Folding DNA into twisted and curved nanoscale shapes

Abstract

We demonstrate the ability to engineer complex shapes that twist and curve at the nanoscale from DNA. Through programmable self-assembly, strands of DNA are directed to form a custom-shaped bundle of tightly cross-linked double helices, arrayed in parallel to their helical axes. Targeted insertions and deletions of base pairs cause the DNA bundles to develop twist of either handedness or to curve. The degree of curvature could be quantitatively controlled, and a radius of curvature as tight as 6 nanometers was achieved. We also combined multiple curved elements to build several different types of intricate nanostructures, such as a wireframe beach ball or square-toothed gears.

Figure 1

Design principles for controlling twist and curvature in DNA bundles. (A) Double helices are constrained to a honeycomb arrangement by staple-strand crossovers. Semi-transparent crossover planes mark the locations of strand crossovers between neighboring helices, which are spaced at 7 bp intervals along the helical axis. From left to right, each plane contains a class of crossovers rotated in-plane by 240° clockwise with respect to the preceding plane. The crossover planes divide the bundle conceptually into helix fragments that can be viewed as residing in array cells (one cell is highlighted). (B) Array cell with default content of 7 bp, which exerts no stress on its neighbors. (C) Above, array cell with content of 5 bp, which is under strain and therefore exerts a left-handed torque and a pull on its neighbors. Below, array cell with content of 9 bp, which is under strain and therefore exerts a right-handed torque and a push on its neighbors. Force vectors are shown on only two of the four strand ends of the array-cell fragment for clarity. (D) Left (right), site-directed deletions (insertions) installed in selected array cells indicated in orange (blue) result in global left-handed (right-handed) twisting with cancellation of compensatory global bend contributions. (E) Site-directed base-pair deletions, indicated in orange, and base-pair insertions, indicated in blue, can be combined to induce tunable global bending of the DNA bundle with cancellation of compensatory global twist contributions.

Figure 2

Deviations from 10.5 bp/turn twist density induce global twisting. (A–C) Top left: models of a 10 by 6-helix DNA bundle (red) with 10.5, 10, and 11 bp/turn average double-helical twist density, respectively, and models of ribbons when polymerized (silver). Bottom left: monomeric particles as observed by negative-stain transmission electron microscopy. Scale bars: 20 nm. Right: polymeric ribbons as observed by TEM. Scale bars: 50 nm. (D, E) Tilt-pair images of twisted ribbons polymerized from 11 bp/turn (D) and 10 bp/turn (E) 10 by 6-helix bundles, recorded at goniometer angles 40° and −40°. Arrows indicate the observed upward (for 11 bp/turn) or downward (for 10 bp/turn) direction of movement of the twisted-ribbon nodes. Dashed line provides reference point (ends of ribbons remain stationary upon goniometer rotation). (F) Ethidium-bromide-stained 2% agarose gel, comparing migration of unpurified folded bundles. (G) Histograms of the observed node-to-node distance in twisted ribbons as observed in negative-stain TEM micrographs. Left-handed and right-handed ribbons undergo half-turns every 235±(32 s.d.) nm (N=62) and 286±(48 s.d.) nm (N=197), respectively. (H) Plot of observed global compensatory twist per turn versus double-helical twist density initially imposed by design. A value of 0.335 nm/bp was used to calculate global twist per turn from values obtained in (G).

Figure 3

Combining site-directed insertions and deletions induces globally bent shapes. (A–G) Models of seven 3 by 6-helix-bundle versions programmed to different degrees of bending and typical particles as observed by negative-stain TEM. Scale bars: 20 nm. (H) Ethidium-bromide-stained 2% agarose gel comparing migration of unpurified folding products of the seven differently bent bundles. (I, J) Low-magnification TEM micrographs of the bundle versions programmed to bend by 30° and 150°, respectively. Scale bars: 100 nm. (K) Histograms of bend angles as observed in individual particles for the seven different bundle versions. Average bend angles were determined to be 0°±(3° s.d.) (N=74); 30.7°±(5.4° s.d.) (N=212); 62.4°±(5.9° s.d.) (N=208); 91.3°±(5.2° s.d.) (N=206); 121°±(8.4° s.d.) (N=212); 143.4°±(9° s.d.) (N=131); 166°±(9° s.d.) (N=106). Insets: Plot of average double-helical twist density through the cross section of the bent segment that results from the pattern of insertions and deletions installed to induce bending.

Figure 4

Bending enables design of intricate nonlinear shapes. Scale bars: 20 nm. (A) Model of a 3 by 6-helix DNA-origami bundle designed to bend into a half-circle with a 25 nm radius that bears three non-bent teeth. Monomers were folded in separate chambers, purified, and mixed with connector staple strands to form six-toothed gears. Typical monomer and dimer particles visualized by negative-stain transmission electron microscopy (TEM). (B) 3 by 6-helix bundle as in (A), modified to bend into a quarter circle with a 50 nm radius. Hierarchical assembly of monomers yields 12-tooth gears. (C) A single scaffold strand designed to fold into a 50-nm-wide spherical wireframe capsule resembling a beach ball and four typical particles representing different projections of the beach ball. The design folds as six bent crosses (inset) connected on a single scaffold. (D) A concave triangle that is folded from a single scaffold strand. The design can be conceptualized as three 3 by 6 bundles with internal segments designed to bend by 60°. (E) A convex triangle assembled hierarchically from three 3 by 6 bundles designed with a 120° bend (Fig. 3E). (F) A six-helix bundle programmed with varying degrees of bending folds into a spiral-like object.