Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “Bacterial DNA replication”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 811 records · Page 45Linked to original sources
PubMed1992

Cloning of an autonomously replicating sequence (ars) from the Bacillus subtilis chromosome.

Cloning of an autonomously replicating sequence (ars) from the origin region of Bacillus subtilis was previously unsuccessful because of the strong incompatibility exerted by sequences located within the oriC region. Using an ars searching vector which would be selective for drug resistance even at one copy per cell, and by cloning large fragments covering as much as possible of the oriC region, we have succeeded in isolating ars fragments from the origin region of the chromosome. The minimum essential fragment contains two DnaA-box regions (non-translatable regions containing multiple repeats of DnaA-box) separated by the dnaA gene. Neither one of the DnaA-box regions by itself showed ars activity. When constructed as oriC plasmids, the dnaA coding region could be removed without affecting ars activity. The minimum distance between the two DnaA-box regions obtained so far is 274 bp. The copy number of the oriC plasmid is estimated as one per replicating chromosome. These plasmids are unstable and tend to be lost or integrated into chromosome.

Amino Acid Sequence↗
PubMed2006

Systematic localisation of proteins fused to the green fluorescent protein in Bacillus subtilis: identification of new proteins at the DNA replication factory.

Construction and microscopic imaging of protein fusions to green fluorescent protein (GFP) have revolutionised our understanding of bacterial structure and function. We have undertaken a systematic study of the localisation of over 100 Bacillus subtilis proteins, following the development of high-throughput construction and analysis procedures. We focused on proteins linked in various ways to the DNA replication machinery, as well as on proteins exemplifying a range of other cellular functions and structures. The results validate the approach as a way of obtaining systematic protein localisation information. They also provide a range of novel biological insights, particularly through the identification of a number of proteins not previously known to be associated with the DNA replication factory.

Bacillus subtilis↗
PubMed2005

A study on the correlation of nucleotide skews and the positioning of the origin of replication: different modes of replication in bacterial species.

Deviations from Chargaff's 2nd parity rule, according to which A approximately T and G approximately C in single stranded DNA, have been associated with replication as well as with transcription in prokaryotes. Based on observations regarding mainly the transcription-replication co-linearity in a large number of prokaryotic species, we formulate the hypothesis that the replication procedure may follow different modes between genomes throughout which the skews clearly follow different patterns. We draw the conclusion that multiple functional sites of origin of replication may exist in the genomes of most archaea and in some exceptional cases of eubacteria, while in the majority of eubacteria, replication occurs through a single fixed origin.

Archaea↗
PubMed1996

The localized melting of mini-F origin by the combined action of the mini-F initiator protein (RepE) and HU and DnaA of Escherichia coli.

Replication of mini-F plasmids requires the initiator protein RepE, which binds specifically to four iterons within the origin (ori2), as well as some host factors that are involved in chromosomal DNA replication. To understand the role of host factors and RepE in the early steps of mini-F DNA replication, we examined the effects of RepE and the Escherichia coli proteins DnaA and HU on the localized melting of ori2 DNA in a purified in vitro system. We found that the binding of RepE to an iteron causes a 50 degrees bend at or around the site of binding. RepE and HU exhibited synergistic effects on the localized melting within the ori2 region, as detected by sensitivity to the single-strand specific P1 endonuclease. This opening of duplex DNA occurred around the 13mer of ori2, whose sequence closely resembles the set of 13mers found in the chromosomal origin oriC. Further addition of DnaA to the reaction mixture increased the efficiency of melting and appeared to extend melting to the adjacent AT-rich region. Moreover, DNA melting with appreciably higher efficiencies was observed with mutant forms of RepE that were previously shown to be hyperactive both in DNA binding in vitro and in initiator activity in vivo. We propose that the binding of RepE to four iterons of ori2 causes bending at the sites of RepE binding and, with the assistance of HU, induces a localized melting in the 13mer region. The addition of DnaA extends melting to the AT-rich region, which could then serve as the entry site for the DnaB-DnaA complex, much as has been documented for oriC dependent replication.

