Spatially-Interactive Biomolecular Networks Organized by Nucleic Acid Nanostructures

Abstract

Conspectus Living systems have evolved a variety of nanostructures to control the molecular interactions that mediate many functions including the recognition of targets by receptors, the binding of enzymes to substrates, and the regulation of enzymatic activity. Mimicking these structures outside of the cell requires methods that offer nanoscale control over the organization of individual network components. Advances in DNA nanotechnology have enabled the design and fabrication of sophisticated one-, two- and three-dimensional (1D, 2D and 3D) nanostructures that utilize spontaneous and sequence specific DNA hybridization. Compared to other self-assembling biopolymers, DNA nanostructures offer predictable and programmable interactions, and surface features to which other nanoparticles and bio-molecules can be precisely positioned. The ability to control the spatial arrangement of the components while constructing highly-organized networks will lead to various applications of these systems. For example, DNA nanoarrays with surface displays of molecular probes can sense noncovalent hybridization interactions with DNA, RNA, and proteins and covalent chemical reactions. DNA nanostructures can also align external molecules into well-defined arrays, which may improve the resolution of many structural determination methods, such as X-ray diffraction, cryo-EM, NMR, and super-resolution fluorescence. Moreover, by constraining target entities to specific conformations, self-assembled DNA nanostructures can serve as molecular rulers to evaluate conformation-dependent activities. This Account describes the most recent advances in the DNA nanostructure directed assembly of biomolecular networks and explores the possibility of applying this technology to other fields of study. Recently, several reports have demonstrated the DNA nanostructure directed assembly of spatially-interactive biomolecular networks. For example, researchers have constructed synthetic multi-enzyme cascades by organizing the position of the components using DNA nanoscaffolds in vitro, or by utilizing RNA matrices in vivo. These structures display enhanced efficiency compared to the corresponding unstructured enzyme mixtures. Such systems are designed to mimic cellular function, where substrate diffusion between enzymes is facilitated and reactions are catalyzed with high efficiency and specificity. In addition, researchers have assembled multiple choromophores into arrays using a DNA nanoscaffold that optimizes the relative distance between the dyes and their spatial organization. The resulting artificial light harvesting system exhibits efficient cascading energy transfers. Finally, DNA nanostructures have been used as assembly templates to construct nanodevices that execute rationally-designed behaviors, including cargo loading, transportation and route control.

Multi-enzyme complexes on DNA scaffolds capable of substrate channelling with an artificial swinging arm

“…In Fig. 3a, the activity of this complete two-enzyme nanostructure is compared to that of partially assembled structures including a G6pDH-MDH assembly with freely diffusing NAD + , a G6pDH-NAD + arm assembly with freely diffusing 6 MDH, and an MDH-NAD + arm assembly with freely diffusing G6pDH. For the same total NAD + and enzyme concentrations (100 nM each), the activity of the complete swinging arm structure is ~90-fold higher than that obtained using the same two-enzyme complex but freely diffusing NAD + .…”

Magnetic metal–organic frameworks as scaffolds for spatial co-location and positional assembly of multi-enzyme systems enabling enhanced cascade biocatalysis

“…Several attractive materials and successful strategies have been proposed leading to the design of an arsenal of multienzyme systems for mimicking the natural metabolic pathways in vitro. Examples are the preparations of hydrogels, 15,22 hourglass shaped nanochannel reactor, 23 mesoporous silica nanoparticles, 18,24 formation of inorganic nanocrystal-protein complexes 25 and self-assembled crystals, 26 microbeads, 27,28 DNA nanostructures, 4,5,11,[29][30][31] polymersomes, 14,32 nanoparticles, 33 nanobers, 13,16,34 graphene, 9,35 and metal-organic frameworks (MOFs). 17,36,37 In contrast to other scaffolds, MOFs, which are formed by the self-assembly of metal ions and organic linkers, feature ultrahigh surface area and porosity, uniform pores with tunable sizes, surfaces with variable chemistries, and structural diversity.…”

A DNA tweezer-actuated enzyme nanoreactor

“…Scientists are currently interested in finding ways to mimic enzyme regulatory circuitry outside of the cell 4,5 , not only to increase our knowledge of cellular metabolism but also so that we may create man-made nanoreactors that have potential utility in applications ranging from diagnostics to the production of high-value chemicals [6][7][8] and smart materials 9 . DNA nanostructures are promising scaffolds for use in the organization of molecules on the nanoscale because they can be engineered to site-specifically incorporate functional elements in precise geometries [10][11][12] and to enable nanomechanical control capabilities 13,14 . Examples of such structures include autonomous walkers 15,16 , nanotweezers [17][18][19][20] and nanocages for controlled encapsulation and payload release 21,22 .…”