scholarly journals Unstable Escherichia coli L Forms Revisited: Growth Requires Peptidoglycan Synthesis

2007 ◽  
Vol 189 (18) ◽  
pp. 6512-6520 ◽  
Author(s):  
Danièle Joseleau-Petit ◽  
Jean-Claude Liébart ◽  
Juan A. Ayala ◽  
Richard D'Ari
Keyword(s):  
Escherichia Coli ◽  
Cell Division ◽  
High Performance ◽  
Specific Inhibitor ◽  
Hypertonic Medium ◽  
Penicillin Binding Proteins ◽  
E Coli ◽  
Chromatography Analysis ◽  
Peptidoglycan Synthesis ◽  
K 12

ABSTRACT Growing bacterial L forms are reputed to lack peptidoglycan, although cell division is normally inseparable from septal peptidoglycan synthesis. To explore which cell division functions L forms use, we established a protocol for quantitatively converting a culture of a wild-type Escherichia coli K-12 strain overnight to a growing L-form-like state by use of the β-lactam cefsulodin, a specific inhibitor of penicillin-binding proteins (PBPs) 1A and 1B. In rich hypertonic medium containing cefsulodin, all cells are spherical and osmosensitive, like classical L forms. Surprisingly, however, mutant studies showed that colony formation requires d-glutamate, diaminopimelate, and MurA activity, all of which are specific to peptidoglycan synthesis. High-performance liquid chromatography analysis confirmed that these L-form-like cells contain peptidoglycan, with 7% of the normal amount. Moreover, the β-lactam piperacillin, a specific inhibitor of the cell division protein PBP 3, rapidly blocks the cell division of these L-form-like cells. Similarly, penicillin-induced L-form-like cells, which grow only within the agar layers of rich hypertonic plates, also require d-glutamate, diaminopimelate, and MurA activity. These results strongly suggest that cefsulodin- and penicillin-induced L-form-like cells of E. coli—and possibly all L forms—have residual peptidoglycan synthesis which is essential for their growth, probably being required for cell division.

Effect of 5-bromouracil on the growth of a thymine auxotroph of Escherichia coli K-12

1971 ◽  
Vol 17 (1) ◽  
pp. 87-93 ◽  
Author(s):  
Roosevelt J. Jones ◽  
Roger R. Hewitt
Keyword(s):  
Escherichia Coli ◽  
Cell Division ◽  
Solid Medium ◽  
Cell Number ◽  
Bacteriostatic Effect ◽  
E Coli ◽  
Static Phase ◽  
Two Phases ◽  
K 12 ◽  
Mass Increase

An atypical viability response to 5-bromouracil has been observed in a thymine auxotroph of E. coli K-12. The response occurs in two phases, the first reflecting tolerance to the analogue during continued exponential growth and cell division. The second is a static phase during which viable number remains constant, while cell number and mass increase at a diminishing rate.During the latter phase filamentous cells increase in number and length. Examination of the cloning potential of cells after 10 h of growth in 5-bromouracil indicated that filamentous cells continue extension on solid medium into non-septate coils that are sterile. Other cells, presumably static when plated, readily form microcolonies free of defective members.Observed responses to penicillin, potentially stabilizing media, or added thymine suggest that 5-bromouracil evokes a bimodal response in this strain. The analogue exerts a bacteriostatic effect on some cells which remain viable for several hours. The bacteriocidal effect, presumably on cells continuing growth, interferes with cell division by preventing septation.

scholarly journals TolC-Dependent Exclusion of Porphyrins in Escherichia coli

2008 ◽  
Vol 190 (18) ◽  
pp. 6228-6233 ◽  
Author(s):  
Ryoko Tatsumi ◽  
Masaaki Wachi
Keyword(s):  
Escherichia Coli ◽  
Uv Irradiation ◽  
High Performance ◽  
Aminolevulinic Acid ◽  
Wild Type ◽  
E Coli ◽  
Chromatography Analysis ◽  
Increased Sensitivity ◽  
Mutant Cells ◽  
Reddish Brown Color

ABSTRACT We found that Escherichia coli tolC mutants showed increased sensitivity to 5-aminolevulinic acid (ALA), a precursor of porphyrins. The tolC mutant cells grown in the presence of ALA showed a reddish brown color under visible light and a strong red fluorescence under near-UV irradiation. Fluorescence spectrometry and high-performance liquid chromatography analysis showed that the tolC mutant cells grown in the presence of ALA accumulated a large amount of coproporphyrin(ogen) intracellularly. In contrast, the wild-type cells produced coproporphyrin extracellularly. The tolC mutant cells grown in the presence of ALA, which were capable of surviving in the dark, were killed by near-UV irradiation, suggesting that the intracellular coproporphyrin(ogen) renders these cells photosensitive. These results suggest that the TolC-dependent efflux system is involved in the exclusion of porphyrin(ogen)s in E. coli.

