DNA scaffolds, rafts and switchable supported lipid membranes studied by x-rays

The project A07 aims at resolving the nanoscale structure of DNA-origami, origami crystals and photoswitchable lipid membranes using x-ray scattering. In our first experiments we determined the so-far unexplored intramolecular spacing, structural changes and high-order self-assembly of well dispersed DNA constructs. In the current funding period, we will extend our scattering approach towards single crystal studies in micro-focused x-ray beams using state-of-the art 4th generation synchrotrons (ESRF, Petra III) and free electron x-ray lasers (X-FEL). The new x-ray sources will also allow for studying photo-induced processes with higher precision and time resolution.

Initially we studied basic DNA origami constructs such as bricks and rods, and their swelling and melting as function of ionic strength and temperature, respectively [Fischer 2016]. We probed the relative placement of nanoparticles at DNA-origami binding sites with nm resolution for different geometries such as dimer, trimer, and helix [Hartl 2018]. Larger 3D DNA crystals were characterized and their rhombohedral unit cell geometry was quantified. We are now in the position to push small angle x-ray scattering (SAXS) further to resolve even larger 3D unit cells developed for placement of proteins in defined spatial relations and for photonic structures. To this end we explored the efficiency and structural integrity of encapsulation of DNA origami in silica which protects and stabilizes the structures. We will move towards structure information from individual DNA origami crystals by “serial” or “single molecule” x-ray crystallography. Such experimental schemes became available with coherent high flux X-ray sources and are still developing. In this context, regular DNA based structures, that are highly scattering, provide expedient and challenging samples.
Furthermore, x-rays will be used to probe how molecular switching of nanoagents translates into a mesoscopic structural and mechanical response. Changes in bending rigidity of bilayers in response to molecular photo switching will be extracted from x-ray WAXS scattering data of multistacks within the Caillé theory. Structural properties of unilamelar vesicles, especially bilayer thickness changes, will be determined from SAXS measurements. For this purpose, we will develop high x-ray energy SAXS and WAXS experiments. High energy x-rays are only weakly absorbed in water and therefore x-ray induced photodamage and electrochemical backswitching is prevented. High energy SAXS and WAXS will be used to probe the structural influence of photoactive molecules, which disorder membrane packing in response to visible light.