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Gigadalton-scale shape-programmable DNA assemblies

Abstract

Natural biomolecular assemblies such as molecular motors, enzymes, viruses and subcellular structures often form by self-limiting hierarchical oligomerization of multiple subunits1,2,3. Large structures can also assemble efficiently from a few components by combining hierarchical assembly and symmetry, a strategy exemplified by viral capsids4. De novo protein design5,6,7,8,9 and RNA10,11 and DNA nanotechnology12,13,14 aim to mimic these capabilities, but the bottom-up construction of artificial structures with the dimensions and complexity of viruses and other subcellular components remains challenging. Here we show that natural assembly principles can be combined with the methods of DNA origami15,16,17,18,19,20,21,22,23,24 to produce gigadalton-scale structures with controlled sizes. DNA sequence information is used to encode the shapes of individual DNA origami building blocks, and the geometry and details of the interactions between these building blocks then control their copy numbers, positions and orientations within higher-order assemblies. We illustrate this strategy by creating planar rings of up to 350 nanometres in diameter and with atomic masses of up to 330 megadaltons, micrometre-long, thick tubes commensurate in size to some bacilli, and three-dimensional polyhedral assemblies with sizes of up to 1.2 gigadaltons and 450 nanometres in diameter. We achieve efficient assembly, with yields of up to 90 per cent, by using building blocks with validated structure and sufficient rigidity, and an accurate design with interaction motifs that ensure that hierarchical assembly is self-limiting and able to proceed in equilibrium to allow for error correction. We expect that our method, which enables the self-assembly of structures with sizes approaching that of viruses and cellular organelles, can readily be used to create a range of other complex structures with well defined sizes, by exploiting the modularity and high degree of addressability of the DNA origami building blocks used.

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Figure 1: Shape-programmable self-limiting ring oligomerization in solution.
Figure 2: Assembly of self-limiting ring oligomers into tubes.
Figure 3: Shape-programmed hierarchical assembly and cryo-EM analysis of a three-fold self-complementary vertex.
Figure 4: Self-limiting hierarchical assembly of closed polyhedra from the reactive vertex.
Figure 5: Single-particle tomographic validation of correct polyhedral assembly by cryo-TEM.

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Acknowledgements

We thank A. Neuner and V. Hechtl for technical help, M. Kube and J. Funke for computational assistance, D. van Sinten and S. Welsch from FEI for their support with the Titan Krios, F. Praetorius and B. Kick for scaffold preparation, S. Fraden for discussions, and S. Barth for collecting auxiliary data. We also thank A. Holleitner and M. Altschner for access to the helium-ion microscope. This work was supported by a European Research Council Starting Grant to H.D. (grant agreement number 256270) and the Deutsche Forschungsgemeinschaft through grants provided within the Gottfried-Wilhelm-Leibniz Program, the Excellence Clusters CIPSM (Center for Integrated Protein Science Munich), NIM (Nanosystems Initiative Munich) and the Technische Universität München (TUM) Institute for Advanced Study. K.F.W. and H.D. are grateful for additional support from the Bosch Forschungsstiftung.

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Contributions

K.F.W. and C.S. performed research, H.D. designed the research. K.F.W. and H.D. wrote the manuscript. All authors analysed and discussed data and commented on the manuscript.

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Correspondence to Hendrik Dietz.

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The authors declare no competing financial interests.

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Reviewer Information Nature thanks M. Kostiainen, T. LaBean and H. Yan for their contribution to the peer review of this work.

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Supplementary information

Supplementary Information

This file contains Supplementary Materials and Methods, Supplementary Figures 1-35, Supplementary Notes 1-3 and Supplementary references. (PDF 32136 kb)

Tomogram of the tetrahedron object

Top: cryo-EM micrographs acquired from different tilt angles of the sample goniometer. Bottom: slices through the tomogram. (MOV 6740 kb)

Tomogram of the hexahedron object

Top: cryo-EM micrographs acquired from different tilt angles of the sample goniometer. Bottom: slices through the tomogram. (MOV 7166 kb)

Tomogram of the dodecahedron object

Top: negative-stained micrographs acquired from different tilt angles of the sample goniometer. Bottom: slices through the tomogram. (MOV 5656 kb)

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Wagenbauer, K., Sigl, C. & Dietz, H. Gigadalton-scale shape-programmable DNA assemblies. Nature 552, 78–83 (2017). https://doi.org/10.1038/nature24651

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