Alignment of Gold Nanoparticle-Decorated DNA Origami Nanotubes: Substrate Prepatterning versus Molecular Combing

DNA origami has become an established technique for designing well-defined nanostructures with any desired shape and for the controlled arrangement of functional nanostructures with few nanometer resolution. These unique features make DNA origami nanostructures promising candidates for use as scaffolds in nanoelectronics and nanophotonics device fabrication. Consequently, a number of studies have shown the precise organization of metallic nanoparticles on various DNA origami shapes. In this work, we fabricated large arrays of aligned DNA origami decorated with a high density of gold nanoparticles (AuNPs). To this end, we first demonstrate the high-yield assembly of high-density AuNP arrangements on DNA origami adsorbed to Si surfaces with few unbound background nanoparticles by carefully controlling the concentrations of MgCl₂ and AuNPs in the hybridization buffer and the hybridization time. Then, we evaluate two methods, i.e., hybridization to prealigned DNA origami and molecular combing in a receding meniscus, with respect to their potential to yield large arrays of aligned AuNP-decorated DNA origami nanotubes. Because of the comparatively low MgCl₂ concentration required for the efficient immobilization of the AuNPs, the prealigned DNA origami become mobile and displaced from their original positions, thereby decreasing the alignment yield. This increased mobility, on the other hand, makes the adsorbed origami susceptible to molecular combing, and a total alignment yield of 86% is obtained in this way

Multilayer DNA Origami Packed on Hexagonal and Hybrid Lattices

“Scaffolded DNA origami” has been proven to be a powerful and efficient approach to construct two-dimensional or three-dimensional objects with great complexity. Multilayer DNA origami has been demonstrated with helices packing along either honeycomb-lattice geometry or square-lattice geometry. Here we report successful folding of multilayer DNA origami with helices arranged on a close-packed hexagonal lattice. This arrangement yields a higher density of helical packing and therefore higher resolution of spatial addressing than has been shown previously. We also demonstrate hybrid multilayer DNA origami with honeycomb-lattice, square-lattice, and hexagonal-lattice packing of helices all in one design. The availability of hexagonal close-packing of helices extends our ability to build complex structures using DNA nanotechnology

A DNA-Mediated Homogeneous Binding Assay for Proteins and Small Molecules

Optical detection of molecular targets typically requires immobilization, separation, or chemical or enzymatic processing. An important exception is aptamers that allow optical detection in solution based on conformational changes. This method, however, requires the laborious selection of aptamers with high target specificity and affinity, and the ability to undergo the required conformational changes. Here we report on an alternative generic scheme for detecting small molecules and proteins in solution based on a shift in the equilibrium of DNA-based strand displacement competition reaction. The shift occurs upon binding of a protein, for example, an antibody to its target. We demonstrate nanomolar detection of small molecules such as biotin, digoxigenin, vitamin D, and folate, in buffer and in plasma. The method is flexible, and we also show nanomolar detection of the respective antibodies or protein targets of these molecules. The detection scheme provides a generic alternative to aptamers for detection of analytes

Probing Electron-Induced Bond Cleavage at the Single-Molecule Level Using DNA Origami Templates

Low-energy electrons (LEEs) play an important role in nanolithography, atmospheric chemistry, and DNA radiation damage. Previously, the cleavage of specific chemical bonds triggered by LEEs has been demonstrated in a variety of small organic molecules such as halogenated benzenes and DNA nucleobases. Here we present a strategy that allows for the first time to visualize the electron-induced dissociation of single chemical bonds within complex, but well-defined self-assembled DNA nanostructures. We employ atomic force microscopy to image and quantify LEE-induced bond dissociations within specifically designed oligonucleotide targets that are attached to DNA origami templates. In this way, we use a highly selective approach to compare the efficiency of the electron-induced dissociation of a single disulfide bond with the more complex cleavage of the DNA backbone within a TT dinucleotide sequence. This novel technique enables the fast and parallel determination of DNA strand break yields with unprecedented control over the DNA’s primary and secondary structure. Thus the detailed investigation of DNA radiation damage in its most natural environment, <i>e</i>.<i>g</i>., DNA nucleosomes constituting the chromatin, now becomes feasible

Single Molecule FRET Analysis of the 11 Discrete Steps of a DNA Actuator

DNA hybridization allows the design and assembly of dynamic DNA-based molecular devices. Such structures usually accomplish their function by the addition of fuel strands that drive the structure from one conformation to a new one or by internal changes in DNA hybridization. We report here on the performance and robustness of one of these devices by the detailed study of a dynamic DNA actuator. The DNA actuator was chosen as a model system, as it is the device with most discrete states to date. It is able to reversibly slide between 11 different states and can in principle function both autonomously and nonautonomously. The 11 states of the actuator were investigated by single molecule Förster Resonance Energy Transfer (smFRET) microscopy to obtain information on the static and dynamic heterogeneities of the device. Our results show that the DNA actuator can be effectively locked in several conformations with the help of well-designed DNA lock strands. However, the device also shows pronounced static and dynamic heterogeneities both in the unlocked and locked modes, and we suggest possible structural models. Our study allows for the direct visualization of the conformational diversity and movement of the dynamic DNA-based device and shows that complex DNA-based devices are inherently heterogeneous. Our results also demonstrate that single molecule techniques are a powerful tool for structural dynamics studies and provide a stringent test for the performance of molecular devices made out of DNA

