During the first and second reporting period (RP1, RP2) multiple generations of rotary mechanisms with ratchet implementations and also translational mechanisms were being designed. We also discovered an attractive possibility to covalently stabilize DNA origami nanostructures. This is highly advantageous for this project, as it allows us to build and test machine-like objects for a much wider range of conditions. The additional covalent bonds may be used in a sequence-programmable fashion to link free strand termini, to bridge strand breaks at crossover sites, and to create additional interhelical connections. We built several new DNA origami prototypes and characterized them using single particle cryo-EM. We started to carry out single-molecule fluorescence experiments to study the motion of these mechanisms. A prototype of a translational mechanism with long-range travel along a one-dimensional track was successfully prepared. We also developed design tools to construct scaffold sequences de novo based on strand diagrams, and we developed scalable production methods for creating design-specific scaffold strands with fully user-defined sequences.
In the third funding period (RP3) we successfully accomplished and published the construction and study of synthetic molecular mechanisms with rotational and translational degrees of freedom. These systems, although still being passive mechanisms displaying only undirected random Brownian motions, were important stepping stones for us to establish synthesis methods and analytical methods for studying the dynamics and mechanical interplay of the components in these systems. Yet, we also succeeded constructing several motor prototypes with rotary degrees of freedom that we tested for motor activity in response of different stimuli, notably ion currents through solid state nanopores, and by alternating AC fields, which we could successfully drive directionally. We thus now have two prototypes of actual molecular motors, a nanoscale turbine, and a nanoscale electrical motor that both show directional motion and sustained motion against viscous drag. The motors are capable of doing work on the environment. Much of RP3 was then consumed by the process of completing the in depth study of these new motors prototypes.
The last funding period (RP4) covered only a short time frame (6 months), but we completed demonstration of three novel and distinct DNA-origami based molecular motor systems. These three systems are capable of directed motions and doing work on the environment, as evidenced by their sustained directional motion against viscous friction. We also directly show the ability to do work by winding a molecular spindle with our electrically driven DNA origami nanomotor. With angular speeds up to 250 revolutions per minute and torques up to 10 pN nm, the motors achieve rotational speeds and torques that are approaching those known from powerful natural molecular machines, such as the ATP synthase. The motors move directionally owing to intrinsic mechanistic properties, powered by a simple external energy modulation that does not need any feedback or information supplied by the user to direct the motors. Our motors also afford control options that one is familiar with from macroscale motors: the user can turn them on and off at will, they respond quickly, and the speed and direction of rotation can be regulated. The motors can be produced and operated by anyone having access to standard wet-lab equipment. It suffices to transmit the sequence information to enable other users to replicate and build their own motors using DNA molecules obtained, for example, from commercial sources. Owing to the modularity of the DNA origami components, we expect that the motors can also be modified, adapted and integrated into other contexts. A natural next frontier would be to explore causing a barrier modulation through a chemical reaction and to exploit the directed motor motion to drive uphill chemical synthesis using a more elaborate synthetic mechanism featuring coordinated reciprocal motion—much like F1F0-ATP synthase mechanically synthesizes ATP driven by rotary motion.
In addition to this core work, we also pursued several other avenues of inquiry, such as logic-gated switching of DNA nanodevices; and in-depth structural analysis of DNA nanostructures by cryo-EM. The experimental laboratory activities were naturally affected somewhat through lockdowns and social distancing measures in response to the COVID-19 pandemics.
