title: Purification and partial characterization of the OmpA family of proteins of Pasteurella haemolytica.
abstract: This study was conducted to partially characterize and identify the purity of two major outer membrane proteins (OMPs) (with molecular weights of 32,000 and 35,000 [32K and 35K, respectively]) of Pasteurella haemolytica. The 35K and 32K major OMPs, designated Pasteurella outer membrane proteins A and B (PomA and PomB, respectively), were extracted from P. haemolytica by solubilization in N-octyl polyoxyl ethylene. The P. haemolytica strain used was a mutant serotype A1 from which the genes expressing the 30-kDa lipoproteins had been deleted. PomA and PomB were separated and partially purified by anion-exchange chromatography. PomA but not PomB was heat modifiable. The N-terminal amino acid sequences of the two proteins were determined and compared with reported sequences of other known proteins. PomA had significant N-terminal sequence homology with the OmpA protein of Escherichia coli and related proteins from other gram-negative bacteria. Moreover, polyclonal antiserum raised against the E. coli OmpA protein reacted with this protein. PomA was surface exposed, was conserved among P. haemolytica biotype A serotypes, and had porin activity in planar bilayers. No homology between the N-terminal amino acid sequence of PomB and those of other known bacterial proteins was found. Cattle vaccinated with live P. haemolytica developed a significant increase in serum antibodies to partially purified PomA, as shown by enzyme-linked immunosorbent assays, and to purified PomA and PomB, as detected on Western blots and by densitometry.
authors.affiliation: Department of Anatomy, Pathology, and Pharmacology, College of Veterinary Medicine, Oklahoma State University, Stillwater 74078, USA.
authors.forename: P J
authors.initials: PJ
authors.lastname: Mahasreshti
authors.forename: G L
authors.initials: GL
authors.lastname: Murphy
authors.forename: J H
authors.initials: JH
authors.lastname: Wyckoff
authors.forename: S
authors.initials: S
authors.lastname: Farmer
authors.forename: R E
authors.initials: RE
authors.lastname: Hancock
authors.forename: A W
authors.initials: AW
authors.lastname: Confer
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000907:Antibodies, Bacterial / Q000096:biosynthesis; D001425:Bacterial Outer Membrane Proteins / Q000737:chemistry* / Q000235:genetics / Q000276:immunology; D002417:Cattle; D003902:Detergents; D008051:Lipid Bilayers; D008074:Lipoproteins / Q000737:chemistry* / Q000276:immunology; D008168:Lung / Q000473:pathology; D016978:Mannheimia haemolytica / Q000737:chemistry* / Q000145:classification / Q000276:immunology; D008969:Molecular Sequence Data; D010326:Pasteurella Infections / Q000276:immunology / Q000473:pathology; D011092:Polyethylene Glycols; D017421:Sequence Analysis; D017386:Sequence Homology, Amino Acid; D012703:Serotyping; D012995:Solubility
chemical_list: D000907:Antibodies, Bacterial; D001425:Bacterial Outer Membrane Proteins; D003902:Detergents; D008051:Lipid Bilayers; D008074:Lipoproteins; C103352:PomA protein, bacteria; C103353:PomB protein, bacteria; C072375:n-octyl-polyoxyethylene; D011092:Polyethylene Glycols