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Enzymatic reactions can be coupled together by carefully organizing the enzymes on DNA scaffolds.

Most previously reported methods for purifying DNA-origami nanostructures rely on agarose-gel electrophoresis (AGE) for separation. Although AGE is routinely used to yield 0.1–1 µg purified DNA nanostructures, obtaining >100 µg of purified DNA-origami structure through AGE is typically laborious because of the post-electrophoresis extraction, desalting and concentration steps. Here, we present a readily scalable purification approach utilizing rate-zonal centrifugation, which provides comparable separation resolution as AGE. The DNA nanostructures remain in aqueous solution throughout the purification process. Therefore, the desired products are easily recovered with consistently high yield (40–80%) and without contaminants such as residual agarose gel or DNA intercalating dyes. Seven distinct three-dimensional DNA-origami constructs were purified at the scale of 0.1–100 µg (final yield) per centrifuge tube, showing the versatility of this method. Given the commercially available equipment for gradient mixing and fraction collection, this method should be amenable to automation and further scale up for preparation of larger amounts (e.g. milligram quantities) of DNA nanostructures.

We report a novel stepwise layer-by-layer synthesis strategy to achieve multi-component barcode nanospheres that contain magnetic nanoparticles (MNPs) as the core and quantum dots (QDs) of different emission colors in spatially separated silica layers as the shells, with QD-free silica layers as the insulation layers. This strategy offers the following unique features: (1) the location of the MNPs and the QDs in the silica spheres are separated spatially, so that no interference of the QD photoluminescence (PL) by the magnetic particles is observed; (2) the PL spectra of barcode nanospheres can be easily tuned through the ratio of different QDs loaded in each layer; (3) the size of the silica nanospheres can range from submicron (∼100 nm) to micrometers depending on the number of layers and the thickness of each layer; (4) QD stability is preserved by embedding the QDs covalently in the silica matrix; (5) fluorescence resonance energy transfer (FRET) between different colored QDs is avoided by isolating them into separated layers with a silica spacer layer.

The need for a renewable and sustainable energy source has become apparent, and focusing on a bioinspired approach emphasizing the water splitting mechanism in photosynthesis is a challenging, yet practical and attainable strategy. In this work, a stable framework consisting of a three-dimensional DNA tetrahedron has been used for the design of a biomimic of the Oxygen-Evolving Complex (OEC) found in natural Photosystem II (PSII). In nature, one of Photosystem II's core proteins, D1, is degraded every half hour in the presence of sunlight. D1's sensitivity photodamage resides in triplet state formation of chlorophyll P680+, the primary donor of PSII. Our project aims to build the heart of the OEC, including the Mn4CaCl metal cluster and its protein environment in stable DNA nanocages, which can be connected to a photostable artificial reaction center that performs light-induced charge separation. The peptide sequences responsible for coordinating the cluster have been identified through x-ray structure analysis. RTruncated regions of the peptide sequences containing Mn4CaCl ligation sites are implemented in the design of the aOEC and are attached to sites within the tetrahedron to facilitate assembly. Crystals of the tetrahedron have been obtained, and X-ray crystallography has been used for characterization. As a proof of concept, metal-containing porphyrins, specifically Fe(III) meso-Tetra(4-sulfonatophenyl) porphine chloride, have been implemented for testing of the encapsulation of a metal-containing molecule inside of the DNA cage. Upon reduction of the metal through coordination of two orthogonally oriented peptides covalently attached to the DNA tetrahedron which contain terminal histidine residues, iron (III) becomes EPR active and the assembly can be analyzed electrochemically. Upon assembly of the complete metal-containing center (both porphyrin molecule and aOEC), x-ray crystallography, EPR, and electrochemistry will be used to test functionality in the stable DNA framework.