scholarly journals Regulation of Cell Division, Biofilm Formation, and Virulence by FlhC in Escherichia coli O157:H7 Grown on Meat

2011 ◽  
Vol 77 (11) ◽  
pp. 3653-3662 ◽  
Author(s):  
Preeti Sule ◽  
Shelley M. Horne ◽  
Catherine M. Logue ◽  
Birgit M. Prüß
Keyword(s):  
Escherichia Coli ◽  
Cell Division ◽  
Real Time ◽  
Real Time Pcr ◽  
Biofilm Formation ◽  
Parent Strain ◽  
Cellular Functions ◽  
Content Type ◽  
E Coli ◽  
K 12

ABSTRACTTo understand the continuous problems thatEscherichia coliO157:H7 causes as food pathogen, this study assessed global gene regulation in bacteria growing on meat. Since FlhD/FlhC ofE. coliK-12 laboratory strains was previously established as a major control point in transducing signals from the environment to several cellular processes, this study compared the expression pattern of anE. coliO157:H7 parent strain to that of its isogenicflhCmutant. This was done with bacteria that had been grown on meat. Microarray experiments revealed 287 putative targets of FlhC. Real-time PCR was performed as an alternative estimate of transcription and confirmed microarray data for 13 out of 15 genes tested (87%). The confirmed genes are representative of cellular functions, such as central metabolism, cell division, biofilm formation, and pathogenicity. An additional 13 genes from the same cellular functions that had not been hypothesized as being regulated by FlhC by the microarray experiment were tested with real-time PCR and also exhibited higher expression levels in theflhCmutant than in the parent strain. Physiological experiments were performed and confirmed that FlhC reduced the cell division rate, the amount of biofilm biomass, and pathogenicity in a chicken embryo lethality model. Altogether, this study provides valuable insight into the complex regulatory network of the pathogen that enables its survival under various environmental conditions. This information may be used to develop strategies that could be used to reduce the number of cells or pathogenicity ofE. coliO157:H7 on meat by interfering with the signal transduction pathways.

scholarly journals Unexpected Phytostimulatory Behavior for Escherichia coli and Agrobacterium tumefaciens Model Strains

2013 ◽  
Vol 26 (5) ◽  
pp. 495-502 ◽  
Author(s):  
Vincent Walker ◽  
Maxime Bruto ◽  
Floriant Bellvert ◽  
René Bally ◽  
Daniel Muller ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Agrobacterium Tumefaciens ◽  
Nitrite Reductase ◽  
Azospirillum Brasilense ◽  
High Performance ◽  
Positive Control ◽  
Shoot Biomass ◽  
E Coli ◽  
K 12 ◽  
The Impact

Plant-beneficial effects of bacteria are often underestimated, especially for well-studied strains associated with pathogenicity or originating from other environments. We assessed the impact of seed inoculation with the emblematic bacterial models Agrobacterium tumefaciens C58 (plasmid-cured) or Escherichia coli K-12 on maize seedlings in nonsterile soil. Compared with the noninoculated control, root biomass (with A. tumefaciens or E. coli) and shoot biomass (with A. tumefaciens) were enhanced at 10 days for ‘PR37Y15’ but not ‘DK315’, as found with the phytostimulator Azospirillum brasilense UAP-154 (positive control). In roots as well as in shoots, Agrobacterium tumefaciens and E. coli triggered similar (in PR37Y15) or different (in DK315) changes in the high-performance liquid chromatography profiles of secondary metabolites (especially benzoxazinoids), distinct from those of Azospirillum brasilense UAP-154. Genome sequence analysis revealed homologs of nitrite reductase genes nirK and nirBD and siderophore synthesis genes for Agrobacterium tumefaciens, as well as homologs of nitrite reductase genes nirBD and phosphatase genes phoA and appA in E. coli, whose contribution to phytostimulation will require experimental assessment. In conclusion, the two emblematic bacterial models had a systemic impact on maize secondary metabolism and resulted in unexpected phytostimulation of seedlings in the Azospirillum sp.-responsive cultivar.

scholarly journals Chlamydial MreB Directs Cell Division and Peptidoglycan Synthesis in Escherichia coli in the Absence of FtsZ Activity

mBio ◽  
2020 ◽  
Vol 11 (1) ◽  
Author(s):  
Dev K. Ranjit ◽  
George W. Liechti ◽  
Anthony T. Maurelli
Keyword(s):  
Escherichia Coli ◽  
Cell Division ◽  
Morphological Changes ◽  
Content Type ◽  
E Coli ◽  
Obligate Intracellular ◽  
Peptidoglycan Synthesis ◽  
Division Site ◽  
A Cell ◽  
Cell Division Protein

