Antijunctions and mesojunctions are new classes of multistranded DNA complexes. They represent a generalization of DNA branched junction complexes, such as the Holliday recombination intermediate. Each strand of a conventional branched junction participates in two different double helices, and this is also true for mesojunctions and antijunctions. The helix axes of conventional branched junction complexes may be drawn to converge at a point, but this convergence occurs for lines drawn perpendicular to the helix axes of antijunctions. Mesojunctions are complexes that mix these features of junctions and antijunctions. Antijunction complexes require an even number of strands. We have synthesized the mesojunction containing three strands, the two mesojunctions containing four strands, and the antijunction containing four strands; we compare them with branched junctions containing three and four strands, derived by permutations of the same sequences. Each double helix is designed to contain 1.5 turns of DNA. A tendency to oligomerize makes it difficult to capture antijunctions and mesojunctions in stable discrete complexes, in contrast to conventional branched junctions. For both three-strand and four-strand complexes, Tm is highest for conventional branched junctions. Ferguson analysis indicates similarities in the occluded surface area of junctions, antijunctions, and one four-strand mesojunction, but the other four-strand mesojunction has a much lower apparent surface area. Hydroxyl radical cleavage patterns suggest that the four-strand antijunction and the low-surface-area four-strand mesojunction form stacking domains, analogous to the behavior of conventional branched junctions. These new structures are related to replicational and recombinational intermediates and to single-stranded nucleic acid knots.

The construction of knotted topologies is a key goal of stereochemistry. In order to measure the chiral properties of knotted molecules, it is necessary to produce both enantiomers of a knot from the same molecule. A molecule containing the same backbone structure that is an amphichiral knot can provide a useful control molecule for such measurements. In the case Of molecules with chiral backbones, configurational chirality, exclusive of the chirality due to knotting, must be measured from the circle of the same sequence. Trefoil knots of both chiralities, an amphichiral knot, and an unknotted circular molecule have all been constructed by enzymatic closute of the same linear DNA molecule. The molecule contains two double helical domains that can be induced to assume the righthanded B conformation or the left-handed Z conformation under selected solution conditions. The molecules expected to contain left-handed DNA have been shown to bind an anti-Z-DNA antibody in gel-retention assays.Knotted topologies have a long history in both biology1-5 and chemistry,6-9 but it is only recently that it has been possible to construct particular knotted molecules. Sauvage and his colleagues have reported the synthesis of a mixture of both chiralities of trefoil knots10'11 from small molecules, and we have reported the synthesis of trefoil12-14 and figure-815 knots from single-stranded DNA. The physical properties of knots are of great interest.16 However, they are not readily available from chemical species, because of the difficulties associated with their syntheses. In order to use physical means to probe chirality resulting from knots of opposite handedness, it is necessary to produce and isolate both enantiomers of the same molecule. Liang and Mislow17 have used the term amphicheiral, to describe knots that are topologically achiral; in discussing knots constructed from chiral components (D-nucleotides), we use the term "cheirality" here to describe the chirality that refers

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Recently, we have invested a great deal of effort to construct molecular building blocks from unusual DNA motifs. DNA is an extremely favorable construction medium. The sticky-ended association of DNA molecules occurs with high specificity, and it results in the formation of B-DNA, whose structure is well known. The use of stable-branched DNA molecules permits one to make stick-figures. We have used this strategy to construct a covalently closed DNA molecule whose helix axes have the connectivity of a cube, and a second molecule, whose helix axes have the connectivity of a truncated octahedron. In addition to branching topology, DNA also yields control of linking topology, because double helical half-turns of B-DNA or Z-DNA can be equated, respectively, with negative or positive crossings in topological objects. Consequently, we have been able to use DNA to make trefoil knots of both signs and figure of 8 knots. By making RNA knots, we have discovered the existence of an RNA topoisomerase. DNA-based topological control has also led to the construction of Borromean rings, which could be used in DNA-based computing applications. The key feature previously lacking in DNA construction has been a rigid molecule. We have discovered that DNA double crossover molecules can provide this capability. We have incorporated these components in DNA assemblies that use this rigidity to achieve control on the geometrical level, as well as on the topological level. Some of these involve double crossover molecules, and others involve double crossovers associated with geometrical figures, such as triangles and deltahedra.