Covalent Linkage of One-Dimensional DNA Arrays Bonded by Paranemic Cohesion

The construction of DNA nanostructures from branched DNA motifs, or tiles, typically relies on the use of sticky-ended cohesion, owing to the specificity and programmability of DNA sequences. The stability of such constructs when unligated is restricted to a specific range of temperatures, owing to the disruption of base pairing at elevated temperatures. Paranemic (PX) cohesion was developed as an alternative to sticky ends for the cohesion of large topologically closed species that could be purified reliably on denaturing gels. However, PX cohesion is also of limited stability. In this work, we added sticky-ended interactions to PX-cohesive complexes to create interlocked complexes by functionalizing the sticky ends with psoralen, which can form cross-links between the two strands of a double helix. We were able to reinforce the stability of the constructs by creating covalent linkages between the 3′-ends and 5′-ends of the sticky ends; the sticky ends were added to double crossover domains via 3′-3′ and 5′-5′ linkages. Catenated arrays were obtained either by enzymatic ligation or by UV cross-linking. We have constructed finite-length one-dimensional arrays linked by interlocking loops and have positioned streptavidin-gold particles on these constructs

Regulation of 2D DNA Nanostructures by the Coupling of Intrinsic Tile Curvature and Arm Twist

The overwinding and underwinding of duplex segments between junctions have been used in designing both left-handed and right-handed DNA origami nanostructures. For a variety of DNA tubes obtained from self-assembled tiles, only a theoretical approach of the intrinsic curvature of the DNA tile (specified as the intrinsic tile curvature) has been previously used to explain their formation. Details regarding the quantitative and structural descriptions of the tile curvature and its evolution in DNA tubes by the coupling of the twist of the inter-tile arm (specified as the arm twist) have never been addressed. In this work, we designed three types of tile cores built around a circular 128 nucleotide scaffold by using longitudinal weaving (LW), bridged longitudinal weaving (bLW) and transverse weaving (TW). Joining the tiles with inter-tile arms having the length of an odd number of DNA half-turns (termed O-tiling) almost resulted into planar 2D lattices, whereas joining the tiles with the arms having the length of an even number of DNA half-turns (termed E-tiling) nearly generated tubes. Streptavidin bound to biotin was used as a labeling technique to characterize the inside and outside surfaces of the E-tiling tubes and thereby the conformations of their component tiles with addressable concave and convex curvatures. When the arms have the normal winding at the relaxed B-form of DNA, the intrinsic tile curvature deter-mines the chirality of the E-tiling tubes. By regulating the arm length and the sticky end length of the bLW-Ep/q (E-tiling of the bLW cores with the arm length of p-bp and the sticky end length of q-nt) assemblies, the arm can be overwound, resulting in a left-handed twist, and can also be underwound, resulting in a right-handed twist. Chiral bLW-Ep/q tubes with either a right-handed curvature or a left-handed curvature can also be formed by the coupling of the intrinsic tile curvature and the arm twist. We were able to assign the chiral indices (n,m) to each tube using high-resolution AFM images, and therefore were able to estimate the tile curvature using a regular polygon model that approximated the transverse section of the tube. A deeper understanding of the integrated actions of dif-ferent types of twisting forces on the DNA tubes will be extremely helpful in engineering more elaborate DNA nanostructures in the future

Stabilisation of self-assembled DNA crystals by triplex-directed photo-cross-linking

The tensegrity triangle is a robust DNA motif that can self-assemble to generate macroscopic three-dimensional crystals. However, the stability of these crystals is dependent on the high ionic conditions used for crystal growth. Here we demonstrate that a triplex-forming oligonucleotide can be used to direct the specific intercalation, and subsequent photo-cross-linking, of 4,5?,8-trimethylpsoralen to single or multiple loci within or between the tiles of the crystal. Cross-linking between the tiles of the crystal improves their thermal stability. Such an approach is likely to facilitate the removal of crystals from their mother liquor and may prove useful for applications that require greater crystal stability

Topological Linkage of DNA Tiles Bonded by Paranemic Cohesion

Catenation is the process by which cyclic strands are combined like the links of a chain, whereas knotting changes the linking properties of a single strand. In the cell, topoisomerases catalyzing strand passage operations enable the knotting and catenation of DNA so that single- or double-stranded segments can be passed through each other. Here, we use a system of closed DNA structures involving a paranemic motif, called PX-DNA, to bind double strands of DNA together. These PX-cohesive closed molecules contain complementary loops whose linking by Escherichia coli topoisomerase 1 (Topo 1) leads to various types of catenated and knotted structures. We were able to obtain specific DNA topological constructs by varying the lengths of the complementary tracts between the complementary loops. The formation of the structures was analyzed by denaturing gel electrophoresis, and the various topologies of the constructs were characterized using the program Knotilus

Covalent Linkage of One-Dimensional DNA Arrays Bonded by Paranemic Cohesion

The construction of DNA nanostructures from branched DNA motifs, or tiles, typically relies on the use of sticky-ended cohesion, owing to the specificity and programmability of DNA sequences. The stability of such constructs when unligated is restricted to a specific range of temperatures, owing to the disruption of base pairing at elevated temperatures. Paranemic (PX) cohesion was developed as an alternative to sticky ends for the cohesion of large topologically closed species that could be purified reliably on denaturing gels. However, PX cohesion is also of limited stability. In this work, we added sticky-ended interactions to PX-cohesive complexes to create interlocked complexes by functionalizing the sticky ends with psoralen, which can form cross-links between the two strands of a double helix. We were able to reinforce the stability of the constructs by creating covalent linkages between the 3′-ends and 5′-ends of the sticky ends; the sticky ends were added to double crossover domains <i>via</i> 3′–3′ and 5′–5′ linkages. Catenated arrays were obtained either by enzymatic ligation or by UV cross-linking. We have constructed finite-length one-dimensional arrays linked by interlocking loops and have positioned streptavidin–gold particles on these constructs

Construction and Structure Determination of a Three-Dimensional DNA Crystal

Structural DNA nanotechnology combines branched DNA junctions with sticky-ended cohesion to create self-assembling macromolecular architectures. One of the key goals of structural DNA nanotechnology is to construct three-dimensional (3D) crystalline lattices. Here we present a new DNA motif and a strategy that has led to the assembly of a 3D lattice. We have determined the X-ray crystal structures of two related constructs to 3.1 Å resolution using bromine-derivatized crystals. The motif we used employs a five-nucleotide repeating sequence that weaves through a series of two-turn DNA duplexes. The duplexes are tied into a layered structure that is organized and dictated by a concert of four-arm junctions; these in turn assemble into continuous arrays facilitated by sequence-specific sticky-ended cohesion. The 3D X-ray structure of these DNA crystals holds promise for the design of new structural motifs to create programmable 3D DNA lattices with atomic spatial resolution. The two arrays differ by the use of four or six repeats of the five-nucleotide units in the repeating but statistically disordered central strand. In addition, we report a 2D rhombuslike array formed from similar components