Enzymatic Ligation of Large Biomolecules to DNA

The ability to synthesize, characterize, and manipulate DNA forms the foundation of a range of advanced disciplines including genomics, molecular biology, and biomolecular engineering. In particular for the latter field, DNA has proven useful as a structural or functional component in nanoscale self-assembled structures, antisense therapeutics, microarray diagnostics, and biosensors. Such applications frequently require DNA to be modified and conjugated to other macromolecules, including proteins, polymers, or fatty acids, in order to equip the system with properties required for a particular application. However, conjugation of DNA to large molecular components using classical chemistries often suffers from suboptimal yields. Here, we report the use of terminal deoxynucleotidyl transferase (TdT) for direct enzymatic ligation of native DNA to nucleotide triphosphates coupled to proteins and other large macromolecules. We demonstrate facile synthesis routes for a range of NTP-activated macromolecules and subsequent ligation to the 3′ hydroxyl group of oligodeoxynucleotides using TdT. The reaction is highly specific and proceeds rapidly and essentially to completion at micromolar concentrations. As a proof of principle, parallelly labeled oligonucleotides were used to produce nanopatterned DNA origami structures, demonstrating rapid and versatile incorporation of non-DNA components into DNA nanoarchitectures

On-Surface Azide–Alkyne Cycloaddition on Cu(111): Does It “Click” in Ultrahigh Vacuum?

Using scanning tunneling microscopy, we demonstrate that the 1,3-dipolar cycloaddition between a terminal alkyne and an azide can be performed under solvent-free ultrahigh vacuum conditions with reactants adsorbed on a Cu(111) surface. XPS shows significant degradation of the azide upon adsorption, which is found to be the limiting factor for the reaction

Chiral Induction with Chiral Conformational Switches in the Limit of Low “Sergeants to Soldiers” Ratio

Molecular-level insights into chiral adsorption phenomena are highly relevant within the fields of asymmetric heterogeneous catalysis or chiral separation and may contribute to understand the origins of homochirality in nature. Here, we investigate chiral induction by the “sergeants and soldiers” mechanism for an oligo(phenylene ethynylene) based chiral conformational switch by coadsorbing it with an intrinsically chiral seed on Au(111). Through statistical analysis of scanning tunneling microscopy (STM) data, we demonstrate successful chiral induction with a very low concentration of seeding molecules down to 3%. The microscopic mechanism for the observed chiral induction is suggested to involve nucleation of the intrinsically chiral seeds, allowing for effective transfer and amplification of chirality to large numbers of soldier target molecules

Construction of a Polyhedral DNA 12-Arm Junction for Self-Assembly of Wireframe DNA Lattices

A variety of different tiles for the construction of DNA lattices have been developed since the structural DNA nanotechnology field was born. The majority of these are designed for the realization of close-packed structures, where DNA helices are arranged in parallel and tiles are connected through sticky ends. Assembly of such structures requires the use of cation-rich buffers to minimize repulsion between parallel helices, which poses limits to the application of DNA nanostructures. Wireframe structures, on the other hand, are less susceptible to salt concentration, but the assembly of wireframe lattices is limited by the availability of tiles and motifs. Herein, we report the construction of a polyhedral 12-arm junction for the self-assembly of wireframe DNA lattices. Our approach differs from traditional assembly of DNA tiles through hybridization of sticky ends. Instead, the assembly approach presented here uses small polyhedral shapes as connecting points and branch points of wires in a lattice structure. Using this design principle and characterization techniques, such as transmission electron microscopy, single-particle reconstruction, patterning of gold nanoparticles, dynamic light scattering, UV melting analyses, and small-angle X-ray scattering among others, we demonstrated formation of finite 12-way junction structures, as well as 1D and 2D short assemblies, demonstrating an alternative way of designing polyhedral structures and lattices

Supramolecular Corrals on Surfaces Resulting from Aromatic Interactions of Nonplanar Triazoles

Interaction forces between aromatic moieties, often referred to as π–π interactions, are an important element in stabilizing complex supramolecular structures. For supramolecular self-assembly occurring on surfaces, where aromatic moieties are typically forced to adsorb coplanar with the surface, the possible role of intermolecular aromatic interactions is much less explored. Here, we report on unusual, ring-shaped supramolecular corral surface structures resulting from adsorption of a molecule with nonplanar structure, allowing for intermolecular aromatic interactions. The discrete corral structures are observed using high-resolution scanning tunneling microscopy, and the energetic driving forces for their formation are elucidated using density functional theory calculations and Monte Carlo simulations. The individual corrals involve between 11 and 18 molecules bound through triazole moieties to a ring-shaped ensemble of bridge site positions on (111) surfaces of copper, silver, or gold. The curvature required to form the corrals is identified to result from the angle dependence of aromatic interactions between molecular phenanthrene moieties. The study provides detailed quantitative insights into triazole−surface and aromatic interactions and illustrates how they may be used to drive surface supramolecular self-assembly