Dr Jon Bath

Abstract:

Nanoscale manipulation and patterning usually require costly and sensitive top-down techniques such as those used in scanning probe microscopies or in semiconductor lithography. DNA nanotechnology enables exploration of bottom-up fabrication and has previously been used to design self-assembling components capable of linear and rotary motion. In this work, we combine three independently controllable DNA origami linear actuators to create a nanoscale robotic printer. The two-axis positioning mechanism comprises a moveable gantry, running on parallel rails, threading a mobile sleeve. We show that the device is capable of reversibly positioning a write head over a canvas through the addition of signaling oligonucleotides. We demonstrate “write” functionality by using the head to catalyze a local DNA strand–exchange reaction, selectively modifying pixels on a canvas. This work demonstrates the power of DNA nanotechnology for creating nanoscale robotic components and could find application in surface manufacturing, biophysical studies, and templated chemistry.

Abstract:

DNA origami is a robust assembly technique that folds a single-stranded DNA template into a target structure by annealing it with hundreds of short 'staple' strands. Its guiding design principle is that the target structure is the single most stable configuration. The folding transition is cooperative and, as in the case of proteins, is governed by information encoded in the polymer sequence. A typical origami folds primarily into the desired shape, but misfolded structures can kinetically trap the system and reduce the yield. Although adjusting assembly conditions or following empirical design rules can improve yield, well-folded origami often need to be separated from misfolded structures. The problem could in principle be avoided if assembly pathway and kinetics were fully understood and then rationally optimized. To this end, here we present a DNA origami system with the unusual property of being able to form a small set of distinguishable and well-folded shapes that represent discrete and approximately degenerate energy minima in a vast folding landscape, thus allowing us to probe the assembly process. The obtained high yield of well-folded origami structures confirms the existence of efficient folding pathways, while the shape distribution provides information about individual trajectories through the folding landscape. We find that, similarly to protein folding, the assembly of DNA origami is highly cooperative; that reversible bond formation is important in recovering from transient misfoldings; and that the early formation of long-range connections can very effectively enforce particular folds. We use these insights to inform the design of the system so as to steer assembly towards desired structures. Expanding the rational design process to include the assembly pathway should thus enable more reproducible synthesis, particularly when targeting more complex structures. We anticipate that this expansion will be essential if DNA origami is to continue its rapid development and become a reliable manufacturing technology.

Abstract:

We are learning to build synthetic molecular machinery from DNA. This research is inspired by biological systems in which individual molecules act, singly and in concert, as specialized machines: our ambition is to create new technologies to perform tasks that are currently beyond our reach. DNA nanomachines are made by self-assembly, using techniques that rely on the sequence-specific interactions that bind complementary oligonucleotides together in a double helix. They can be activated by interactions with specific signalling molecules or by changes in their environment. Devices that change state in response to an external trigger might be used for molecular sensing, intelligent drug delivery or programmable chemical synthesis. Biological molecular motors that carry cargoes within cells have inspired the construction of rudimentary DNA walkers that run along self-assembled tracks. It has even proved possible to create DNA motors that move autonomously, obtaining energy by catalysing the reaction of DNA or RNA fuels.

Abstract:

The SpoIIIE protein of Bacillus subtilis is required for chromosome segregation during spore formation. The COOH-terminal cytoplasmic part of SpoIIIE was shown to be a DNA-dependent adenosine triphosphatase (ATPase) capable of tracking along DNA in the presence of ATP, and the NH(2)-terminal part of the protein was found to mediate its localization to the division septum. Thus, during sporulation, SpoIIIE appears to act as a DNA pump that actively moves one of the replicated pair of chromosomes into the prespore. The presence of SpoIIIE homologs in a broad range of bacteria suggests that this mechanism for active transport of DNA may be widespread.

Abstract:

Fundamental investigations of ice nucleation, a key process in fields from environmental science to cryobiology, require model systems with chemical and physical structures that are well defined and easily varied. DNA origami is an especially promising model because of the exquisite control that it offers over the physical geometry of the nucleating agent at the nano-scale. Here we compare ice nucleation by solutions of a rectangular DNA origami tile, formed by annealing a 2.6 kbase single-stranded DNA scaffold with ninety shorter ‘staple’ oligonucleotides, to ice nucleation when these components are mixed at the same concentrations but not annealed. Isothermal measurements show that the molecular conformation has a dramatic effect on the ice nucleating efficiency. For an array of droplets containing annealed, well-folded tiles the freezing rate is constant, whereas for unannealed DNA the freezing rate decreases with time. Despite the freezing rate measured at low temperature being higher for the annealed DNA origami samples than for a significant proportion of the unannealed ones, in slow temperature-ramp measurements the latter generally freeze at higher temperatures. We show that this behaviour is consistent with the formation of small numbers of highly efficient nucleating agents in the unannealed samples, likely through molecular aggregation.