DELETION OF THE CENTRAL PROLINE-RICH REPEAT DOMAIN RESULTS IN ALTEREDANTIGENICITY AND LACK OF SURFACE EXPRESSION OF THE STREPTOCOCCUS-MUTANS P1 ADHESIN MOLECULE

Members of the family of surface adhesins of oral streptococci, includ ing P1 of Streptococcus mutans, contain two highly conserved repeat do mains, one rich in alanine! (A region) and the other rich in proline ( ]P region). To assess the contribution of the P region to the biologic al properties of P1, an internal deletion in spaP was engineered. In a ddition, the P region was subcloned and expressed as a fusion partner with the maltose binding protein of Escherichia coli and liberated by digestion with factor Xa, Results of Western blot experiments in which recombinant polypeptides were probed with a panel of 11 monoclonal an tibodies indicated that the P region is a necessary component of confo rmational epitopes within the central portion of pi. Antibodies reacti ve with the P region were detected in a polyclonal rabbit antiserum ge nerated against whole S. mutans cells but not in two rabbit antisera g enerated against purified pi (M-r similar to 185,000), suggesting that this domain is immunogenic on the surface of intact bacteria but not as past of a soluble full-length molecule. Finally, transformation of a spaP-negative mutant with a shuttle vector containing an internally deleted spaP lacking P-region DNA resulted in a complete absence of su rface-localized P1 and substantially less P1 in sonicated cells compar ed to the case for the mutant complemented with the full-length gene, These results suggest that the P region is an integral component contr ibuting to the conformation of the central region of P1 and indicate t hat its presence is necessary for surface expression of the molecule o n S. mutans.