BROADCASTS.com

In the emerging field of nanobiotechnology, deoxyribonucleic acid (DNA) has already established as a key building block with versatile properties. In the present thesis, self-organising and controllable nanoactuators and a functional "nanomachine", consisting of artificially synthesised short DNA strands, have been designed and characterised. The techniques employed for the construction and analysis of the DNA nanodevices are from the field of physics and biochemistry.\n\nThe essential results of this work are:\n\n\u2022 Characterisation of the DNA tweezers in their open and closed configuration by the means of single-molecule fluorescence measurements. Hence the distribution of the transfer efficiency could be determined for both states and the influence of salt concentration on the distribution in the open state. Based on these results, the distributions of the distances between the dye molecules (and therefore the distances between both arms of the tweezers) could be calculated. For the closed state three subpopulations have been found. \n\u2022 The successful integration of gold nanoparticles into switchable DNA actuators. Here, the fluorescence emission of dyes was measured whose distance to a gold nanoparticle was varied by the DNA actuator. The bulk experiments showed that gold nanoparticles can be effectually used as quenchers for the fluorescent dyes in DNA nanoactuators to monitor distance changes \u2265 10 nm. \n\u2022 The existing concepts for the operation of nanomechanical DNA machines could be employed for the construction of the first switchable aptamer. Switchable aptamers fulfil the function of binding and releasing a protein or other molecules reversibly in a controlled way. In the present project, a molecular machine based on a DNA aptamer was constructed that can cyclically bind to the human protein thrombin and release it upon addition of fuel strands. In fact, one can imagine the developed aptamer machine as a specific "molecular claw". The mode of operation of this switchable aptamer was verified and characterised by fluorescence resonance energy transfer and fluorescence anisotropy measurements. To study the kinetics of the release of thrombin in detail the binding constant of the aptamer machine has been determined using fluorescence correlation spectroscopy measurements. A model of the single reaction steps has been put up, simulated numerically and fitted to the experimentally obtained curves. \n\nThe combination of the operation principle of DNA based nanomechanical devices with the binding properties of DNA aptamers greatly expands the scope for the design and construction of further functional DNA nanodevices. It is assumed that in the near future single-molecule measurements will be increasingly employed within the development of DNA nanomachines and actuators to gain insight in the actually existing structures of these devices.