Mre11 and Rad50 from Pyrococcus furiosus: Cloning and biochemical characterization reveal an evolutionarily conserved multiprotein machine

The processing of DNA double-strand breaks is a critical event in nucleic a cid metabolism. This is evidenced by the severity of phenotypes associated with deficiencies in this process in multiple organisms. The core component involved in double-strand break repair in eukaryotic cells is the Mre11-Ra d50 protein complex, which includes a third protein, p95, in humans and Xrs 2 in yeasts. Homologues of Mre11 and Rad50 have been identified in all king doms of life, while the Nbs1 protein family is found only in eukaryotes. In eukaryotes the Mre11-Rad50 complex has nuclease activity that is modulated by the addition of ATP. We have isolated the Mre11 and Rad50 homologues fr om the thermophilic archaeon Pyrococcus furiosus and demonstrate that the t wo proteins exist in a large, heat-stable complex that possesses single-str and endonuclease activity and ATP-dependent double-strand-specific exonucle ase activity. These findings verify the identification of the P. furiosus R ad50 and Mre11 homologues and demonstrate that functional homologues with s imilar biochemical properties exist in all kingdoms of life.