Role of the ruvB gene in homologous and homeologous recombination in Rhizobium etli

The Rhizobium etli ruvA and ruvB genes were cloned through a PCR-based appr oach, using degenerate primers matching conserved sectors in the amino acid sequences of RuvB from eight bacterial species. Comparative analysis of th e predicted polypeptides for RuvA and RuvB of R. etli showed highly conserv ed blocks with the corresponding homologs in other bacteria; RuvB depicts c haracteristic motifs for DNA helicases (ATP-binding and DEXH-box motifs). A n R. etli ruvB::loxP Sp mutant was constructed by interposon mutagenesis. T his mutant was highly sensitive to DNA-damaging agents, such as methyl meth anesulfonate and nitrofurantoin, implying a deficiency in DNA repair. Homol ogous and homeologous conjugational recombination was reduced almost tenfol d in the ruvB::loxP Sp mutant; a recombination defect was also observed in assays employing recombination between small plasmids, albeit at a smaller magnitude. Although the ruvA and ruvB genes are contiguous in R. etli, comp lementation studies suggest that they are expressed independently. (C) 2000 Elsevier Science B.V. All rights reserved.