SdrG, a fibrinogen-binding bacterial adhesin of the microbial surface components recognizing adhesive matrix molecules subfamily from Staphylococcus epidermidis, targets the thrombin cleavage site in the Bbeta chain.
Staphylococcus epidermidis is an important opportunistic pathogen and is a major cause of foreign body infections. We have characterized the ligand binding activity of SdrG, a fibrinogen-binding microbial surface component recognizing adhesive matrix molecules from S. epidermidis. Western ligand blot analysis showed that a recombinant form of the N-terminal A region of SdrG bound to the native Bbeta chain of fibrinogen (Fg) and to a recombinant form of the Bbeta chain expressed in Escherichia coli. By analyzing recombinant truncates and synthetic peptide mimetics of the Fg Bbeta chain, the binding site for SdrG was localized to residues 6-20 of this polypeptide. Recombinant SdrG bound to a synthetic 25-amino acid peptide (beta1-25) representing the N terminus of the Fg Bbeta chain with a KD of 1.4 x 10(-7) m as determined by fluorescence polarization experiments. This was similar to the apparent K(D) (0.9 x 10(-7) m) calculated from an enzyme-linked immunosorbent assay where SdrG bound immobilized Fg in a concentration-dependent manner. SdrG could recognize fibrinopeptide B (residues 1-14), but with a substantially lower affinity than that observed for SdrG binding to synthetic peptides beta1-25 and beta6-20. However, SdrG does not bind to thrombin-digested Fg. Thus, SdrG appears to target the thrombin cleavage site in the Fg Bbeta chain. In fact, SdrG was found to inhibit thrombin-induced fibrinogen clotting by interfering with fibrinopeptide B release.
Davis
S L
SL
Center for Extracellular Matrix Biology, Institute of Biosciences and Technology, Texas A & M University Health Science Center, Houston, Texas 77030-3303, USA.
Gurusiddappa
S
S
McCrea
K W
KW
Perkins
S
S
Höök
M
M
D018829:Adhesins, Bacterial / Q000737:chemistry* / Q000378:metabolism*; D001665:Binding Sites; D001667:Binding, Competitive; D015153:Blotting, Western; D002352:Carrier Proteins / Q000737:chemistry* / Q000378:metabolism*; D002448:Cell Adhesion; D018076:DNA, Complementary / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004797:Enzyme-Linked Immunosorbent Assay; D004926:Escherichia coli / Q000378:metabolism; D005340:Fibrinogen / Q000378:metabolism; D005345:Fibrinopeptide B / Q000737:chemistry; D007700:Kinetics; D008024:Ligands; D008856:Microscopy, Fluorescence; D010455:Peptides / Q000737:chemistry; D016133:Polymerase Chain Reaction; D011485:Protein Binding; D011993:Recombinant Fusion Proteins / Q000737:chemistry; D011994:Recombinant Proteins / Q000378:metabolism; D013212:Staphylococcus epidermidis / Q000378:metabolism*; D013917:Thrombin / Q000378:metabolism
D018829:Adhesins, Bacterial; D002352:Carrier Proteins; D018076:DNA, Complementary; D008024:Ligands; D010455:Peptides; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D005345:Fibrinopeptide B; D005340:Fibrinogen; D013917:Thrombin