Programming crystallization kinetics of self-assembled DNA crystals with 5-methylcytosine modification.

Self-assembled DNA crystals offer a precise chemical platform at the ångström-scale for DNA nanotechnology, holding enormous potential in material separation, catalysis, and DNA data storage. However, accurately controlling the crystallization kinetics of such DNA crystals remains challenging. Herein, we found that atomic-level 5-methylcytosine (5mC) modification can regulate the crystallization kinetics of DNA crystal by tuning the hybridization rates of DNA motifs. We discovered that by manipulating the axial and combination of 5mC modification on the sticky ends of DNA tensegrity triangle motifs, we can obtain a series of DNA crystals with controllable morphological features. Through DNA-PAINT and FRET-labeled DNA strand displacement experiments, we elucidate that atomic-level 5mC modification enhances the affinity constant of DNA hybridization at both the single-molecule and macroscopic scales. This enhancement can be harnessed for kinetic-driven control of the preferential growth direction of DNA crystals. The 5mC modification strategy can overcome the limitations of DNA sequence design imposed by limited nucleobase numbers in various DNA hybridization reactions. This strategy provides a new avenue for the manipulation of DNA crystal structure, valuable for the advancement of DNA and biomacromolecular crystallography.

Atomically precise photothermal nanomachines.

Interfacing molecular machines to inorganic nanoparticles can, in principle, lead to hybrid nanomachines with extended functions. Here we demonstrate a ligand engineering approach to develop atomically precise hybrid nanomachines by interfacing gold nanoclusters with tetraphenylethylene molecular rotors. When gold nanoclusters are irradiated with near-infrared light, the rotation of surface-decorated tetraphenylethylene moieties actively dissipates the absorbed energy to sustain the photothermal nanomachine with an intact structure and steady efficiency. Solid-state nuclear magnetic resonance and femtosecond transient absorption spectroscopy reveal that the photogenerated hot electrons are rapidly cooled down within picoseconds via electron-phonon coupling in the nanomachine. We find that the nanomachine remains structurally and functionally intact in mammalian cells and in vivo. A single dose of near-infrared irradiation can effectively ablate tumours without recurrence in tumour-bearing mice, which shows promise in the development of nanomachine-based theranostics.

Welded Gold Nanoparticle Assemblies Defined Plasmonic Coupling.

Nanoparticle assemblies with interparticle ohmic contacts are crucial for nanodevice fabrication. Despite tremendous progress in DNA-programmable nanoparticle assemblies, seamlessly welding discrete components into welded continuous three-dimensional (3D) configurations remains challenging. Here, we introduce a single-stranded DNA-encoded strategy to customize welded metal nanostructures with tunable morphologies and plasmonic properties. We demonstrate the precise welding of gold nanoparticle assemblies into continuous metal nanostructures with interparticle ohmic contacts through chemical welding in solution. We find that the welded gold nanoparticle assemblies show a consistent morphology with welded efficiency over 90%, such as the rod-like, triangular, and tetrahedral metal nanostructures. Next, we show the versatility of this strategy by welding gold nanoparticle assemblies of varied sizes and shapes. Furthermore, the experiment and simulation show that the welded gold nanoparticle assemblies exhibit defined plasmonic coupling. This single-stranded DNA encoded welding system may provide a new route for accurately building functional plasmonic nanomaterials and devices.

Programmed Remodeling of the Tumor Milieu to Enhance NK Cell Immunotherapy Combined with Chemotherapy for Pancreatic Cancer.

Natural killer (NK) cell-based adoptive immunotherapy has demonstrated encouraging therapeutic effects in clinical trials for hematological cancers. However, the effectiveness of treatment for solid tumors remains a challenge due to insufficient recruitment and infiltration of NK cells into tumor tissues. Herein, a programmed nanoremodeler (DAS@P/H/pp) is designed to remodel dense physical stromal barriers and for dysregulation of the chemokine of the tumor environment to enhance the recruitment and infiltration of NK cells in tumors. The DAS@P/H/pp is triggered by the acidic tumor environment, resulting in charge reversal and subsequent hyaluronidase (HAase) release. HAase effectively degrades the extracellular matrix, promoting the delivery of immunoregulatory molecules and chemotherapy drugs into deep tumor tissues. In mouse models of pancreatic cancer, this nanomediated strategy for the programmed remodeling of the tumor microenvironment significantly boosts the recruitment of NK92 cells and their tumor cell-killing capabilities under the supervision of multiplexed near-infrared-II fluorescence.

