scholarly journals The Moraxella catarrhalis Porin-Like Outer Membrane Protein CD Is an Adhesin for Human Lung Cells

2004 ◽  
Vol 72 (4) ◽  
pp. 1906-1913 ◽  
Author(s):  
Melissa M. Holm ◽  
Serena L. Vanlerberg ◽  
Ian M. Foley ◽  
Darren D. Sledjeski ◽  
Eric R. Lafontaine
Keyword(s):  
Membrane Protein ◽  
Outer Membrane ◽  
Gene Product ◽  
Outer Membrane Protein ◽  
Moraxella Catarrhalis ◽  
Human Lung ◽  
A549 Cells ◽  
Vaccine Antigen ◽  
Lung Cells ◽  
Human Lung Cells

ABSTRACT The outer membrane protein CD (OMPCD) of Moraxella catarrhalis is an outer membrane protein with several attributes of a potential vaccine antigen. We isolated four transposon mutants of strain O35E on the basis of their reduced binding to A549 human lung cells in microcolony formation assays, and we determined that they contain a transposon in ompCD. We also found that these transposon insertions had pleiotropic effects: mutants grew slower, became serum sensitive, bound ∼10-fold less to A549 cells, and appeared transparent when grown on solid medium. We confirmed that these various phenotypes could be attributed solely to disruption of ompCD by constructing the isogenic strain O35E.CD1. O35E-ompCD was cloned, and recombinant Escherichia coli bacteria expressing the gene product exhibited a 10-fold increase in adherence to A549 cells. This is the first report of M. catarrhalis ompCD mutants, and our findings demonstrate that this gene product is an adhesin for human lung cells.

scholarly journals Identification of aFrancisella tularensisLVS outer membrane protein that confers adherence to A549 human lung cells

2006 ◽  
Vol 263 (1) ◽  
pp. 102-108 ◽  
Author(s):  
Amanda Melillo ◽  
Darren D. Sledjeski ◽  
Serena Lipski ◽  
Ronald Mark Wooten ◽  
Venkatesha Basrur ◽  
...  
Keyword(s):  
Membrane Protein ◽  
Outer Membrane ◽  
Outer Membrane Protein ◽  
Human Lung ◽  
Lung Cells ◽  
Human Lung Cells

scholarly journals Moraxella catarrhalis Outer Membrane Protein CD Elicits Antibodies That Inhibit CD Binding to Human Mucin and Enhance Pulmonary Clearance of M. catarrhalis in a Mouse Model

2007 ◽  
Vol 75 (6) ◽  
pp. 2818-2825 ◽  
Author(s):  
Dai-Fang Liu ◽  
John C. McMichael ◽  
Steven M. Baker
Keyword(s):  
Membrane Protein ◽  
Outer Membrane ◽  
Outer Membrane Protein ◽  
Moraxella Catarrhalis ◽  
Signal Sequence ◽  
Vaccine Antigen ◽  
Enzyme Linked Immunosorbent Assay ◽  
Dependent Manner ◽  
Bacterial Clearance ◽  
Vitro Assay

ABSTRACT The outer membrane protein CD of Moraxella catarrhalis is considered to be a potential vaccine antigen against Moraxella infection. We purified the native CD from isolate O35E, administered it to mice, and detected considerable titers of anti-CD antibodies. Anti-CD sera were cross-reactive towards six different M. catarrhalis isolates and promoted bacterial clearance of O35E in a pulmonary challenge model. To circumvent the difficulty of generating large quantities of CD from M. catarrhalis for vaccine use, the CD gene from O35E was cloned into Escherichia coli, and the recombinant CD, expressed without a signal sequence or fusion tags, represented ∼70% of the total E. coli proteins. The recombinant CD formed inclusion bodies that were solubilized with 6 M urea and then purified by ion-exchange chromatography, a procedure that produced soluble CD of high purity and yield. Mice immunized with the purified recombinant CD had significant titers of anti-CD antibodies that were cross-reactive towards 24 different M. catarrhalis isolates. Upon challenge, these mice showed enhanced bacterial clearance of both O35E and a heterologous M. catarrhalis isolate, TTA24. In an in vitro assay, antisera to either the native or the recombinant CD inhibited the binding activity of CD to human tracheobronchial mucin in a serum concentration-dependent manner, and the extent of inhibition appeared to correlate with the corresponding anti-CD antibody titer and whole-cell enzyme-linked immunosorbent assay titer. Our results demonstrate that the recombinant CD is a promising vaccine candidate for preventing Moraxella infection.

