THE PORIN PROTEIN OF THE OUTER MEMBRANE OF ESCHERICHIA COLI: REACTIVITY IN IMMUNOBLOTTING, ANTIBODY-BINDING BY THE NATIVE PROTEIN, AND CROSS-REACTIVITY WITH OTHER ENTERIC BACTERIA

2009 ◽  
Vol 95B (1-6) ◽  
pp. 315-321
Author(s):  
ARNE Z. Henriksen ◽  
JOHAN A. MaeLand
Keyword(s):  
Escherichia Coli ◽  
Outer Membrane ◽  
Enteric Bacteria ◽  
Cross Reactivity ◽  
Antibody Binding ◽  
Native Protein ◽  
Porin Protein

scholarly journals Outer Membrane Permeability Barrier in Escherichia coli Mutants That Are Defective in the Late Acyltransferases of Lipid A Biosynthesis

1999 ◽  
Vol 43 (6) ◽  
pp. 1459-1462 ◽  
Author(s):  
Martti Vaara ◽  
Marjatta Nurminen
Keyword(s):  
Escherichia Coli ◽  
Membrane Permeability ◽  
Outer Membrane ◽  
Lipid A ◽  
Enteric Bacteria ◽  
Normal Function ◽  
Permeability Barrier ◽  
Core Oligosaccharide ◽  
Hydrophobic Solutes ◽  
Outer Membrane Permeability

ABSTRACT The tight packing of six fatty acids in the lipid A constituent of lipopolysaccharide (LPS) has been proposed to contribute to the unusually low permeability of the outer membrane of gram-negative enteric bacteria to hydrophobic antibiotics. Here it is shown that theEscherichia coli msbB mutant, which elaborates defective, penta-acylated lipid A, is practically as resistant to a representative set of hydrophobic solutes (rifampin, fusidic acid, erythromycin, clindamycin, and azithromycin) as the parent-type control strain. The susceptibility index, i.e., the approximate ratio between the MIC for the msbB mutant and that for the parent-type control, was maximally 2.7-fold. In comparison, the rfa mutant defective in the deep core oligosaccharide part of LPS displayed indices ranging from 20 to 64. The lpxA and lpxD lipid A mutants had indices higher than 512. Furthermore, the msbBmutant was resistant to glycopeptides (vancomycin, teicoplanin), whereas the rfa, lpxA, and lpxDmutants were susceptible. The msbB htrB double mutant, which elaborates even-more-defective, partially tetra-acylated lipid A, was still less susceptible than the rfa mutant. These findings indicate that hexa-acylated lipid A is not a prerequisite for the normal function of the outer membrane permeability barrier.

scholarly journals Immunization with the Recombinant PorB Outer Membrane Protein Induces a Bactericidal Immune Response against Neisseria meningitidis

2002 ◽  
Vol 70 (8) ◽  
pp. 4028-4034 ◽  
Author(s):  
J. Claire Wright ◽  
Jeannette N. Williams ◽  
Myron Christodoulides ◽  
John E. Heckels
Keyword(s):  
Recombinant Protein ◽  
Neisseria Meningitidis ◽  
Outer Membrane ◽  
Lipid A ◽  
Meningococcal Infection ◽  
Native Protein ◽  
Monophosphoryl Lipid A ◽  
Porin Protein ◽  
Life Threatening ◽  
Porin Proteins

ABSTRACT Infections with Neisseria meningitidis are characterized by life-threatening meningitis and septicemia. The meningococcal porin proteins from serogroup B meningococci have been identified as candidates for inclusion in vaccines to prevent such infections. In this study, we investigated the vaccine potential of the PorB porin protein free of other meningococcal components. The porB gene from a strain of Neisseria meningitidis expressing the class 3 outer membrane porin protein (PorB3) was cloned into the pRSETB vector, and the protein was expressed at high levels in a heterologous host Escherichia coli. The recombinant protein was purified to homogeneity by affinity chromatography and used for immunization after incorporation into liposomes and into micelles composed either of zwitterionic detergent or nondetergent sulfobetaine. The immunogenicity of these preparations was compared to recombinant PorB protein adsorbed to Al(OH)3 adjuvant as a control. Although sera raised against the protein adsorbed to Al(OH)3 reacted with the purified recombinant protein, sera raised against liposomes and micelles showed greater activity with native protein, as measured by enzyme immunoassay with outer membranes and by whole-cell immunofluorescence. Reactivity with native protein was considerably enhanced by incorporation of the adjuvant monophosphoryl lipid A into the liposome or micelle preparations. Recognition of the native protein was in a serotype-specific manner and was associated with the ability of the antisera to promote high levels of serotype-specific complement-mediated killing of meningococci. These results demonstrate that the PorB protein should be considered as a component of a vaccine designed to prevent serogroup B meningococcal infection.

