Learn More
Since most archaea are extremophilic and difficult to cultivate, our current knowledge of their biology is confined largely to comparative genomics and biochemistry. Haloferax volcanii offers great promise as a model organism for archaeal genetics, but until now there has been a lack of a wide variety of selectable markers for this organism. We describe(More)
ruvC mutants of Escherichia coli appear to lack an activity that resolves Holliday intermediates into recombinant products. Yet, these strains produce close to normal numbers of recombinants in genetic crosses. This recombination proficiency was found to be a function of recG. A "mini-kan" insertion in recG was introduced into ruvA, ruvB, and ruvC strains.(More)
Chromosome duplication normally initiates through the assembly of replication fork complexes at defined origins. DNA synthesis by any one fork is thought to cease when it meets another travelling in the opposite direction, at which stage the replication machinery may simply dissociate before the nascent strands are finally ligated. But what actually happens(More)
The RuvA and RuvB proteins of Escherichia coli play important roles in the post-replicational repair of damaged DNA, genetic recombination and cell division. In this paper, we describe the construction of over expression vectors for RuvA and RuvB and detail simple purification schemes for each protein. The purified 22 kDa RuvA polypeptide forms a tetrameric(More)
Faithful duplication of the genome relies on the ability to cope with an imperfect template. We investigated replication of UV-damaged DNA in Escherichia coli and found that ongoing replication stops for at least 15-20 min before resuming. Undamaged origins of replication (oriC) continue to fire at the normal rate and in a DnaA-dependent manner. UV(More)
The RuvAB and RuvC enzymes of Escherichia coli define a molecular pathway for the resolution of Holliday intermediates in recombination and DNA repair. They bind specifically to Holliday junctions, and catalyse their branch migration and cleavage, respectively. In a RuvA(B)-junction complex, the Holliday structure is held in an open (square planar)(More)
The RecG protein of Escherichia coli is needed for normal levels of recombination and for repair of DNA damaged by ultraviolet light, mitomycin C and ionizing radiation. The true extent of its involvement in these processes is masked to a large degree by what appears to be a functional overlap with the products of the three ruv genes. RuvA and RuvB act(More)
The RuvAB, RuvC and RecG proteins of Escherichia coli process intermediates in recombination and DNA repair into mature products. RuvAB and RecG catalyse branch migration of Holliday junctions, while RuvC resolves these structures by nuclease cleavage around the point of strand exchange. The overlap between RuvAB and RecG was investigated using synthetic X-(More)
In previous studies, Holliday junctions generated during RecA-mediated strand-exchange reactions were resolved by fractionated Escherichia coli extracts. We now report the specific binding and cleavage of synthetic Holliday junctions (50 base pairs long) by a fraction purified by chromatography on DEAE-cellulose, phosphocellulose, and single-stranded(More)
Organisms rely on close interplay between DNA replication, recombination, and repair to secure transmission of the genome. In rapidly dividing cells, there is also great pressure for transcription, which may induce conflict with replication. We investigated the potential for conflict in bacterial cells, where there is no temporal separation of these(More)