Designed two-dimensional DNA Holliday junction arrays visualized by atomic force microscopy

A two-dimensional DNA crystal has been designed and constructed from Holliday junction analogues that contain two helical domains twisted relative to each other. The Holliday junction is not an inherently rigid system, but it can be made less flexible if it is combined into a larger construct. We have fused four junctions into a rhombus-like molecule consisting of four six-turn helices, two on-an upper layer and two on a lower layer; the branch points, which define vertices, are separated by four double helical turns each. Ligation of the rhombus-like motifs produces no cyclic species, when assayed by ligation-closure experiments. Self-assembly of the rhombuses in one dimension leads to a linear pattern. The rhombuses can be directed to self-assemble by hydrogen bonding into a two-dimensional periodic array, whose spacing is six turns in each direction. The expected spacing is seen when the array is observed by atomic force microscopy (AFM). Variation of the dimensions of the repeat unit from six turns x six turns to six turns x eight turns results in the expected increase in unit cell dimensions. Hence,it is possible to assemble periodic arrays with tunable cavities using these components. This system also provides the opportunity to measure directly the angles or torsion angles between the arms of branched junctions; here we measure the torsion angle between the helical domains of the Holliday junction analogue. We find by AFM that the torsion angle between helices is 63.5 degrees, in good agreement with previous estimates.