligand led to diastereoselective product generation. These results show how geometric complementarity between different ligands impacts upon the degree of hydrogen-bonding within the assembly, stabilizing specific low-symmetry architectures from among many possible structural outcomes.

Fluorometric Recognition of Nucleotides within a Water-Soluble Tetrahedral Capsule.

The design of aqueous probes and binders for complex, biologically relevant anions presents a key challenge in supramolecular chemistry. Herein, a tetrahedral assembly with cationic faces and corners is reported that is capable of discriminating between anionic and neutral guests in water. Electrostatic repulsion between subcomponents can be overcome by the addition of an anionic template, or generating a robust covalent framework by incorporating tris(2-aminoethyl)amine (TREN). The resultant TREN-capped, water-soluble, fluorescent cage binds mono- and poly-phosphoric esters, including nucleotides. Its covalent skeleton renders it stable at micromolar concentrations in water, enabling the fluorometric detection of biologically relevant guests in an aqueous environment. Selective supramolecular encapsulants, such as 1, could enable new sensing applications, such as recognition of toxins and drugs, under biological conditions.