Cloning and expression in Escherichia coli of a recA-like gene from Vibrio cholerae

Paul, Kalidas; Ghosh, Subrata; Das, Jyotirmoy

doi:10.1007/bf00330384

Cited by 19 publications

(7 citation statements)

References 38 publications

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“…We also concluded that homologous-dependent recombination proficiency was restored, since the cloned recA gene efficiently promoted recombination between two mutant lacZ genes, resulting in the production of Lac' papillae. These data are in agreement with those of Keener et al (15) and others (3,5,7,9,12,16,17,19,(21)(22)(23)25), who have shown that cloned, heterologous recA genes are capable of functionally complementing various E. coli recA mutations.…”

Section: Discussionsupporting

confidence: 93%

“…We have identified the product of the recA gene of serovar patoc as a polypeptide with an Mr of 43,000 by maxicell analysis. This Mr is similar to those reported for the RecA proteins of several other bacterial species (5,9,12,15,17,19,(21)(22)(23)25). Anti-E. coli RecA serum efficiently precip-itated the serovar patoc RecA protein, providing evidence of the structural conservation of this gene product with the RecA protein of E. coli.…”

Section: Discussionsupporting

confidence: 88%

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

Stamm

Parrish

Gherardini

1991

Appl Environ Microbiol

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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.

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Section: Discussionsupporting

confidence: 93%

Section: Discussionsupporting

confidence: 88%

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

Stamm

Parrish

Gherardini

1991

Appl Environ Microbiol

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“…The labeled cells were harvested, washed, suspended in electrophoresis sample buffer and analyzed by SDS-polyacrylamide (10%) gel electrophoresis followed by autoradiography of dried gels as described previously (30). DNA RESULTS AND DISCUSSION mutS and mutL genes of V. cholerae Genomic libraries of V. cholerae DNA were constructed by cloning either EcoRI or Pstl restriction fragments into the plasmid pUC8 as described previously (21). These libraries were maintained in E. coli strain JM101 and used to search for the mut genes of V. cholerae.…”

Section: Protein Labelling In Maxicellsmentioning

confidence: 99%

“…MutL and mutS gene products are also involved in very short patch repair (16,17). Vibrio cholerae, a highly pathogenic gram negative bacterium and the causative agent of cholera, is inefficient in repairing UV-induced DNA damage (18)(19)(20)(21)(22). The spontaneous mutation frequency for any given marker, however, is comparable to that of other organisms.…”

Section: Introductionmentioning

confidence: 99%

Cloning and characterization ofmutLandmutSgenes ofVibrio cholerae: nucleotide sequence of themutLgene

Bera¹,

Ghosh²,

Das³

1989

Nucl Acids Res

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The mutL and mutS genes of Vibrio cholerae have been identified using interspecific complementation of Escherichia coli mutL and mutS mutants with plasmids containing the gene bank of V. cholerae. The recombinant plasmid pJT470, containing a 4.7 kb fragment of V. cholerae DNA codes for a protein of molecular weight 92,000. The product of this gene reduces the spontaneous mutation frequency of the E. coli mutS mutant. The plasmid, designated pJT250, containing a 2.5 kb DNA fragment of V. cholerae and coding for a protein of molecular weight 62,000, complements the mutL gene function of E. coli mutL mutants. These gene products are involved in the repair of mismatches in DNA. The complete nucleotide sequence of mutL gene of V. cholerae has been determined.

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“…Analysis of DNA restriction fragments was done with agarose (0.8%) horizontal slab gels or polyacrylamide vertical slab gels as described previously (28). For electrophoresis of RNA, formaldehyde-agarose gels were used (22).…”

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confidence: 99%

A 14-kilodalton inner membrane protein of Vibrio cholerae biotype e1 tor confers resistance to group IV choleraphage infection to classical vibrios

1992

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Choleraphage +149 differentiates the two biotypes, classical and el tor, of Vibrio cholerae. This phage cannot replicate in V. cholerae biotype el tor cells because the concatemeric DNA intermediates produced are unstable and cannot be chased to mature phage DNA. A V. cholerae biotype el tor gene coding for a 14,000-Da inner membrane protein which destabilizes the concatemeric DNA intermediates by hindering their binding to the cell membrane has been identified. Presumably, a 22,000-Da V. cholerae biotype el tor protein might also have a role in conferring phage +149 resistance to cells belonging to the biotype el tor. A nucleotide sequence homologous to the 1.2-kb V. cholerae biotype el tor DNA coding for both the 14,000-and 22,000-Da proteins is present in all strains of classical vibrios but is not transcribed. The nucleotide sequence of the gene coding for the 14,000-Da protein has been determined.Vibrio cholerae strains of serotype 01 comprise two different biotypes, classical and el tor. Of the seven pandemics of cholera recorded in recent times, V. cholerae biotype el tor has been identified as the causative agent of the seventh pandemic, while the previous ones were due to the classical strains. Recently, there has been a resurgence of classical vibrios in certain areas of endemicity (33). In spite of broad similarities in the nature of infection by the classical and el tor vibrios, important differences exist in the epidemiology of el tor compared with that of classical vibrios. The ratio of asymptomatic infection to clinical illness is greater with the el tor biotype than with the classical vibrios (2), and the el tor vibrios survive longer under unfavorable conditions. Both the classical and the el tor strains of V. cholerae react with V. cholerae antisera, and taxonomic studies have shown that these two biotypes are the same species (12). They are isogenic, and tests to distinguish them often give ambiguous results (16). One of the most reliable criteria differentiating between these two biotypes is their susceptibility to group IV choleraphages. This group of choleraphages can infect and lyse all strains of classical vibrios but none of the el tor biotype (24). Phage 4149 adsorbs irreversibly to the biotype el tor cells, and 50% of the injected phage DNA binds to the cell membrane. Synthesis of monomeric phage DNA continued, similar to that observed for the permissive host. However, the concatemeric DNA intermediates produced were unstable and could not be chased to mature phage DNA (9). Although most of the early proteins are made, only some of the late proteins were transiently synthesized for infection in el tor cells. Formation and stabilization of concatemeric DNA structures essential for the synthesis of mature phage DNA and their packaging into phage heads are not hindered in V. cholerae biotype el tor cells for the replication of choleraphage 4138 belonging to serological group II. This phage also contains a linear double-stranded circularly permuted DNA molecule and can replicate in both V....

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Cloning and expression in Escherichia coli of a recA-like gene from Vibrio cholerae

Cited by 19 publications

References 38 publications

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

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

Cloning and characterization ofmutLandmutSgenes ofVibrio cholerae: nucleotide sequence of themutLgene

A 14-kilodalton inner membrane protein of Vibrio cholerae biotype e1 tor confers resistance to group IV choleraphage infection to classical vibrios

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