Crystal structures of fibronectin-binding sites from Staphylococcus aureus FnBPA in complex with fibronectin domains

Abstract

Staphylococcus aureus can adhere to and invade endothelial cells by binding to the human protein fibronectin (Fn). FnBPA and FnBPB, cell wall-attached proteins from S. aureus, have multiple, intrinsically disordered, high-affinity binding repeats (FnBRs) for Fn. Here, 30 years after the first report of S. aureus/Fn interactions, we present four crystal structures that together comprise the structures of two complete FnBRs, each in complex with four of the N-terminal modules of Fn. Each approximately 40-residue FnBR forms antiparallel strands along the triple-stranded beta-sheets of four sequential F1 modules ((2-5)F1) with each FnBR/(2-5)F1 interface burying a total surface area of approximately 4,300 A(2). The structures reveal the roles of residues conserved between S. aureus and Streptococcus pyogenes FnBRs and show that there are few linker residues between FnBRs. The ability to form large intermolecular interfaces with relatively few residues has been proposed to be a feature of disordered proteins, and S. aureus/Fn interactions provide an unusual illustration of this efficiency.

Fig. 1.

FnBR peptides from S. aureus FnBPA in complex with NTD F1 module pairs from Fn. (A) FnBPA contains 11 FnBRs (4), 6 of which bind the NTD of Fn with high affinity (5) (indicated with asterisks, an orange asterisk indicates an FnBR for which structural data are presented). The fibrinogen-binding A domain and the C-terminal bacterial cell-wall attachment site (LPETG) are indicated. (B) Sequence alignment of FnBRs from A (residues 508–874 of Swiss-Prot entry P14738) highlighting residues (orange) conserved between FnBRs. Peptide sequences are indicated by a solid bar, and the first and last residue in each peptide that undergoes a >2-Ų change in accessible surface area on complex formation are underlined. (C) Ribbon diagram for the structures of ²F1³F1/STATT1 and ⁴F1⁵F1/STAFF1. Fn modules are shown in cyan, and the FnBR is shown in gray. Strand naming for F1 modules is shown on ²F1. (D) Ribbon diagrams for the structures of ²F1³F1/STATT5 and ⁴F1⁵F1/STAFF5. The ²F1³F1/STATT1, ⁴F1⁵F1/STAFF1, ²F1³F1/STATT5, and ⁴F1⁵F1/STAFF5 structures were solved to resolutions of 2, 1.8, 1.7, and 2 Å, respectively. The dotted lines indicate that the orientation between ³F1 and ⁴F1 was approximated based on the other F1 module interfaces and the short length on the intermodule linker.

Fig. 2.

Specificity of ²F1-peptide and ⁴F1-peptide interactions. The conserved hydroxyl-containing side-chain threonines ⁴F1:STAFF1 (A) and ²F1:STATT5 (B), serine ⁴F1:STAFF5 (C), and tyrosine ²F1:STATT1 (D) of the FnBR (gray) forms hydrogen bonds with backbone atoms of homologous residues in ²F1 and ⁴F1 (cyan). The positions of conserved acidic and hydrophobic FnBR residues (gray) with respect to the electrostatic surface of ²F1 or ⁴F1 are shown below with FnBR sequences.

Fig. 3.

Role of conserved residues in ³F1 binding and ⁵F1 binding. (A) Structure of ²F1³F1/STATT1 focusing on a conserved glycine residue in the ³F1-binding motif (gray) and a tryptophan in the E strand of ³F1 (cyan). (B) Homologous region of ⁴F1⁵F1/STAFF1 showing that the different orientation of the tryptophan side chain accommodates a glutamine residue in the bound FnBR. (C and D) Structures of ⁴F1⁵F1/STAFF1 (C) and ⁴F1⁵F1/STAFF5 (D) highlighting the hydrogen bond formed by the conserved asparagine residue of the FnBR (gray) with the backbone of ⁵F1 (cyan).

Fig. 4.

Efficiency of S. aureus binding to Fn. Schematic (not to scale) showing how nine copies of Fn (cyan) might cluster on FnBPA (gray) and illustrating the efficiency of the binding. The five F1 modules from the N-terminal domain are shown with ^2–5F1 (FnBPA-1,5,9,10,11) or ^3–5F1(FnBPA-2,3,4,8) bound to FnBRs through tandem β-zippers, and the remainder of each Fn molecule and FnBPA are shown as cyan and gray ovals, respectively.