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At least 19 recordsLinked to original sources
PubMed1977

Origin of replication, oriC, of the Escherichia coli chromosome: mapping of genes relative to R.EcoRI cleavage sites in the oriC region.

A precise genetic-physical map of the tna-ilv region at 82 min on the genetic map of E. coli is obtained through deletion mapping and analysis by restriction endonuclease EcoRI of plasmids, derived from an F' carrying the genes between aroE and ilv. A locus, designated het, which in its diploid state results in slow growth and heterogeneity of cell size due to distorted cell division, maps between bglB and asn, 30-45 kb counterclockwise of ilv. The pattern of R.EcoRI cleavage sites in the het region is identical with the pattern obtained by Marsh and Worcel (1977) who analyzed DNA labeled preferentially in the region of the DNA replication origin (oriC). We suggest that oriC is identical with the het site and that it can be allocated to a position 32 kb counterclockwise of the ilv operon.

Chromosome Mapping↗
PubMed1979

Deoxyribonucleic acid and outer membrane: strains diploid for the oriC region show elevated levels of deoxyribonucleic acid-binding protein and evidence for specific binding of the oriC region to outer membrane.

We have recently reported that part of the chromosomal deoxyribonucleic acid (DNA) of Escherichia coli is associated with the outer membrane fraction and that an outer membrane protein having a molecular weight of 31,000 probably is involved in this association (H. Wolf-Watz and A. Norqvist, J. Bacteriol. 140:43-49, 1979). We have now found that F' merodiploid strains containing two copies of the DNA between bglB and ilv have increased levels of this protein and an increased amount of DNA in their outer membranes. Increased levels of the protein are also found when lambda asn phage, containing at 1.5-megadalton fragment of DNA located to the right of the uncA uncB genes but to the left of oriC, are induced. It therefore seems that this 1.5-megadalton fragment of DNA either codes for or binds to the 31,000-dalton outer membrane protein. Hybridization studies utilizing DNA found to be bound to outer membrane and DNA isolated from a specialized transducing phage lambda asn 132 revealed that at least 5 to 10% of outer membrane DNA has a DNA sequence homologous with a chromosomal segment carried by this oriC-containing phage.

Bacterial Proteins↗
PubMed1992

HU, the major histone-like protein of E. coli, modulates the binding of IHF to oriC.

HU is one of the most abundant DNA binding proteins of bacteria. Unlike IHF, integration host factor of Escherichia coli, with which HU shares many properties, including a strong sequence homology and similar predicted structure, HU seems to bind non-specifically to DNA whereas IHF binds to specific sites. In this work we compare the binding characteristics of HU and IHF to a DNA fragment containing the minimal origin of replication of E. coli (oriC) and we analyse the effect of HU on the binding capacity of IHF to this oriC fragment. We show that HU interacts randomly and non-specifically with oriC as opposed to the specific binding of IHF to this same DNA sequence. In addition, we show that HU can modulate the binding of IHF to its specific oriC site. Depending on the relative concentrations of HU and IHF, HU is able either to activate or to inhibit the binding of IHF to oriC.

Autoradiography↗
PubMed1992

Characterization of the Escherichia coli membrane domain responsible for binding oriC DNA.

It has previously been shown that hemimethylated DNA from the Escherichia coli replication origin (oriC) binds with high specificity to membrane fractions isolated from disrupted cells. In this article, the membrane localization of oriC-binding activity was studied by subjecting crude membrane preparations to successive cycles of sedimentation and flotation gradient analysis. This revealed that approximately two-thirds of the membrane-associated oriC-binding activity of the cell was not associated with the outer membrane fraction as previously suggested but was recovered instead in a unique membrane fraction (OCB1) whose buoyant density and protein profile differed from those of both inner and outer membranes. The specific activity of oriC binding in OCB1 was approximately fivefold higher than the activity of the isolated outer membrane peak. It is likely that membrane fraction OCB1 includes the membrane domain responsible for the binding of hemimethylated oriC to the cell envelope in intact cells.

Bacterial Outer Membrane Proteins↗
PubMed1978

Origin of replication, oriC, or the Escherichia coli chromosome on specialized transducing phages lambda asn.

