The recA gene of Borrelia burgdorferi

Dew-Jager, K; Yu, Weng-Qing; Huang, Wai Mun

doi:10.1016/0378-1119(95)00646-x

Cited by 5 publications

(5 citation statements)

References 13 publications

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“…Interestingly, B. burgdorferi RecA contains both K39 and K174 in the wild-type sequence. These naturally occurring amino acid changes could explain the enhanced coprotease activity observed, as first suggested by Dew-Jager et al (10).…”

Section: Discussionsupporting

confidence: 51%

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Functional Properties ofBorrelia burgdorferi recA

Liveris¹,

Mulay²,

Schwartz

2004

J Bacteriol

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Functions of the Borrelia burgdorferi RecA protein were investigated in Escherichia coli recA null mutants. Complementation with B. burgdorferi recA increased survival of E. coli recA mutants by 3 orders of magnitude at a UV dose of 2,000 J/cm 2 . The viability at this UV dose was about 10% that provided by the homologous recA gene. Expression of B. burgdorferi recA resulted in survival of E. coli at levels of mitomycin C that were lethal to noncomplemented hosts. B. burgdorferi RecA was as effective as E. coli RecA in mediating homologous recombination in E. coli. Furthermore, E. coli phage lysogens complemented with B. burgdorferi recA produced phage even in the absence of UV irradiation. The level of phage induction was 55-fold higher than the level in cells complemented with the homologous recA gene, suggesting that B. burgdorferi RecA may possess an enhanced coprotease activity. This study indicates that B. burgdorferi RecA mediates the same functions in E. coli as the homologous E. coli protein mediates. However, the rapid loss of viability and the absence of induction in recA expression after UV irradiation in B. burgdorferi suggest that recA is not involved in the repair of UV-induced damage in B. burgdorferi. The primary role of RecA in B. burgdorferi is likely to be a role in some aspect of recombination.The spirochete Borrelia burgdorferi, the causative agent of Lyme disease, has a unique genomic organization consisting of a 910,725-bp linear chromosome and at least 21 linear and circular plasmids (6, 13). Less than one-half of the 1,689 annotated open reading frames have an identifiable database match (6, 13). In addition, very few genes have been characterized biochemically. Functional studies of B. burgdorferi genes in the postgenomic era have been hampered by this spirochete's slow growth in culture and by the limited availability of genetic tools. A potentially useful strategy for circumventing these difficulties is complementation of mutants in well-characterized bacteria, such as Escherichia coli. This strategy has been employed to elucidate the functions of a number of B. burgdorferi gene products (1,5,7,8,15,20,26,32,44,49,50).Genetic recombination may play a role in B. burgdorferi persistence by generation of antigenic variation (2,31,37,45,47,52,56,57). RecA is the key protein linking genetic recombination to DNA replication and repair in bacteria (23,24). In E. coli, RecA promotes pairing and strand exchange between a single-stranded DNA molecule and a homologous doublestranded DNA molecule through a complex multistep pathway that requires a DNA-dependent ATPase function (29). In addition to this recombinase activity, an ATP-dependent coprotease activity has also been ascribed to RecA. In enterobacteria and Bacillus sp., the latter activity is part of a specialized and regulated form of DNA repair designated SOS repair (3). In E. coli, the SOS response involves the transcriptional repressor LexA, whose autocatalytic cleavage is enhanced by activated RecA following DNA damage (19). This res...

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Section: Discussionsupporting

confidence: 51%

“…The B. burgdorferi recA gene was originally described from isolate Sh-2-82 by Dew-Jager et al (10). An orthologue of this gene from type strain B31MI has been annotated in the B. burgdorferi genome as BB0131 (13).…”

mentioning

confidence: 99%

Functional Properties ofBorrelia burgdorferi recA

Liveris¹,

Mulay²,

Schwartz

2004

J Bacteriol

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“…PCR-based measurement of B. burgdorferi in tissues requires that the starting copy of the bacterial DNA target sequence accurately indicate the number of bacteria within a sample. For this study, the recA gene was selected (6). The recA gene is located on the chromosome and therefore should be present in every bacterium (8).…”

