Chromosome partitioning in Escherichia coli: novel mutants producing anucleate cells

Hiraga, Sota; Niki, Hironori; Ogura, Teru; Ichinose, Chiyome; Mori, Hirotada; Ezaki, Bunichi; Jaffé, A

doi:10.1128/jb.171.3.1496-1505.1989

Cited by 356 publications

(275 citation statements)

References 41 publications

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“…In addition, six so-called par genes involved in decatenation of interlinked daughter chromosomes have been characterized, as well as a number of genes thought to be involved in chromosome movement and partitioning (reviewed in reference 43). Among the latter group are the muk, minD, and ftsK genes of E. coli (12,23,49) and the smc, spoOJ, and spoIIIE genes of B. subtilis (6,13,26,45). Finally, recent work by Lemon and Grossman (19) indicates that the process of DNA replication itself may contribute to the movement and separation of daughter chromosomes.…”

Section: Discussionmentioning

confidence: 99%

Penicillin-Binding Protein-Related Factor A Is Required for Proper Chromosome Segregation in Bacillus subtilis

Pedersen

Setlow

2000

J Bacteriol

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Previous work has shown that the ponA gene, encoding penicillin-binding protein 1 (PBP1), is in a two-gene operon with prfA (PBP-related factor A) (also called recU), which encodes a putative 206-residue basic protein (pI ‫؍‬ 10.1) with no significant sequence homology to proteins with known functions. Inactivation of prfA results in cells that grow slower and vary significantly in length relative to wild-type cells. We now show that prfA mutant cells have a defect in chromosome segregation resulting in the production of ϳ0.9 to 3% anucleate cells in prfA cultures grown at 30 or 37°C in rich medium and that the lack of PrfA exacerbates the chromosome segregation defect in smc and spoOJ mutant cells. In addition, overexpression of prfA was found to be toxic for and cause nucleoid condensation in Escherichia coli.Penicillin-binding proteins (PBPs), which catalyze the polymerization and cross-linking of bacterial peptidoglycan, can be divided into three classes based on their amino acid sequence: the low-molecular-weight PBPs and the high-molecular-weight (HMW) class A and class B PBPs (8). The HMW class B PBPs are monofunctional transpeptidases, some of which have essential functions in septation and maintenance of cell shape (8), while the HMW class A PBPs have both transglycosylase and transpeptidase activities (14, 42) and appear to be somewhat functionally redundant (17, 34). The Bacillus subtilis ponA gene, coding for the HMW class A PBP1, is transcribed predominantly during log-phase growth (34). Previous work with B. subtilis mutants lacking one or several of the three known HMW class A PBPs (PBP1, PBP2c, and PBP4) showed that (i) lack of PBP1 results in slower growth, increased cell length, and decreased cell diameter and (ii) PBP1 is functionally more important than PBP2c and PBP4 (34). It was also recently demonstrated that PBP1 localizes to cell division sites and plays an important role in the formation of the peptidoglycan division septum in vegetative cells of B. subtilis (30).The ponA gene is part of a two-gene operon that also includes prfA (PBP-related factor A [note that in the B. subtilis genome database, prfA is called recU]), which is located immediately upstream of and cotranscribed with ponA (33). prfA codes for a putative 206 residue, basic protein (pI of 10.1), which has no significant sequence homology to proteins with known functions. However, DNA sequencing has indicated that genes encoding similar proteins are present in a large number of gram-positive bacteria: a BLAST search (2) of prfA against completed and unfinished microbial genomes (preliminary sequence data were obtained from The Institute of Genomic Research Website at http://www.tigr.org) produced sequences with significant homology from 10 different grampositive organisms, while no prfA homologs were detected in gram-negative bacteria by this analysis. The former organisms include Enterococcus faecalis (54% identity in a 192-aminoacid overlap), Staphylococcus aureus (58% identity in a 167-amino-acid overlap), Streptococcus p...

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Section: Discussionmentioning

confidence: 99%

Penicillin-Binding Protein-Related Factor A Is Required for Proper Chromosome Segregation in Bacillus subtilis

Pedersen

Setlow

2000

J Bacteriol

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“…Bacterial cell membrane preparation E. coli BL21 carrying the plasmid pAX629 (pACYC184 containing the tolC gene; Hiraga et al, 1989) was grown in 2× TY ( Tryptone-Yeast) medium (Oxoid), 20 g ml ¹1 chloramphenicol, in a shaker at 37ЊC. At OD 600 ¼ 1.5, cells were harvested and to prepare outer membrane, cells were resuspended in a minimum volume of 10 mM Tris-HCl, pH 7.4, 5 mM MgCl 2 .…”

Section: Methodsmentioning

confidence: 99%

“…TolC was obtained from BL21 carrying pAX629 that expresses intact tolC from its own Escherichia coli promotor (Hiraga et al, 1989). Cells were broken in a French pressure cell, and separation of outer membrane (OM) from inner membrane (IM) was achieved by centrifugation onto a 1.6 M sucrose cushion.…”

Section: Tolc Purification From Membranes Yields Oligomers In Solutionmentioning

confidence: 99%

Structure of TolC, the outer membrane component of the bacterial type I efflux system, derived from two‐dimensional crystals

Koronakis

et al. 1997

Molecular Microbiology

122

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SummaryTolC is an outer membrane protein required for the export of virulence proteins and toxic compounds without a periplasmic intermediate. We show that TolC is an integral part of the translocator, interacting with inner membrane components, by demonstrating a need for TolC in protein export not only from intact cells but also from sphaeroplasts. To establish the structure of TolC, and thus gain information on how this might be achieved, the protein was purified from the Escherichia coli outer membrane, as a trimer, and crystallized in two-dimensional lattices by reconstitution in phospholipid bilayers. The projection structure at 12 Å resolution showed a threefold symmetric molecule of 58 Å outer diameter, and a single pool of stain filling its centre. Side views parallel to the membrane plane revealed an additional domain outside the membrane. Eighteen membranespanning ␤-strands were predicted for the 51.5 kDa monomer, excluding a 7 kDa C-terminal segment, and this segment was shown to contain a proteinase K-sensitive site that was exposed in reconstituted membranes and sphaeroplasts, but which was protected in intact cells. The combined data suggest that TolC is a trimeric outer membrane protein with each monomer comprising a membrane domain, predicted to be ␤-barrel, and a C-terminal periplasmic domain. The latter could form part of the bridge to the energized inner membrane component of the translocation complex.

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“…L medium (Hiraga et al 1989) was used for growth of YK1100 and CM748. L medium supplemented with thymine (50 mg/mL) was used for growth of PC2.…”

Section: Bacterial Strainsmentioning

confidence: 99%

Bidirectional migration of SeqA‐bound hemimethylated DNA clusters and pairing of oriC copies in Escherichia coli

et al. 2000

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Chromosome partitioning in Escherichia coli: novel mutants producing anucleate cells

Cited by 356 publications

References 41 publications

Penicillin-Binding Protein-Related Factor A Is Required for Proper Chromosome Segregation in Bacillus subtilis

Penicillin-Binding Protein-Related Factor A Is Required for Proper Chromosome Segregation in Bacillus subtilis

Structure of TolC, the outer membrane component of the bacterial type I efflux system, derived from two‐dimensional crystals

Bidirectional migration of SeqA‐bound hemimethylated DNA clusters and pairing of oriC copies in Escherichia coli

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