The apparent DNA site size obtained from an assay monitoring the ATPase activity of Escherichia coli recA protein (n = 3.5) differs from that determined from a direct DNA binding assay (n = 7) done under identical conditions. Investigation of this discrepancy indicates that at a DNA:protein ratio of 3.5:1, one-half of the recA protein population is less sensitive to ATPase activity inhibition b...

Genetic recombination in Escherichia coli requires recA protein, the product of the recA+ gene. In this paper we show that purified recA protein, which binds strongly to denatured DNA, cooperatively recognizes DNA containing short single-stranded regions. The interaction of varying amounts of recA protein with DNA molecules was investigated by measuring its DNA-dependent ATPase activity. In 3mM...

The mutation of Pro67 to Trp (P67W) in the Escherichia coli RecA protein results in reduced recombination and constitutive coprotease phenotypes. We examined the biochemical properties of this mutant in an effort to understand these altered behaviors. We find that RecA P67W protein can access single-stranded DNA (ssDNA) binding sites within regions of secondary structure more effectively than w...

Mutations in recA, such as recA801(Srf) (suppressor of RecF) or recA441(Tif) (temperature-induced filamentation) partially suppress the deficiency in postreplication repair of UV damage conferred by recF mutations. We observed that spontaneous recA(Srf) mutants accumulated in cultures of recB recC sbcB sulA::Mu dX(Ap lac) lexA51 recF cells because they grew faster than the parental strain. We s...

In all organisms, RecA-family recombinases catalyze homologous joint formation in homologous genetic recombination, which is essential for genome stability and diversification. In homologous joint formation, ATP-bound RecA/Rad51-recombinases first bind single-stranded DNA at its primary site and then interact with double-stranded DNA at another site. The underlying reason and the regulatory mec...

We have investigated the structural, biochemical and cellular roles of the two single-stranded (ss) DNA-binding proteins from Bacillus subtilis, SsbA and SsbB. During transformation, SsbB localizes at the DNA entry pole where it binds and protects internalized ssDNA. The 2.8-Å resolution structure of SsbB bound to ssDNA reveals a similar overall protein architecture and ssDNA-binding surface to...

The recA gene of Synechococcus sp. strain PCC 7002 was detected and cloned from a lambda gtwes genomic library by heterologous hybridization by using a gene-internal fragment of the Escherichia coli recA gene as the probe. The gene encodes a 38-kilodalton polypeptide which is antigenically related to the RecA protein of E. coli. The nucleotide sequence of a portion of the gene was determined. T...

Photochemical cross-linking has been used to identify residues in the Escherichia coli RecA protein that are proximal to and may directly mediate binding of DNA. Ultraviolet irradiation promotes specific and efficient cross-linking of the RecA protein to poly(deoxythymidylic) acid. Cross-linked peptides remaining covalently attached to the polynucleotide following proteolytic digestion with try...

The renaturation of complementary DNA strands by the recA protein of Escherichia coli has been found to exhibit the following features. (i) Optimal renaturation occurs at recA protein levels below that required to saturate the DNA strands; saturating amounts of recA protein significantly reduce the rate of reaction. (ii) The reaction proceeds in the absence of a nucleotide cofactor but is marke...

OBJECTIVES In eubacteria, RecA is essential for recombinational DNA repair and for stalled replication forks to resume DNA synthesis. Recent work has implicated a role for RecA in the development of antibiotic resistance in pathogenic bacteria. Consequently, our goal is to identify and characterize small-molecule inhibitors that target RecA both in vitro and in vivo. METHODS We employed ATPas...