Ca2+-induced permeabilization of the Escherichia coli outer membrane: comparison of transformation and reconstitution of binding-protein-dependent transport

Bukau, Bernd; Brass, J M; Boos, Winfried

doi:10.1128/jb.163.1.61-68.1985

Cited by 22 publications

(3 citation statements)

References 35 publications

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“…In addition to the role of glycine in structural impairment, Kurkdjian et al (1993) reported that glycine acts as an osmoticum and increases the permeability of the membrane. Hence, the addition of glycine to the transformation mixture helps the uptake of exogenous DNA present in the transformation mixture (Bukau et al, 1985). Thus, the increased transformation efficiencies observed in presence of sucrose and glycine, as shown in Table 2, might be due to structural impairments of the bacterial cell wall; furthermore, the osmotic agents might have facilitated DNA uptake.…”

Section: Discussionmentioning

confidence: 99%

“…The combination of chemical-chemical, chemicalphysical, and physical-physical methods improves the transformation manifolds and is used in laboratories worldwide. Bukau et al (1985) suggested that growth medium supplemented with sucrose facilitates DNA uptake. Transformation efficiency of E. coli may also be enhanced by the use of the simplest amino acid, glycine (1% w/v), in growth medium, as reported by Akhtar et al (1999).…”

Section: Introductionmentioning

confidence: 99%

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Optimization of E. coli culture conditions for efficient DNA uptake by electroporation

Ahmad¹,

Rubbab

Deeba³

et al. 2014

Turk J Biol

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Bacterial transformation is a significant breakthrough in the field of molecular biology for cloning purposes. A variety of chemical and physical methods are utilized for enhancing bacterial competence. The present study was designed, via the consolidation of chemical and physical methods, to improve competence of cells for high transformation efficiency up to 10 9 transformants/µg of DNA. LB and SOC media and their supplemented combinations with osmotic agents (sucrose and glycine) were used to prepare electrocompetent cells of E. coli DH5α at temperatures of 25 and 37 °C, which were then subjected to electroporation. The results showed that cells grown in SOC medium showed high growth rates and increased transformation efficiencies as compared to LB, while media supplemented with osmotic agents also showed higher transformation proficiency as compared to the controls. In all combinations tested, the highest transformation efficiencies were observed in the cells grown in SOC-SG at 25 °C, i.e. 3.56 × 10 9 cfu/µg of DNA. This study provides valuable knowledge for low-budget biotechnology labs to prepare competent cells with efficiency comparable to commercially available competent cells.

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Optimization of E. coli culture conditions for efficient DNA uptake by electroporation

Ahmad¹,

Rubbab

Deeba³

et al. 2014

Turk J Biol

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show abstract

“…The procedure developed in this report was based upon the ability of certain agents to alter the permeability properties of the outer membrane of gram-negative bacteria. Agents such as calcium ions, chelators, and certain antibiotics and heat or osmotic shock treatments have been used to alter the physical characteristics of the outer membrane (12,33). Molecular mechanisms to explain the loss of the membrane barrier are based upon the disruption of bacterial lipopolysaccharide (25,31,32), as reviewed by Vaara (43), but these explanations remain speculative.…”

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confidence: 99%

C9-mediated killing of bacterial cells by transferred C5b-8 complexes: transferred C5b-9 complexes are nonbactericidal

Blanchard

Dankert

1994

Infect Immun

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The formation of the CSb-9 complex on the outer membrane of complement-sensitive cells ofEscherichia coli results in inhibition of inner membrane function and the death of the cell. Cells bearing a precursor of the C5b-9 site, the C5b-8 complex, suffer no loss in viability. Antibiotic-sensitive, complement-sensitive donor cells bearing precursor C5b-8 complexes were incubated with equal numbers of antibiotic-resistant, complementsensitive acceptor cells that had not been exposed to a complement source. This cell mixture was incubated with 5 mM EDTA for 5 min and then with calcium chloride (20 mM) for various times. The excess calcium ion concentration was effectively reduced with additional EDTA, and the cell mixture was washed and resuspended in buffer. The viability of the acceptor cells was assayed by plating on antibiotic-containing media. C9 was added to the mixture, and the mixture was incubated for 10 min at 3rC and then plated as described above.It was found that the acceptor cells were killed by the addition of purified C9 only after incubation with donor cells bearing C5b-8 sites during the transfer procedure. This indicates that precursor CSb-8 sites that support C9-mediated killing could be transferred between cells. No loss in viability was detected for acceptor cells subjected to the procedure described above in the presence of donor cells bearing complete C5b-9 complexes, formed prior to mixing with acceptor cells for the transfer procedure.

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Calcium and the biological activities of two Streptomyces species isolated from the rhizosphere of soybean plants

Kamel

Khalil

Shalaby

1989

J. Basic Microbiol.

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Careful examination of the characteristics of both organisms revealed that we are dealing with Streptomyces violaceochromogenes and Streptomyces glaucescens. Calcium chloride decreased the permeability of both organisms as indicated by the low total nitrogen content of their media compared to the controls. Small doses of calcium nitrate suppressed the permeability of S. violaceochromogenes and increased that of S. glaucescens whereas the larger doses were stimulatory for both organisms. Calcium nitrate had similar effects on S. violaceochromogenes only without affecting S. glaucescens. The stimulatory effects of calcium gradually faded with increased concentration. Calcium chloride increased the proteolytic activity in S. violaceochromogenes media ; a phenomenon that was apparent in S. glaucescens media only in the case of the larger doses. Calcium nitrate suppressed the proteolytic activity in S. glaucescens media. The cellulolytic activity of both organisms was suppressed by calcium but amylases were initiated in S. glaucescens media. Those of S. violaceochromogenes were arrested with the lower levels of calcium but the larger doses restored or stimulated their activity.The metabolic activities of rhizospheric microflora play an important role in the growth of plants through supplying them with the mineralization products of organic matter. They may produce degrading enzymes, growth promoting or growth retarding substances for higher as well as lower plants (RAo 1977, HEcco 1984.The role of rhizospheric actinomycetes has received little attention. Streptomyces species are relatively good decomposers of organic matter (ALEXANDER 1977). KUSTER (1979) reported that a large number of actinomycetes, exhibiting different kinds of unusual physio-

show abstract

Ca2+-induced permeabilization of the Escherichia coli outer membrane: comparison of transformation and reconstitution of binding-protein-dependent transport

Cited by 22 publications

References 35 publications

Optimization of E. coli culture conditions for efficient DNA uptake by electroporation

Optimization of E. coli culture conditions for efficient DNA uptake by electroporation

C9-mediated killing of bacterial cells by transferred C5b-8 complexes: transferred C5b-9 complexes are nonbactericidal

Calcium and the biological activities of two Streptomyces species isolated from the rhizosphere of soybean plants

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