Exploring the speed limit of toehold exchange with a cartwheeling DNA acrobat

Dynamic DNA nanotechnology has yielded nontrivial autonomous behaviours such as stimulus-guided locomotion, computation and programmable molecular assembly. Despite these successes, DNA-based nanomachines suffer from slow kinetics, requiring several minutes or longer to carry out a handful of operations. Here, we pursue the speed limit of an important class of reactions in DNA nanotechnology-toehold exchange-through the single-molecule optimization of a novel class of DNA walker that undergoes cartwheeling movements over a field of complementary oligonucleotides. After optimizing this DNA 'acrobat' for rapid movement, we measure a stepping rate constant approaching 1 s^-1, which is 10- to 100-fold faster than prior DNA walkers. Finally, we use single-particle tracking to demonstrate movement of the walker over hundreds of nanometres within 10 min, in quantitative agreement with predictions from stepping kinetics. These results suggest that substantial improvements in the operating rates of broad classes of DNA nanomachines utilizing strand displacement are possible.