Bacterial Proteins↗
PubMed1998

Bacillus subtilis cell cycle as studied by fluorescence microscopy: constancy of cell length at initiation of DNA replication and evidence for active nucleoid partitioning.

Fluorescence microscopic methods have been used to characterize the cell cycle of Bacillus subtilis at four different growth rates. The data obtained have been used to derive models for cell cycle progression. Like that of Escherichia coli, the period required by B. subtilis for chromosome replication at 37 degrees C was found to be fairly constant (although a little longer, at about 55 min), as was the cell mass at initiation of DNA replication. The cell cycle of B. subtilis differed from that of E. coli in that changes in growth rate affected the average cell length but not the width and also in the relative variability of period between termination of DNA replication and septation. Overall movement of the nucleoid was found to occur smoothly, as in E. coli, but other aspects of nucleoid behavior were consistent with an underlying active partitioning machinery. The models for cell cycle progression in B. subtilis should facilitate the interpretation of data obtained from the recently introduced cytological methods for imaging the assembly and movement of proteins involved in cell cycle dynamics.

Bacillus subtilis↗
PubMed1987

Coordinate initiation of chromosome and minichromosome replication in Escherichia coli.

Escherichia coli minichromosomes harboring as little as 327 base pairs of DNA from the chromosomal origin of replication (oriC) were found to replicate in a discrete burst during the division cycle of cells growing with generation times between 25 and 60 min at 37 degrees C. The mean cell age at minichromosome replication coincided with the mean age at initiation of chromosome replication at all growth rates, and furthermore, the age distributions of the two events were indistinguishable. It is concluded that initiation of replication from oriC is controlled in the same manner on minichromosomes and chromosomes over the entire range of growth rates and that the timing mechanism acts within the minimal oriC nucleotide sequence required for replication.

Cell Cycle↗
PubMed1993

Replicatively active complexes of DnaA protein and the Escherichia coli chromosomal origin observed in the electron microscope.

DnaA protein and the Escherichia coli chromosomal origin (oriC) form an initial complex at an early stage in the initiation of DNA replication. We have used electron microscopy to determine which structure among the several formed in the reconstitution of this multicomponent system is the replicatively active complex. One distinctive structure could be correlated with activity and localized to oriC, whilst several others could not. Formation of an open complex in the next stage of initiation was accompanied by the presence of a structure similar in size and shape to that of the functional initial complex. Whereas the initial complex was observed with either ATP or the ADP-forms of DnaA protein, only the ATP-form was effective in producing the open complex. Mutagenesis of several DNA sequence elements in oriC, known to be important for replication, was employed to determine the effects of these alterations on formation of the initial complex. As judged by electron microscopy and by functional assays, the region containing the four 9-mer dnaA boxes proved to be essential for the formation of the initial complex, while the three contiguous AT-rich 13-mers, known sites for opening of oriC, were not.

Adenosine Diphosphate↗
PubMed1989

Negative control of plasmid R6K replication: possible role of intermolecular coupling of replication origins.

The gamma origin binding sites of the replication initiator pi protein, composed of seven 22-base-pair (bp) direct repeats and previously shown to be essential for replication of plasmid R6K, can also act as an inhibitor of R6K replication in Escherichia coli cells if provided in trans. Inhibition is dependent upon the ability of these repeats to bind the R6K-encoded pi protein but is not overcome by increasing the intracellular pi level. The insertion of a second repeat cluster in close proximity to the gamma origin also can markedly inhibit replication. The severity of this effect is dependent upon the position, orientation, and number of repeats present in the extra cluster. As few as six extra repeats can result in a completely nonfunctional gamma origin. However, this inactive gamma origin plasmid containing the six extra repeats is functional when placed in a strain that underproduces the wild-type pi protein or when placed in the presence of any of several copy-up mutant pi proteins. On the basis of these observations, we propose that the nucleoprotein structures formed by the binding of pi protein to the seven 22-bp direct repeats at the gamma origin are capable of coupling with each other in vivo and that replication initiation is prevented at such coupled origins. In support of this model of replication control, we demonstrate by electron microscopy analysis that the pi protein has the ability to associate two DNA molecules containing gamma origin sequences and also show that pi enhances the DNA ligase-catalyzed multimerization of a DNA fragment containing the gamma origin.

