Protein F, a fibronectin-binding protein of Streptococcus pyogenes, also binds human fibrinogen: isolation of the protein and mapping of the binding region

Abstract

Summary: During screening of a gene library of Streptococcus pyogenes type M15 for fibrinogen-binding material, a protein of approximately 100 kDa, encoded outside the vir region, was found. DNA sequencing revealed this component to be identical to protein F, a fibronectin-binding protein. Isolation of the recombinant protein, termed F15, was performed by the use of fibrinogen affinity chromatography. The affinity constant (K a) of protein F15 for fibrinogen, 1.25 x 107 mol−1, was lower than that for fibronectin, 1.8 x 108 mol−1. The fibrinogen-binding domain was located in the N-terminal part of the molecule, while the fibronectin-binding domains, as previously determined, were in the C-terminal portion of protein F. To examine the amino acid sequence heterogeneity of protein F, the 5′ part of the prtF gene, corresponding to the N-terminal variable region of the protein, was amplified by PCR from 12 strains of S. pyogenes belonging to six different M-types. Alignment of these nucleotide sequences indicated that the 5′ portion of the prtF gene had probably undergone a number of intragenic recombination and horizontal gene transfer events, allowing a pattern of structural diversity of protein F observed earlier for some other streptococcal virulence factors. There was no strict correlation between M-type and nucleotide sequence of the variable region of the prtF gene and, compared to streptococcal M protein, the overall variation observed for protein F appeared more limited.

Site-Specific Manifestations of Invasive Group A Streptococcal Disease: Type Distribution and Corresponding Patterns of Virulence Determinants

“…Sequences specific for speA (32), speB (34), speC (24), speF (28), speG (56), speH (56), speJ (56), smeZ (21), ssa (59), cpa (55), cpa-1 (T. Miyoshi-Akiyama, N. Wakisaka, J. Zhao, and T. Uchiyama, unpublished data), fba (75), fbp-54 (Miyoshi-Akiyama et al, unpublished), pfbp (61), prtf-1 (66), prtf-2 (29), prtf-15 (35), sciA (58), sciB (78), sfb (72), and sfb-II (40) were detected by PCR on a model 9600 thermocycler (Perkin-Elmer, Gouda, The Netherlands) with the primers listed in Table 1. Amplification of all genes was carried out under the following conditions: an initial 5-min denaturation step at 96°C, followed by 30 cycles of denaturation at Table 1, and 70 s of extension at 72°C, with a final extension step at 72°C for 5 min.…”

Function of the Fibronectin-Binding Serum Opacity Factor ofStreptococcus pyogenesin Adherence to Epithelial Cells

“…A variable number of repetitive units and one nonrepetitive module with a unique sequence mediating fibronectin binding are located in the C-terminal region of the molecule (17,(44)(45)(46). The large N-terminal domain of the molecule was shown to bind to the extracellular plasma protein fibrinogen (23). The gene encoding SfbI is present in about 50 to 80% of the isolates tested (3,16,25,27,34,50).…”

Evolution ofsfbIEncoding Streptococcal Fibronectin-Binding Protein I: Horizontal Genetic Transfer and Gene Mosaic Structure

“…The biological function of the aromatic amino acid-rich domain is not known. To date, the reported binding functions of this domain include fibrinogen binding (20) and a putative carbohydrate-binding motif (40) which, from our data, appears to be highly conserved (Fig. 3A).…”

“…In addition, while the distribution of sfbI within M types (28) or Vir types (VTs) (13) is generally consistent, there is evidence of heterogeneity of sfbI species within M types, as the number of fibronectinbinding repeats detected in M8 and M28 serotype strains is variable (28). Significant heterogeneity in sfbI sequences encoding the amino-terminal half of the aromatic amino acid-rich domain from M4, M12, M15, and M18 strains has also been reported (20).…”

“…Identified simultaneously in two independent laboratories (15,39), SfbI is a membraneanchored surface protein shown to be a major adhesin of GAS (13,15,28,29,39,42). In addition, SfbI binds to fibrinogen via a domain in the amino terminus (20) and human immunoglobulin (IgG) via the carboxy-terminal fibronectin-binding repeat domain (25). SfbI is protective in mice (14,34) and a mucosal adjuvant (26).…”