Structural mechanism of ATP-dependent DNA binding and DNA end bridging by eukaryotic Rad50

Seifert, Florian Ulrich; Lammens, Katja; Stoehr, Gabriele; Kessler, Brigitte and Hopfner, Karl-Peter (2016): Structural mechanism of ATP-dependent DNA binding and DNA end bridging by eukaryotic Rad50. In: Embo Journal, Vol. 35, No. 7: pp. 759-772

Abstract

The Mre11-Rad50-Nbs1 (MRN) complex is a central factor in the repair of DNA double-strand breaks (DSBs). The ATP-dependent mechanisms of how MRN detects and endonucleolytically processes DNA ends for the repair by microhomology-mediated end-joining or further resection in homologous recombination are still unclear. Here, we report the crystal structures of the ATPS-bound dimer of the Rad50(NBD) (nucleotide-binding domain) from the thermophilic eukaryote Chaetomium thermophilum (Ct) in complex with either DNA or CtMre11(RBD) (Rad50-binding domain) along with small-angle X-ray scattering and cross-linking studies. The structure and DNA binding motifs were validated by DNA binding experiments invitro and mutational analyses in Saccharomyces cerevisiae invivo. Our analyses provide a structural framework for the architecture of the eukaryotic Mre11-Rad50 complex. They show that a Rad50 dimer binds approximately 18 base pairs of DNA along the dimer interface in an ATP-dependent fashion or bridges twoDNA ends with a preference for 3' overhangs. Finally, our results may provide a general framework for the interaction of ABC ATPase domains of the Rad50/SMC/RecN protein family with DNA.

• Item Type: Journal article
• Faculties: Chemistry and Pharmacy > Department of Biochemistry
• Subjects: 500 Science > 540 Chemistry
• ISSN: 0261-4189
• Language: English