Lysis of Escherichia coli by induction of cloned ϕX174 genes

Henrich, Bernhard; Lubitz, Werner; Plapp, Roland

doi:10.1007/bf00334146

Cited by 123 publications

(90 citation statements)

References 18 publications

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“…The same injury then would allow the release of autolytic enzymes to the cell wall and initiate lysis of the bacteria. This model is formally analogous to models proposed for the mechanism of lysis by bacteriophages in which the release of phage progeny is preceded by membrane injury through the insertion of a holin protein into the bacterial membrane (32,33). Our model also is analogous to the proposed mode of action of nisin at the level of lipid II (34).…”

Section: Discussionmentioning

confidence: 52%

The murMN operon: A functional link between antibiotic resistance and antibiotic tolerance in Streptococcus pneumoniae

Filipe

Severina²,

Tomasz³

2002

Proc. Natl. Acad. Sci. U.S.A.

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Inactivation of the recently identified murMN operon in penicillinresistant strains of Streptococcus pneumoniae was shown already to cause two major effects: elimination of branched-structured muropeptides from the cell wall and complete loss of penicillin resistance. We now show that cells with inactivated murMN also have a third phenotype: an increased susceptibility to lysis when exposed to low concentrations of fosfomycin, D-cycloserine, vancomycin, and nisin, indicating a wide-spectrum hypersensitivity to inhibitors of both early and late stages of cell wall biosynthesis. Mutants of murMN also lysed faster than the parental strain when treated with the detergent deoxycholate. Several different alleles of murM cloned in plasmid pLS578 and introduced into a murM deletion mutant of the penicillin-resistant strain Pen6 were able to reconstitute each one of the three mutant phenotypes: the highly branched cell wall structure, original high level of penicillin resistance, and normal sensitivity to lysis. In a penicillin-susceptible strain the same experiments caused increased concentration of cell wall branched peptides and suppression of sensitivity to antibiotic induced lysis. The observations suggest that the murMN operon plays a key role in the regulation of a stress-response pathway that can be triggered by perturbation of cell wall biosynthesis in S. pneumoniae. S everal penicillin-resistant clones of Streptococcus pneumoniaeproduce abnormal cell walls in which a high proportion of the stem peptide lysine residues carry short (seryl alanine or alanyl alanine) branches (1). The genetic determinants involved with the branching process, the murMN operon, were identified recently (2). Penicillin-resistant isolates with highly branched cell walls were shown to carry murM alleles that contained divergent sequences as compared with the murM genes of penicillin-susceptible pneumococci (3). The two proteins, MurM (407 amino acids) and MurN (411 amino acids), are involved with the addition of the first and second amino acid residues, respectively, to the cell wall precursor stem peptide at the lipid II stage of biosynthesis (4, 5). Inactivation of murMN by insertion-duplication mutagenesis did not interfere with growth of the bacteria but caused two phenotypes: elimination of branched peptides from the cell wall and complete loss of penicillin resistance (2). The purpose of the studies described in this communication was to follow up these earlier findings by the use of a deletion mutant of murM and complementation experiments. The surprising observation made was that inactivation of murM also caused a third phenotype: increased sensitivity of the bacteria to lysis by cell wall inhibitors. Experimental ProceduresStrains and Growth Conditions. All strains and plasmids used in this study are listed in Table 1. S. pneumoniae strains were grown in a casein-based semisynthetic medium (C ϩ Y) at 37°C without aeration as described (11). S. pneumoniae strains containing pLS578 (10) or its derivatives were grown in the presen...

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Section: Discussionmentioning

confidence: 52%

The murMN operon: A functional link between antibiotic resistance and antibiotic tolerance in Streptococcus pneumoniae

Filipe

Severina²,

Tomasz³

2002

Proc. Natl. Acad. Sci. U.S.A.

