Involvement of outer membrane proteins in freeze–thaw resistance of Escherichia coli

1984 ◽  
Vol 30 (3) ◽  
pp. 339-344 ◽  
Author(s):  
Peter H. Calcott ◽  
Katherine N. Calcott
Keyword(s):  
Escherichia Coli ◽  
Outer Membrane ◽  
Outer Membrane Proteins ◽  
Membrane Damage ◽  
Wild Type ◽  
Freezing And Thawing ◽  
Freeze Thaw ◽  
Porin Proteins ◽  
Protein Components ◽  
Type Strains

Two families of Escherichia coli mutants with altered outer membrane protein components were examined for sensitivity to freezing and thawing and other stresses. A mutant unable to make the lipoprotein (lpo) was extremely sensitive to freezing and thawing in water or saline and to challenge with detergent, while the mutant unable to make the porin proteins (ompB) was more resistant than the isogenic wild type; strains unable to make the tsx and ompA proteins were slightly more sensitive to the stresses. Similarly, the lpo deficient strain exhibited more and the ompB less wall and membrane damage than the wild-type strains. Little difference in the extent of wall damage, but more membrane damage, was seen for the two tsx and the ompA strains when compared with the wild-type strain. The roles of the specific proteins in determining sensitivity to freeze–thaw are discussed.

Extrusion of actin-positive strands from HEp-2 and Int 407 cells caused by outer membrane preparations of enteropathogenicEscherichia coliand specific attachment of wild type bacteria to the strands

2001 ◽  
Vol 47 (8) ◽  
pp. 727-734 ◽  
Author(s):  
Sukumaran Sunil Kumar ◽  
Vasantha Malladi ◽  
Krishnan Sankaran ◽  
Richard Haigh ◽  
Peter Williams ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Membrane Proteins ◽  
Outer Membrane ◽  
Outer Membrane Proteins ◽  
Three Dimensional ◽  
Secretory Proteins ◽  
Enteropathogenic Escherichia Coli ◽  
Wild Type ◽  
Cytopathic Effects ◽  
Cytoskeletal Elements

Enteropathogenic Escherichia coli (EPEC) causes persistent infantile diarrhoea. This nontoxigenic E. coli exhibits a complicated pathogenic mechanism in which its outer membrane proteins and type III secretory proteins damage intestinal epithelium and cause diarrhoea. In accordance with this, our previous study using HEp-2 cells demonstrated cytopathic effects caused by cell-free outer membrane preparations of EPEC. In this study, we report the extrusion of actin-positive strands from HEp-2 and Int 407 cells when treated with outer membrane preparations. An interesting observation of this work, perhaps relevant to the characteristic localized three-dimensional colony formation of EPEC, is the attachment of a wild type EPEC strain to these actin-positive strands.Key words: enteropathogenic Escherichia coli, actin, outer membrane proteins, cytoskeletal elements.

scholarly journals Effect on solute size on diffusion rates through the transmembrane pores of the outer membrane of Escherichia coli.

1981 ◽  
Vol 77 (2) ◽  
pp. 121-135 ◽  
Author(s):  
H Nikaido ◽  
E Y Rosenberg
Keyword(s):  
Escherichia Coli ◽  
Outer Membrane ◽  
Specific Class ◽  
Wild Type ◽  
Intact Cells ◽  
E Coli ◽  
Low Concentrations ◽  
Transmembrane Pores ◽  
Diffusion Rates ◽  
Type Strains

Nutrients usually cross the outer membrane of Escherichia coli by diffusion through water-filled channels surrounded by a specific class of protein, porins. In this study, the rates of diffusion of hydrophilic nonelectrolytes, mostly sugars and sugar alcohols, through the porin channels were determined in two systems, (a) vesicles reconstituted from phospholipids and purified porin and (b) intact cells of mutant strains that produce many fewer porin molecules than wild-type strains. The diffusion rates were strongly affected by the size of the solute, even when the size was well within the "exclusion limit" of the channel. In both systems, hexoses and hexose disaccharides diffused through the channel at rates 50-80% and 2-4%, respectively, of that of a pentose, arabinose. Application of the Renkin equation to these data led to the estimate that the pore radius is approximately 0.6 nm, if the pore is assumed to be a hollow cylinder. The results of the study also show that the permeability of the outer membrane of the wild-type E. coli cell to glucose and lactose can be explained by the presence of porin channels, that a significant fraction of these channels must be functional or "open" under our conditions of growth, and that even 10(5) channels per cell could become limiting when E. coli tries to grow at a maximal rate on low concentrations of slowly penetrating solutes, such as disaccharides.

scholarly journals Additive and Synergistic Bactericidal Activity of Antibodies Directed against Minor Outer Membrane Proteins of Neisseria meningitidis

2007 ◽  
Vol 75 (11) ◽  
pp. 5434-5442 ◽  
Author(s):  
Vincent E. Weynants ◽  
Christiane M. Feron ◽  
Karine K. Goraj ◽  
Martine P. Bos ◽  
Philippe A. Denoël ◽  
...  
Keyword(s):  
Neisseria Meningitidis ◽  
Outer Membrane ◽  
Bactericidal Activity ◽  
Outer Membrane Proteins ◽  
Critical Density ◽  
Membrane Vesicles ◽  
Outer Membrane Vesicles ◽  
Wild Type ◽  
Major Variable ◽  
Type Strains

ABSTRACT Neisseria meningitidis serogroup B is a major cause of bacterial meningitis in younger populations. The available vaccines are based on outer membrane vesicles obtained from wild-type strains. In children less than 2 years old they confer protection only against strains expressing homologous PorA, a major, variable outer membrane protein (OMP). We genetically modified a strain in order to eliminate PorA and to overproduce one or several minor and conserved OMPs. Using a mouse model mimicking children's PorA-specific bactericidal activity, it was demonstrated that overproduction of more than one minor OMP is required to elicit antibodies able to induce complement-mediated killing of strains expressing heterologous PorA. It is concluded that a critical density of bactericidal antibodies needs to be reached at the surface of meningococci to induce complement-mediated killing. With minor OMPs, this threshold is reached when more than one antigen is targeted, and this allows cross-protection.