scholarly journals Conservation of Outer Membrane Protein E among Strains of Moraxella catarrhalis

2001 ◽  
Vol 69 (6) ◽  
pp. 3576-3580 ◽  
Author(s):  
Timothy F. Murphy ◽  
Aimee L. Brauer ◽  
Norine Yuskiw ◽  
Erin R. McNamara ◽  
Charmaine Kirkham
Keyword(s):  
Monoclonal Antibodies ◽  
Membrane Protein ◽  
Respiratory Tract ◽  
Outer Membrane ◽  
Outer Membrane Protein ◽  
Moraxella Catarrhalis ◽  
Vaccine Antigen ◽  
Effective Vaccine ◽  
Human Respiratory Tract ◽  
The Stability

ABSTRACT Outer membrane protein E (OMP E) is a 50-kDa protein ofMoraxella catarrhalis which has several features that suggest that the protein may be an effective vaccine antigen. To assess the conservation of OMP E among strains of M. catarrhalis,22 isolates were studied with eight monoclonal antibodies which recognize epitopes on different regions of the protein. Eighteen of 22 strains were reactive with all eight antibodies. The sequences ofompE from 16 strains of M. catarrhalis were determined, including the 4 strains which were nonreactive with selected monoclonal antibodies. Analysis of sequences indicate a high degree of conservation among strains, with sequence differences clustered in limited regions of the gene. To assess the stability ofompE during colonization of the human respiratory tract, the sequences of ompE of isolates collected from patients colonized with the same strain for 3 to 9 months were determined. The sequences remained unchanged. These results indicate that OMP E is highly conserved among strains of M. catarrhalis, and preliminary studies indicate that the gene which encodes OMP E remains stable during colonization of the human respiratory tract.

scholarly journals Intranasal Vaccination with Recombinant Outer Membrane Protein CD and Adamantylamide Dipeptide as the Mucosal Adjuvant Enhances Pulmonary Clearance of Moraxella catarrhalis in an Experimental Murine Model

2006 ◽  
Vol 75 (4) ◽  
pp. 1778-1784 ◽  
Author(s):  
Pablo D. Becker ◽  
Gustavo M. Bertot ◽  
David Souss ◽  
Thomas Ebensen ◽  
Carlos A. Guzmán ◽  
...  
Keyword(s):  
Membrane Protein ◽  
Outer Membrane ◽  
Outer Membrane Protein ◽  
Moraxella Catarrhalis ◽  
Interleukin 2 ◽  
Immunoglobulin A ◽  
Acute Otitis Media ◽  
Vaccine Antigen ◽  
Mucosal Adjuvant ◽  
Tract Infections

ABSTRACT Moraxella catarrhalis causes acute otitis media in children and lower respiratory tract infections in adults and elderly. In children the presence of antibodies against the highly conserved outer membrane protein CD correlates with protection against infection, suggesting that this protein may be useful as a vaccine antigen. However, native CD is difficult to purify, and it is still unclear if recombinant CD (rCD) is a valid alternative. We performed a side-by-side comparison of the immunogenicities and efficacies of vaccine formulations containing native CD and rCD with adamantylamide dipeptide as the mucosal adjuvant. Intranasal vaccination of mice stimulated the production of high CD-specific antibody titers in sera and of secretory immunoglobulin A in mucosal lavages, which cross-recognized both antigens. While vaccination with native CD increased the number of interleukin-2 (IL-2)- and gamma interferon-producing cells, rCD mainly stimulated IL-4-secreting cells. Nevertheless, efficient bacterial clearance was observed in the lungs of challenged mice receiving native CD and in the lungs of challenged mice receiving rCD (96% and 99%, respectively). Thus, rCD is a promising candidate for incorporation in vaccine formulations for use against M. catarrhalis.