Bacterial Proteins↗
PubMed1995

The interaction of RepC initiator with iterons in the replication of the broad host-range plasmid RSF1010.

The replication origin of the broad host-range plasmid RSF1010 contains 3.5 copies of a 20mer iteron sequence that bind specifically to the plasmid-encoded initiator, RepC. Here we demonstrated that even a single iteron was bent upon binding of RepC. Moreover, the bending angle seems to become larger along with the increment of the number of iterons. In a mutational analysis of the iteron sequence, we isolated seven kinds of base-substitution mutants of iterons, and estimated the replication activity of these mutants in vivo. We found that each of the subsections in the 20mer iteron sequence made a distinct contribution to the initiation of RSF1010 DNA replication. With the binding assay of RepC and mutated iterons in vitro, we found that the formation of a productive RepC-iteron complex was required for the initiation of plasmid DNA replication.

Bacterial Proteins↗
PubMed1983

Two pathways of division inhibition in UV-irradiated E. coli.

We have investigated the mechanism of division inhibition in E. coli following UV-irradiation or nalidixic acid treatment. After UV, two separate mechanisms, both dependent upon recA+, appear to block division. One mechanism is dependent upon sfiA and sfiB, is inhibited by low levels (4 micrograms/ml) of rifamycin and is expressed in tif mutants at 42 degrees C. The second mechanism is independent of sfiA, and sfiB, is resistant to rifamycin and does not occur in cells lacking DNA replication forks. We suggest that this second mechanism is the result of the failure to terminate DNA replication in inhibited cells. Nalidixic acid inhibition of cell division also appears to involve both mechanisms but as found previously replication forks are also necessary to induce the sfi pathway.

Bacterial Proteins↗
PubMed2000

FtsZ ring formation without subsequent cell division after replication runout in Escherichia coli.

In this report, we have investigated cell division after inhibition of initiation of chromosome replication in Escherichia coli. In a culture grown to the stationary phase, cells containing more than one chromosome were able to divide some time after restart of growth, under conditions not allowing initiation of chromosome replication. This shows that there is no requirement for cell division to take place within a certain time after initiation of chromosome replication. Continued growth without initiation of replication resulted in filamented cells that generally did not have any constrictions. Interestingly, FtsZ rings were formed in a majority of these cells as they reached a certain cell length. These rings appeared and were maintained for some time at the cell quarter positions on both sides of the centrally localized nucleoid. These results confirm previous findings that cell division sites are formed independently of chromosome replication and indicate that FtsZ ring assembly is dependent on cell size rather than on the capacity of the cell to divide. Disruption of the mukB gene caused a significant increase in the region occupied by DNA after the replication runout, consistent with a role of MukB in chromosome condensation. The aberrant nucleoid structure was accompanied by a shift in FtsZ ring positioning, indicating an effect of the nucleoid on the positioning of the FtsZ ring. A narrow cell length interval was found, under and over which primarily central and non-central FtsZ rings, respectively, were observed. This finding correlates well with the previously observed oscillatory movement of MinC and MinD in short and long cells.

Bacterial Proteins↗
PubMed1978

Map location of the Escherichia coli origin of replication.

