Cloning, expression, purification and ligand binding studies of novel fibrinogen-binding protein FbsB of Streptococcus agalactiae.
Fibrinogen (Fg) is often a common site for bacterial recognition. In Streptococcus agalactiae, two surface proteins that recognize Fg are FbsA and FbsB. FbsA and the N-terminal region of FbsB have been shown to bind to human Fg, while the C-terminal region of FbsB [FbsB(C)] has been speculated to bind to bovine Fg. This C-terminal region which is conserved in many of the S. agalactiae strains was tested for binding to bovine Fg. For this, FbsB(C) was cloned, expressed and purified. Dot blot, Western blot and ELISA experiments carried out with the purified protein showed that FbsB(C) has the ability to bind to bovine Fg. It was also observed that other than binding to the native form of Fg, FbsB(C) also has the ability to bind to the Fg subunits when reduced. On studying the influence of Ca(2+) on the FbsB(C)-bovine Fg binding it was observed that the addition of Ca(2+) in the assay experiment greatly stimulated the binding. When the primary structure of FbsB(C) was analyzed, it was seen that other than similarities with strains of the same organism, it does not have any similarity with any protein characterized so far. In addition to this, its secondary structure component analysis by circular dichroism revealed that it is composed mainly of alpha helices and random coils unlike other Fg-binding surface proteins where beta sheets are dominant. FbsB(C) indeed is a novel protein and understanding the mechanism of its interaction with Fg would be useful in developing strategies to fight against infections by Streptococcus.
Devi
Aribam Swarmistha
AS
Centre of Advanced Study in Crystallography and Biophysics, University of Madras, Guindy Campus, Chennai, India.
Ponnuraj
Karthe
K
D000595:Amino Acid Sequence; D000818:Animals; D002352:Carrier Proteins / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification* / Q000378:metabolism; D002417:Cattle; D003001:Cloning, Molecular; D005340:Fibrinogen / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification* / Q000378:metabolism; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D016415:Sequence Alignment; D013292:Streptococcus agalactiae / Q000378:metabolism*
D002352:Carrier Proteins; C487204:FbsB protein, Streptococcus agalactiae; D011994:Recombinant Proteins; D005340:Fibrinogen