A plasmonic nanorod that walks on DNA origami

In nano-optics, a formidable challenge remains in precise transport of a single optical nano-object along a programmed and routed path toward a predefined destination. Molecular motors in living cells that can walk directionally along microtubules have been the inspiration for realizing artificial molecular walkers. Here we demonstrate an active plasmonic system, in which a plasmonic nanorod can execute directional, progressive and reverse nanoscale walking on two or three-dimensional DNA origami. Such a walker comprises an anisotropic gold nanorod as its ‘body' and discrete DNA strands as its ‘feet'. Specifically, our walker carries optical information and can in situ optically report its own walking directions and consecutive steps at nanometer accuracy, through dynamic coupling to a plasmonic stator immobilized along its walking track. Our concept will enable a variety of smart nanophotonic platforms for studying dynamic light–matter interaction, which requires controlled motion at the nanoscale well below the optical diffraction limit.

triggers a series of conformational changes of the system as well as activates subsequent nearfield interaction changes with the stator, thus giving rise to immediate spectral response changes that can be read out optically. As a result, locomotion on the order of several nanometers, which is far below the optical resolution limit, can be optically discriminated in real time.

Results
As shown in Figure 1, a walker, gold nanorod (AuNR) in yellow and a stator (AuNR in red) are organized in a chiral geometry. Specifically, the walker and the stator are placed on two opposite surfaces of a two-dimensional (2D) rectangular DNA origami platform, forming a 90° cross configuration. A chiral geometry is chosen in that circular dichroism (CD), that is, differential absorption of left-and right-handed circularly polarized light, of three-dimensional (3D) chiral structures are markedly sensitive on their conformational changes. [15][16][17][18] The stator is immobilized on one surface through the capture strands on the origami, whereas on the other surface the walker can execute stepwise movements by programmably attaching (detaching) its feet on (from) the track through hybridization (de-hybridization) with the footholds (coded A-F in Fig. 1). In particular, double-layer DNA origami is utilized to achieve a rigid and robust track. The DNA origami (58 nm×42 nm×7 nm) was prepared by folding a long single-stranded DNA scaffold with staple strands and specific capture strands, following a self-assembly process (see Supplementary Note 1) 19,20 .
In contrast to previous tiny DNA walkers, 1-11 our walker comprises an anisotropic AuNR, which is as large as 35 nm×10 nm. On one hand, a large metal nanoparticle is essential for plasmonic probing, as it yields distinct and pronounced optical response. On the other hand, the anisotropic nature of the AuNR also brings about substantial challenges to implement directional and progressive walking.
To impose directional walking, the feet of the walker and the footholds on the track are specifically designed. The walker AuNR is fully covered with identical foot strands, which contain a 9-nt segment for hybridization and four thymine bases as spacer. Along the track, six parallel rows of footholds A-F, are utilized to establish five walking stations I-V, which are evenly separated by 7 nm. This also defines the step size of the walker. At each station, the walker's feet step on two rows of the footholds to accommodate its transverse dimension as well as to ensure stable binding. In each row, five binding sites with identical footholds are extended from the origami. Each foothold consists of two parts: a binding segment (9-nt, black) for hybridization with a foot strand of the walker as well as a toehold segment (8-nt, colored), which is differently sequenced in different foothold rows for achieving programmable reactions. The walker enters the right-handed configuration region. As shown in Fig. 2b, when the walker strides from station III to IV and subsequently to V, the CD response strengthens successively. At station V, the CD response reaches approximately -200 mdeg, exhibiting a dip-to-peak line shape, which is nearly a mirror image of the CD spectrum at station I. This importantly indicates that in the solution the walkers that were directed to walk from station I have nearly all successfully reached station V, demonstrating the high fidelity of the walking process. In short, the individual steps of the walker which are well below the optical diffraction limit can be optically discriminated in real time.
For comparison, theoretical calculations 24 of the CD spectra were carried out (for details, see Supplementary Note 5) and are presented in Fig. 2c. In the calculations, the right-handed preference at station III was not included. Overall, the experimental spectra agree well with To in situ monitor the dynamic walking process, CD spectra of the sample were recorded using a time-scan function of the CD spectrometer at a fixed wavelength of 685 nm. As shown in Fig. 4, the CD intensity displays a successive decrease when the walker executes discrete steps from station I to station V. In average, the transition between different steps takes approximately 25 min to complete. Previous DNA walkers based on 'burnt bridge' render impossible reverse walking along the same track. 4,8,11 To demonstrate the switchable directionality of our walker, reverse walking is carried out after the walker reaches station V.