Working with restriction fragments obtained directly from the Escherichia coli K12 chromosome, the EcoRI-HindIII restriction map of the section of the chromosome containing the replication origin has been extended by 14 kilobase pairs (kb) to cover 56 kb. Within this newly mapped portion, the ilv and rrnC cistrons have been identified by (1) hybridization of individual restriction fragmanents to the ilv-transducing phage lambdadilv5 and (2) a comparison of the restriction map of this region with the EcoRI map of lambda dilv5 and the Hind III map of the plasmid pJC110, a ColEl-ilv hybrid. The replication origin is located approximately 30 kb from the ilvE gene and 20 kb from the rrnC 16S rRNA cistron. This places the origin near 82.7 min on the genetic map, close to uncA.

Chromosome Mapping↗
PubMed1988

A model for initiation at origins of DNA replication.

Many prokaryotic origins resemble E. coli oriC in possessing essential AT-rich sequences, tandemly repeated. The role of these repeats may be in the initial opening of the duplex by the initiator protein, as has been found for the 13-mers in oriC and is implied for the 11-mers of the lambda origin. Regulatory influences on the effective action of the initiator protein of E. coli (dnaA protein) include transcriptional activation of the origin, nucleotide binding and membrane attachment of the protein, and interactions leading to the introduction of helicases to start replication forks.

Adenosine Triphosphate↗
PubMed2001

Feedback controls restrain the initiation of Escherichia coli chromosomal replication.

In Escherichia coli, initiation of chromosomal replication is activated by a nucleoprotein complex formed primarily between the DnaA protein and oriC (replication origin) DNA. After replicational initiation, this complex has to be inactivated in order to repress the appearance of initiation events until the next scheduled round of initiation. Studies of the mechanisms responsible for this repression have recently revealed direct coupling between these mechanisms and key elements of the replication process, suggesting that feedback-type regulatory loops exist between the factors implicated in initiation and the elements yielded by the replication process. The loading of the ring-shaped beta-subunit of DNA polymerase III onto DNA plays a key role in the inactivation of the DnaA protein. Duplication of oriC DNA results in hemimethylated DNA, which is inert for reinitiation. Titration of large amounts of DnaA protein to a non-oriC locus can repress untimely initiations, and timely duplication of this locus is required for this repression in rapidly growing cells. All these systems functionally complement one another to ensure the maintenance of the interinitiation interval between two normal DNA replication cycles. The mechanisms that link the replication cycle to the progression of the cell cycle are also discussed.

Bacterial Proteins↗
PubMed1996

The nucleoid protein H-NS facilitates chromosome DNA replication in Escherichia coli dnaA mutants.

Growth inhibition of the dnaA(Cs) mutant, which overinitiates chromosome replication, was shown to be dependent upon the nucleoid protein H-NS. [3H]thymine incorporation experiments indicated that the absence of H-NS inhibited overreplication by the dnaA(Cs) mutant. In addition, the temperature-sensitive phenotype of a dnaA46 mutant was enhanced by disruption of H-NS. These observations suggest that H-NS directly or indirectly facilitates the initiation of chromosome replication.

Bacterial Outer Membrane Proteins↗
PubMed1988

Location of sites that inhibit progression of replication forks in the terminus region of Escherichia coli.

We used a Southern hybridization assay to locate precisely the sites at which DNA replication is arrested in the terminus region of the Escherichia coli chromosome. The assay was based on the properties of restriction fragments that contain stalled replication forks. Replication forks that entered the terminus from the clockwise direction with respect to the genetic map were inhibited near manA at a site called T2, which we located at kilobase 442 on the physical map of Bouché (J. P. Bouché, J. Mol. Biol. 154:1-20, 1982). Those that entered the terminus region traveling in the counterclockwise direction were inhibited near pyrF at a site called T1, which we located at kilobase 90. In each case we found only a single, precise site of arrest. Inhibition at T1 was not detectable in our assay in strains lacking the trans-acting locus tus, which is located near T2 and is required for T1 to function. We demonstrated that the sites of inhibition are also used during termination of replication in exponentially growing, wild-type cells. In all previous studies on the terminus of E. coli, inhibition has only been detected in strains that were modified so that the origin used was placed near the terminus to force the use of the sites of inhibition.

Chromosome Mapping↗