Evelyn Auyeung

Spherical Nucleic Acids

A historical perspective of the development of spherical nucleic acid (SNA) conjugates and other three-dimensional nucleic acid nanostructures is provided. This Perspective details the synthetic methods for preparing them, followed by a discussion of their unique properties and theoretical and experimental models for understanding them. Important examples of technological advances made possible by their fundamental properties spanning the fields of chemistry, molecular diagnostics, gene regulation, medicine, and materials science are also presented.

DNA-mediated nanoparticle crystallization into Wulff polyhedra

Crystallization is a fundamental and ubiquitous process much studied over the centuries. But although the crystallization of atoms is fairly well understood, it remains challenging to predict reliably the outcome of molecular crystallization processes that are complicated by various molecular interactions and solvent involvement. This difficulty also applies to nanoparticles: high-quality three-dimensional crystals are mostly produced using drying and sedimentation techniques that are often impossible to rationalize and control to give a desired crystal symmetry, lattice spacing and habit (crystal shape). In principle, DNA-mediated assembly of nanoparticles offers an ideal opportunity for studying nanoparticle crystallization: a well-defined set of rules have been developed to target desired lattice symmetries and lattice constants, and the occurrence of features such as grain boundaries and twinning in DNA superlattices and traditional crystals comprised of molecular or atomic building blocks suggests that similar principles govern their crystallization. But the presence of charged biomolecules, interparticle spacings of tens of nanometres, and the realization so far of only polycrystalline DNA-interconnected nanoparticle superlattices, all suggest that DNA-guided crystallization may differ from traditional crystal growth. Here we show that very slow cooling, over several days, of solutions of complementary-DNA-modified nanoparticles through the melting temperature of the system gives the thermodynamic product with a specific and uniform crystal habit. We find that our nanoparticle assemblies have the Wulff equilibrium crystal structure that is predicted from theoretical considerations and molecular dynamics simulations, thus establishing that DNA hybridization can direct nanoparticle assembly along a pathway that mimics atomic crystallization.

Mechanism for the endocytosis of spherical nucleic acid nanoparticle conjugates.

Intracellular delivery of nucleic acids as gene regulation agents typically requires the use of cationic carriers or viral vectors, yet issues related to cellular toxicity or immune responses hamper their attractiveness as therapeutic candidates. The discovery that spherical nucleic acids (SNAs), polyanionic structures comprised of densely packed, highly oriented oligonucleotides covalently attached to the surface of nanoparticles, can effectively enter more than 50 different cell types presents a potential strategy for overcoming the limitations of conventional transfection agents. Unfortunately, little is known about the mechanism of endocytosis of SNAs, including the pathway of entry and specific proteins involved. Here, we demonstrate that the rapid cellular uptake kinetics and intracellular transport of SNAs stem from the arrangement of oligonucleotides into a 3D architecture, which supports their targeting of class A scavenger receptors and endocytosis via a lipid-raft–dependent, caveolae-mediated pathway. These results reinforce the notion that SNAs can serve as therapeutic payloads and targeting structures to engage biological pathways not readily accessible with linear oligonucleotides.

Nucleic acid-metal organic framework (MOF) nanoparticle conjugates

Nanoparticles of a metal-organic framework (MOF), UiO-66-N3 (Zr6O4OH4(C8H3O4-N3)6), were synthesized. The surface of the MOF was covalently functionalized with oligonucleotides, utilizing a strain promoted click reaction between DNA appended with dibenzylcyclooctyne and azide-functionalized UiO-66-N3 to create the first MOF nanoparticle-nucleic acid conjugates. The structure of the framework was preserved throughout the chemical transformation, and the surface coverage of DNA was quantified. Due to the small pore sizes, the particles are only modified on their surfaces. When dispersed in aqueous NaCl, they exhibit increased stability and enhanced cellular uptake when compared with unfunctionalized MOF particles of comparable size.

Polyvalent Nucleic Acid Nanostructures

Polyvalent oligonucleotide-nanoparticle conjugates possess several unique emergent properties, including enhanced cellular uptake, high antisense bioactivity, and nuclease resistance, which hypothetically originate from the dense packing and orientation of oligonucleotides on the surface of the nanoparticle. In this Communication, we describe a new class of polyvalent nucleic acid nanostructures (PNANs), which are comprised of only cross-linked and oriented nucleic acids. We demonstrate that these particles are capable of effecting high cellular uptake and gene regulation without the need of a cationic polymer co-carrier. The PNANs also exhibit cooperative binding behavior and nuclease resistance properties.