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“…2,3 Later the temperature-sensitive λ-system (λp L /λp R -cI857) has proven to be more suitable for quick and efficient lysis without the need of any addition of chemical inducers. 18 Since the λ repressor cI857 shows incipient expression of downstream gene E at temperatures above 30°C the temperature sensitivity of the system was optimized to meet more favorable fermentation temperatures of 35°C or higher.…”

Section: Bg-production Processmentioning

confidence: 99%

“…1 More than 16 years later, when genetic engineering had been developed, it could be shown that its sole expression after cloning is sufficient to cause subsequent lysis of E. coli. 2,3 E was the first lethal gene for bacteria which could be silenced on plasmids. When established in non-host range bacteria of the phage expression of E converts Gram-negative bacteria into BGs whereas Gram-positive bacteria are killed without lysis.…”

Section: Introductionmentioning

confidence: 99%

The bacterial ghost platform system

Langemann¹,

et al. 2010

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The Bacterial Ghost (BG) platform technology is an innovative system for vaccine, drug or active substance delivery and for technical applications in white biotechnology. BGs are cell envelopes derived from Gram-negative bacteria. BGs are devoid of all cytoplasmic content but have a preserved cellular morphology including all cell surface structures. Using BGs as delivery vehicles for subunit or DNA-vaccines the particle structure and surface properties of BGs are targeting the carrier itself to primary antigenpresenting cells. Furthermore, BGs exhibit intrinsic adjuvant properties and trigger an enhanced humoral and cellular immune response to the target antigen. Multiple antigens of the native BG envelope and recombinant protein or DNA antigens can be combined in a single type of BG. Antigens can be presented on the inner or outer membrane of the BG as well as in the periplasm that is sealed during BG formation. Drugs or supplements can also be loaded to the internal lumen or periplasmic space of the carrier. BGs are produced by batch fermentation with subsequent product recovery and purification via tangential flow filtration. For safety reasons all residual bacterial DNA is inactivated during the BG production process by the use of staphylococcal nuclease A and/or the treatment with β-propiolactone. After purification BGs can be stored long-term at ambient room temperature as lyophilized product. The production cycle from the inoculation of the pre-culture to the purified BG concentrate ready for lyophilization does not take longer than a day and thus meets modern criteria of rapid vaccine production rather than keeping large stocks of vaccines. The broad spectrum of possible applications in combination with the comparably low production costs make the BG platform technology a safe and sophisticated product for the targeted delivery of vaccines and active agents as well as carrier of immobilized enzymes for applications in white biotechnology.

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“…[28][29][30] In this concept study the ice nucleation protein Z (InaZ) of Pseudomonas syringae was expressed in E. coli which were further processed to Bacterial Ghosts (BGs). BGs are empty cell envelopes of Gram-negative bacteria produced by the expression of cloned gene E of bacteriophage Phix174 [31][32][33][34][35] The conversion of a single living bacterium into a BG is a rapid process in the range of milliseconds 35 and can be characterized by puncturing the bacterium cell envelope from the inside to the outside expelling its cytoplasmic content to the surrounding medium. Electron micrographs of this process can be depicted from the original description in 1990 33 and a more recent one using this Emediated lysis of E. coli for cryo-electron tomography of membrane protein complexes within the native cell envelope complex remaining intact after E-mediated lysis.…”

Section: Introductionmentioning

confidence: 99%

Functional display of ice nucleation protein InaZ on the surface of bacterial ghosts

Kassmannhuber

Rauscher²,

Schöner³

et al. 2017

Bioengineered

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In a concept study the ability to induce heterogeneous ice formation by Bacterial Ghosts (BGs) from Escherichia coli carrying ice nucleation protein InaZ from Pseudomonas syringae in their outer membrane was investigated by a droplet-freezing assay of ultra-pure water. As determined by the median freezing temperature and cumulative ice nucleation spectra it could be demonstrated that both the living recombinant E. coli and their corresponding BGs functionally display InaZ on their surface. Under the production conditions chosen both samples belong to type II ice-nucleation particles inducing ice formation at a temperature range of between ¡5.6 C and ¡6.7 C, respectively. One advantage for the application of such BGs over their living recombinant mother bacteria is that they are non-living native cell envelopes retaining the biophysical properties of ice nucleation and do no longer represent genetically modified organisms (GMOs).

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Lysis of Escherichia coli by induction of cloned ϕX174 genes

Cited by 123 publications

References 18 publications

The murMN operon: A functional link between antibiotic resistance and antibiotic tolerance in Streptococcus pneumoniae

The murMN operon: A functional link between antibiotic resistance and antibiotic tolerance in Streptococcus pneumoniae

The bacterial ghost platform system

Functional display of ice nucleation protein InaZ on the surface of bacterial ghosts

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