Computer‐Aided Production of Scaffolded DNA Nanostructures from Flat Sheet Meshes

Abstract

The use of DNA as a nanoscale construction material has been a rapidly developing field since the 1980s, in particular since the introduction of scaffolded DNA origami in 2006. Although software is available for DNA origami design, the user is generally limited to architectures where finding the scaffold path through the object is trivial. Herein, we demonstrate the automated conversion of arbitrary two‐dimensional sheets in the form of digital meshes into scaffolded DNA nanostructures. We investigate the properties of DNA meshes based on three different internal frameworks in standard folding buffer and physiological salt buffers. We then employ the triangulated internal framework and produce four 2D structures with complex outlines and internal features. We demonstrate that this highly automated technique is capable of producing complex DNA nanostructures that fold with high yield to their programmed configurations, covering around 70 % more surface area than classic origami flat sheets.

Spatial Regulation of T-Cell Signaling by Programmed Death-Ligand 1 on Wireframe DNA Origami Flat Sheets

“…As a platform for assembling the protein–oligo conjugates, we used a wireframe DNA origami flat sheet design ( Figure S3 ) which yields nanostructures that have the capacity to fold and remain stable under physiological salt conditions. 43 The flat sheets were first folded using short ssDNA oligos (staples) which bind complementarily to regions of the p8064 phage ssDNA scaffold. Protein–oligo conjugates were then assembled onto flat sheets via hybridization with complementary 5′ ends of staples at designated positions that protrude out of the nanostructure ( Figure S3b ).…”

“…Staple strand sequences for folding the 3-tessellation flat sheet were described previously. 43 The 216 core staples were obtained from Integrated DNA Technologies at a concentration of 100 μM in water in three 96-well plates. Staple mixes for folding each flat sheet design were prepared to a final concentration of 463 nM based on the pipetting schemes outlined in Supporting Information Figure S25 with some core staples replaced by protruding/modified staples ( Tables S3 and S4 ).…”

“…The flat sheets were produced as previously described with the following minor modifications. 43 In the folding reactions, the p8064 scaffold (Tilibit) was mixed at concentration of 20 with 200 nM of staples in 1 × PBS, pH 7.4. After removal of excess staples with 100 kDa MWCO Amicon centrifugal filters (Milipore), the concentration of flat sheets was adjusted to a concentration of 20 nM in 1 × PBS, pH 7.4.…”

“… 39 − 41 Here, we used the wireframe-scaffolded DNA origami strategy to produce DNA origami flat sheets that remain stable under physiological ionic strengths, unlike DNA nanostructures composed of densely packed DNA helices. 42 , 43 These DNA origami flat sheets were functionalized to present either agonistic anti-CD3 antibodies (FS-α-CD3), antibodies against CD3 and CD28 (FS-α-CD3-CD28), or PD-L1 proteins separated by varying nanoscale distances. As a readout of T-cell activation, we analyzed the activity of a luciferase reporter driven by nuclear factor of activated T cells (NFAT) response elements in PD-1-expressing Jurkat T cells.…”

DNA origami

“…vHelix 15 is the most widely used software in this category and also contains an integrated simulation platform that can predict the folding of the designed structures in standard DNA origami folding buffers. Other software such as DAEDALUS 47 and TALOS 9 for 3D origami and vHelix-BSCOR 48 , PERDIX 49 and METIS 10 for 2D origami are also available. Other automated design tools such as MagicDNA 50 have been reported in preprints.…”