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Current Research Highlights

Optical Tweezers: At the Institute of Physics, a new type of combined optical tweezers instrument has been setup in our group (2, 3, 7). The implementation of confocal optical spectroscopy with single photon sensitivity allows combining force measurement in the subpiconewton regime with optical detection of nanometer distances between single fluorophores (e.g. fluorescence resonance energy transfer, FRET) on interacting biomolecules or detection of local motion on a biomolecule. In figure 1 part of the current setup is shown.

Figure 1: The biomolecule of interest is placed inbetween two polystyrene spheres (size ~2 µm diameter). The lower bead is held by a micropipette by suction and the top bead is held by laser tweezers that allow measuring sub-pN forces. On one end, the molecule is covalenty attached and on the other end, the molecule is bound via a strong biomolecular bond. Optical spectroscopy is performed by illuminating part of the field of view by an excitation laser (488 nm) and the optical response (fluorescence) is read out by EM-CCD or APDs.

By fluorescently labeling molecules (e.g. dsDNA with SYBR^® green, or single fluorophores at specific sites) the mechanical response can be evaluated with pN precision and in parallel visual inspection can take place by single molecule fluorescence imaging (see figure 2). Our current measurements suggest that dsDNA undergoes a structural transition since the fluorescence response does not change during the transition at pulling forces between 65 and 75pN.

Figure 2: A single dsDNA molecule subjected to an external force of ~80 pN. The dsDNA molecule is clamped in between two polystyrene spheres and pulled by a piezo electric element. One question remaining is: Does the dsDNA dehybridize (dsDNA to two ssDNA) during the overstretch transition or does it undergo a structural transition into S-form (as shown in the inset: data from molecular modeling Lebrun A et al. NAR (1996)).

Additional projects are focused on the genetic transfer from bacteria to plant cells and at influences on DNA structure/mechanics of substances used for gene transfer in eukaryotes. Here we study binding kinetics or mechanical changes induced by binding the molecules of interest to DNA molecules and then correlate our physical measurement to structure and function of the molecules involved in the process (collaborations with the Engel group and Giese group.).