(189e) Stay Double, Stay Homologous: Combined Computational/Esperimental Approaches to RAD51/ssDNA Interactions in DNA Damage Repair

Authors: 
Marson, D., University of Trieste
Laurini, E., University of Trieste
Aulic, S., University of Trieste
Fermeglia, M., University of Trieste
Pricl, S., University of Trieste
DNA replication, repair, and recombination proteins form complex and agile networks. These networks organize the participating proteins into molecular machines that act on different substrates and channel them to different outcomes. Some of these machines display the capacity to accurately repair DNA damage or reestablish damage DNA replication forks without the loss of genetic information. Under other circumstances, action of the same molecular machines destabilizes the genome, leading to cancer or to the accumulation of toxic repair intermediates resulting in cell death. The central player in homologous recombination (HR) is the RAD51 DNA strand exchange protein (aka recombinase); its inactive conformation is tightly controlled by a double-heptameric doughnut-like assembly while its active form is a nucleoprotein filament assembled on the single-strand DNA generated at the site of DNA damage. In order to get a comprehensive structural and thermodynamic perspective of RAD51/ssDNA interaction, in this work we will present an application of large scale, parallel atomistic and coarse-grained (CG) simulations, coupled with high resolution imaging techniques and biophysical methods (ITC and CD) to i) dissect the docking mode of RAD51 to ssDNA, ii) deriving the energetics of the interface in the protein/nucleic acid interface, iii) uncovering the role of DNA and protein flexibility in their assembly formation, and iv) determining the mechanism of RAD51 binding to ssDNA.