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At least 217 records · Page 12Linked to original sources
PubMed1976

Suppression of an Escherichia coli dnaA mutation by the integrated R factor R.100.1: Change of chromosome replication origin in synchronized cultures.

We have followed, by deoxyribonucleic acid-deoxyribonucleic acid hybridization, the order of replication of three chromosomal markers during a synchronous round of replication in three strains of Escherichia coli carrying a dnaAts mutation: one strain in which the F-like R factor R.100.1 was established as a plasmid and two strains in which the dnaA mutation was suppressed by the integration of R.100.1 into the chromosome. In the R+ strain at 30C, replication of the plasmid took place simultaneously with the initiation of chromosome replication at the normal origin. In the integratively suppressed Hfr strains, at 42.5 C, chromosome replication was initiated preferentially from the integrated plasmid; little or no initiation occurred at the normal origin. Similar results were obtained for the one strain tested at 30 C. For both Hfr strains at 42.5 C, the data suggest that at least part of the population replicated bidirectionally. This conclusion had been confirmed using an autoradiographic procedure. Both types of experiment indicate a wide variation in the rate of travel of individual replication forks within the population.

Cell Division↗
PubMed1968

Effect of p-fluorophenylalanine on chromosome replication in Escherichia coli.

The effect of p-fluorophenylalanine (FPA) on deoxyribonucleic acid (DNA) synthesis and chromosome replication was studied in a thymine-requiring mutant of Escherichia coli. The rate and extent of chromosome replication were followed by labeling the DNA with isotopic thymine and a density marker, bromouracil. The DNA was extracted and analyzed by CsCl gradient centrifugation. The block in chromosome replication caused by high concentrations of FPA occurred at the same point on the chromosome as that caused by amino acid starvation. In a random culture, DNA in cells treated with FPA replicated only slightly slower than the DNA from cells that were not exposed to the analogue. In cultures which had been previously starved for thymine, however, the DNA from the cells treated with FPA showed a marked decrease in the rate and extent of replication. It was concluded that the E. coli cell is most sensitive to FPA when a new cycle of chromosome replication is being initiated at the beginning of the chromosome.

Amino Acids↗
PubMed1990

Escherichia coli molecular genetic map (1000 kbp): update I.

The sequenced genes from Escherichia coli that are available in the EMBL library (release 21) have been localized on an updated and corrected version of the restriction map of the chromosome generated by Kohara et al. (1987). One thousand kbp of sequenced DNA are incorporated in this update; this is equivalent to 23% of the total genome. The accuracy of the map is assessed, and it is corrected and updated where appropriate. A significant number of sites were missing from the original map, mainly involving two of the eight enzymes used by Kohara et al. (1987), ie. PvuII and EcoRV. The nucleotide environment of such missing sites was examined and, using an Artificial Intelligence approach, it appears that the site for these enzymes is sensitive to context effects. Several genes of known position on the E. coli chromosome could not be placed on the restriction map; this suggests that additional gaps are likely to exist on the restriction map, in addition to the original seven identified by Kohara et al. We have also obtained information about the probable direction of transcription of chromosomal genes with respect to the map. Most genes are transcribed in the same direction as the replication forks, particularly around oriC at 84 min.

Artificial Intelligence↗
PubMedDate not supplied

E. coli minichromosome replication: regulation of initiation at oriC.

The initiation of Escherichia coli DNA replication is a highly regulated event with many parameters exerting positive and negative effects. The activity of the dnaA protein (the initiator protein) is profoundly influenced by the tight binding of the adenine nucleotides ATP and ADP. Further regulation of dnaA protein activity may occur through dnaA protein-cell membrane associations. A replicatively inactive form of dnaA protein is found aggregated with phospholipids; enzymatic treatment of the aggregates with phospholipase A2 or dnaK protein liberates dnaA protein with restored replication activity. Proper DNA structure is essential for replication. The energy stored in the DNA's supercoiling is crucial for dnaA protein's ability to initiate replication. Under conditions where strand-opening by dnaA protein is inhibited, such as low free superhelicity, an R-loop formed by RNA polymerase activates the origin at a distance by aiding strand-opening. A novel protein has been identified as a specific inhibitor of the initiation of DNA replication. This 33-kDa protein binds to the AT rich region of oriC and inhibits strand-opening by dnaA protein.

Bacterial Proteins↗
PubMed1979

Membrane enrichment of genetic markers close to the origin and terminus during the deoxyribonucleic acid replication cycle in Bacillus subtilis.

A temperature-sensitive Bacillus subtilis initiation mutant was used to achieve one cycle of synchronized deoxyribonucleic acid (DNA) replication. Markers near the origin of replication and the terminus were assayed for association with the cell membrane at intervals during the DNA replication cycle. DNA near the origin and terminus was found to be enriched in the membrane fraction throughout the DNA replication cycle. The magnitude of membrane enrichment or origin and terminus markers varied coincidentally, possibly as a consequence of incubating the cells at 45 degrees C.

Bacillus subtilis↗
PubMed1987

OriC plasmids do not affect timing of chromosome replication in Escherichia coli K12.

The variability of the time interval between successive rounds of chromosome replication was estimated by density-shift experiments, by measuring the conversion of heavy DNA to hybrid density and light DNAs upon transfer of a steady-state culture growing in medium with [13C]glucose and 15NH4Cl to medium with light isotopes. The coefficient of variation (CV%) for the interreplication time of the Escherichia coli K12 chromosome was found to be 17%, i.e. similar to that for interdivision time. The presence of additional copies of oriC in the cell on a high copy number plasmid did not increase the CV of interreplication time. It is concluded that a single rate-limiting event is unlikely to time the initiation of chromosome replication. The regulation of initiation at oriC and the coordination with cell division is discussed.

