Genetic analysis of an archaeal Holliday junction resolvase in Escherichiacoli

The study of genes and proteins in heterologous model systems provides a po werful approach to the analysis of common processes in biology. Here, we sh ow how the bacterium Escherichia coil can be exploited to analyse genetical ly and biochemically the activity and function of a Holliday junction resol ving enzyme from an archaeal species. We have purified and characterised a member of the newly discovered Holliday junction cleaving (Hjc) family of r esolvases from the moderately thermophilic archaeon Methanobacterium thermo autotrophicum and demonstrate that it promotes DNA repair in resolvase-defi cient ruv mutants of E. coil. The data presented provide the first direct e vidence that such archaeal enzymes can promote DNA repair in vivo, and supp ort the view that formation and resolution of Holliday junctions are key to the interplay between DNA replication, recombination and repair in all org anisms. We also show that Hjc promotes DNA repair in E. coil in a manner th at requires the presence of the RecG branch migration protein. These result s support models in which RecG acts at a replication fork stalled at a lesi on in the DNA, catalysing fork regression and forming a Holliday junction t hat can then be acted upon by Hjc. (C) 2001 Academic Press.