Highlights 2014

About the center

Center for DNA Nanotechnology (CDNA) was established in March 2007 by Kjems, Besenbacher and Gothelf in collaboration with the American researchers Yan and LaBean. In 2012 the center was extended to 2017 by which four scientists, Ferapontova, Dong, Birkedal and Andersen at iNANO were included as senior members, and William Shih at Harvard University became associated with CDNA. The purpose of the research at CDNA is to explore fundamental aspects of DNA as a programmable tool for directing the assembly of molecules and materials into nanoarchitectures and functional structures. The highlights for CDNA in 2014 are described in the following.

RNA Nanotechnology

Artificial DNA and RNA structures have been used as scaffolds for a variety of nanoscale devices. In comparison to DNA structures, RNA structures have been limited in size, but they also have advantages: RNA can fold during transcription and thus can be genetically encoded and expressed in cells. In a collaboration between Cody Geary and Ebbe Andersen from CDNA and Paul Rothemund at Caltech an architecture has been introduced for designing artificial RNA structures that fold from a single strand, in which arrays of antiparallel RNA helices are precisely organized by RNA tertiary motifs and a new type of crossover pattern. They constructed RNA tiles that assemble into hexagonal lattices and demonstrated that lattices can be made by annealing and/or cotranscriptional folding (Figure 1). Tiles can be scaled up to 660 nucleotides in length, reaching a size comparable to that of large natural ribozymes. The study was published in Science.[1]

Figure 1. In vitro expression and folding of RNA nanostructures.

DNA-Templated conjugation of DNA to proteins

A new method for site selective conjugation DNA to proteins was developed by CDNA researchers and published in Nature Chemistry. DNA-protein conjugates are important in bioanalytical chemistry, molecular diagnostics and bionanotechnology, where the DNA provides a unique handle to identify, functionalize or manipulate proteins. Site-selective conjugation is frequently required to maintain protein activity. However, the preparation of such high quality conjugates most often requires genetically engineered proteins, which is a laborious and technically challenging approach. In the paper a novel and simpler method for creating site-selective DNAprotein conjugates was demonstrated. The method applies concepts from affinity probe technology and DNA directed chemistry to guide a DNA strand to the vicinity of a metalbinding site of the protein (Figure 2). The method is general for his6-tagged proteins and wild-type metal-binding proteins such as serotransferrin. Notably, the method also facilitates conjugation via a histidine cluster in the constant domain of IgG1 antibodies. The technique, DNA-templated protein conjugation (DTPC), offers a more straightforward route to produce site-selective protein conjugates for applications including immuno-PCR, DNA-sensor technology and RNAi delivery.

Figure 2. Site selective conjugation of DNA to an antibody by coordination of a first DNA strand containing a metal-ligand complex to a histidine cluster at an antibody, followed by DNA templated coupling of another DNA strand to a lysine in the proximity of the metal binding site.

Large DNA brick lattices on surfaces

Another important contribution to structural DNA nanotechnology was made by the group of Mingdong Dong at CDNA in collaboration with William Shih and Peng Yin at Harvard University. In this work a general framework for constructing two-dimensional crystals with prescribed depths and sophisticated three-dimensional features is described.[3] The crystals are self-assembled from single-stranded DNA components called DNA bricks (Figure 3). It is demonstrated in the Nature Chemistry paper how the experimental construction of DNA brick crystals that can grow to micrometre size in their lateral dimensions with precisely controlled depths up to 80 nm.

Figure 3. Models and TEM images of DNA brick crystals. The dark areas in the TEM image is the DNA  attice and the local structure is seen in the zoom in in the insets.

References


[1] Geary, C.; Rothemund, P. W. K.; Andersen, E. S. A singlestranded architecture for cotranscriptional folding of RNA nanostructures. Science, 2014, 345, 799.

[2] Rosen, C. B.; Kodal, A. L.; Nielsen, J. S.; Schaffert, D. H.; Scavenius, C.; Okholm, A. H.; Voigt, N. V.; Enghild, J. J.; Kjems, J.; Tørring, T.; Gothelf, K. V. Template-directed covalent conjugation of DNA to native antibodies, transferrin and other metal-binding proteins. Nat. Chem. 2014, 6, 804-809

[3] Ke, Y. G.; Ong, L. L.; Sun, W.; Song, J.; Dong, M. D.; Shih, W. M.; Yin, P. DNA brick crystals with prescribed depths. Nat. Chem. 2014, 6, 994.