Upon addition of blocking strands f and removal strands ̅ , the walker changes its walking direction and executes one step back toward station VI, stepping on rows D and E. This gives rise to an instant CD intensity increase as shown in Fig. 4. When the walker executes one more step backward, the CD intensity shows a further increase to the level at station III.
Subsequently, the walker makes a new turn at station III and undergoes another reverse walking toward station IV. As shown Fig. 4, the CD intensity changes approximately back to the level at station IV. Overall, the walker has successfully carried out directed movements along the track, following a regulated route of I-II-III-IV-V-IV-III-IV.
To demonstrate the capability to perform more complex behavior, stepwise walking of the plasmonic walker on a 3D origami platform is examined. Fig. 5a shows the schematic of the walker system, in which triangle-prism origami is utilized as the walking track. Its length is 35 nm. The three side-lengths of the triangular cross-section are approximately 29 nm, 26 nm, and 38 nm. The stator (AuNR in red) is immobilized on one side-surface of the triangle prism (see Fig. 5a). Seven parallel rows of footholds are extended from the other two side-surfaces to establish six walking stations I-VI. At each station, the feet of the walker (AuNR in yellow) step on two rows of the footholds. Detailed design information can be found in Supplementary Note S6. The stepwise walking starts from station I, where the walker and the stator form a right-handed configuration. Following the same walking strategy described in Fig. 2, the walker first carries out two discrete steps along the track, reaching stations II and III, respectively. Subsequently, the walker approaches the vertex of the triangle prism at station IV. It then makes a turn, entering the left-handed configuration region. By executing two more discrete steps, the walker eventually reaches station VI. Fig. 5b shows the experimental CD spectra at different walking stations. It is evident that the individual steps along the track on the 3D origami can be correlated with distinct CD spectral changes. Representative TEM images of the AuNRs on the 3D origami at different stations are also presented in Fig. 5b. The corresponding calculated CD spectra can be found in Supplementary note S6. The experimental and theoretical results show an overall good agreement.
The powerful combination between the precise control of nanoscale motion enabled by DNA nanotechnology and the rich spectral information offered by plasmonics [25][26][27] suggests a new generation of artificial synthetic machines, which can in situ report their own structural dynamics using a noninvasive, stable, and all-optical approach. This will render profound significance in dual disciplines. First, the realization of advanced plasmonic walkers that can stride along multidirectional footpaths and perform different taskson 2D or 3D prescriptive landscapes can be promptly envisioned. 8,28,29 Second, our walker concept will expand the functional scope of DNA-based devices as well as enrich the category of the state-of-the-art characterization methods for practical applications. Intriguing light-matter interaction studies, for example, distance-dependent interaction between single emitters and plasmonic nanoparticles will no longer be restricted to a static picture. [30][31][32] The plasmonic nanoparticle can be transformed to a walker by proper functionalization and prods the emitter with fully coordinated motion at the nanoscale accuracy. Finally, our walker concept also outlines an exciting prospect of generating programmable large-scale nanocircuits that incorporate biochemical, electrical, and optical components for active transport and information processing. Two gold nanorods (AuNRs) are assembled perpendicularly to one another on a double-layer DNA origami template, forming a left-handed configuration at station I. The yellow AuNR on the top surface represents the "walker" and the red AuNR on the bottom surface represents the "stator". The walking track comprises six rows of footholds (A-F) extended from the origami surface to define five walking stations (I-V). The distance between the neighboring stations is 7 nm, which also corresponds to the step size. In each row, there are five binding sites with identical footholds. Only the footholds in the front line are colored to highlight the different strand segments. The walker AuNR is fully functionalized with foot strands. To enable robust binding, the walker steps on two neighboring footholds at each station. The red beam indicates the incident circularly polarized light.
Figure 2 | Walking mechanism, measured and simulated CD spectra at each station. a, walking mechanism. Initially, the walker resides at station I. Upon addition of blocking strands a and removal strands ̅ , two toehold-mediated strand-displacement reactions occur simultaneously. Blocking strands a trigger the dissociation of the walker's feet from footholds A. Row A is then site blocked. Meanwhile, removal strands ̅ release blocking strands c from footholds C. Row C is therefore site activated to bind the feet of the walker. Subsequently, the walker carries out one step forward, reaching station II. For simplicity, only the front line of the associated strands is shown. b, Measured CD spectra at each station. c, Simulated CD spectra at each station. The right-handed preference at station III was not included in the calculation.