ABSTRACT Cell division is the ultimate process for the propagation of bacteria, and FtsZ is an essential protein used by nearly all bacteria for this function. Chlamydiae belong to a small group of bacteria that lack the universal cell division protein FtsZ but still divide by binary fission. Chlamydial MreB is a member of the shape-determining MreB/Mbl family of proteins responsible for rod shape morphology in Escherichia coli. Chlamydia also encodes a homolog of RodZ, an MreB assembly cytoskeletal protein that links MreB to cell wall synthesis proteins. We hypothesized that MreB directs cell division in Chlamydia and that chlamydial MreB could replace FtsZ function for cell division in E. coli. Overexpression of chlamydial mreB-rodZ in E. coli induced prominent morphological changes with production of large swollen or oval bacteria, eventually resulting in bacterial lysis. Low-level expression of chlamydial mreB-rodZ restored viability of a lethal ΔmreB mutation in E. coli, although the bacteria lost their typical rod shape and grew as rounded cells. When FtsZ activity was inhibited by overexpression of SulA in the ΔmreB mutant of E. coli complemented with chlamydial mreB-rodZ, spherical E. coli grew and divided. Localization studies using a fluorescent fusion chlamydial MreB protein indicated that chlamydial RodZ directs chlamydial MreB to the E. coli division septum. These results demonstrate that chlamydial MreB, in partnership with chlamydial RodZ, acts as a cell division protein. Our findings suggest that an mreB-rodZ-based mechanism allows Chlamydia to divide without the universal division protein FtsZ. IMPORTANCE The study of Chlamydia growth and cell division is complicated by its obligate intracellular nature and biphasic lifestyle. Chlamydia also lacks the universal division protein FtsZ. We employed the cell division system of Escherichia coli as a surrogate to identify chlamydial cell division proteins. We demonstrate that chlamydial MreB, together with chlamydial RodZ, forms a cell division and growth complex that can replace FtsZ activity and support cell division in E. coli. Chlamydial RodZ plays a major role in directing chlamydial MreB localization to the cell division site. It is likely that the evolution of chlamydial MreB and RodZ to form a functional cell division complex allowed Chlamydia to dispense with its FtsZ-based cell division machinery during genome reduction. Thus, MreB-RodZ represents a possible mechanism for cell division in other bacteria lacking FtsZ.

scholarly journals MhpA Is a Hydroxylase Catalyzing the Initial Reaction of 3-(3-Hydroxyphenyl)Propionate Catabolism in Escherichia coli K-12

2019 ◽  
Vol 86 (4) ◽  
Author(s):  
Ying Xu ◽  
Ning-Yi Zhou
Keyword(s):  
Escherichia Coli ◽  
Liquid Chromatography ◽  
High Performance ◽  
Initial Reaction ◽  
Functional Identification ◽  
Bacterial Strains ◽  
Content Type ◽  
E Coli ◽  
C Terminus ◽  
K 12

ABSTRACT Escherichia coli K-12 and some other strains have been reported to be capable of utilizing 3-(3-hydroxyphenyl)propionate (3HPP), one of the phenylpropanoids from lignin. Although other enzymes involved in 3HPP catabolism and their corresponding genes from its degraders have been identified, 3HPP 2-hydroxylase, catalyzing the first step of its catabolism, has yet to be functionally identified at biochemical and genetic levels. In this study, we investigated the function and characteristics of MhpA from E. coli strain K-12 (MhpAK-12). Gene deletion and complementation showed that mhpA was vital for its growth on 3HPP, but the mhpA deletion strain was still able to grow on 3-(2,3-dihydroxyphenyl)propionate (DHPP), the hydroxylation product transformed from 3HPP by MhpAK-12. MhpAK-12 was overexpressed and purified, and it was likely a polymer and tightly bound with an approximately equal number of moles of FAD. Using NADH or NADPH as a cofactor, purified MhpAK-12 catalyzed the conversion of 3HPP to DHPP at a similar efficiency. The conversion from 3HPP to DHPP by purified MhpAK-12 was confirmed using high-performance liquid chromatography and liquid chromatography-mass spectrometry. Bioinformatics analysis indicated that MhpAK-12 and its putative homologues belonged to taxa that were phylogenetically distant from functionally identified FAD-containing monooxygenases (hydroxylases). Interestingly, MhpAK-12 has approximately an extra 150 residues at its C terminus in comparison to its close homologues, but its truncated versions MhpAK-12400 and MhpAK-12480 (with 154 and 74 residues deleted from the C terminus, respectively) both lost their activities. Thus, MhpAK-12 has been confirmed to be a 3HPP 2-hydroxylase catalyzing the conversion of 3HPP to DHPP, the initial reaction of 3HPP degradation. IMPORTANCE Phenylpropionate and its hydroxylated derivatives resulted from lignin degradation ubiquitously exist on the Earth. A number of bacterial strains have the ability to grow on 3HPP, one of the above derivatives. The hydroxylation was thought to be the initial and vital step for its aerobic catabolism via the meta pathway. The significance of our research is the functional identification and characterization of the purified 3HPP 2-hydroxylase MhpA from Escherichia coli K-12 at biochemical and genetic levels, since this enzyme has not previously been expressed from its encoding gene, purified, and characterized in any bacteria. It will not only fill a gap in our understanding of 3HPP 2-hydroxylase and its corresponding gene for the critical step in microbial 3HPP catabolism but also provide another example of the diversity of microbial degradation of plant-derived phenylpropionate and its hydroxylated derivatives.

