Membrane-mediated protein-protein and protein-lipid interactions, membrane protein localization, and related dynamics, modulate membrane protein function [1]. So far membrane structure and dynamics could not be studied altogether lacking the technique that analyses unlabelled proteins at submolecular lateral and high temporal resolution. Here we used high-speed atomic force microscopy (HS-AFM, [2]) to characterize the movements and interactions of unlabelled porin OmpF [3] and aquaporins [4] in native membranes. First an introduction to AFM and its use in membrane biology will be given, followed by an introduction to the membrane proteins and the membrane structure as a current challenge in biology. Using HS-AFM, we are able to describe essential novel aspects that govern membrane protein assembly and membrane superstructure. Protein motion scales roughly with membrane crowding. However molecules display individuality of diffusion behaviour ranging from fast moving to immobile molecules trapped by favourable protein-protein associations. We derive the molecular interaction probability landscapes and assembly rationales that we compare with coarse-grained molecular dynamics and Monte Carlo simulations. Most recently, we were able to monitor for the first time individual unlabelled membrane proteins directly on cells [5]. HS-AFM may open a novel research avenue that bridges structure of individual membrane proteins and supramolecular membrane architecture. Finally, high-speed force spectroscopy (HS-FS) allows the characterization of interactions within and between molecules over a wide dynamic range [6].High-speed Atomic Force Microscopy (HS-AFM) analysis of the lateral and rotational displacement of outer membrane protein F (OmpF). HS-AFM allows to study non-labeled membrane protein dynamics. Furthermore, not only a single molecule of interest is seen, but also its molecular environment, giving access to completely novel parameters such as how crowding alters the diffusion properties and the encounter probability. (adapted from: Nature Nano, 2012, 7: 525-529). To visualize the movie, see: http://www.nature.com/nnano/journal/v7/n8/extref/nnano.2012.109-s2.aviReferences: [1] D.M. Engelman, Nature 438, 578 (2005) [2] T. Ando, et al., Proceedings of the National Academy of Sciences 98, 12468 (2001) [3] I. Casuso, et al., Nature Nanotechnology, 7, 525 (2012) [4] A. Colom, et al., Journal of Molecular Biology, 423, 249 (2012) [5] A. Colom, et al., Nature Communications, DOI:10.1038/ncomms3155 (2013) [6] F. Rico, et al., Science, 342, 741 (2013)

High-speed and high resolution atomic force microscopy monitors dynamic membrane-mediated membrane protein interactions

248th National Meeting (2014)

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