Molecular studies of Ssa1, a serotype-specific antigen of Pasteurella haemolytica A1

Lo, Reggie Y.C.; Strathdee, Craig A.; Shewen, Patricia E.; Cooney, B. J.

doi:10.1128/iai.59.10.3398-3406.1991

Cited by 31 publications

(22 citation statements)

References 29 publications

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“…The serotype-specific antigen Ssa1 is reportedly an outer membrane-associated protein that may be sloughed off during growth or cell lysis of P. haemolytica cultures (17,23). This antigenic peptide has an apparent molecular mass of 103 kDa as determined by SDS-PAGE analysis (23) and may correspond to the band with a molecular mass similar to that of leukotoxin detected in culture supernatants of strains 59B0071 and 59B0072. Alternatively, the band may correspond to the neutrophil chemoattractant detected by Brunner et al (6) in culture supernatants of P. haemolytica serotype 1.…”

Section: Discussionmentioning

confidence: 99%

Antigenic and virulence properties of Pasteurella haemolytica leukotoxin mutants

et al. 1995

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Antigenic properties of two mutants of Pasteurella haemolytica, strains 59B0071 and 59B0072, that do not produce detectable leukotoxin were investigated. Western blot (immunoblot) analysis with a number of polyclonal sera from animals recovering from pasteurellosis revealed that both mutants secreted a variety of antigens that were also present in cultures of several wild-type strains. These antigens ranged from about 100 to 15 kDa. Mutant strain 59B0071 was found to be totally deficient in leukotoxin, as judged not only by Western blotting but also by cytotoxicity assays with bovine lymphoma (BL-3) cells or bovine polymorphonuclear cells as targets. The mutant strain 59B0071 had normal levels of a secreted sialylglycoprotease, however. When strains were tested for virulence in goat and cattle challenge experiments, a reduction in mortality and lung lesions was observed with the mutant 59B0071 in comparison with results obtained with wild-type strains. These results are consistent with an important role for leukotoxin in P. haemolytica virulence and suggest that leukotoxin-negative mutants may be useful tools in the investigation of other virulence properties involved in P. haemolytica infections.

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Section: Discussionmentioning

confidence: 99%

Antigenic and virulence properties of Pasteurella haemolytica leukotoxin mutants

et al. 1995

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“…After SDS‐PAGE, the proteins were stained with Commassie brilliant blue. For Western immunoblot, the proteins were transferred to a nitrocellulose membrane as described (Lo et al , 1991), and blocked by immersion in a 3% gelatin solution in Tris‐HCl‐buffered saline containing 0.05% Tween 20 (TTBS). The Lkt neutralizing monoclonal antibody 601 (Gentry & Srikumaran, 1991) was used at a dilution of 1/2000 in antibody solution (1% gelatin in TTBS).…”

Section: Methodsmentioning

confidence: 99%

Identification of putative two-component regulatory systems in the bovine pathogen Mannheimia haemolytica A1, and preliminary characterization of the NarQ/P system

Inamoto

2010

FEMS Microbiology Letters

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The complete genome sequence of the bovine pathogen Mannheimia haemolytica A1 was analyzed by blast searches for the presence of two-component regulatory system proteins. Five complete sets of putative two-component systems were identified, and the NarQ/P system was further investigated. in silico analysis of the NarQ and NarP proteins showed features that are typical of the sensor and response regulator proteins. A narP knock-out mutant was constructed. The narP mutant has lost its ability to respond to NaNO(3) in the media and fail to alter the expression of several proteins. One of the proteins that showed increased production in the parent strain in response to NaNO(3) was analyzed by matrix-assisted laser desorption ionization time-of-flight MS. Unexpectedly, the protein was identified to be FbpA, a periplasmic component of the iron transporter system. Sequence analysis of the promoter region of fbpA identified motifs typical for NarP-regulated genes. The expression of the leukotoxin gene was also altered in the narP mutant as shown by Western immunoblot analysis and reverse transcription-PCR.

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“…Inner and outer membrane proteins of M. haemolytica A1 were prepared by sucrose gradient centrifugation according to laboratory procedure (Lo et al, 1991). The proteins were separated by SDS-PAGE, transferred onto a nitrocellulose membrane and immunostained with antibodies as described (Lo et al, 1991). For Far-Western immunoblot analysis, the outer and inner membrane proteins were separated by SDS-PAGE and transferred onto a nitrocellulose membrane as described.…”

Section: Sodium Dodecyl Sulfate-polyacrylamide Gel Electrophoresis (Smentioning

confidence: 99%

The Outer membrane protein OmpA ofMannheimia haemolyticaA1 is involved in the binding of fibronectin

Lo¹,

Sorensen²

2007

FEMS Microbiology Letters

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An enzyme-linked immunosorbent assay using bovine fibronectin as the substrate was used to demonstrate that Mannheimia haemolytica A1 binds to fibronectin. This binding to fibronectin was specific as no binding was observed with bovine fibrinogen. The binding to fibronectin was not observed if the M. haemolytica A1 cells were pretreated with trypsin or proteinase K, suggesting that it involved a protein molecule on the cell surface. Interestingly, the fibronectin-binding activity was found to be higher in an acapsular mutant compared with its parent strain. The fibronectin-binding protein was shown to be present in the outer membrane fraction of M. haemolytica A1. A 45 kDa outer membrane protein that binds to fibronectin was identified by Far-Western immunoblot analysis. This protein was purified and subjected to MS matrix-assisted laser desorption ionization time-of-flight analysis. The results identified it to be outer membrane OmpA based on comparison with the M. haemolytica A1 genomic sequence.

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Molecular studies of Ssa1, a serotype-specific antigen of Pasteurella haemolytica A1

Cited by 31 publications

References 29 publications

Antigenic and virulence properties of Pasteurella haemolytica leukotoxin mutants

Antigenic and virulence properties of Pasteurella haemolytica leukotoxin mutants

Identification of putative two-component regulatory systems in the bovine pathogen Mannheimia haemolytica A1, and preliminary characterization of the NarQ/P system

The Outer membrane protein OmpA ofMannheimia haemolyticaA1 is involved in the binding of fibronectin

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