Aufbau nanoskopischer Hybridstrukturen am Beispiel von MrgA und Purpurmembran
Aufbau nanoskopischer Hybridstrukturen am Beispiel von MrgA und Purpurmembran.
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Table of Contents: Biological compounds display a rich variety of features which have implications for technical applications, such as catalysis and selective high affinity. The utility of biological compounds as building blocks for device fabrication, however, is severely hampered by two general restrictions: Firstly, biomolecules tend to lose their native biological integrity and function in other than near-physiological conditions, and they usually have poor long term stabilities. Secondly, many of the well estabished state-of-the-art techniques for building up nanostructured surfaces, such as methods used in silicon wafer based semiconductor fabrication, are not suited for the procession of biological building blocks. In order to tie in with these techniques new assembly strategies have to be implemented, such as bottom-up and templated self-assembly strategies. In this work, techniques to overcome these problems were elaborated in the context of two biological model substances: MrgA and purple membranes. Firstly, the dodecameric iron storage protein MrgA was used to develop methods for generating nanostructured surfaces. By a three-step procedure (adsorption from an incubation solution, rinsing to remove excess salt and protein, and drying), dry monolayers of MrgA on various solid substrates were constructed, and their morphologies were analyzed by means of AFM. Hexagonal, two-dimensional crystalline monolayers on hydrophilic surfaces were obtained upon adsorption under aqueous conditions at low supersaturation. It was found that the formation of such two-dimensional crystals is heavily dependent on the pH and the salinity of the incubation solution as well as on the surface properties. The correlation of surface coverage with substrate charge, ionic strength, and pH indicated the dominance of electrostatic effects in adsorption – due to the competing interactions of intermolecular repulsion and protein-substrate attraction. In this work, it is shown that adsorption of MrgA leading to the formation of two-dimensional crystals is favoured under conditions close to the isoelectric point of the biomolecule. These results were used to further develop techniques for the guided self-assembly of MrgA nanostructures. To this end, substrates which are nanostructured in regard of their affinities to the protein were generated on template-stripped gold by micro-contact printing of functionalized thiols. Taking advantage of the fine-tunable affinity and a phenomenon usually regarded as an undesired artifact in micro-contact printing, the edge-dominant ink transfer, the surface-templated self-assembly of MrgA in nanometer patterns was achieved. In addition, the protein-shell was removed by pyrolysis in a low pressure oxygen plasma. After this procedure, the metaloxide cores of the holo-protein were found to remain at the location where the proteins had originally been adsorbed on the surface. Therefore, the system developed in this work fulfils all requirements for being a useful tool for structured nanoparticle deposition. Both the discovered novel adsorption morphologies on non-structured substrates and the guided self-assembly of protein nanostructures, represent promising starting points for further scientific studies, such as solid-supported co-crystallization with DNA, and developments aiming at technological applications like the mesostructured deposition of MrgA-caged nanoparticles. Secondly, a method for improving the stability of biological compounds far beyond the limits of natural conditions was developed based on purple membrane as a model substance. Generally, embedding such targets in solid host material increases the stability and the range of possible applications. In medicine and biotechnology, there is a great interest in such composite hybrid materials in form of micro- or nanosized particles. In this work, a biomimetic approach for the encapsulation of biomolecules with silica is presented. It was inspired by the remarkable biomineralisation performed by some eucaryotic algae, the so-called diatoms. In the method elaborated here, two generic techniques were combined: In a first step, the surface of the biomolecule was modified via polyelectrolyte layer adsorption. It was shown that the polyamines used in the first step allow for the surface-templated mineralization of silicic acid and silica nano-particles on the modified surface in the second step of preparation. Both preparative steps were optimized towards mild conditions as required by many biomolecules. Application of this method to the model substance purple membrane resulted in a defined hybrid material with a nano-scale protective encapsulating silica shell. It was demonstrated that while the purple membrane retains its biological function, it is also shielded from otherwise detrimental solvents or molecules of low molecular weight. The method presented here not only allows for the production of significant amounts of encapsuled material, but it was furthermore shown to be adaptable to a variety of substances, such as single-walled carbon nanotubes and polymer microparticles – emphasizing the wide range of application for this method.