Cell 2008: Mre11 Dimers Coordinate DNA End Bridging and Nuclease Processing in Double-Strand-Break Repair .

R . Williams* , G . Moncalian* , J . Williams , Y . Yamada , O . Limbo , D . Shin , L . Groocock , D . Cahill , C . Hitomi , G . Guenther. (* these authors contribute equally)

Mre11 forms the core of the multifunctional Mre11-Rad50-Nbs1 (MRN) complex that detects DNA double-strand breaks (DSBs), activates the ATM checkpoint kinase, and initiates homologous recombination (HR) repair of DSBs. To define the roles of Mre11 in both DNA bridging and nucleolytic processing during initiation of DSB repair, we combined small-angle X-ray scattering (SAXS) and crystal structures of Pyrococcus furiosus Mre11 dimers bound to DNA with mutational analyses of fission yeast Mre11. The Mre11 dimer adopts a four-lobed U-shaped structure that is critical for proper MRN complex assembly and for binding and aligning DNA ends. Further, mutations blocking Mre11 endonuclease activity impair cell survival after DSB induction without compromising MRN complex assembly or Mre11-dependant recruitment of Ctp1, an HR factor, to DSBs. These results show how Mre11 dimerization and nuclease activities initiate repair of DSBs and collapsed replication forks, as well as provide a molecular foundation for understanding cancer-causing Mre11 mutations in ataxia telangiectasia-like disorder (ATLD). http://dx.doi.org/10.1016/j.cell.2008.08.017

Diario Médico

Europa press

Journal Bacteriology, 2008 ATPase activity and oligomeric state of TrwK, the VirB4 homologue of plasmid R388 Type IV secretion system.

Ignacio Arechaga, Alejandro Peña, Sandra Zunzunegui, María del Carmen Fernández-Alonso, Germán Rivas and Fernando de la Cruz.

Type IV secretion systems (T4SS) mediate the transfer of DNA and protein substrates to target cells. TrwK, encoded by the conjugative plasmid R388, is a member of the VirB4 family, the largest and most conserved protein of T4SS. VirB4 was suggested to be an ATPase involved in energizing pilus assembly and substrate transport. However, conflicting experimental evidence concerning VirB4 ATP hydrolase activity was reported. Here, we demonstrate that TrwK is able to hydrolyze ATP in vitro in the absence of its potential macromolecular substrates and other T4SS components. The kinetic parameters of its ATPase activity have been characterized. TrwK oligomerization state was investigated by analytical ultracentrifugation and electron microscopy, and its effects on the ATPase activity were analyzed. The results suggest that the hexameric form of TrwK is the catalytically active state, much like the structurally related protein TrwB, the conjugative coupling protein. http://dx.doi.org/10.1128/JB.00321-08