Cloning of the recA gene from a free-living leptospire and distribution of RecA-like protein among spirochetes

Abstract

A recombinant plasmid carrying the recA gene of Leptospira bfifexa serovar patoc was isolated from a cosmid library of genomic DNA by complementation of an Escherichia coli recA mutation. The cloned serovar patoc recA gene efficiently restored resistance to UV radiation and methyl methanesulfonate. Recombination proficiency was also restored, as measured by the formation of Lac' recombinants from duplicated mutant lacZ genes. Additionally, the cloned recA gene increased the spontaneous and mitomycin C-induced production of lambda phage in lysogens of an E. coli recA mutant. The product of the cloned recA gene was identified in maxicells as a polypeptide with an Mr of 43,000. Antibodies prepared against the E. coli RecA protein cross-reacted with the serovar patoc RecA protein, indicating structural conservation. Southern hybridization data showed that the serovar patoc recA gene has diverged from the recA gene of L. interrogans, Leptonema illini, and E. coli. With the exception of the RecA protein of L. interrogans serovar hardjo, the RecA protein of the Leptospira serovars and L. illini were synthesized at elevated levels following treatment of cells with nalidixic acid. The level of detectable RecA correlated with previous studies demonstrating that free-living cells of L. biflexa serovars and L. illini were considerably more resistant to DNA-damaging agents than were those of parasitic L. interrogans serovars. RecA protein was not detected in cells of virulent Treponema pallidum or Borrelia burgdorferi.

First evidence for gene replacement in Leptospira spp. Inactivation of L. biflexa flaB results in non‐motile mutants deficient in endoflagella

“…It should be noted that repair-deficient mutants, such as uvrA and recA mutants, of E. coli and B. subtilis are dramatically more sensitive to UV than the wild-type strains (Howard-Flanders, 1968;Friedman and Yasbin, 1983). In addition, the recA gene was identified previously in L. biflexa and was found to complement recA mutations of E. coli functionally (Stamm et al, 1991). Taken together, these results indicated that L. biflexa may possess an efficient DNA repair system.…”

Polypeptides of Treponema pallidum: progress toward understanding their structural, functional, and immunologic roles. Treponema Pallidum Polypeptide Research Group.

“…Stamm et al (200) were unable to detect the recombination-mediating protein RecA in T. pallidum and B. burgdorfeni by immunoprecipitation with antibodies against E. coli RecA, although the same antiserum detected RecA in Leptospira interrogans. Further studies are needed to substantiate this unusual finding.…”

Laboratory diagnosis and interpretation of tests for syphilis

“…Whereas variation in membrane and surface-associated proteins is common among many species of bacteria, this variation has never been shown with T. pallidum; even the different subspecies show essentially identical patterns by one-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis (196). This extreme genetic stability is probably related to the apparent lack of significant DNA repair capacity in T. pallidum (193) and the probable absence of the recA gene product (192), which would indicate little or no recombinational capacity in this spirochete.…”