Spacer-Programmed Two-Dimensional DNA Origami Assembly.

Two-dimensional (2D) DNA origami assembly represents a powerful approach to the programmable design and construction of advanced 2D materials. Within the context of hybridization-mediated 2D DNA origami assembly, DNA spacers play a pivotal role as essential connectors between sticky-end regions and DNA origami units. Here, we demonstrated that programming the spacer length, which determines the binding radius of DNA origami units, could effectively tune sticky-end hybridization reactions to produce distinct 2D DNA origami arrays. Using DNA-PAINT super-resolution imaging, we unveiled the significant impact of spacer length on the hybridization efficiency of sticky ends for assembling square DNA origami (SDO) units. We also found that the assembly efficiency and pattern diversity of 2D DNA origami assemblies were critically dependent on the spacer length. Remarkably, we realized a near-unity yield of ∼98% for the assembly of SDO trimers and tetramers via this spacer-programmed strategy. At last, we revealed that spacer lengths and thermodynamic fluctuations of SDO are positively correlated, using molecular dynamics simulations. Our study thus paves the way for the precision assembly of DNA nanostructures toward higher complexity.

Twisted DNA Origami-Based Chiral Monolayers for Spin Filtering.

DNA monolayers with inherent chirality play a pivotal role across various domains including biosensors, DNA chips, and bioelectronics. Nonetheless, conventional DNA chiral monolayers, typically constructed from single-stranded DNA (ssDNA) or double-stranded DNA (dsDNA), often lack structural orderliness and design flexibility at the interface. Structural DNA nanotechnology has emerged as a promising solution to tackle these challenges. In this study, we present a strategy for crafting highly adaptable twisted DNA origami-based chiral monolayers. These structures exhibit distinct interfacial assembly characteristics and effectively mitigate the structural disorder of dsDNA monolayers, which is constrained by a limited persistence length of ∼50 nm of dsDNA. We highlight the spin-filtering capabilities of seven representative DNA origami-based chiral monolayers, demonstrating a maximal one-order-of-magnitude increase in spin-filtering efficiency per unit area compared with conventional dsDNA chiral monolayers. Intriguingly, our findings reveal that the higher-order tertiary chiral structure of twisted DNA origami further enhances the spin-filtering efficiency. This work paves the way for the rational design of DNA chiral monolayers.

Complex silica composite nanomaterials templated with DNA origami.

Genetically encoded protein scaffolds often serve as templates for the mineralization of biocomposite materials with complex yet highly controlled structural features that span from nanometres to the macroscopic scale1-4. Methods developed to mimic these fabrication capabilities can produce synthetic materials with well defined micro- and macro-sized features, but extending control to the nanoscale remains challenging5,6. DNA nanotechnology can deliver a wide range of customized nanoscale two- and three-dimensional assemblies with controlled sizes and shapes7-11. But although DNA has been used to modulate the morphology of inorganic materials12,13 and DNA nanostructures have served as moulds14,15 and templates16,17, it remains challenging to exploit the potential of DNA nanostructures fully because they require high-ionic-strength solutions to maintain their structure, and this in turn gives rise to surface charging that suppresses the material deposition. Here we report that the Stöber method, widely used for producing silica (silicon dioxide) nanostructures, can be adjusted to overcome this difficulty: when synthesis conditions are such that mineral precursor molecules do not deposit directly but first form clusters, DNA-silica hybrid materials that faithfully replicate the complex geometric information of a wide range of different DNA origami scaffolds are readily obtained. We illustrate this approach using frame-like, curved and porous DNA nanostructures, with one-, two- and three-dimensional complex hierarchical architectures that range in size from 10 to 1,000 nanometres. We also show that after coating with an amorphous silica layer, the thickness of which can be tuned by adjusting the growth time, hybrid structures can be up to ten times tougher than the DNA template while maintaining flexibility. These findings establish our approach as a general method for creating biomimetic silica nanostructures.

DNA-mediated regioselective encoding of colloids for programmable self-assembly.

