Evaluating the effect of two-dimensional molecular layout on DNA origami-based transporters

Cellular transport systems are sophisticated and efficient. Hence, one of the ultimate goals of nanotechnology is to design artificial transport systems rationally. However, the design principle has been elusive, because how motor layout affects motile activity has not been established, partially owing to the difficulty in achieving a precise layout of the motile elements. Here, we employed a DNA origami platform to evaluate the two-dimensional (2D) layout effect of kinesin motor proteins on transporter motility. We succeeded in accelerating the integration speed of the protein of interest (POI) to the DNA origami transporter by up to 700 times by introducing a positively charged poly-lysine tag (Lys-tag) into the POI (kinesin motor protein). This Lys-tag approach allowed us to construct and purify a transporter with high motor density, allowing a precise evaluation on the 2D layout effect. Our single-molecule imaging showed that the densely packed layout of kinesin decreased the run length of the transporter, although its velocity was moderately affected. These results indicate that steric hindrance is a critical parameter to be considered in the design of transport systems.

Automated summary

Cellular transport systems are complex and efficient, and there is a need to design artificial transport systems using nanotechnology. However, the design principles have been difficult to establish due to the lack of understanding on how motor layout affects motile activity. In this study, a DNA origami platform was used to evaluate the effect of motor protein layout on transporter motility. The densely packed layout of kinesin motor protein was found to decrease the run length of the transporter, highlighting the importance of considering steric hindrance in transport system design.