Section: Resultsmentioning

confidence: 99%

Rapid and Sensitive Quantification of Borrelia burgdorferi -Infected Mouse Tissues by Continuous Fluorescent Monitoring of PCR

et al. 1999

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The quantity of Borrelia burgdorferi organisms in tissue samples is an important determinant for infection studies in the mouse model of Lyme disease. This report presents the development of a rapid and sensitive external-standard-based PCR assay for the absolute quantification of B. burgdorferi in mouse tissue samples. The assay uses a double-stranded DNA dye to continuously monitor product formation and in less than an hour was able to quantify samples ranging up to 6 log units in concentration. The PCR efficiencies of the sample and the standard were matched by using a standard composed of purified B. burgdorferi chromosome mixed with tissue-matched mouse genome lacking bacterial DNA. Normalization ofB. burgdorferi quantities to the mouse nidogengene allowed comparison of B. burgdorferi numbers in samples isolated from different tissues and strains. PCR analysis of the chromosomal gene recA in cultured B. burgdorferi was consistent with a single recA per bacterium. The parameters defined in this assay should be applicable to quantification of other organisms, even infectious agents for which no ready source of DNA standard is available. In summary, this report presents a rapid external-standard-based PCR method for the quantification of B. burgdorferi in mouse DNA samples.

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“…In that study, immunoprecipitation of cellular extracts of B. burgdorferi and T. pallidum with rabbit antiserum to E. coli RecA failed to detect RecA protein in these spirochetes. A recA gene of B. burgdorferi was later found by Dew-Jager et al, who showed that the deduced sequence of B. burgdorferi's RecA protein had several substitutions, which, when individually present in E. coli RecA, altered or eliminated the function in that organism (11).…”

Section: Discussionmentioning

confidence: 95%

“…The RecA protein of B. burgdorferi was identified previously and was shown to have many of the conserved motifs of other bacterial RecA proteins (11). To assess the role of RecA in the frequency of recombination at the VlsE locus in B. burgdorferi (54), we attempted to produce a knockout of the recA gene of this species by insertional inactivation (4).…”

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confidence: 99%

Transgenic Expression of RecA of the SpirochetesBorrelia burgdorferiandBorrelia hermsiiinEscherichia coliRevealed Differences in DNA Repair and Recombination Phenotypes

2004

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After unsuccessful attempts to recover a viable RecA-deficient mutant of the Lyme borreliosis agent Borrelia burgdorferi, we characterized the functional activities of RecA of B. burgdorferi, as well as RecA of the relapsing fever spirochete Borrelia hermsii and the free-living spirochete Leptospira biflexa, in a recA mutant of Escherichia coli. As a control, E. coli RecA was expressed from the same plasmid vector. DNA damage repair activity was assessed after exposure of the transgenic cells to UV light or the radiomimetic chemicals methyl methanesulfonate and mitomycin C. Recombination activity in the cells was assessed by using an assay for homologous recombination between repeats in the chromosome and by measuring the ability of the cells to foster lytic growth by red gam mutant bacteriophage . Overall, we found that transgenic cells with recA genes of B. burgdorferi, B. hermsii, and L. biflexa had approximately equivalent activities in promoting homologous recombination in the lacZ duplication assay, but cells with B. burgdorferi recA and, most notably, B. hermsii recA were significantly less capable than cells with L. biflexa recA or E. coli recA in responding to DNA damage or in facilitating plaque formation in the phage assay. The comparatively poor function of Borrelia recA in the latter set of assays may be the consequence of impaired coordination in the loading of the transgenic RecA by RecBCD and/or RecFOR in E. coli.

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The recA gene of Borrelia burgdorferi

Cited by 5 publications

References 13 publications

Functional Properties ofBorrelia burgdorferi recA

Functional Properties ofBorrelia burgdorferi recA

Rapid and Sensitive Quantification of Borrelia burgdorferi -Infected Mouse Tissues by Continuous Fluorescent Monitoring of PCR

Transgenic Expression of RecA of the SpirochetesBorrelia burgdorferiandBorrelia hermsiiinEscherichia coliRevealed Differences in DNA Repair and Recombination Phenotypes

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