Janshoff, Andreas - Georg-August-Universitäten Göttingen

研究领域

Mechanics of small systems The behavior of small systems has attracted great interest in biology, chemistry and physics since their display striking properties as a direct result of their small size. The physics of small systems is strongly governed by fluctuations that produce significant deviations from the behavior of large ensembles. The ultimate small device is a single molecule, where fluctuations can be considered to be large and stochasticity dominates its thermal behavior. We are interested in the mechanical, optical and electrical properties of small systems ranging from single molecules to living cells. Nanotechnology is not a novel conception as it has been used by nature for a long time giving rise to complex and highly efficient machines on the nanometer scale, e.g. ATPase or flagella. Hence, artificial replications of biological concepts are envisioned in many scientific branches. For example, in materials science, the development of self‐cleaning surfaces was inspired by the lotus plant that is in Buddhism a symbol for purity. Although the “lotus effect” is a famous mimic of nature’s nanotech toolbox, it is merely one out of many examples from biomimetic engineering. Mimicking detailed features of complex natural systems, like living cells, remains a challenge in actual research that is engaged in different strategies. Quantitative, systematic and reliable studies of individual biological phenomena are often only feasible by usage of simplified systems that focus exclusively on the subject of investigation, but with limited degree of complexity that still permits an authentic representation of biological activity. Hence, in actual research there is an ongoing demand for new and innovative biomimetic model systems enabling the investigation and understanding of fundamental biological processes. Mechanics of single molecules Chemical reactions and structural transitions of supramolecular systems require a comprehensive understanding of the stochasticity of transformations on small length scales. Particularly, mechanically driven transformations as carried out by single molecule stretching experiments offer a unique way to study fundamental theorems of statistical mechanics as recently shown for the unzipping of RNA hairpins. In our research group, we design single-molecule experiments that allow to address fundamental statistical mechanics in an unprecedented way employing a molecular design based on oligo calix[4]arene catenanes. These catenanes permit the control of the boundaries for separation and recombination of hydrogen bridges by mechanically locking the unbound state with intramolecularly entangled loops. This mechanically locked structure tunes the energy landscape of calixarene dimers, thus permitting the reversible rupture and rejoining of individual nanocapsules. Stochastic modeling of hydrogen bonds under external load allows reconstruction of the energy landscape. Local Mechanics and Dynamics of Cells and Membranes Eukaryotic cells are supramolecular assemblies of vastly different composition and topology. A structural model for the adherent cell has been developed by drawing analogy to the so-called “tensegrity” ( from tension integrity) structures found in modern architecture. A key feature of tensegrity structures is that they require tension in some of their elements to resist shape distortion due to an external load. Elasticity of cells largely determines fundamental cell behavior such as migration, differentiation, and interaction with each other. On the single cell level force can initiate cell protrusion, alters cell motility and affect biochemical pathways that regulate cell function division and death. Resolving cell mechanics on various length scales is therefore pivotal to understand how cells respond to mechanical stress and how the entity of plasma membrane and cytoskeleton framework interact with each other on a supramolecular level. Atomic force microscopy allows not only visualizing the topography of submicrometer structures, it also permits to study material properties using phase imaging, pulsed force mode microscopy or force volume measurements. Crucial information about the cell membrane’s bending resistance, however, remains inaccessible due to the complex elastic response of whole cells originating from interplay of cytoskeleton, cytoplasm, osmotic pressure and cell membrane. Our approach towards elasticity mapping of free standing cellular membranes relies on the combination of AFM as a scanning device with a substrate topology that allows bending membranes at locally defined positions using highly ordered porous substrates with holes in the nano- to micrometer regime. The dynamic response of mammalian cells to nanoparticles-exposure is measured by thickness shear mode resonators that pick up harmful substances with unprecedented sensitivity in a non-invasive and label free fashion. The figure above shows cell slices of MDCK II cells exposed to gold nanoparticles (40 nm in diameter) in dark-field (plasmon resonance) and electron microscopy. Cellular dynamics or motility is measured by means of electric cell-substrate impedance sensing which allows to measure minuscule shape changes of cells as well as changes of the cleft between cell and substrate. The figure shows a sequence of optical micrographs together with the corresponding impedance changes (normalized to the impedance of the uncovered electrode). The circular gold electrode is visible in the center and seeding of amoebae (7500 cells/mm²) produces a significant increase in impedance. Keeping the cells in the absence of food (buffer), they started to oscillate after 4 - 5 h with a time period of 12 – 20 min. Mimicking Membrane Docking and Fusion Membrane-Membrane interaction plays a pivotal role in many essential biological processes such as fusion, exocytosis, endocatosis and formation of cell-cell contacts. Fusion with the host-cell plasma membrane is a crucial stage in the life cycle of all enveloped viruses as it enables intracellular deposition of the viral genome before replication. The mechanics of viral fusion is still a matter of debate and an intricate physical and biological problem. Retroviruses, such as HIV, or human T cell leukemia virus 1 (HTLV-1), enter cells by receptor mediated attachment and subsequent membrane fusion of virions with target cells. This membrane fusion is based on virally encoded envelope glycoproteins (Env), which are presented on the surface of the virus or infected cell. Structural analysis of these trimers of surface glycoprotein subunits anchored to a trimer of transmembrane glycoproteins has provided insight into the mechanism of Env-catalyzed membrane fusion. A variety of different approaches have been proposed to interfere with fusion at the crucial step of formation of the trimer-of-hairpin conformation. However, no unifying assay concept has yet been suggested that stringently allows quantification and screening of potential antagonists. As a consequence, we suggest and in principle realized a sensing scheme for fusion inhibitors based on lipopeptides that present the N-peptides of SIV or HIV in the required prehairpin intermediate conformation as a binding receptor unit for potential antagonists. Besides designing tailored sensors mimicking the natural situation as closely as possible we are also interested in measuring the force between functionalized lipid bilayers to model the cell-cell contact or docking event in vesicle fusion. The figure above shows a colloidal probe coated with a lipid bilayer that displays embedded receptors. The corresponding force distance curve shows the formation of membrane tethers upon retraction from a surface bearing the respective ligands in a matrix of lipids

