Regulation of Biological Activity
DNA Nanoarchitectures

 

Regulation of Biological Activity

One of our research interests is the spatiotemporal control of the activity of biologically active compounds.[15] Our aim is to control nucleic acid activity with light as trigger signal. Nucleic acids and their derivatives and analogues can be used for a variety of applications ranging from gene regulation over the modulation of protein function to molecular diagnostics, molecular probing and beyond. Light is an ideal trigger signal which can be used for addressing single cells in thin tissues or small organisms. With (confocal) microscopes it is possible to both visualize the sample of interest and irradiate certain areas with spatiotemporal and dosage control. There are two different strategies to control the functionality of nucleic acids with light.

The first strategy relies on the installation of a photoswitch which upon irradiation switches between an active and an inactive state. We are using two different approaches to reversibly control the activity of biological systems with light. Either the photoswitch is installed covalently into nucleic acids or modified photoswitches are used as interaction partners. With both approaches we want to generate reliable photoswitches applicable in biological contexts. Two compound classes are of special interest, azobenzene and spiropyran derivatives. The synthesis and modification of these two photoswitch species are our main interests in this field. In first attempts we were able to synthesize a spiropyran compound that is a promising candidate for our further studies.[55]

The second strategy relies on the temporal protection of one or several moieties with a photolabile protecting group (caging group) which renders the whole molecule inactive. Upon irradiation with light the caging groups are removed irreversibly and the “native activity” is restored.
With this strategy we were able to control transcription[11], RNA interference[22], MicroRNA activity[62] and protein activity with light[16,35,52,56]. Furthermore we were able to develop tools (so-called Light-inducible Molecular Beacons) to observe intracellular transport of mRNA to specific cell compartments.[50]

 

DNA Nanoarchitectures

About half a century ago G. E. Moore (cofounder of Intel) predicted an exponential growth of the transistor density in integrated circuits. While the empiric ‘law’ still holds amazingly well it is becoming more and more difficult to build even smaller objects: In the majority of cases photolithographic methods are applied and the wavelength of the light which is used represents a natural lower limit for the smallest possible feature. In a complementary ‘bottom up’ approach it should be possible to build up functional architectures with molecules.
Among all possible solutions to realize this DNA is a very interesting candidate: It can be easily synthesized and manipulated, its structure is well known and architectures can be “programmed” using the Watson-Crick interaction. Yet there are even more interaction principles one can use: For example DNA-binding polyamides as a sequence-specific glue to produce defined assemblies or Hoogsteen hydrogen bonding that occurs for example in G-quadruplexes.
In most cases, hybridization and thus the assembly of nanostructures can only be controlled by designing the sequence and the length of the oligonucleotides and changing the melting temperatures, or by sequential addition of oligonucleotides to an experiment. Therefore, the application of photoreactive compounds provides a very potent tool for the whole field of DNA nanotechnology, since they offer the possibility to control the assembly of DNA strands by light. The application of DNA based nanoarchitectures in future devices depends on the ability to functionalize them. Modifying DNA to obtain strands with desired physical properties and incorporation of such modified strands in DNA nanoarchitectures is of great interest for the development of nanoscale biosensors, molecular wires or other nanoscale machines. In combination with the ability to control processes by light, we want to develop light-inducible nanomachines. The aforementionend principles have been used in our group i.e. to build double-stranded DNA catenanes[44] and light-triggered DNA minicircles[46]

To characterize connectivity and structural integrity of structures on the dimension of nanoscale appropriately, imaging techniques with powerful instruments obtaining high spatial resolution like AFM (atomic force microscopy) are used.