"We have managed to create multiple orthogonal bonds between DNA nanostructures without using hybridization of 'sticky ends'," explained team member Sungwook Woo. "Instead, we programmed bonds using 'blunt-end' stacking interactions and encoded bond types using binary codes and shape complementarity along the edge of a DNA origami rectangle."

DNA strands are long chains composed of four different bases, A, C, G and T. A always sticks to, or pairs up with T and C always pairs up with G, so that a strand of DNA with a given sequence, say ATGC, pairs strongly only to a complementary sequence GCAT (in an antiparallel fashion). While this base pairing is crucial for specific binding between DNA strands, another type of interaction, known as base stacking seems to be even more important for stabilizing DNA duplexes. Stacking involves pairs of bases, say AT sticking strongly to GC.

Twisted ladder
"If we then visualize DNA as a twisted ladder rather than a twisted helix, we notice that it has a top rung and a bottom rung that are not joined to other bases," explained team leader Paul Rothemund. "Such an end, which is like the sawn-off end of a log, is called a blunt-end. Without partners to stick to, these blunt-ends can stick to each other."

The California researchers exploited this phenomenon to create DNA shapes essentially made up of parallel DNA helices that resemble the walls of a log cabin. The secret to their technique was to give the log walls a sort of jigsaw-puzzle shape (or "orthogonality"), rather than just a straight shape. The team designed their jigsaw-puzzle shapes using a computer model and then created five different real DNA structures in the lab. Each structure was actually a folded-up DNA "origami".

"When we mixed the five different structures together, they assembled to form exactly the five-structure chains that we intended," Rothemund told nanotechweb.org. "In other words, the five distinct DNA origami were connected by placing mutually orthogonal stacking bond pairs (stacking sequences and their respective complementary sequences) on the edges of each origami," added Woo.

According to the researchers, the technique could be used as an additional way, together with the sticky-end method, to assemble DNA structures into larger, more complex structures. The stacking-bond method also offers some advantages compared with using sticky ends. For example, researchers do not need to design and synthesize new sequences for each new origami-origami interaction and the binary coding approach allows each bond type to be reprogrammed easily and cheaply following synthesis.

Lower activation energies
"Looking ahead, we believe that stacking bonds will offer a new and important way to rearrange bonds using forces that are difficult or impossible to implement with DNA hybridization," said Woo. "For example, for structures linked by DNA hybridization to rearrange, the DNA helices must unwind and then rewind. This could involve high activation energies depending on the number and strength of the links involved. Stacking bonds, on the other hand, may have low activation energies."

If this happens to be true, it may come in handy for making more sophisticated nanomachines in the future. Indeed, the team says that it hopes to exploit stacking bonds between the mechanical parts on devices that must both self-assemble and then slide past each other (while remaining in contact) under the shear forces applied by molecular motors. "Many biological nanomachines, such as ATP synthase and the rotary motors of bacterial flagella, already use such a mechanism and being able to mimic this action is fundamental if we are to build truly complex machines on the nanoscale," added Rothemund.

The work was reported in Nature Chemistry.