Proteolytic digestion and labelling studies of the organization of the proteins in the outer membrane of Escherichia coli

1977 ◽  
Vol 55 (10) ◽  
pp. 1082-1090 ◽  
Author(s):  
Reinhart A. F. Reithmeier ◽  
Philip D. Bragg
Keyword(s):  
Escherichia Coli ◽  
Outer Membrane ◽  
Hydrophobic Interactions ◽  
Proteolytic Digestion ◽  
Wild Type ◽  
Terminal Portion ◽  
Intact Cells ◽  
Protein Components ◽  
Pronase Treatment ◽  
Protein B

The arrangement of the proteins in the outer membrane of Escherichia coli was examined by treating intact cells and isolated membrane preparations with fluorescamine and with pronase. Intact wild-type cells, or those of a mutant in which the core region of the lipopolysaccharide was absent, were equally resistant to pronase treatment. The protein components of isolated outer membrane preparations varied in their rate of digestion and labelling with fluorescamine. The N-terminal portion of protein B was removed by pronase to yield a fragment (protein Bp) still embedded in the membrane. Protein Bp was not significantly enriched in nonpolar amino acids, suggesting that protein B may not be held in the membrane primarily by hydrophobic interactions. This was confirmed by reconstitution experiments in which protein B could be reassociated with itself, without lipopolysaccharide or phospholipid, in the presence of a divalent cation such that pronase digestion of the reassociated material gave protein Bp.

scholarly journals Effects of bfp Mutations on Biogenesis of Functional Enteropathogenic Escherichia coli Type IV Pili

2000 ◽  
Vol 182 (9) ◽  
pp. 2498-2506 ◽  
Author(s):  
Ravi P. Anantha ◽  
Kelly D. Stone ◽  
Michael S. Donnenberg
Keyword(s):  
Escherichia Coli ◽  
Outer Membrane ◽  
Signal Sequence ◽  
Outer Membrane Proteins ◽  
Laboratory Strain ◽  
Type Iv Pili ◽  
Host Cells ◽  
Wild Type ◽  
Type Iv ◽  
The Individual

ABSTRACT Enteropathogenic Escherichia coli expresses a type IV fimbria known as the bundle-forming pilus (BFP) that is required for autoaggregation and localized adherence (LA) to host cells. A cluster of 14 genes is sufficient to reconstitute BFP biogenesis in a laboratory strain of E. coli. We have undertaken a systematic mutagenesis of the individual genes to determine the effect of each mutation on BFP biogenesis and LA. Here we report the construction and analysis of nonpolar mutations in six genes of thebfp cluster, bfpG, bfpB,bfpC, bfpD, bfpP, andbfpH, as well as the further analysis of a previously described bfpA mutant strain that is unable to express bundlin, the pilin protein. We found that mutations inbfpB, which encodes an outer membrane protein;bfpD, which encodes a putative nucleotide-binding protein; and bfpG and bfpC, which do not have sequence homologues in other type IV pilus systems, do not affect prebundlin expression or processing but block both BFP biogenesis and LA. The mutation in bfpP, the prepilin peptidase gene, does not affect prebundlin expression but blocks signal sequence cleavage of prebundlin, BFP biogenesis, and LA. The mutation in bfpH, which is predicted to encode a lytic transglycosylase, has no effect on prebundlin expression, prebundlin processing, BFP biogenesis, or LA. For each mutant for which altered phenotypes were detected, complementation with a plasmid containing the corresponding wild-type allele restored the wild-type phenotypes. We also found that association of prebundlin or bundlin with sucrose density flotation gradient fractions containing both inner and outer membrane proteins does not require any accessory proteins. These studies indicate that many bfp gene products are required for biogenesis of functional type IV pili but that mutations in the individual genes do not lead to the identification of new phases of pilus assembly.

The survival of Escherichia coli from freeze–thaw damage: the relative importance of wall and membrane damage

1975 ◽  
Vol 21 (12) ◽  
pp. 1960-1968 ◽  
Author(s):  
P. H. Calcott ◽  
R. A. MacLeod
Keyword(s):  
Escherichia Coli ◽  
Cell Survival ◽  
Membrane Damage ◽  
Tween 80 ◽  
Relative Importance ◽  
Freezing And Thawing ◽  
Freeze Thaw

When Escherichia coli is frozen rapidly in saline and thawed slowly, survival is very low; however, the inclusion of 3% glycerol or 1% Tween 80 in the saline freezing menstruum results in near complete survival. The release of material from, and penetration of, substances into the cell indicate that both membrane and wall damage occur during freezing and thawing. Glycerol, under these conditions, is able to reduce severely both the damage to the wall and membrane, whereas Tween 80 prevents only membrane damage. This indicates that freezing and thawing in saline results in membrane damage which is lethal to the cell whereas wall damage which occurs is not detrimental to cell survival.

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