Highlights 2013

About the center

Centre for DNA Nanotechnology (CDNA) was established in March 2007 by Kjems, Besenbacher and Gothelf in collaboration with the two 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 2013 are described in the following.

Enzymatic coupling of macromolecules to DNA

Complex DNA nanostructures such as DNA origami are highly useful for organization materials other than DNA at the nanoscale, however it requires that the material is attached to a DNA strand. Another aspect of this is that the price of chemically modified DNA strands is generally much higher than unlabelled synthetic DNA strands. Therefore we have explored the opportunities of enzymatic coupling of small and large biomolecules and polymers to DNA. We have shown that the enzyme Terminal deoxynucleotidyl Transferase (TdT) is able to accept nucleoside triphosphates coupled to macromolecules in an enzymatic reaction[1]. The method enables rapid functionalization of multiple DNA-strands for functionalized nanostructures with polymers, dendrimers and proteins and is a major asset for covering a complete surface of a DNA structure e.g. with polyethylene glycol chains.

Figure 1. Enzymatic conjugation of a protein, here streptavidin, to a DNA strand by linking a triphosphate to the protein and subsequent enzymatic labeling.


Assembly of DNA Nanostructures at room temperature

One of the fundamental requirements for self-assembly of DNA nanostructures is a thermal annealing step, which limits the choice of experimental conditions, pathway designs for the formation process and the integration of nanostructures with other materials. In 2013 we reported on a new and general method to assemble DNA nanostructures at room temperature in one step without any use of heating[2]. The method involves incubation of the DNA strands with 30-40% formamide in the assembly buffer. The method has proven efficient for assembly of 2D and 3D origami, SST tile assembly and for a combination of the two. The ability to form complex DNA nanostructures opens a range of new opportunities, in particular for the integration of biomaterials, and for stepwise assembly and fusion of origami and SST structures.

A DNA tweezer controlling enzyme cascades

In collaboration with Hao Yan at Arizona State University we explored the coupling of enzyme cascades in DNA nanostructures[3]. In a tweezer-like DNA nanodevice the activity of an enzyme/cofactor pair was accurately controlled (Figure 2). A dehydrogenase and NAD+ cofactor are attached to different arms of the DNA tweezer structure and actuation of enzymatic function is achieved by switching the tweezers
between open and closed states. The conformational state of the DNA tweezer is controlled by the addition of specific oligonucleotides that serve as the thermodynamic driver (fuel) to trigger the change. Using this approach, several cycles of externally controlled enzyme inhibition and activation are successfully demonstrated. This principle of responsive enzyme nanodevices may be used to regulate other types of enzymes and to introduce feedback control loops.

Figure 2. Schematic illustration of the mechanics of the DNA tweezer-regulated enzyme nanoreactor.

Instigation of EScoDNA

The European School of DNA Nanotechnology (EScoDNA), an Initial Training Network (ITN) under the European Commission’s Marie Curie Actions research fellowship programme (FW7), was instigated in February 2013 with a total funding of EUR 4 million. Kurt Gopthelf is the Coordinator of the network Dr. Lise R. L. Pedersen was employed as administrator at both CDNA and EScoDNA. The first out of 14 international PhD students and 2 post docs were enrolled at the Partner institutions Ludwig Maximilian University of Munich, Technical University of Munich, Karolinska Institutet, University of Oxford, Vipergen ApS (Denmark), baseclick GmbH (Germany) and Aarhus University in Spring 2013 and in October 2013 the enrolement was completed. Three PhD students were enrolled at CDNA.

Figure 3. Students and supervisors at the EScoDNA kick-off meeting in Aarhus in November 2013

In November 2013 the EScoDNA kick off meeting was organized at Aarhus University where all students and the partners Tim Liedl (LMU), Fritz Simmel (TUM), Björn Högbjerg (Karolinska) and Andrew Turberfield (Oxford) were present (Figure 3).

References


[1] Sørensen, R. S.; Okholm, A. H.; Schaffert, D.; Kodal, A. L. B.; Gothelf, K. V.; Kjems, J. K. Enzymatic Ligation of Large Biomolecules to DNA. ACS Nano, 2013, 7, 8098-8104.

[2] Zhang, Z.; Song, J.; Besenbacher, F.; Dong, M.; Gothelf, K. V. Self-Assembly of DNA Origami and Single-Stranded Tile Structures at Room Temperature. Angew. Chem. Int. Ed. 2013, 52, 9219.

[3] Liu, M.; Fu, J.; Hejesen, C.; Yang, Y.; Woodbury, N. W.; Gothelf, K. V.; Liu, Y.;Yan, H. A DNA-tweezer-actuated enzyme nanoreactor. Nat. Comm. 2013, 4, 2127.