Tuesday, 30 January 2018

Recent Developments in DNA Origami

Javier Cabello – PhD student

Using both technique and imagination, origami allows the production of many shapes with only a folded sheet of paper. After the birth of DNA nanotechnology, the potential of this folding art inspired Paul Rothermund to develop a technique that allows the creation of complex nanoscale objects out of DNA. A single DNA strand is a flexible material that can be folded by hundreds of smaller strands (“staples”). These staples bind to specific non-consecutive regions of the larger strand (“template”) by Watson-Crick base-pairing, folding the template into a DNA structure with a particular shape.

DNA origami structures have a wide variety of potential uses that greatly differ from the original biological role of the molecule. These range from drug delivery to the production of electric circuits, with the biocompatible nature of DNA suggesting application in living organisms. However, the structures created are still limited by the size of the template, which becomes harder to fold when longer. Another problem with scaling up the technology is the cost of synthesising large numbers of staple strands.

Last November, four publications in Nature focused on overcoming the limitations of DNA origami. Three of the publications proposed new approaches for the assembly of bigger DNA structures. All three involved combining blocks of different sizes and complexity to form bigger structures.

Gigadalton-scale shape-programmable DNA assemblies
The first approach, suggested by Wagenbauer, Sigl and Dietz, involves making origami structures that can then self-polymerize. The resultant structures depend on the geometry of the individual origami building blocks. This approach, heavily inspired by viral capsid assembly, was used to form DNA rings from V-shaped DNA components. Afterwards, these rings were assembled to form DNA tubes. The method was generalised to produce different figures, as dodecahedra, by varying V-block angles and adding blocks with more than two binding sites.

Fractal assembly of micrometre-scale DNA origami arrays with arbitrary patterns
Tikhomirov, Petersen and Qian’s work revolves around the design of a robust algorithm to produce ordered assembly of complex patterns in 2D. In this work, individual origami tiles were first folded in isolation. Tiles were then combined in groups of four to produce larger “super-tiles”. Successively, four of these assembled larger tiles were employed to produce even bigger tiles. To achieve the necessary control of this process, the authors used three basic design rules, which they incorporated into a computational tool that they named the “FracTile Compiler”. This compiler allows the systematic design of complex, puzzle-like patterns. The resultant array can show an arbitrary picture, displayed by unique modifications to the surface of each tile, as they do with “La Gioconda” (ed.: the Mona Lisa for English readers) among others (see fig. 1).

Fig. 1 Complex shapes created using the method of Tikhomorov et al., reproduced with permission from Springer Nature, Fractal assembly of micrometre-scale DNA origami arrays with arbitrary patterns, G Tikhomirov, P Petersen, L Qian, Nature 552 (7683), 67.

Programmable self-assembly of three-dimensional nanostructures from 10,000 unique components
In this case, Ong et al improved a previously published variant related to DNA origami: DNA bricks. Instead of using large DNA strands as in previous approaches, the authors design structures built from smaller building blocks, of only 56 nucleotides each. Each of these bricks can bind to three others, forming a dense “lego”-like structure. By using 10,000 different sequences for these DNA bricks, the authors achieve the production of cubes of more than 1 GDa (3000 tunes the mass of the largest proteins). These cubes were proposed to serve as a canvas to produce different structures by simply removing bricks from the design, like chiselling a rock. This approach even allows the production of negative structures by carving cavities inside the DNA cube. To help in the design task, the authors created the software “NanoBricks” and proved its utility by producing different patterns. such as a helix or a teddy bear inside the DNA cube.

Biotechnological mass production of DNA origami
Despite these significant improvements in the production of larger DNA origami structures, all their possible applications are being hindered by the pecuniary costs involved in synthesising the component strands. However, new methods inspired by the current biotechnological industry are being developed to mass produce DNA origami designs.

Praetorius et al. have demonstrated the possibility of biotechnological mass production of a DNA origami design. Their approach involves encoding all the components necessary for the folding of an origami (ie. template and staples) in a single genetic sequence, and incorporating it into bacteria. Of course, this isn’t all. Between the different components they added DNA sequences with enzymatic activity: DNAzymes. The whole DNA strand is then produced in a culture of bacteria, and the DNAzymes, in the presence of zinc, cut themselves out of the sequence, separating it into the desired components. The result is that the staples and template, now separate, can assemble themselves to form the intended DNA origami structure.

This new approach, if escalated, would drastically reduce the production costs of DNA origami designs. However, the production of the initial genetic material would still require an expensive synthesis of the desired sequences for each new design produced. In order to make this solution even more affordable the produced DNA origami should be a very general or standardized one which can be customized with minor changes. Who knows if the answer lies in any of the previous papers? My humble guess is that DNA bricks are good candidates.

Read more:
- Rothemund, P. (2006). Folding DNA to create nanoscale shapes and patterns. Nature, 440(7082), pp.297-302. Link: https://www.nature.com/articles/nature04586
- Wagenbauer, K., Sigl, C. and Dietz, H. (2017). Gigadalton-scale shape-programmable DNA assemblies. Nature, 552(7683), pp.78-83. Link: https://www.nature.com/articles/nature24651
- Tikhomirov, G., Petersen, P. and Qian, L. (2017). Fractal assembly of micrometre-scale DNA origami arrays with arbitrary patterns. Nature, 552(7683), pp.67-71. Link: https://www.nature.com/articles/nature24655
Ong, L., Hanikel, N., Yaghi, O., Grun, C., Strauss, M., Bron, P., Lai-Kee-Him, J., Schueder, F., Wang, B., Wang, P., Kishi, J., Myhrvold, C., Zhu, A., Jungmann, R., Bellot, G., Ke, Y. and Yin, P. (2017). Programmable self-assembly of three-dimensional nanostructures from 10,000 unique components. Nature, 552(7683), pp.72-77. Link: https://www.nature.com/articles/nature24648
- Praetorius, F., Kick, B., Behler, K., Honemann, M., Weuster-Botz, D. and Dietz, H. (2017). Biotechnological mass production of DNA origami. Nature, 552(7683). pp.84-87. Link: https://www.nature.com/articles/nature24650

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