The changing faces of Streptococcus antigen I/II polypeptide family adhesins

Abstract

Streptococcus mutans antigen I/II (AgI/II) protein was one of the first cell wall-anchored adhesins identified in Gram-positive bacteria. It mediates attachment of S. mutans to tooth surfaces and has been a focus for immunization studies against dental caries. The AgI/II family polypeptides recognize salivary glycoproteins, and are also involved in biofilm formation, platelet aggregation, tissue invasion and immune modulation. The genes encoding AgI/II family polypeptides are found among Streptococcus species indigenous to the human mouth, as well as in Streptococcus pyogenes, S. agalactiae and S. suis. Evidence of functionalities for different regions of the AgI/II proteins has emerged. A sequence motif within the C-terminal portion of Streptococcus gordonii SspB (AgI/II) is bound by Porphyromonas gingivalis, thus promoting oral colonization by this anaerobic pathogen. The significance of other epitopes is now clearer following resolution of regional crystal structures. A new picture emerges of the central V (variable) region, predicted to contain a carbohydrate-binding trench, being projected from the cell surface by a stalk formed by an unusual association between an N-terminal alpha-helix and a C-terminal polyproline helix. This presentation mode might be important in determining functional conformations of other Gram-positive surface proteins that have adhesin domains flanked by alpha-helical and proline-rich regions.

Fig. 1. Linear structure of AgI/II family proteins showing localization of specific functional regions

Six structural regions are shown, based upon primary sequence: LP, leader peptide; A, Ala-rich repeats; V, variable or divergent region; P, Pro-rich repeats; C, C-terminal region; CWA, cell-wall anchorage region. Amino acid (aa) residue numbers refer to S. mutans SpaP. Mapped position of adhesintope (Adh2) corresponding to colonization-inhibitory peptide (Kelly et al., 1999) is indicated, as well as defined sequences in S. mutans SpaP involved in binding of salivary glycoproteins (SGPs) and collagen (Sciotti et al., 1997) (black lines). The region within S. gordonii SspB interacting with Porphyromonas gingivalis (Daep et al., 2006) is shown together with the region thought to interact with A. naeslundii T14V (Jakubovics et al., 2005) (grey lines).

Fig. 2. Primary sequence homologies across selected AgI/II family proteins

The structural regions are depicted as in Fig. 1, and percentage aa residue identities to S. mutans SpaP (GenBank: X17390) are indicated. The inferred amino acid sequences from the genes shown are (GenGank accession nos in parentheses): S. gordonii DL1 sspA and sspB (U40027), S. intermedius pas (AB045140.2), S. pyogenes MGAS6180 spy1325 (aspA) (3574034) and S. agalactiae 2603V/R SAG2021(bspD) (1014832). Colour key: red, leader peptide; purple, A-repeats; blue, P-repeats; orange, C-terminal and cell-wall anchorage.

Fig. 3. Crystal structures of AgI/II family protein fragments

Panel A, structure of SspB-C_1061-1413 region showing the BAR motif (in green) recognized by P. gingivalis, Ca²⁺ ions (•) and intramolecular isopeptide bonds (black) between asparagine and lysine; panel B, structure of the SpaP A₃VP₁ fragment showing unique interaction of A and P regions to generate a stalk-like structure presenting a V region head. The homologous site to the core B cell epitope TYEAALKQYEADL (Senpuku et al., 1997), which is located within a larger synthetic peptide reported to contribute to protective immunity against S. mutans, is illustrated in yellow. This core sequence is repeated four times within the A-region of SpaP. The binding motif within the synthetic peptide reported to interact with human MHC class II molecules (Senpuku et al., 1998) falls outside of the crystallized segment. Likewise the homologous site to the minimal consensus sequence (XPX(E/D)PXYXXXPXPP) reported to contribute to recognition by Guy's 13 monoclonal antibody used in passive protection studies to prevent S. mutans re-colonization in humans (van Dolleweerd et al., 2004) is illustrated in light purple. This core sequence is repeated 4 times within the P-region of AgI/II. The binding partner of the P-region consensus sequence necessary to reconstitute the Guy's 13 discontinuous epitope was localized within a 97- aa residue fragment that corresponds to the amino-terminal portion of the A-region, and both partners fall outside of the crystallized fragment.

Fig. 3. Crystal structures of AgI/II family protein fragments

Panel A, structure of SspB-C_1061-1413 region showing the BAR motif (in green) recognized by P. gingivalis, Ca²⁺ ions (•) and intramolecular isopeptide bonds (black) between asparagine and lysine; panel B, structure of the SpaP A₃VP₁ fragment showing unique interaction of A and P regions to generate a stalk-like structure presenting a V region head. The homologous site to the core B cell epitope TYEAALKQYEADL (Senpuku et al., 1997), which is located within a larger synthetic peptide reported to contribute to protective immunity against S. mutans, is illustrated in yellow. This core sequence is repeated four times within the A-region of SpaP. The binding motif within the synthetic peptide reported to interact with human MHC class II molecules (Senpuku et al., 1998) falls outside of the crystallized segment. Likewise the homologous site to the minimal consensus sequence (XPX(E/D)PXYXXXPXPP) reported to contribute to recognition by Guy's 13 monoclonal antibody used in passive protection studies to prevent S. mutans re-colonization in humans (van Dolleweerd et al., 2004) is illustrated in light purple. This core sequence is repeated 4 times within the P-region of AgI/II. The binding partner of the P-region consensus sequence necessary to reconstitute the Guy's 13 discontinuous epitope was localized within a 97- aa residue fragment that corresponds to the amino-terminal portion of the A-region, and both partners fall outside of the crystallized fragment.

Fig. 4. Diagrammatic representation of AgI/II family polypeptide structure showing interactive regions

The predicted domain structures of S. mutans SpaP and S. gordonii SspB are shown. The known crystal structures of the V-regions of SpaP and SspB (PDB:1JMM and PDB:2WD6, respectively) and the crystal structure of the SspB C-terminus (PDB: 2WZA) have been incorporated into the models. The A-P stalk of SspB and the continuation of the A-P stalk of SpaP were modelled based on homology with the A3VP1 fragment of SpaP and the known crystal structure (PDB:31PK). The approximate locations of epitopes recognized by anti-AgI/II monoclonal antibodies, and those that cross-react with SspB, are indicated. The BAR region of SspA/B that interacts with P. gingivalis minor fimbrial protein (Mfa1) is highlighted. Binding of AgI/II family proteins to a range of receptors as indicated has been well documented, but the precise sequences involved in the adhesion-receptor interactions are not yet known.