recA Sequences Research Articles

Molecular cloning and nucleotide sequence of the Rhizobium phaseoli recA gene

A recombinant lambda phage carrying the recA gene of Rhizobium phaseoli was isolated from a R. phaseoli genomic library by complementation of the Fec- phenotype of the recombinant phage in Escherichia coli. When expressed in E. coli, the cloned recA gene was shown to restore resistance to both UV irradiation and the DNA alkylating agent methyl methanesulphonate (MMS). The R. phaseoli recA gene also promoted homologous recombination in E. coli. The cloned recA gene was only weakly inducible in E. coli recA strains by DNA damaging agents. The DNA sequence of the R. phaseoli recA gene was determined and compared with published recA sequences. No LexA-binding site was detected in the R. phaseoli recA upstream region.

Differences in mutagenic and recombinational DNA repair in enterobacteria.

The incidence of recombinational DNA repair and inducible mutagenic DNA repair has been examined in Escherichia coli and 11 related species of enterobacteria. Recombinational repair was found to be a common feature of the DNA repair repertoire of at least 6 genera of enterobacteria. This conclusion is based on observations of (i) damage-induced synthesis of RecA-like proteins, (ii) nucleotide hybridization between E. coli recA sequences and some chromosomal DNAs, and (iii) recA-negative complementation by plasmids showing SOS-inducible expression of truncated E. coli recA genes. The mechanism of DNA damage-induced gene expression is therefore sufficiently conserved to allow non-E. coli regulatory elements to govern expression of these cloned truncated E. coli recA genes. In contrast, the process of mutagenic repair, which uses umuC+ umuD+ gene products in E. coli, appeared less widespread. Little ultraviolet light-induced mutagenesis to rifampicin resistance was detected outside the genus Escherichia, and even within the genus induced mutagenesis was detected in only 3 out of 6 species. Nucleotide hybridization showed that sequences like the E. coli umuCD+ gene are not found in these poorly mutable organisms. Evolutionary questions raised by the sporadic incidence of inducible mutagenic repair are discussed.

Purified lexA protein is a repressor of the recA and lexA genes.

Escherichia coli shows a pleiotropic response (the SOS response) to treatments that damage DNA or inhibit DNA replication. Previous evidence has suggested that the product of the lexA gene is involved in regulating the SOS response, perhaps as a repressor, and that it is sensitive to the recA protease. We show here that lexA protein is a repressor of at least two genes, recA and lexA. Purified protein bound specifically to the regulatory regions of the two genes, as judged by DNase I protection experiments, and it specifically inhibited in vitro transcription of both genes. The binding sites in recA and lexA were found to be about 20 base pairs (bp) and 40 bp long, respectively. The 40-bp sequence in lexA was composed of two adjacent 20-bp sequences, which had considerable homology to one another and to the corresponding recA sequence. These 20-bp sequences, which we term "SOS boxes," show considerable inverted repeat structure as well. These features suggest that each box represents a single repressor binding site. Finally, we found that purified lexA protein was a substrate for the recA protease in a reaction requiring ATP or an analogue, adenosine 5'-[gamma-thio]triphosphate, and denatured DNA.