Regular Nanoscale Protein Patterns via Directed Adsorption through Self-Assembled DNA Origami Masks

Abstract

DNA origami has become a widely used method for synthesizing well-defined nanostructures with promising applications in various areas of nanotechnology, biophysics, and medicine. Recently, the possibility to transfer the shape of single DNA origami nanostructures into different materials via molecular lithography approaches has received growing interest due to the great structural control provided by the DNA origami technique. Here, we use ordered monolayers of DNA origami nanostructures with internal cavities on mica surfaces as molecular lithography masks for the fabrication of regular protein patterns over large surface areas. Exposure of the masked sample surface to negatively charged proteins results in the directed adsorption of the proteins onto the exposed surface areas in the holes of the mask. By controlling the buffer and adsorption conditions, the protein coverage of the exposed areas can be varied from single proteins to densely packed monolayers. To demonstrate the versatility of this approach, regular nanopatterns of four different proteins are fabricated: the single-strand annealing proteins Redβ and Sak, the iron-storage protein ferritin, and the blood protein bovine serum albumin (BSA). We furthermore demonstrate the desorption of the DNA origami mask after directed protein adsorption, which may enable the fabrication of hierarchical patterns composed of different protein species. Because selectivity in adsorption is achieved by electrostatic interactions between the proteins and the exposed surface areas, this approach may enable also the large-scale patterning of other charged molecular species or even nanoparticles.

On the Stability of DNA Origami Nanostructures in Low‐Magnesium Buffers

“…Some applications require the use of potassium-based phosphate buffers instead of sodium-based ones,for instance to ensure the stability of certain proteins when arranging them with DNAo rigami scaffolds. [20] Therefore,w eh ave evaluated DNAo rigami stability also in 10 mm K 2 HPO 4 .A s can be seen in Figure 2( condition 6), as imilar behavior is observed as for Na 2 HPO 4 ,w ith DNAo rigami triangles and 24HBs being almost completely denatured, while the 6HBs remain intact. In contrast to the case of Na + ,h owever, addition of 100 mm KCl already results in perfectly intact DNAo rigami triangles (Figure 2a,c ondition 7).…”

Protein Coating of DNA Nanostructures for Enhanced Stability and Immunocompatibility

“…[10][11][12] The computational tools [11][12][13] for designing such objects have emerged along with these techniques, and this progress has opened up new possibilities for the researchers to effortlessly build their own nanostructures for tailored uses. [14] Recently demonstrated applications based on customized DNA nanostructures include artificial ion channels, [15] optical (plasmonic and photonic) structures, [16,17] high-precision molecular positioning devices, [18] modifiable templates for arranging, e.g., proteins, [19][20][21] polymers, [22] and nanotubes, [23] as well as DNA-assisted techniques for creating arbitrarily shaped metal nanoparticles. [24][25][26] Fully addressable DNA nanostructures, especially DNA origami, possess huge potential to serve as inherently biocompatible and versatile molecular platforms.…”

On the Stability of DNA Origami Nanostructures in Low‐Magnesium Buffers

“…[20] Therefore,w eh ave evaluated DNAo rigami stability also in 10 mm K 2 HPO 4 .A s can be seen in Figure 2( condition 6), as imilar behavior is observed as for Na 2 HPO 4 ,w ith DNAo rigami triangles and 24HBs being almost completely denatured, while the 6HBs remain intact. [20] Therefore,w eh ave evaluated DNAo rigami stability also in 10 mm K 2 HPO 4 .A s can be seen in Figure 2( condition 6), as imilar behavior is observed as for Na 2 HPO 4 ,w ith DNAo rigami triangles and 24HBs being almost completely denatured, while the 6HBs remain intact.…”