The cylinder is put in the center of the motor as an axis supporting the rotation. DNA strands of staples and a scafold are formed into a cylindrical shape using DNA Origami technology. It was designed with cadnano software, which "simplifies and enhances the process of designing three-dimensional DNA origami nanostructures".(Figure 1)
In order to bind footing DNAs on its surface spirally, 10mer long DNA strands which we call footings of footings come up from the cylinder's surface. After the cylinder with the footing of footings are formed, one of the three legs of DNA spiders is connected to a specific strand at the start point on the cylinder's surface. Then the footings get connected to the footing of footings. The footing tracks, each of which consists of three lanes of the footings, enable DNA Spiders to orbit the cylinder. In detail, there are two tracks of DNA footings and two spiders can walk on each track. In addition, in order to make it easy to observe the direction of spiders' walk, a small sub-cylinder is attached at the end of the footing tracks.(Figure 2)
The diameter of the cylinder is 25.4 nm and height is 43.5 nm. This is calculated considering that the cylinder can be observed with an Atomic Force Microscope and that the interval between the two tracks are wide enough for spiders not to jump to next footing track.(Figure 3)
Our DNA screw rotates by using DNA spiders produced by Lund, et al. DNA spider is connected to the DNA ring by a capture leg, and also, DNA spider's legs are connected to strands on the surface of the DNA cylinder. DNA spider consists of a core and three legs. These legs move by cutting strands with DNAzyme. DNAzyme is a DNA which cuts DNA strands like enzymes do. Because the spider's track is a spiral, DNA rings rotates as DNA spiders walk.
In detail, legs and the footings are complementary strands. The footings are attached spirally to the surface of the DNA cylinder and function as a substrate of the DNA spider. DNA spiders normally move one way. Once DNAzyme cuts the tip of the footings, DNA spiders are disconnected to the footings. Next, DNA spiders attach to the next full-length strands.
In this paragraph, how to create DNA spiders is described. This method is the same as the way Lund, et al. did. First, Streptavidin(STV) and capture leg [5’ - GCC GAG AAC CTG ACG CAA GT/iSp18//iSp18//3Bio/ - 3] are connected in a solution. To create one-to-one product (“STV-(C)1” ), ion exchange is used. Second, deoxyribozyme legs [5’ - /5BioTEG//iSp18//iSp18//TCT CTT CTC CGA GCC GGT CGA AAT AGT GAA AA – 3’ ] are attached to the STV-(C) i.e. NICK3.4A+1. Third, Cy3 MONO NHS ester dissolved in DMSO is added in the solution containing NICK3.4A+1.
Fig.4 Image of cadnano
Fig.5 Image of rotary ring
It is the DNA ring that actually rotates. It has a shape of a cylinder short in the axial direction and is also composed with DNA Origami technology. We made the structure using cadnano.(Figure4) Two 10mer long strands come up from the inner side of the ring and are connected to the DNA spiders. The diameter of the ring is 50.9 nm, and so it can hold the cylinder and spiders inside of the ring. The thickness of ring is 10.9 nm in consideration of Atomic Force Microscope visibility.(Figure 5)
Fig.6 Image of DNA screw
The DNA screw is realized by assembling the above four parts: the cylinder, footings, DNA Spiders, and the rotary ring.(Figure 5)