Topotactic Interconversion of Nanoparticle Superlattices

Sticking with DNA One strategy for creating superlattices from nanoparticles is to coat the particles with DNA strands that have sticky ends that can be exploited to control the assembly of the lattice. This method can create binary lattices, but now Macfarlane et al. (p. 1222, published online 22 August) have succeeded in inserting a third type of nanoparticle into a predetermined site by tuning the strength of the relative DNA binding interactions. DNA-coated nanoparticles can reversibly incorporate a third nanoparticle. The directed assembly of nanoparticle building blocks is a promising method for generating sophisticated three-dimensional materials by design. In this work, we have used DNA linkers to synthesize nanoparticle superlattices that have greater complexity than simple binary systems using the process of topotactic intercalation—the insertion of a third nanoparticle component at predetermined sites within a preformed binary lattice. Five distinct crystals were synthesized with this methodology, three of which have no equivalent in atomic or molecular crystals, demonstrating a general approach for assembling highly ordered ternary nanoparticle superlattices whose structures can be predicted before their synthesis. Additionally, the intercalation process was demonstrated to be completely reversible; the inserted nanoparticles could be expelled into solution by raising the temperature, and the ternary superlattice could be recovered by cooling.

Transitioning DNA‐Engineered Nanoparticle Superlattices from Solution to the Solid State

In the assembly and crystallization of nanoparticles into ordered lattices, DNA is a powerful structure-directing ligand because its programmability allows for a priori control over the lattice symmetries and lattice constants of the nanoparticle superstructures. [ 1–8 ] Practically, however, characterization and processing of superlattices made from DNA-modifi ed particles is limited because the morphology and programmability of these structures exhibited in solution are either distorted or lost entirely when they are removed from their assembly medium (aqueous saline solution). Because these superlattices are held together via cooperative DNA duplexes, they rapidly collapse or dissociate where these interactions are unfavorable, such as in distilled water, in common organic solvents, at high temperatures, or under vacuum. Therefore, the development of a method for improving the mechanical stability and solution processability of the nanoparticle lattices is a necessary step as studies of these materials shift from understanding the parameters that govern their assembly to pursuing fundamental properties and useful applications. [ 9–11 ] In this work, we report a method for stabilizing DNA-assembled three-dimensional superlattices in the solid state by silica encapsulation, where both the symmetries and lattice spacings of the solution-phase lattices are preserved. Once encapsulated, superlattice morphologies are no longer dictated by DNA interactions, and as such remain stable against distortion, collapse, or dissociation under many previously inaccessible conditions. Silica encapsulation is a technique commonly used to stabilize inorganic nanoparticles of varying chemical compositions and morphologies against aggregation or oxidation. [ 12–17 ]

Stepwise Evolution of DNA‐Programmable Nanoparticle Superlattices

Colloidal crystals can be assembled using a variety of entropic, depletion, electrostatic, or biorecognition forces and provide a convenient model system for studying crystal growth. Although superlattices with diverse geometries can be assembled in solution and on surfaces, the incorporation of specific bonding interactions between particle building blocks and a substrate would significantly enhance control over the growth process. Herein, we use a stepwise growth process to systematically study and control the evolution of a body-centered cubic (bcc) crystalline thinfilm comprised of nanoparticle building blocks functionalized with DNA on a complementary DNA substrate. We examine crystal growth as a function of temperature, number of layers, and substrate–particle bonding interactions. Importantly, the judicious choice of DNA interconnects allows one to tune the interfacial energy between various crystal planes and the substrate, and thereby control crystal orientation and size in a stepwise fashion using chemically programmable attractive forces. This is a unique approach since prior studies involving superlattice assembly typically rely on repulsive interactions between particles to dictate structure, and those that rely on attractive forces (e.g. ionic systems) still maintain repulsive particle–substrate interactions. In addition to providing a model for crystallization, the field of particle assembly has garnered considerable interest because materials generated from ordered particle arrays can have novel optical, 13–17] electronic, and magnetic properties. These properties can be sensitive to the composition, symmetry, and distance between nanoparticles, in addition to the number of layers and orientation. DNA-mediated nanoparticle crystallization is particularly attractive for preparing these materials because the nanoparticle building blocks can be considered a type of “programmable atom equivalent” with tailorable size, composition, shape, and bonding interactions. This tunability allows one to access a diverse class of crystal symmetries, tailor lattice parameters with sub-nanometer resolution, and create structures that have no known mineral equivalent. Indeed, to date, 17 unique symmetries have been realized and over 100 unique crystal structures have been synthesized, all of which conform to a key hypothesis: these atom equivalents assemble into structures that maximize the total number of hybridized DNA interconnects between particles. While these structures have enormous potential, their use is limited because they are typically formed in solution as polycrystalline aggregates with little control over crystal size or orientation. Consequently, it is difficult to measure their properties or integrate them with other device elements using existing microfabrication techniques. The development of thin-film superlattices is therefore necessary to fully realize the potential of these structures as metamaterials, photonic crystals, and data storage elements. The growth of DNA-mediated monoand multi-layered nanoparticle structures was first examined by our group and later by Niemeyer and co-workers. However, the use of strong DNA interactions prevented nanoparticle crystallization. Herein, we exploit multiple weak DNA interactions for superlattice growth to examine the development of crystal orientation (texture) and control film thickness. Body-centered cubic colloidal crystals composed of spherical nucleic acid gold nanoparticle conjugates (SNA-AuNPs) were used as a model system since these structures require two complementary particle types and therefore allow the stepwise introduction of each layer. Alternatively, other crystal symmetries such as face-centered cubic (fcc) require