Specialized transducing phages lambda asn harboring chromosomal DNA and genetic markers on either side of the asn gene were isolated. Phages carrying chromosomal DNA counterclockwise of the asn gene can upon infection establish themselves as self-replicating plasmids in asn, recA hosts lysogenic for lambda. It is concluded that this bypassing of normal lambda immunity is due to the presence of the chromosomal replication origin, oriC, in this class of phages. Genetic analysis and the determination of restriction endonuclease cleavage patterns of the different lambda asn lead to the allocation of oriC within 1.5 megadaltons of the asn gene towards the uncA, uncB genes at 82 min on the genetic map of E. coli. The clockwise order of genes on the chromosomes is found to be: bglB, (pst, glmS), (uncA, uncB), oriC, asn, trkD, rbs, rrnC, ilv.

Chromosome Mapping↗
PubMed1992

The 55-kilodalton protein in an oriC complex fraction is glycogen synthase.

Three proteins with molecular masses of 35, 55, and 75 kDa were found in an oriC complex fraction after purification through CsCl density gradient centrifugation (W. G. Hendrickson, T. Kusano, H. Yamaki, R. Balakrishnan, M. King, J. Murchie, and M. Schaechter, Cell 30:915-923, 1982). Of these three proteins, the 55-kDa protein was determined to be glycogen synthase on the basis of the N-terminal amino acid sequence and the molecular weight. The oriC complex was formed in glgA mutant cells, which produce no detectable glycogen, as well as in wild-type cells. None of the 35-, 55-, and 75-kDa proteins were detected in the fraction from this mutant. The results indicate that these proteins were not constituents of the oriC complex.

Bacterial Proteins↗
PubMed1992

Physical distance between the site of type II DNA binding to the membrane and oriC on the Bacillus subtilis 168 chromosome.

The precise physical locations of the oriC region and the region for type II DNA binding to the membrane on the Bacillus subtilis 168 chromosome were determined. The DNA regions were physically mapped by creating new restriction sites (NotI and SfiI) within these regions. The physical distance between oriC and the type II DNA-binding region was verified with the creation of a novel sequence cleaved by endonuclease I-SceI in each of the above regions. Complete removal of the defined type II membrane-binding region produced no noticeable phenotype.

Bacillus subtilis↗
PubMed1992

Association of oriC region of Escherichia coli chromosome with outer membrane: effects of culture condition.

We isolated complexes containing oriC region DNA and outer membrane, named origin complex heavy and origin complex light, from the cells of Escherichia coli cultured in media with poor or rich of nutrients, and found the different nature of association between origin DNA and outer membrane. The ratio of origin complex light to origin complex heavy prepared from the cells cultured in rich media was lower than that of those from minimal medium culture. Outer membrane preparations from the cells grown in nutritious media had high abilities of association with origin complex light in the presence of magnesium. These results indicated that the number of binding sites on outer membrane with origin region DNA increase, or the binding between outer membrane and origin region DNA become more rigid, when cells grow faster and DNA replication initiate more frequently in a nutritious medium.

Binding Sites↗
PubMed1992

The chromosome origin of Escherichia coli stabilizes DnaA protein during rejuvenation by phospholipids.

DnaA protein (the initiator protein) binds and clusters at the four DnaA boxes of the Escherichia coli chromosomal origin (oriC) to promote the strand opening for DNA replication. DnaA protein activity depends on the tight binding of ATP; the ADP form of DnaA protein, generated by hydrolysis of the bound ATP, is inactive. Rejuvenation of ADP-DnaA protein, by replacement with ATP, is catalyzed by acidic phospholipids in a highly fluid bilayer. We find that interaction of DnaA protein with oriC DNA is needed to stabilize DnaA protein during this rejuvenation process. Whereas DnaA protein bound to oriC DNA responds to phospholipids, free DnaA protein is inactivated by phospholipids and then fails to bind oriC. Furthermore, oriC DNA facilitates the high affinity binding of ATP to DnaA protein during treatment with phospholipids. A significant portion of the DnaA protein associated with oriC DNA can be replaced by the ADP form of the protein, suggesting that all of the DnaA protein bound to oriC DNA need not be rejuvenated between rounds of replication.

Adenosine Diphosphate↗
PubMed1992

Novel Escherichia coli mutant, dnaR, thermosensitive in initiation of chromosome replication.