Chromosomes, Bacterial↗
PubMed1987

Synthesis of linear multimers of OriC and pBR322 derivatives in Escherichia coli K-12: role of recombination and replication functions.

Inactivation of RecBCD nuclease (exonuclease V) and SbcB nuclease (exonuclease I) in Escherichia coli K-12 diverts most of plasmid replication activity from circular monomer production to the synthesis of linear multimers. Linear multimer synthesis has been demonstrated in plasmids of diverse origins and copy numbers, including E. coli minichromosomes. The effect of dnaA, dnaB, recF, and recJ mutations on the rate of linear multimer synthesis in sbcB cells after gam inactivation of RecBCD nuclease was investigated. Results are consistent with the hypothesis that homologous recombination, but not activities at the plasmid origin of replication, is involved in initiation of linear multimer synthesis.

Bacterial Proteins↗
PubMed1990

The E. coli cell surface specifically prevents the initiation of DNA replication at oriC on hemimethylated DNA templates.

A particular outer membrane fraction previously defined as possessing specific affinity for the hemimethylated form of the origin of replication of the E. coli chromosome (oriC) is shown to inhibit the initiation of DNA synthesis at this site on hemimethylated DNA templates in vitro. The replication of fully methylated or unmethylated DNA templates is not affected. Also, no inhibition is observed if initiation takes place at random sites on the hemimethylated template. The key inactivation step appears to be membrane inhibition of DnaA initiator protein binding to oriC. Remethylation of the membrane-bound hemimethylated DNA results in reactivation. Our results demonstrate direct involvement of the membrane in the control of DNA replication. We propose that association/dissociation of the origin from the cell membrane is one of the control elements governing interinitiation times in E. coli.

Bacterial Proteins↗
PubMed2000

Tn7 transposes proximal to DNA double-strand breaks and into regions where chromosomal DNA replication terminates.

We report that the bacterial transposon Tn7 can preferentially transpose into regions where chromosomal DNA replication terminates. DNA double-strand breaks are associated with the termination of chromosomal replication; therefore, we directly tested the effect of DNA breaks on Tn7 transposition. When DNA double-strand breaks are induced at specific sites in the chromosome, Tn7 transposition is stimulated and insertions are directed proximal to the induced break. The targeting preference for the terminus of replication and DNA double-strand breaks is dependent on the Tn7-encoded protein TnsE. The results presented in this study could also explain the previous observation that Tn7 is attracted to events associated with conjugal DNA replication during plasmid DNA transfer.

Bacterial Proteins↗
PubMed1970

Origin and direction of replication of the chromosome of E. coli B-r.

The origin and direction of replication of the E. coli B/r chromosome has been determined by comparing gene frequencies in P1-transducing lysates prepared on cultures growing at different rates. The gene frequencies found are consistent with the idea that replication of the chromosome is dichotomous in rapidly growing B/r. The origin was found to be between 40 and 55 min on the E. coli genetic map with replication proceeding in a clockwise direction. Markers near the origin behaved anomalously.

Chromosome Mapping↗
PubMed1972

Bidirectional replication of the chromosome in Escherichia coli.

E. coli cells initiating chromosome replication after release from amino-acid starvation were incubated in [(3)H]thymine of moderate specific activity, followed by incubation in [(3)H]thymine plus [(3)H]thymidine of very high specific activity. The grain tracks produced in autoradiographs of chromosomes were denser on both ends than in the middle. The autoradiographic patterns are, therefore, evidence that replication of the chromosome in E. coli is bidirectional.

Autoradiography↗
PubMed1981

Cell wall synthesis and initiation of deoxyribonucleic acid replication in Bacillus subtilis.

We have observed a connection between cell wall synthesis and the initiation of chromosome replication in Bacillus subtilis. Initiation of chromosome replication was prevented in synchronous cultures in the presence of the cell wall synthesis inhibitor vancomycin. When vancomycin was added to the cultures after initiation of chromosome replication, one round of replication was completed but no reinitiation occurred. Similar results were obtained when cell wall synthesis was inhibited by ristocetin, cycloserine, cloxacillin, or cephaloridine. When sucrose was added to the medium, initiation of deoxyribonucleic acid replication occurred in the presence of vancomycin, to an extent which allowed replication of no more than approximately one-half of the deoxyribonucleic acid of the culture. The same was found in cultures of spheroplasts of B. subtilis. However, initiation of chromosome replication in spheroplasts was completely insensitive to cloxacillin.

Bacillus subtilis↗
PubMed1978

Relationship of R6K replicating forms to the folded chromosome of Escherichia coli.

An examination of the relationship of both nonreplicating and replicating forms of R6K plasmid DNA to the Escherichia coli folded chromosome showed that both forms cosediment with the chromosome in neutral sucrose gradients. Approximately 20% of the nonreplicatin molecules was found as freely sedimenting forms when the folded-configuration of the chromosomes was preserved. However, under the same conditions negligible amounts of the replicating forms were found as freely sedimenting molecules. Thus, it is concluded that the replicating forms, when compared with nonreplicating molecules, are preferentially associated with the folded chromosomal structure. Exposure of the folded chromosomal structure to RNase resulted in an unfolding of the chromosome and a concomitant increase in the amount of freely sedimenting replicating and nonreplicating forms of R6K DNA. Analyses of the single-stranded length of RNase-released nascent molecules suggest that they replicate in continuous association with the folded chromsome complex. Nonenzymatic unfolding of the chromosomes by progressively lowering the sodium ion concentration during lysis resulted in a progressive increase in the release of nonreplicating molecules. Replicating molecules wer not released by unfolding the chromosome in this fashion.

Centrifugation, Density Gradient↗