DNA-mediated engineering of multicomponent enzyme crystals

Significance Due to their unique structures and diverse catalytic functionalities, proteins represent a nearly limitless set of precursors for constructing functional supramolecular materials. However, programming the assembly of even a single protein into ordered superlattices is a difficult task, and a generalizable strategy for coassembling multiple proteins with distinct surface chemistries, or proteins and inorganic nanoparticles, does not currently exist. Here, we use the high-fidelity interactions characteristic of DNA–DNA “bonds” to direct the assembly of two proteins into six unique superlattices composed of either a single protein, multiple proteins, or proteins and gold nanoparticles. Significantly, the DNA-functionalized proteins retain their native catalytic functionalities both in the solution and crystalline states. The ability to predictably control the coassembly of multiple nanoscale building blocks, especially those with disparate chemical and physical properties such as biomolecules and inorganic nanoparticles, has far-reaching implications in catalysis, sensing, and photonics, but a generalizable strategy for engineering specific contacts between these particles is an outstanding challenge. This is especially true in the case of proteins, where the types of possible interparticle interactions are numerous, diverse, and complex. Herein, we explore the concept of trading protein–protein interactions for DNA–DNA interactions to direct the assembly of two nucleic-acid–functionalized proteins with distinct surface chemistries into six unique lattices composed of catalytically active proteins, or of a combination of proteins and DNA-modified gold nanoparticles. The programmable nature of DNA–DNA interactions used in this strategy allows us to control the lattice symmetries and unit cell constants, as well as the compositions and habit, of the resulting crystals. This study provides a potentially generalizable strategy for constructing a unique class of materials that take advantage of the diverse morphologies, surface chemistries, and functionalities of proteins for assembling functional crystalline materials.

Controlling Structure and Porosity in Catalytic Nanoparticle Superlattices with DNA

Herein, we describe a strategy for converting catalytically inactive, highly crystalline nanoparticle superlattices embedded in silica into catalytically active, porous structures through superlattice assembly and calcination. First, a body-centered cubic (bcc) superlattice is synthesized through the assembly of two sets of 5 nm gold nanoparticles chemically modified with DNA bearing complementary sticky end sequences. These superlattices are embedded in silica and calcined at 350 °C to provide access to the catalytic nanoparticle surface sites. The calcined superlattice maintains its bcc ordering and has a surface area of 210 m(2)/g. The loading of catalytically active nanoparticles within the superlattice was determined by inductively coupled plasma mass spectrometry, which revealed that the calcined superlattice contained approximately 10% Au by weight. We subsequently investigate the ability of supported Au nanoparticle superlattices to catalyze alcohol oxidation. In addition to demonstrating that calcined superlattices are effective catalysts for alcohol oxidation, electron microscopy reveals preservation of the crystalline structure of the bcc superlattice following calcination and catalysis. Unlike many bulk nanoparticle catalysts, which are difficult to characterize and susceptible to aggregation, nanoparticle superlattices synthesized using DNA interactions offer an attractive bottom-up route to structurally defined heterogeneous catalysts, where one has the potential to independently control nanoparticle size, nanoparticle compositions, and interparticle spacings.