A newly isolated Escherichia coli mutant thermosensitive in DNA synthesis had an allele named dnaR130, which was located at 26.3 minutes on the genetic map. The mutant was defective in initiation of chromosome replication but not in propagation at a high temperature. This mutant was capable of growing in the absence of the rnh function at the high temperature by means of a dnaA-independent replication mechanism. In the mutant exposed to the high temperature, an oriC plasmid was able to replicate, although at a lower rate than at the low temperature. The plasmid replication at the high temperature depended on the dnaA function essential for the initiation of replication from oriC. The mutant lacking the rnh function persistently maintained the oriC plasmid at the high temperature in a dnaA-dependent manner. Thus, the dnaR function was required for initiation of replication of the bacterial chromosome from oriC but not the oriC plasmid. This result reveals that a dnaR-dependent initiation mechanism that is dispensable for oriC plasmid replication operates in the bacterial chromosome replication.

Bacterial Proteins↗
PubMed1992

The complex for replication initiation of Escherichia coli.

We probed the complex between oriC and DnaA protein using two types of mutants in oriC. Base changes in the DnaA binding sites, DnaA boxes, had little effect on origin function. Mutations which change the distance between DnaA boxes R3 and R4, on the other hand, inactivated oriC unless the mutation deleted or inserted one complete helical turn. Origins with other 10 base pair insertions in the interval between DnaA boxes R2 and R3 were functional, but not insertions in the R1-R2 interval. FIS protein binds to a bipartite site in oriC between DnaA boxes R2 and R3. A model for the oriC/DnaA complex based on these results suggests an array of DnaA monomers with a 34 A spacing upon which oriC is arranged.

Bacterial Proteins↗
PubMed2026

Ori-Finder-Arch: An Updated Web Server for the Annotation and Visualization of Archaeal Replication Origins.

Archaea are promising chassis organisms in biotechnology, and the accurate annotation of their chromosomal replication origins (oriCs) is the key to unlocking their full potential. However, the existing Ori-Finder 2 web server suffers from low accuracy, slow speed, and limited scalability. In this study, we present Ori-Finder-Arch, an updated web server for high-performance oriC prediction in archaea. This pipeline integrates HMMER-based replication initiation protein (RIP) annotation, refined consensus motif recognition, and GC profile-based DNA unwinding element (DUE) detection. On a benchmark set of experimentally validated oriCs, Ori-Finder-Arch achieved a recall of 95.6% and a precision of 86.0%, substantially outperforming Ori-Finder 2 (62.2% and 63.6%, respectively), while running 4.75 times faster and supporting diverse assembly levels. When applied to the available archaeal assemblies, it successfully annotated 17,472 oriCs. Meanwhile, the web server provides interactive visualizations at different levels. In conclusion, Ori-Finder-Arch offers an efficient, accurate, and user-friendly platform for advanced studies of archaeal DNA replication initiation and synthetic biology applications, and is freely available at https://tubic.org/Ori-Finder-Arch/ and https://tubic.tju.edu.cn/Ori-Finder-Arch/.

Archaea↗
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↗
PubMed1992

DNA damage-inducible origins of DNA replication in Escherichia coli.

Upon induction of the SOS response in Escherichia coli, the mode of initiation of DNA replication is altered such that it can occur in the absence of normally required protein synthesis. This type of DNA replication has been termed induced stable DNA replication (iSDR). We examined the origin usage during iSDR and found that the initiation of iSDR occurs primarily in the oriC and terC regions of the chromosome in a manner completely independent of transcription, translation and DnaA protein. Minichromosomes (oriC plasmids) pOC23 and pOC81 were induced to replicate in the absence of DnaA protein and transcription after SOS induction. The results localized one of the iSDR origin activities in a 596 bp region which includes the minimal oriC.

Bacterial Proteins↗
PubMed1992

Mutations in the DnaA binding sites of the replication origin of Escherichia coli.

Mutations (base changes) were introduced into the four DnaA binding sites (DnaA boxes) of the Escherichia coli replication origin, oriC. Mutations in a single DnaA box did not impair the ability of these origins to replicate in vivo and in vitro. A combination of mutations in two DnaA boxes, R1 and R4, resulted in slower growth of the oriC plasmid-bearing host cells. DnaA protein interaction with mutant and wild-type DnaA boxes was analyzed by DNase I footprinting. Binding of DnaA protein to a mutated DnaA box R1 was not affected by a mutation in DnaA box R4 and vice versa. Mutations in DnaA boxes R1 and R4 did not modify the ability of the DnaA protein to bind to other DnaA boxes in oriC.

Bacterial Proteins↗