Phosphoenolpyruvate-dependent sugar phosphotransferase activity in Megasphaera elsdenii

1981 ◽  
Vol 27 (9) ◽  
pp. 949-952 ◽  
Author(s):  
Steven S. Dills ◽  
Catherine A. Lee ◽  
Milton H. Saier Jr.
Keyword(s):  
Escherichia Coli ◽  
Stationary Phase ◽  
Enteric Bacteria ◽  
Cross Reactivity ◽  
Physical Characteristics ◽  
Sugar Uptake ◽  
Phosphotransferase System ◽  
Substrate Specificities ◽  
Megasphaera Elsdenii

Megasphaera elsdenii was found to possess an inducible phosphoenolpyruvate:sugar phosphotransferase system for glucose and fructose but not for other hexoses, hexosamines, or hexitols. The complexity of the Megasphaera phosphoenolpyruvate:sugar phosphotransferase system lies intermediate to systems found in photoautotrophic bacteria and enteric bacteria. Megasphaera elsdenii phosphotransferase proteins exhibited enzymatic cross-reactivity with those from Escherichia coli; however, differences were found in substrate specificities and the physical characteristics of the proteins from these organisms. Sugar uptake was reduced in M. elsdenii stationary-phase as compared with log-phase cells and this loss correlated with a reduction in enzyme II function.

scholarly journals Identification of Murine B-Cell and T-Cell Epitopes of Escherichia coli Outer Membrane Protein F with Synthetic Polypeptides

2000 ◽  
Vol 68 (5) ◽  
pp. 2535-2545 ◽  
Author(s):  
Kristina M. Williams ◽  
Elmer C. Bigley ◽  
Richard B. Raybourne
Keyword(s):  
Escherichia Coli ◽  
T Cell ◽  
B Cell ◽  
Outer Membrane ◽  
Synthetic Peptides ◽  
Outer Membrane Proteins ◽  
T Cell Epitopes ◽  
Native Protein ◽  
Cell Responses ◽  
Synthetic Polypeptides

ABSTRACT The major pore-forming outer membrane proteins (Omps) of gram-negative bacteria demonstrate numerous immunomodulating properties and are involved in the virulence of pathogenic strains. BecauseEscherichia coli OmpF is the best-characterized porin in terms of structural and functional characteristics, in vitro B-cell and T-cell responses to this porin in six different strains of mice were analyzed. Mice were immunized with purified OmpF trimers or overlapping synthetic polypeptides (20-mers) spanning the entire 340-amino-acid sequence of the OmpF monomer. T-cell proliferative responses and immunoglobulin G antibody responses to native OmpF and the peptide analogues were determined. For each strain, patterns of T-cell proliferation were similar regardless of whether native OmpF or synthetic peptides were inoculated, although all strains recognized one or more cryptic determinants. Mice exhibited several haplotype-specific responses, but genetically permissive epitopes were also identified. Four peptides (75-94, 265-284, 295-314, and 305-324) elicited strong T-cell proliferative responses from all strains of mice when mice were presensitized with native OmpF or a homologous peptide. In general, 10 or fewer peptides were recognized by sera from mice immunized with native OmpF or synthetic peptides, and most sera from peptide-immunized mice reacted poorly with the native protein. Four peptides spanning amino acids 45 to 64, 95 to 114, 115 to 134, and 275 to 294 were recognized by sera from all strains immunized with native OmpF but not by sera from peptide-immunized mice. Peptides 245-264 and 305-324 were universally recognized by sera from peptide-immunized mice, but these sera reacted weakly or were negative when tested against the native protein. Based on the pattern of cytokine secretion by proliferating T cells, immunization with native OmpF polarizes T helper cells toward development of a TH1 response. T-cell and B-cell responses have been investigated based on the assumption that differences in epitope specificity could influence protective or pathologic host reactions. Because of the high level of structural homology of OmpF to porins isolated from other enteric pathogens, the identification of T- and B-cell-stimulatory determinants of E. coli OmpF may have broader application.

scholarly journals Expression in Escherichia coli and function of Pseudomonas aeruginosa outer membrane porin protein F.

1986 ◽  
Vol 167 (2) ◽  
pp. 473-479 ◽  
Author(s):  
W A Woodruff ◽  
T R Parr ◽  
R E Hancock ◽  
L F Hanne ◽  
T I Nicas ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Pseudomonas Aeruginosa ◽  
Outer Membrane ◽  
Expression In Escherichia Coli ◽  
Porin Protein ◽  
Outer Membrane Porin ◽  
Protein F ◽  
And Function

scholarly journals Association of tetrapyrrole intermediates in the bacteriochlorophyll a biosynthetic pathway with the major outer-membrane porin protein of Rhodobacter capsulatus

1992 ◽  
Vol 282 (2) ◽  
pp. 471-476 ◽  
Author(s):  
D W Bollivar ◽  
C E Bauer
Keyword(s):  
Outer Membrane ◽  
Electrophoretic Mobility ◽  
Biosynthetic Pathway ◽  
Rhodobacter Capsulatus ◽  
Cross Reactivity ◽  
Bacteriochlorophyll A ◽  
Early Model ◽  
Porin Protein ◽  
Outer Membrane Porin ◽  
Polyclonal Antisera

Rhodobacter capsulatus regulates synthesis of bacteriochlorophyll a in response to changes in oxygen partial pressure and light intensity. One early model proposed that this regulation involved a carrier polypeptide that functions to tether tetrapyrrole intermediates to the membrane. In the present study we isolated tetrapyrrole intermediates accumulated in three strains of R. capsulatus that contain mutations which block bacteriochlorophyll a biosynthesis at different steps of the magnesium branch of the pathway. Each of the tetrapyrrole intermediates was shown to be associated with the same 32 kDa polypeptide, as indicated by similar electrophoretic mobility and antigenic cross-reactivity with polyclonal antisera. The 32 kDa pigment-associated protein was further found to have an electrophoretic mobility, antigenic cross-reactivity and N-terminal sequence identical with those of the previously characterized major outer-membrane porin protein of R. capsulatus.

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