Affinity purification and characterization of a fibrinogen-binding protein complex which protects mice against lethal challenge with Streptococcus equi subsp. equi.
Cell-wall-associated proteins from Streptococcus equi subsp. equi, the causative agent of strangles, were analysed with a view to identifying a potential protective antigen. Preparations of these proteins, isolated from mutanolysin extracts of cell walls, were shown to contain one major high-M(r) protein species (apparent M(r) 220,000 and 550,000 when analysed by SDS-PAGE and gel-filtration chromatography, respectively). The high-M(r) protein bound horse fibrinogen and was purified under non-denaturing conditions using fibrinogen affinity chromatography. The fibrinogen-binding protein (FgBP) reacted with serum taken from horses recovering from strangles and protected mice against lethal challenge from S. equi subsp. equi. The sequence of the corresponding gene (fbp) was determined and shown to encode a mature protein (M(r) 54,597) with predicted coiled-coil structure. An FgBP truncate, lacking the C-terminal cell wall/membrane anchor domain, was overexpressed in and purified from Escherichia coli and was shown to behave in an analogous fashion to the wild-type product in terms of M(r) estimation, fibrinogen binding and seroreactivity.
Meehan
Mary
M
Department of Microbiology, Moyne Institute of Preventive MedicineTrinity College, Dublin 2, Ireland.|National Pharmaceutical Biotechnology Centre, BioResearch IrelandTrinity College, Dublin 2, Ireland.
Nowlan
Peter
P
Bioresources UnitTrinity College, Dublin 2, Ireland.
Owen
Peter
P
Department of Microbiology, Moyne Institute of Preventive MedicineTrinity College, Dublin 2, Ireland.
D000595:Amino Acid Sequence; D000818:Animals; D001425:Bacterial Outer Membrane Proteins / Q000737:chemistry / Q000276:immunology / Q000302:isolation & purification*; D001426:Bacterial Proteins; D015153:Blotting, Western; D002352:Carrier Proteins / Q000737:chemistry / Q000276:immunology / Q000302:isolation & purification*; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D006736:Horses; D051379:Mice; D008969:Molecular Sequence Data; D017421:Sequence Analysis; D018502:Streptococcus equi / Q000737:chemistry*
D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; D002352:Carrier Proteins; C112406:fibrinogen-binding protein, Streptococcus equi