scholarly journals Antigenic Structure of Outer Membrane Protein E ofMoraxella catarrhalis and Construction and Characterization of Mutants

2000 ◽  
Vol 68 (11) ◽  
pp. 6250-6256 ◽  
Author(s):  
Timothy F. Murphy ◽  
Aimee L. Brauer ◽  
Norine Yuskiw ◽  
Thomas J. Hiltke
Keyword(s):  
Monoclonal Antibodies ◽  
Membrane Protein ◽  
Outer Membrane ◽  
Outer Membrane Protein ◽  
Vaccine Antigen ◽  
Antigenic Structure ◽  
Effective Vaccine ◽  
Fusion Peptides ◽  
Bacterial Surface ◽  
Normal Human

ABSTRACT Outer membrane protein E (OMP E) is a 50-kDa protein ofMoraxella catarrhalis which possesses several characteristics indicating that the protein will be an effective vaccine antigen. To study the antigenic structure of OMP E, eight monoclonal antibodies were developed and characterized. Three of the antibodies recognized epitopes which are present on the bacterial surface. Fusion peptides corresponding to overlapping regions of OMP E were constructed, and immunoblot assays were performed to localize the areas of the molecule bound by the monoclonal antibodies. These studies identified a surface-exposed epitope in the region of amino acids 80 through 180. To further study the protein, two mutants which lack OMP E were constructed. In bactericidal assays, the mutants were more readily killed by normal human serum compared to the isogenic parent strains. These results indicate that OMP E is involved in the expression of serum resistance of M. catarrhalis.

scholarly journals The Hag Protein of Moraxella catarrhalis Strain O35E Is Associated with Adherence to Human Lung and Middle Ear Cells

2003 ◽  
Vol 71 (9) ◽  
pp. 4977-4984 ◽  
Author(s):  
Melissa M. Holm ◽  
Serena L. Vanlerberg ◽  
Darren D. Sledjeski ◽  
Eric R. Lafontaine
Keyword(s):  
Cell Lines ◽  
Middle Ear ◽  
Moraxella Catarrhalis ◽  
Human Lung ◽  
Primary Cultures ◽  
A549 Cells ◽  
Pcr Analysis ◽  
Nucleotide Sequence Data ◽  
Human Lung Cells ◽  
Conjunctival Cells

ABSTRACT Previous studies have demonstrated that the Moraxella catarrhalis surface antigen UspA1 is an adhesin for Chang human conjunctival cells. The present report demonstrates that lack of UspA1 expression does not affect the adherence of strain O35E to A549 human lung cells or primary cultures of human middle ear epithelial (HMEE) cells. These results imply that another molecule mediates the adherence of M. catarrhalis to these two cell lines. To identify this adhesin, strain O35E was mutagenized with a transposon and 1,000 mutants were screened in a microcolony formation assay using A549 cells. Nine independent isolates exhibited an 8- to 19-fold reduction in adherence and contained a transposon in the same locus. Nucleotide sequence data and PCR analysis indicated that the transposons were inserted in different locations in the gene encoding the surface protein Hag. Quantitative assays using one representative transposon mutant, O35E.TN2, showed considerably decreased binding to A549 as well as HMEE cells. However, this mutant adhered at wild-type levels to Chang conjunctival cells. These findings suggest that the M. catarrhalis Hag protein is an adhesin for cell lines derived from human lung and middle ear tissues.

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