Adherence of Streptococcus sanguis to conformationally specific determinants in fibronectin

Abstract

The adherence of Streptococcus sanguis to specific receptors exposed or deposited at the site of endothelial damage may play an important role in the development of infective endocarditis. Adherence of the Challis strain of S. sanguis to gelatin (or collagen) and gelatin-binding components of plasma was examined with an enzyme-linked immunosorbent assay. S. sanguis adhered poorly to immobilized gelatin and to molecular or fibrillar collagen. However, in the presence of fresh human plasma, the adherence of S. sanguis to all three substrates increased as much as eightfold. Removal of gelatin-binding proteins eliminates the ability of plasma to enhance adherence of S. sanguis to the substrates. Addition of purified human plasma fibronectin (Fn) to the absorbed plasma restored the adherence-promoting ability in a dose-dependent manner. A similar dose-dependent increase in S. sanguis adherence was observed when increasing concentrations of Fn alone were added to the gelatin-coated assay wells. S. sanguis adherence to immobilized fibronectin could not be inhibited by preincubating either the bacteria or the gelatin-coated assay wells with Fn or by including excess soluble Fn in the assay mixture. Studies with peptides purified from trypsin digests of Fn indicated that the 160- to 180-kilodalton (kDa) fragments which retain both the gelatin-binding and the cell-binding regions of the intact molecule support adherence of S. sanguis to gelatin. The 160- to 180-kDa fragments inhibited the interaction of S. sanguis with immobilized Fn. In contrast, intact Fn and the 31-kDa amino-terminal fragment were unable to inhibit the adherence when used in equivalent or greater molar amounts. These in vitro results suggest that in the presence of whole plasma, S. sanguis binds to immobilized gelatin or collagen via Fn bound to the immobilized substrates. Our finding that adherence of S. sanguis to immobilized Fn can occur in the presence of large concentrations of Fn, whether in plasma or purified, indicates that a S. sanguis-binding domain is cryptic in the Fn molecule while in solution and is exposed by a conformational change when the Fn becomes bound to gelatin-coated plastic. The ability of peptide fragments of Fn to inhibit S. sanguis adherence is consistent with this hypothesis.