Anchoring, Sliding, And Rolling: Visualizing The Three-dimensional Nano-motion And Orientation Of A Single Virus As It Diffuses On A Flat Membrane

Abstract

The localization of objects within the cell and the accurate measurement of relative positions on the molecular level are essential to an understanding of the function of macromolecular complexes. As a consequence, much effort has been directed towards developing imaging techniques that allow the temporal and spatial resolution of events on the nanoscopic scale. Due to their non-invasive nature, optical techniques are particularly suited for studying live samples and several methods have recently demonstrated subdiffraction resolution. However, all these novel methods are based on detecting fluorescence and thereby face strict limitations in accuracy, time-resolution and dimensionality. Here, we show how the combination of label-free detection of nano-objects and single molecule fluorescence detection allows one to map the center of mass motion and the absolute orientation of a single virus with nanometer resolution in real time. We use interferometric scattering detection to resolve the position of individual virions of Simian Virus 40 (a 45 nm DNA tumor virus) with 2 nm accuracy while bound to its cellular receptor GM1 in supported membrane bilayers. At the same time, we detect the fluorescence of a single fluorescent quantum dot attached to the virus via streptavidin-biotin linkage and determine its position with 4 nm accuracy. By overlapping the fluorescence and scattering trajectories, we can resolve the absolute three-dimensional nano-motion of the virus as it diffuses on a two-dimensional membrane. We find that membrane-bound virions exhibit different modes of motion that are strongly influenced by the concentration of the GM1 receptor in the membrane. Besides Brownian motion in the plane of the membrane, we also observe rolling motion on the sub-20 nm scale and periods of apparent standstill in both two and three dimensions.