How far we can push chemical self-assembly is one of the most important scientific questions of the century. Colloidal self-assembly is a bottom-up technique for the rational design of functional materials with desirable collective properties. Due to the programmability of DNA base pairing, surface modification of colloidal particles with DNA has become fundamental for programmable material self-assembly. However, there remains an ever-lasting demand for surface regioselective encoding to realize assemblies that require specific, directional, and orthogonal interactions. Recent advances in surface chemistry have enabled regioselective control over the formation of DNA bonds on the particle surface. In particular, the structural DNA nanotechnology provides a simple yet powerful design strategy with unique regioselective addressability, bringing the complexity of colloidal self-assembly to an unprecedented level. In this review, we summarize the state-of-art advances in DNA-mediated regioselective surface encoding of colloids, with a focus on how the regioselective encoding is introduced and how the regioselective DNA recognition plays a crucial role in the self-assembly of colloidal structures. This review highlights the advantages of DNA-based regioselective modification in improving the complexity of colloidal assembly, and outlines the challenges and opportunities for the construction of more complex architectures with tailored functionalities.

Single-Stranded RNA Origami-Based Epigenetic Immunomodulation.

The integration of functional modules at the molecular level into RNA nanostructures holds great potential for expanding their applications. However, the quantitative integration of nucleoside analogue molecules into RNA nanostructures and their impact on the structure and function of RNA nanostructures remain largely unexplored. Here, we report a transcription-based approach to controllably integrate multiple nucleoside analogues into a 2000 nucleotide (nt) single-stranded RNA (ssRNA) origami nanostructure. The resulting integrated ssRNA origami preserves the morphology and biostability of the original ssRNA origami. Moreover, the integration of nucleoside analogues introduced new biomedical functions to ssRNA origamis, including innate immune recognition and regulation after the precise integration of epigenetic nucleoside analogues and synergistic effects on tumor cell killing after integration of therapeutic nucleoside analogues. This study provides a promising approach for the quantitative integration of functional nucleoside analogues into RNA nanostructures at the molecular level, thereby offering valuable insights for the development of multifunctional ssRNA origamis.

DNA Framework-Templated Fabrication of Ultrathin Electroactive Gold Nanosheets.

Generally, two-dimensional gold nanomaterials have unique properties and functions that offer exciting application prospects. However, the crystal phases of these materials tend to be limited to the thermodynamically stable crystal structure. Herein, we report a DNA framework-templated approach for the ambient aqueous synthesis of freestanding and microscale amorphous gold nanosheets with ultrathin sub-nanometer thickness. We observe that extended single-stranded DNA on DNA nanosheets can induce site-specific metallization and enable precise modification of the metalized nanostructures at predefined positions. More importantly, the as-prepared gold nanosheets can serve as an electrocatalyst for glucose oxidase-catalyzed aerobic oxidation, exhibiting enhanced electrocatalytic activity (~3-fold) relative to discrete gold nanoclusters owing to a larger electrochemical active area and wider band gap. The proposed DNA framework-templated metallization strategy is expected to be applicable in a broad range of fields, from catalysis to new energy materials.

Micron-Scale Fabrication of Ultrathin Amorphous Copper Nanosheets Templated by DNA Scaffolds.

Two-dimensional (2D) amorphous materials could outperform their crystalline counterparts toward various applications because they have more defects and reactive sites and thus could exhibit a unique surface chemical state and provide an advanced electron/ion transport path. Nevertheless, it is challenging to fabricate ultrathin and large-sized 2D amorphous metallic nanomaterials in a mild and controllable manner due to the strong metallic bonds between metal atoms. Here, we reported a simple yet fast (10 min) DNA nanosheet (DNS)-templated method to synthesize micron-scale amorphous copper nanosheets (CuNSs) with a thickness of 1.9 ± 0.4 nm in aqueous solution at room temperature. We demonstrated the amorphous feature of the DNS/CuNSs by transmission electron microscopy (TEM) and X-ray diffraction (XRD). Interestingly, we found that they could transform to crystalline forms under continuous electron beam irradiation. Of note, the amorphous DNS/CuNSs exhibited much stronger photoemission (∼62-fold) and photostability than dsDNA-templated discrete Cu nanoclusters due to the elevation of both the conduction band (CB) and valence band (VB). Such ultrathin amorphous DNS/CuNSs hold great potential for practical applications in biosensing, nanodevices, and photodevices.

Edge Length-Programmed Single-Stranded RNA Origami for Predictive Innate Immune Activation and Therapy.

