DNA strand exchange and hydrophobic interactions between biomolecules

University dissertation from Chalmers University of Technology

Abstract: The role of hydrophobic interactions in DNA strand exchange has been studied using fluorescence-labeled DNA oligomers in a FRET assay. Strand exchange was found to be accelerated in the presence of polyethylene glycol, which provides a crowded and hydrophobic environment possibly mimicking that of the catalytically active recombinase-DNA complexes. Circular dichroism spectroscopy shows that B-DNA conformation is conserved, so the increased rate of exchange is not simply caused by melting of DNA duplexes. A hydrophobic environment increases the base pairing accuracy of DNA strand exchange, which causes mismatched duplexes to quickly be replaced in the presence of matching strands. It is inferred that these effects are caused by a decrease in water activity which weakens the DNA stacking forces, and by favorable hydrophobic interactions between PEG and DNA chains, with the result that DNA breathing and subsequent strand invasion is facilitated. Linear dichroism and dynamic light scattering were also used to study some other biomolecular systems where hydrophobic interactions are important: lipid membranes, DNA-protein complex, DNA nanoconstructs anchored to membrane surface, and to study fusion of liposomes induced by shearing forces. A DNA hexagon construct was found to adopt different orientations at the membrane surface depending on the number of attached anchors, but the construct itself was inferred to have a metastable shape due to internal flexibility. Finally, an example of assembly of protein subunits to a membrane surface was considered in shape of the ATP synthase system for which we propose that the activation energy of ATP synthesis may be reduced through coupled reactions between three active sites. The results are interesting in more general contexts of methodological improvements for studying biomolecular assembly, including linear dichroism spectroscopy of transmembrane proteins.