Professor Andrew Turberfield

Abstract:

We describe the triggered assembly of a bio-inspired DNA origami meshwork on a lipid membrane. DNA triskelia, three-armed DNA origami nanostructures inspired by the membrane-modifying protein clathrin, are bound to lipid mono- and bi-layers using cholesterol anchors. Polymerization of triskelia, triggered by the addition of DNA staples, links triskelion arms to form a mesh. Using transmission electron microscopy, we observe nanoscale local deformation of a lipid monolayer induced by triskelion polymerization that is reminiscent of the formation of clathrin-coated pits. We also show that the polymerization of triskelia bound to lipid bilayers modifies interactions between them, inhibiting the formation of a synapse between giant unilamellar vesicles and a supported lipid bilayer.

Abstract:

In nature, co-assembly of polypeptides, nucleic acids, and polysaccharides is used to create functional supramolecular structures. Here, we show that DNA nanostructures can be used to template interactions between peptides and to enable the quantification of multivalent interactions that would otherwise not be observable. Our functional building blocks are peptide–oligonucleotide conjugates comprising de novo designed dimeric coiled-coil peptides covalently linked to oligonucleotide tags. These conjugates are incorporated in megadalton DNA origami nanostructures and direct nanostructure association through peptide–peptide interactions. Free and bound nanostructures can be counted directly from electron micrographs, allowing estimation of the dissociation constants of the peptides linking them. Results for a single peptide–peptide interaction are consistent with the measured solution-phase free energy; DNA nanostructures displaying multiple peptides allow the effects of polyvalency to be probed. This use of DNA nanostructures as identifiers allows the binding strengths of homo- and heterodimeric peptide combinations to be measured in a single experiment and gives access to dissociation constants that are too low to be quantified by conventional techniques. The work also demonstrates that hybrid biomolecules can be programmed to achieve spatial organization of complex synthetic biomolecular assemblies.

Abstract:

The rational design of complementary DNA sequences can be used to create nanostructures that self-assemble with nanometer precision. DNA nanostructures have been imaged by atomic force microscopy and electron microscopy. Small-angle X-ray scattering (SAXS) provides complementary structural information on the ensemble-averaged state of DNA nanostructures in solution. Here we demonstrate that SAXS can distinguish between different single-layer DNA origami tiles that look identical when immobilized on a mica surface and imaged with atomic force microscopy. We use SAXS to quantify the magnitude of global twist of DNA origami tiles with different crossover periodicities: these measurements highlight the extreme structural sensitivity of single-layer origami to the location of strand crossovers. We also use SAXS to quantify the distance between pairs of gold nanoparticles tethered to specific locations on a DNA origami tile and use this method to measure the overall dimensions and geometry of the DNA nanostructure in solution. Finally, we use indirect Fourier methods, which have long been used for the interpretation of SAXS data from biomolecules, to measure the distance between DNA helix pairs in a DNA origami nanotube. Together, these results provide important methodological advances in the use of SAXS to analyze DNA nanostructures in solution and insights into the structures of single-layer DNA origami.

Abstract:

We report the design and assembly of chiral DNA nanotubes with well‐defined and addressable inside and outside surfaces. We demonstrate that the outside surface can be functionalised with a chiral arrangement of gold nanoparticles to create a plasmonic device and that the inside surface can be functionalised with a track for a molecular motor allowing transport of a cargo within the central cavity.