1. Field of the Invention
The present invention relates to nucleic acid structures and to symmetrical and asymmetrical two dimensional and three dimensional polynucleic acid structures with symmetrical intermolecular contacts formed from joining antiparallel double crossover molecules. In addition, the present invention relates to the method for producing such polynucleic acid structures.
2. Description of the Related Art
A key aim of biotechnology and nanotechnology (Feynman et al., Miniaturization, 282-296, 1961 and Drexler, Proc. Nat. Acad. Sci. (USA) 78: 5275-5278, 1981) is a rational approach to the construction of new biomaterials, including individual geometrical objects and nanomechanical devices, and extended constructions, particularly periodic matter with control of the molecule architecture such that it would permit the fabrication of intricate arrangements of atoms to serve many practical purposes (Robinson et al. Prot. Eng. 1: 295-300, 1987; Seeman, DNA & Cell Biol. 10: 475-486, 1991; Seeman, Nanotechnol. 2: 149-159, 1991). The informational macromolecules of biological systems, proteins and nucleic acids, are believed to have the potential to serve as building blocks for these constructions, because they are used for similar purposes in the cell. For instance, nanometer-scale circuitry and robotics could accomplish many tasks that are impossible today. One can envision improvements in the storage and retrieval of information, directed attacks on the molecular basis of medical problems, and the assembly of very smart materials as possible end products of the ability to control the structure of matter on the nanometer scale.
The laboratory of the present inventors has been engaged in the nanoscale construction of stick figures, using branched DNA molecules as building blocks. The edges of these figures consist of double helical DNA, and the vertices correspond to the branch points of stable DNA branched junctions (Seeman, J. Theor. Biol. 99: 237-247, 1982; Seeman, J. Biomol. Str. & Dyns. 3: 11-34, 1985). The molecules can be assembled in solution or on solid supports. The molecules reported previously contain helix axes that have the connectivities of a quadrilateral (Chen et al., J. Am. Chem. Soc. 111: 6402-6407, 1989; U.S. Pat. Nos. 5,278,051 and 5,468,851) a Platonic polyhedron, a cube (Chen et al., Nature (London) 350: 631-633, 1991; U.S. Pat. No. 5,386,020) an Archimedean polyhedron, i.e., a truncated octahedron (Zhang et al., J. Am. Chem. Soc. 116: 1661-1669, 1994). If the edges of DNA polyhedra are designed to contain an integral number of double helical turns, every face corresponds to a cyclic strand of DNA, which is linked to each of its neighbors. Thus, the cube is a hexacatenane, and the truncated octahedron is a 14-catenane of DNA; the extent of linking between faces is equal to the number of turns in the edge where they meet. Both the polyhedra contain two turns per edge, so each of their cyclic strands is doubly linked to each of its neighbors.
The construction of discrete closed structural entities, such as polyhedra, can be controlled readily, because the sequence symmetry of the molecules can be limited by the molecular design. Thus, the ligation of identical DNA sticky end pairs (to yield an edge) can be separated from each other in time by protection techniques (Zhang et al., J. Am. Chem. Soc. 114, 2656-2663, 1992 and U.S. Pat. Nos. 5,278,051 and 5,468,851; the use of sticky ends with unique sequences also provides control over the assembly of finite objects (Chen et al., 1989, supra). This situation does not apply to the construction of periodic matter (crystals), where translational symmetry is an inherent characteristic of the system, because the contacts between all unit cells are identical. It is possible to envision deprotection schemes to unmask, successively, individual polyhedra or polyhedral clusters containing the same sticky ends by means of different restriction enzymes (Seeman et al., Biomolecular Materials: Materials Res. Soc. Symp. Proc. 242: 123-134, 1993). Likewise, one can imagine the construction of `pseudocrystals`, having the same backbone structure and topology in each unit cell, but differing in sequence at key sites. Such schemes may be applicable to DNA computing (Adleman Science 266: 1021-1024, 1994), but they are both cumbersome and expensive and they do not offer a practical route to the assembly of large repetitive constructs, even if one pictures hierarchical assembly of subsections of the target crystal.
There are at least three key elements necessary for the control of three-dimensional structure in molecular construction that involves the high symmetry associated with crystals: (1) the predictable specificity of intermolecular interactions between components; (2) the structural predictability of intermolecular products; and (3) the structural rigidity of the components (Liu et al., Nanobiol. 3: 177-188, 1994). DNA branched junctions are excellent building blocks from the standpoint of the first two requirements, which are also needed for the construction of individual objects, because, (1) ligation directed by Watson-Crick base pairing between sticky ended molecules has been used successfully to direct intermolecular specificity since the early 1970's (Cohen et al., Proc. Natl. Acad. Sci. (USA) 70: 3240-3244, 1973); and (2) the ligated product is double helical B-DNA, whose local structural parameters are well-known (Arnott et al., J. Mol. Biol. 81: 93-105, 1973).
The key problem in working with branched DNA as a construction medium is that branched junctions have been shown to be extremely flexible molecules (Ma et al., Nucl. Acids. Res. 14: 9745-9753, 1986; Petrillo et al., Biopolymers 27: 1337-1352, 1988). The ligation of 3-arm and 4-arm DNA branched junctions leads to many different cyclic products, suggesting that the angles between the arms of the junctions vary on the ligation time-scale; these angles are analogous to valence angles around individual atoms. Likewise, a 5-arm DNA branched junction has been shown to have no well-defined structure, and a 6-arm DNA branched junction has only a single preferred stacking domain (Wang et al., Biochem. 30: 5667-5674). Leontis and his colleagues have shown that a three-arm branched junction containing a loop of two deoxythymidine nucleotides has a preferred stacking direction (Leontis et al., Nucl. Acids Res. 19: 759-766, 1991) and ligation along this direction shows a lower propensity to cyclization (21.3%) than other directions (Liu et al., 1994, supra), but it is not possible to treat the stacking domain in the Leontisian junction as a rigid component (Qi et al. 1996).
To overcome the problem of branched DNA being extremely flexible and therefore unsuitable from the standpoint of structural rigidity of the components as the third key element, DNA structures that fail to cyclize significantly in the course of ligation reactions (a measure of the rigidity of the DNA) were sought by the present inventors. DNA double crossover molecules, which are model systems for structures proposed to be involved in genetic recombination initiated by double strand breaks (Sun et al., Cell 64: 1155-1161, 1991; Thaler et al., Ann. Rev. Genet. 22: 169-197, 1988), as well as meiotic recombination (Schwacha et al., Cell 83: 783-791, 1995), were explored with respect to the structural features of these molecules, and the inventors' laboratory has shown that there are five different isomers of double crossover molecules (Fu et al., Biochem. 32: 3211-3220, 1993). Double crossover molecules were used in the laboratory of the present inventor to establish the sign of the crossover node in the Holliday junction (Fu et al., J. Mol. Biol. 236: 91-105, 1994), to construct symmetric immobile branched junctions (Zhang et al., J. Mol. Biol. 238: 658-668, 1994), and to examine the effect of domain orientation on cleavage by the Holliday junction resolvase, endonuclease VII (Fu et al., Biochemistry 33: 3896-3905, 1994). The helical domains were found to be parallel in three of the five isomers, and antiparallel in the other two. Those with parallel domains are not as well-behaved as those with antiparallel domains (Fu et al., 1993, supra).
Citation of any document herein is not intended as an admission that such document is pertinent prior art, or considered material to the patentability of any claim of the present application. Any statement as to content or a date of any document is based on the information available to applicant at the time of filing and does not constitute an admission as to the correctness of such a statement.