Biological systems use compartmentalisation as a general strategy to control enzymatic reactions by precisely regulating enzyme–substrate interactions. With the advent of DNA nanotechnology, it has become possible to rationally design DNA-based nano-containers with programmable structural and dynamic properties. These DNA nanostructures have been used to cage enzymes, but control over enzyme–substrate interactions using a dynamic DNA nanostructure has not been achieved yet. Here we introduce a DNA origami device that functions as a nanoscale vault: an enzyme is loaded in an isolated cavity and the access to free substrate molecules is controlled by a multi-lock mechanism. The DNA vault is characterised for features such as reversible opening/closing, cargo loading and wall porosity, and is shown to control the enzymatic reaction catalysed by an encapsulated protease. The DNA vault represents a general concept to control enzyme–substrate interactions by inducing conformational changes in a rationally designed DNA nanodevice.

Fluorescent RNA aptamers are useful as markers for tracking RNA molecules inside cells and for creating biosensor devices. Förster resonance energy transfer (FRET) based on fluorescent proteins has been used to detect conformational changes, however, such FRET devices have not yet been produced using fluorescent RNA aptamers. Here we develop an RNA aptamer-based FRET (apta-FRET) system using single-stranded RNA origami scaffolds. To obtain FRET, the fluorescent aptamers Spinach and Mango are placed in close proximity on the RNA scaffolds and a new fluorophore is synthesized to increase spectral overlap. RNA devices that respond to conformational changes are developed, and finally, apta-FRET constructs are expressed in E. coli where FRET is observed, demonstrating that the apta-FRET system is genetically encodable and that the RNA nanostructures fold correctly in bacteria. We anticipate that the RNA apta-FRET system could have applications as ratiometric sensors for real-time studies in cell and synthetic biology.

The DNA origami technique is a recently developed self-assembly method that allows construction of 3D objects at the nanoscale for various applications. In the current study we report the production of a 18 × 18 × 24 nm(3) hollow DNA box origami structure with a switchable lid. The structure was efficiently produced and characterized by atomic force microscopy, transmission electron microscopy, and Förster resonance energy transfer spectroscopy. The DNA box has a unique reclosing mechanism, which enables it to repeatedly open and close in response to a unique set of DNA keys. This DNA device can potentially be used for a broad range of applications such as controlling the function of single molecules, controlled drug delivery, and molecular computing.

Logic gates are devices that can perform logical operations by transforming a set of inputs into a predictable single detectable output. The hybridization properties, structure, and function of nucleic acids can be used to make DNA-based logic gates. These devices are important modules in molecular computing and biosensing. The ideal logic gate system should provide a wide selection of logical operations, and be integrable in multiple copies into more complex structures. Here we show the successful construction of a small DNA-based logic gate complex that produces fluorescent outputs corresponding to the operation of the six Boolean logic gates AND, NAND, OR, NOR, XOR, and XNOR. The logic gate complex is shown to work also when implemented in a three-dimensional DNA origami box structure, where it controlled the position of the lid in a closed or open position. Implementation of multiple microRNA sensitive DNA locks on one DNA origami box structure enabled fuzzy logical operation that allows biosensing of complex molecular signals. Integrating logic gates with DNA origami systems opens a vast avenue to applications in the fields of nanomedicine for diagnostics and therapeutics.