scholarly journals Deficiency in l-Serine Deaminase Interferes with One-Carbon Metabolism and Cell Wall Synthesis in Escherichia coli K-12

2010 ◽  
Vol 192 (20) ◽  
pp. 5515-5525 ◽  
Author(s):  
Xiao Zhang ◽  
Ziad W. El-Hajj ◽  
Elaine Newman
Keyword(s):  
Escherichia Coli ◽  
Cell Wall ◽  
Cell Division ◽  
Amino Acid ◽  
Cell Wall Synthesis ◽  
E Coli ◽  
Amino Acid Mixtures ◽  
Glycine Cleavage ◽  
High Level ◽  
K 12

ABSTRACT Escherichia coli K-12 provided with glucose and a mixture of amino acids depletes l-serine more quickly than any other amino acid even in the presence of ammonium sulfate. A mutant without three 4Fe4S l-serine deaminases (SdaA, SdaB, and TdcG) of E. coli K-12 is unable to do this. The high level of l-serine that accumulates when such a mutant is exposed to amino acid mixtures starves the cells for C1 units and interferes with cell wall synthesis. We suggest that at high concentrations, l-serine decreases synthesis of UDP-N-acetylmuramate-l-alanine by the murC-encoded ligase, weakening the cell wall and producing misshapen cells and lysis. The inhibition by high l-serine is overcome in several ways: by a large concentration of l-alanine, by overproducing MurC together with a low concentration of l-alanine, and by overproducing FtsW, thus promoting septal assembly and also by overexpression of the glycine cleavage operon. S-Adenosylmethionine reduces lysis and allows an extensive increase in biomass without improving cell division. This suggests that E. coli has a metabolic trigger for cell division. Without that reaction, if no other inhibition occurs, other metabolic functions can continue and cells can elongate and replicate their DNA, reaching at least 180 times their usual length, but cannot divide.

Biodecomposition of Phenanthrene and Pyrene by a Genetically Engineered Escherichia coli

2020 ◽  
Vol 14 (2) ◽  
pp. 121-133 ◽  
Author(s):  
Maryam Ahankoub ◽  
Gashtasb Mardani ◽  
Payam Ghasemi-Dehkordi ◽  
Ameneh Mehri-Ghahfarrokhi ◽  
Abbas Doosti ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
High Performance ◽  
Human Cancer ◽  
Genetically Engineered ◽  
Hazardous Substances ◽  
E Coli ◽  
Spiked Soil ◽  
Engineered Escherichia Coli ◽  
Genetically Manipulated ◽  
Hplc Measurement

Background: Genetically engineered microorganisms (GEMs) can be used for bioremediation of the biological pollutants into nonhazardous or less-hazardous substances, at lower cost. Polycyclic aromatic hydrocarbons (PAHs) are one of these contaminants that associated with a risk of human cancer development. Genetically engineered E. coli that encoded catechol 2,3- dioxygenase (C230) was created and investigated its ability to biodecomposition of phenanthrene and pyrene in spiked soil using high-performance liquid chromatography (HPLC) measurement. We revised patents documents relating to the use of GEMs for bioremediation. This approach have already been done in others studies although using other genes codifying for same catechol degradation approach. Objective: In this study, we investigated biodecomposition of phenanthrene and pyrene by a genetically engineered Escherichia coli. Methods: Briefly, following the cloning of C230 gene (nahH) into pUC18 vector and transformation into E. coli Top10F, the complementary tests, including catalase, oxidase and PCR were used as on isolated bacteria from spiked soil. Results: The results of HPLC measurement showed that in spiked soil containing engineered E. coli, biodegradation of phenanthrene and pyrene comparing to autoclaved soil that inoculated by wild type of E. coli and normal soil group with natural microbial flora, were statistically significant (p<0.05). Moreover, catalase test was positive while the oxidase tests were negative. Conclusion: These findings indicated that genetically manipulated E. coli can provide an effective clean-up process on PAH compounds and it is useful for bioremediation of environmental pollution with petrochemical products.

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