Molecular and immunological characterization of Pasteurella multocida serotype A:3 OmpA: evidence of its role in P. multocida interaction with extracellular matrix molecules.
Pasteurella multocida OmpA-like gene (PmOmpA) was cloned and characterized. The mature protein had a molecular mass of 35,075 Da and significant similarity with Escherichia coli (E. coli) OmpA proteins. Membrane topology analyses predict that like E. coli OmpA, the N-terminal half of PmOmpA exists as an eight-stranded transmembrane antiparallel beta-barrel that displays four variable hydrophilic and surface-exposed regions with predicted antigenic peaks that may be involved in serum resistance or adherence. In addition, the Ala-Pro repeat region between the N-terminal beta-barrel and C-terminal periplasmic domains is completely missing in PmOmpA. PmOmpA was expressed in E. coli and immunoblots analysis revealed that the recombinant PmOmpA was immunogenic, and expressed in vivo. The binding of PmOmpA to biotinylated Madin Darby bovine kidney (MDBK) cells surface proteins, fibronectin and heparin was demonstrated. Furthermore, PmOmpA binds MDBK monolayers and pre-treatment of P. multocida whole cells with anti-PmOmpA significantly reduced adherence to fibronectin. Ligand blot analysis revealed that fibronectin binds to the native and heat modified forms of PmOmpA when heated at 37 and 100 degrees C, respectively. Collectively these data indicate that PmOmpA may be involved in P. multocida 232 adherence to host cells via heparin and/or fibronectin bridging.
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Department of Veterinary Pathobiology, Oklahoma State University, Stillwater, OK 74078-2007, USA. mdvpath@okstate.edu
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Quijano-Blas
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D000595:Amino Acid Sequence; D000818:Animals; D001422:Bacterial Adhesion; D001425:Bacterial Outer Membrane Proteins / Q000737:chemistry* / Q000276:immunology* / Q000302:isolation & purification / Q000378:metabolism; D001426:Bacterial Proteins / Q000032:analysis; D015153:Blotting, Western; D002417:Cattle; D002460:Cell Line; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D016326:Extracellular Matrix Proteins / Q000378:metabolism*; D005353:Fibronectins / Q000378:metabolism; D015870:Gene Expression; D006493:Heparin / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism; D008969:Molecular Sequence Data; D016979:Pasteurella multocida / Q000145:classification / Q000235:genetics* / Q000378:metabolism* / Q000472:pathogenicity; D010957:Plasmids; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000737:chemistry / Q000276:immunology / Q000378:metabolism; D017385:Sequence Homology; D012703:Serotyping
D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; D016326:Extracellular Matrix Proteins; D005353:Fibronectins; D008565:Membrane Proteins; D011994:Recombinant Proteins; C078167:OMPA outer membrane proteins; D006493:Heparin