Dokument

Summary:
Bacteriorhodopsin (BR) is a light-driven proton pump and the key protein in halobacterial photosynthesis. In its native host, the archaeon Halobacterium salinarum, BR trimers arrange into a 2-D crystalline lattice, the so-called purple membranes (PMs) which comprise BR and lipids only. Along with the PM assembly BR is astonishingly stable against thermal and chemical stress which makes it an excellent candidate for a variety of technical applications. Many technical applications involve immobilization of PM in a matrix or at a surface. Sugar glasses are frequently used matrixes for the stabilization of biomolecules against thermal stress as well as dehydration. In the following work temperature-dependent interactions between PM and sugar glasses were analyzed. Above T > 60°C a blue membrane is formed the so-called sugar-induced membrane (SIBM). This thermochromism was explained by a model which is based on the release of divalent cations from PM and the chelating properties of carbohydrates. Due to their high mass adsorption is an appropriate method for the immobilization of PMs at surfaces. In this work self-assembled monolayers (SAMs) of alkanethiols on gold were used and analyzed for this purpose. To fabricate selective adsorption sites the alkanethiol-SAMs have to be structured. An appropriate procedure for the patterning of alkanethiol-SAMs has been developed which has been named “submerged laser ablation” (SLAB). Knowledge about the mechanical properties of freely suspended membranes is essential for nanobiotechnological applications of PM. In each PM thousands of BR molecules are arranged in a unidirectional manner and are strongly coupled due to the crystalline assembly. Therefore, collective conformational changes of single BRs should influence the topology of freely suspended PMs. In this work it was demonstrated by direct imaging of freely suspended native PMs via cryogenic high-resolution scanning electron microscopy (cryo-SEM) that the flat disk-like shape of PM changes dramatically as soon as most of the BRs are in the M2 state which is characterized by wedge-shaped BRs due to collective opening of the cytoplasmatic half-channels. Light- as well as pH-induced shape changes are easily observed with mutated BRs which accumulate wedge-shaped BR molecules. These results open the way for further nanobiotechnological applications of PM, i. e. as supramolecular actuator.

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