A 190-kDa surface protein antigen (PAc) of Streptococcus mutans binds to human salivary components. For detection of specific binding of the PAc protein to human salivary components, a simple sandwich assay was used. Microtiter plates precoated with recombinant PAc (rPAc), PAc fragments, or S. mutans whole cells were allowed to react with human whole saliva and then were incubated with biotinylated rPAc. The biotinylated rPAc bound to salivary components was detected by use of alkaline phosphatase-conjugated streptavidin and p-nitrophenylphosphate. In this assay, the binding of whole cells of S. mutans and purified rPAc to salivary components was confirmed. For determination of a saliva-binding region of the PAc molecule, 14 truncated PAc fragments were constructed by use of the polymerase chain reaction and an expression vector, pAX4a+. The binding of these truncated PAc fragments to human salivary components was determined by the sandwich assay. Among the truncated PAc fragments, fragments corresponding to residues 39 to 864 and residues 39 to 1000 of PAc showed a high ability to bind to salivary components. Shorter recombinant fragments corresponding to residues 39 to 217, residues 200 to 481, residues 470 to 749, and residues 688 to 864 did not exhibit any binding ability. The fragment that corresponds to a proline-rich repeating region (residues 828 to 1000) bound directly to the PAc protein. These results suggest that residues 39 to 864 of the PAc molecule are

Fourteen strains of Streptococcus mutans serotype c were examined for their cell-surface protein antigens in terms of hydrophobicity, M , and immunochemical specificities. Thirteen strains were hydrophobic, while strain GS-5 was markedly hydrophilic as compared to the other strains tested. Cell-surface protein antigens were then analysed by sodium dodecyl sulphatepolyacrylamide gel electrophoresis (SDS-PAGE) and Western immunoblotting. A protein antigen of M , 190000 (PAC) was found in cell extracts and culture supernatants of all the hydrophobic strains. Neither culture supernatant nor cell extract of strain GS-5 contained PAC. Strain GS-5, however, produced extracellularly a large amount of a protein of M , 155000 (PAGS-5) which reacted with rabbit anti-PAC serum. Immunodiffusion analysis showed that PAGS-5 lacked a part of the antigenic moieties in the PAC molecule. SDS-PAGE and radioimmunoassay showed a small amount of PAGS-5 on the cell surface of strain GS-5. These findings suggest that PAGS-5 may correspond to PAC which lacks a region participating in binding of PAC to the streptococcal cell. I N T R O D U C T I O NStreptococcus mutans has been strongly implicated as a causative organism of dental caries (Hamada & Slade, 1980; Loesche, 1986). The organism possesses various kinds of cell-surface polymers such as proteinaceous antigens, serotype-specific polysaccharide antigens, lipoteichoic acid and peptidoglycan (Hamada & Slade, 1980). Among these cell-surface polymers, much attention has been focused on a high-M, protein antigen that has been variously designated as antigen B (Russell, 1979), 1/11 (Russell et al., 1980), IF (Hughes et al., 1980) and PI (Forester et al., 1983). Streptococcus sobrinus also possesses a cell-surface protein antigen that is immunologically and biochemically similar to that of S. mutans (Moro & Russell, 1983; Holt et al., 1982; Okahashi et al., 1986).The high-M, protein antigen of S. mutans serotype c (PAC) is an effective vaccine antigen for prevention of dental caries in monkeys (Lehner et al., 1981 ; Russell et al., 1982). Local passive immunization with monoclonal antibodies raised against PAC prevents the colonization by S. mutans of animal and human tooth surfaces (Lehner et al., 1985; Ma et af., 1987). Polyclonal antibodies to PAC are strongly opsonic for S. mutans (Scully et al., 1980). McBride et al. (1984) suggested that location of PAC on the streptococcal surface is important in forming hydrophobic bonds with hydrophobic regions of salivary pellicle. The organization of PAC on the cell surface, however, is poorly understood.

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