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Rother, J.; Büchsenschütz-Göbeler, M.; Nöding, H.; Steltenkamp, S.; Samwer, K.; Janshoff, A. (2015) Cytoskeleton remodelling of confluent epithelial cells cultured on porous substrates. J. R. Soc. Interface 12, 20141057. Milovanovic, D.; Honigmann, A.; Koike, S.; Göttfert, F.; Pähler, G.; Junius, M.; Müllar, S.; Diederichsen, U.; Janshoff, A.; Grubmüller, H.; Risselada, H. J.; Eggeling, C.; Hell, S. W.; van den Bogaart, G.; Jahn, R. (2015) Hydrophobic mismatch sorts SNARE proteins into distinct membrane domains. Nature Communications 6:5984. Breus, V.V.; Pietuch, A.; Tarantola, M.; Basché, T.; Janshoff, A. (2015) The effect of surface charge on nonspecific uptake and cytotoxicity of CdSe/ZnS core/shell quantum dots. Beilstein J. Nanotechnol. 6, 281-292. Pietuch, A.; Brückner, B. R.; Schneider, D.; Tarantola, M.; Rosman, C.; Sönnichsen, C.; Janshoff, A. (2015) Mechanical properties of MDCK II cells exposed to gold nanorods. Beilstein J. Nanotechnol. 6, 223-231. Rosman, C.; Pierrat, S.; Tarantola, M.; Schneider, D.; Sunnick, E.; Janshoff, A.; Sönnichsen, C. (2015) Mammalian cell growth on gold nanoparticle-decorated substrates is influenced by the nanoparticle coating. Beilstein J. Nanotechnol. 5, 2479-2488. Rother, J.; Nöding, H.; Mey, I.; Janshoff, A. (2014) AFM-based microrheology reveals significant differences in the viscoelastic response between malign and benign cell lines. Open Biol. 4: 140046. Fichtner, D.; Lorenz, B.; Engin, S.; Deichmann, C.; Oelkers, M.; Janshoff, A.; Menke, A.; Wedlich, D.; Franz, C. M. (2014) Covalent and Density-Controlled Surface Immobilization of E-Cadherin for Adhesion Force Spectroscopy. PLoS ONE 9(3): e93123. Stephan, M.; Mey, I.; Steinem, C.; Janshoff A. (2014) Combining Reflectometry and Fluorescence Microscopy: An Assay for the Investigation of Leakage Processes across Lipid Membranes. Anal Chem. 86 (3), 1366-1371. Braunger, J. A.; Brueckner, B. R.; Nehls, S.; Pietuch, A.; Gerke, V.; Mey, I.; Janshoff, A.; Steinem, C. (2014) Phosphatidylinositol 4,5-bisphosphate alters the number of attachment sites between ezrin and actin filaments: a colloidal probe study. J. Biol. Chem. 289 (14), 9833-9843. Stephan M.; Kramer C.; SteinemC.; Janshoff A. (2014) Binding Assay for Low Molecular Weight Analytes Based on Reflectometry of Absorbing Molecules in Porous Substrates. Analyst 139, 1987-1992. Chizhik, A.I.; Rother, J.; Gregor, I.; Janshoff, A.; Enderlein, J. (2014) Metal-induced energy transfer for live cell nanoscopy. Nature Photonics 8, 124-127. Vilardi, F.; Stephan, M.; Clancy, A.; Janshoff, A.; Schwappach, B. (2014) WRB and CAML Are Necessary and Sufficient to Mediate Tail-Anchored Protein Targeting to the ER Membrane. PLoS ONE 9(1): e85033.