Traditional robots 1 rely on computing to coordinate sensing and actuating components and to store internal representations of their goals and environment. Any implementation of single-molecule based robotics must overcome the limited ability of individual molecules to store complex programs and, for example, use architectures that obtain complex behaviors from the interaction of simple robots with their environment [2][3][4] . Previous research in DNA walkers 5 focused on transitioning from non-autonomous systems 6, 7 to directed but brief motion on one-dimensional tracks8 -11. Herein, we obtain elementary robotic behaviors from the interaction between a random walker incorporating deoxyribozymes 12 and a precisely defined environment. Using singlemolecule microscopies we demonstrate that such walkers achieve directionality by sensing and modifying their environment, following trails of recognition elements ("bread crumbs") laid out on a two-dimensional DNA origami landscape 13 . These molecular robots autonomously carry out sequences of actions such as "start", "follow", "turn", and "stop", thus laying the foundation for the synthesis of more complex robotic behaviors at the molecular level by incorporating additional layers of control mechanisms. For example, interactions between multiple molecular robots could lead to collective behavior14 , 15, while the ability to read and transform secondary cues on the landscape could provide a mechanism for Turing-universal algorithmic behavior2 ,16,17 .Author Contributions: AFM experiments were performed by K.L. (majority), J.N., and N. D.; analysis was performed by N. D., K.L., J.N., S.T., and supervised by E.W., and H.Y. Fluorescence microscopy and particle tracking analysis were performed by A.J.M., N.M., and A.J.B, supervised by N. G. W. Spiders were synthesized, purified, and their integrity confirmed and monitored by S.T. SPR experiments were performed by R. P. Research coordination by M.N.S., material transfer coordination by S.T., J.N., and K.L. Experimental design and manuscript was done with input from all authors. Author Information: Reprints and permissions information is available at npg.nature.com/reprintsandpermissions. The authors declare no competing financial interests. Correspondence and requests for materials should be addressed to: mns18@columbia.edu, winfree@caltech.edu, nwalter@umich.edu, hao.yan@asu.edu Supplementary Information is linked to the online version of the paper at www.nature.com/nature. NIH Public Access Author ManuscriptNature. Author manuscript; available in PMC 2010 November 1. Restated in a biochemically more intuitive manner: A deoxyribozyme on a site that was previously converted to a product will dissociate faster, whereas it will stick longer on the substrates and eventually cleave them. Because spiders have multiple legs, a single dissociated leg will quickly reattach to nearby product or substrate. It follows that the body of a spider positioned at the interface between products and substrates will move toward the s...

We describe polycatalytic assemblies, comprising one or two streptavidin molecules and two to six attached nucleic acid catalysts (deoxyribozymes), with phosphodiesterase activity. When exposed to a matrix covered at high densities with oligonucleotide substrates, these molecules diffuse through the matrix continuously cleaving the substrate at rates comparable to those of individual catalysts in solution. Rates of diffusion (movement), processivity, and resident times of assemblies can be controlled through the number of catalytic units and the length of substrate/product recognition regions. The assemblies were characterized at the ensemble level using surface plasmon resonance.

Oligonucleotide-based receptors or aptamers can interact with small molecules, but the ability to achieve high-affinity and selectivity of these interactions depends strongly on functional groups or epitopes displayed by the binding targets. Some classes of targets are particularly challenging: for example, monosaccharides have scarce functionalities and no aptamers have been reported to recognize, let alone distinguish from each other, glucose and other hexoses. Here we report aptamers that differentiate low-epitope targets such as glucose, fructose, or galactose by forming ternary complexes with high-epitope organic receptors for monosaccharides. In a follow-up example, we expand this method to isolate high-affinity oligonucleotides against aromatic amino acids complexed in situ with a non-specific organometallic receptor. The method is general and enables broad clinical use of aptamers for detection of small molecules in mix-and-measure assays, as demonstrated by monitoring postprandial waves of phenylalanine in human subjects.