Ligands targeting nucleic acid-sensing receptors activate the innate immune system and play a critical role in antiviral and antitumoral therapy. However, ligand design for in situ stability, targeted delivery, and predictive immunogenicity is largely hampered by the sophisticated mechanism of the nucleic acid-sensing process. Here, we utilize single-stranded RNA (ssRNA) origami with precise structural designability as nucleic acid sensor-based ligands to achieve improved biostability, organelle-level targeting, and predictive immunogenicity. The natural ssRNAs self-fold into compact nanoparticles with defined shapes and morphologies and exhibit resistance against RNase digestion in vitro and prolonged retention in macrophage endolysosomes. We find that programming the edge length of ssRNA origami can precisely regulate the degree of macrophage activation via a toll-like receptor-dependent pathway. Further, we demonstrate that the ssRNA origami-based ligand elicits an anti-tumoral immune response of macrophages and neutrophils in the tumor microenvironment and retards tumor growth in the mouse pancreatic tumor model. Our ssRNA origami strategy utilizes structured RNA ligands to achieve predictive immune activation, providing a new solution for nucleic acid sensor-based ligand design and biomedical applications.

Probing Heterogeneous Folding Pathways of DNA Origami Self-Assembly at the Molecular Level with Atomic Force Microscopy.

A myriad of DNA origami nanostructures have been demonstrated in various intriguing applications. In pursuit of facile yet high-yield synthesis, the mechanisms underlying DNA origami folding need to be resolved. Here, we visualize the folding processes of several multidomain DNA origami structures under ambient annealing conditions in solution using atomic force microscopy with submolecular resolution. We reveal the coexistence of diverse transitional structures that might result in the same prescribed products. Based on the experimental observations and the simulation of the energy landscapes, we propose the heterogeneity of the folding pathways of multidomain DNA origami structures. Our findings may contribute to understanding the high-yield folding mechanism of DNA origami.

Single-Stranded DNA-Encoded Gold Nanoparticle Clusters as Programmable Enzyme Equivalents.

Nanozymes have emerged as a class of novel catalytic nanomaterials that show great potential to substitute natural enzymes in various applications. Nevertheless, spatial organization of multiple subunits in a nanozyme to rationally engineer its catalytic properties remains to be a grand challenge. Here, we report a DNA-based approach to encode the organization of gold nanoparticle clusters (GNCs) for the construction of programmable enzyme equivalents (PEEs). We find that single-stranded (ss-) DNA scaffolds can self-fold into nanostructures with prescribed poly-adenine (polyA) loops and double-stranded stems and that the polyA loops serve as specific sites for seed-free nucleation and growth of GNCs with well-defined particle numbers and interparticle spaces. A spectrum of GNCs, ranging from oligomers with discrete particle numbers (2-4) to polymer-like chains, are in situ synthesized in this manner. The polymeric GNCs with multiple spatially organized nanoparticles as subunits show programmable peroxidase-like catalytic activity that can be tuned by the scaffold size and the inter-polyA spacer length. This study thus opens new routes to the rational design of nanozymes for various biological and biomedical applications.

Water-Dispersible Gold Nanoclusters: Synthesis Strategies, Optical Properties, and Biological Applications.

Atomically precise gold nanoclusters (AuNCs) are an emerging class of quantum-sized nanomaterials. Intrinsic discrete electronic energy levels have endowed them with fascinating electronic and optical properties. They have been widely applied in the fields of optoelectronics, photovoltaics, catalysis, biochemical sensing, bio-imaging, and therapeutics. Nevertheless, most AuNCs are synthesized in organic solvents and do not disperse in aqueous solutions; this restricts their biological applications. In this review, we focus on the recent progress in the preparation of water-dispersible AuNCs and their biological applications. We first review different methods of synthesis, including direct synthesis from hydrophilic templates and indirect phase transfer of hydrophobic AuNCs. We then discuss their photophysical properties, such as emission enhancement and fluorescence lifetimes. Next, we summarize their latest applications in the fields of biosensing, biolabeling, and bioimaging. Finally, we outline the challenges and potential for the future development of these AuNCs.

Sequential Therapy of Acute Kidney Injury with a DNA Nanodevice.

The high demand for acute kidney injury (AKI) therapy calls the development of multifunctional nanomedicine for renal management with programmable pharmacokinetics. Here, we developed a renal-accumulating DNA nanodevice with exclusive kidney retention for longitudinal protection of AKI in different stages in a renal ischemia-reperfusion (I/R) model. Due to the prolonged kidney retention time (>12 h), the ROS-sensitive nucleic acids of the nanodevice could effectively alleviate oxidative stress by scavenging ROS in stage I, and then the anticomplement component 5a (aC5a) aptamer loaded nanodevice could sequentially suppress the inflammatory responses by blocking C5a in stage II, which is directly related to the cytokine storm. This sequential therapy provides durable and pathogenic treatment of kidney dysfunction based on successive pathophysiological events induced by I/R, which holds great promise for renal management and the suppression of the cytokine storm in more broad settings including COVID-19.

Remote Photothermal Control of DNA Origami Assembly in Cellular Environments.

In situ synthesis of DNA origami structures in living systems is highly desirable due to its potential in biological applications, which nevertheless is hampered by the requirement of thermal activation procedures. Here, we report a photothermal DNA origami assembly method in near-physiological environments. We find that the use of copper sulfide nanoparticles (CuS NPs) can mediate efficient near-infrared (NIR) photothermal conversion to remotely control the solution temperature. Under a 4 min NIR illumination and subsequent natural cooling, rapid and high-yield (>80%) assembly of various types of DNA origami nanostructures is achieved as revealed by atomic force microscopy and single-molecule fluorescence resonance energy transfer analysis. We further demonstrate the in situ assembly of DNA origami with high location precision in cell lysates and in cell culture environments.

DNA Origami-Encoded Integration of Heterostructures.

Integrating dissimilar materials at the nanoscale is crucial for modern electronics and optoelectronics. The structural DNA nanotechnology provides a universal platform for precision assembly of materials; nevertheless, heterogeneous integration of dissimilar materials with DNA nanostructures has yet to be explored. We report a DNA origami-encoded strategy for integrating silica-metal heterostructures. Theoretical and experimental studies reveal distinctive mechanisms for the binding and aggregation of silica and metal clusters on protruding double-stranded DNA (dsDNA) strands that are prescribed on the DNA origami template. In particular, the binding energy differences of silica/metal clusters and DNA molecules underlies the accessibilities of dissimilar material areas on DNA origami. By programming the densities and lengths of protruding dsDNA strands on DNA origami, silica and metal materials can be independently deposited at their predefined areas with a high vertical precision of 2 nm. We demonstrate the integration of silica-gold and silica-silver heterostructures with high site addressability. This DNA nanotechnology-based strategy is thus applicable for integrating various types of dissimilar materials, which opens up new routes to bottom-up electronics.

DNA-Encoded Gold-Gold Wettability for Programmable Plasmonic Engineering.

Controlling the deposition and diffusion of adsorbed atoms (adatoms) on the surface of a solid material is vital for engineering the shape and function of nanocrystals. Here, we report the use of single-stranded DNA (oligo-adenine, oligo-A) to encode the wettability of gold seeds by homogeneous gold adatoms to synthesize highly tunable plasmonic nanostructures. We find that the oligo-A attachment transforms the nanocrystal growth mode from the classical Frank-van der Merwe to the Volmer-Weber island growth. Finely tuning the oligo-A density can continuously change the gold-gold contact angle (θ) from 35.1±3.6° to 125.3±8.0°. We further demonstrate the versatility of this strategy for engineering nanoparticles with different curvature and dimensions. With this unconventional growth mode, we synthesize a sub-nanometer plasmonic cavity with a geometrical singularity when θ>90°. Superfocusing of light in this nanocavity produces a near-infrared intraparticle plasmonic coupling, which paves the way to surface engineering of single-particle plasmonic devices.

Programming nanoparticle valence bonds with single-stranded DNA encoders.

Nature has evolved strategies to encode information within a single biopolymer to program biomolecular interactions with characteristic stoichiometry, orthogonality and reconfigurability. Nevertheless, synthetic approaches for programming molecular reactions or assembly generally rely on the use of multiple polymer chains (for example, patchy particles). Here we demonstrate a method for patterning colloidal gold nanoparticles with valence bond analogues using single-stranded DNA encoders containing polyadenine (polyA). By programming the order, length and sequence of each encoder with alternating polyA/non-polyA domains, we synthesize programmable atom-like nanoparticles (PANs) with n-valence that can be used to assemble a spectrum of low-coordination colloidal molecules with different composition, size, chirality and linearity. Moreover, by exploiting the reconfigurability of PANs, we demonstrate dynamic colloidal bond-breaking and bond-formation reactions, structural rearrangement and even the implementation of Boolean logic operations. This approach may be useful for generating responsive functional materials for distinct technological applications.