STRUCTURAL DOMAINS OF PORPHYROMONAS-GINGIVALIS RECOMBINANT FIMBRILLINTHAT MEDIATE BINDING TO SALIVARY PROLINE-RICH PROTEIN AND STATHERIN

Fimbriae (the oligomeric form of fimbrillin) are considered important in the adherence and colonization of Porphyromonas gingivalis in the o ral cavity. In the present study, we have identified the structural do mains of P.gingivalis fimbrillin that mediate the binding to salivary proline-rich protein 1 (PRP1) and statherin. A series of synthetic fim brillin peptides were used to localize the active fimbrillin domains i nvolved in the binding to PRP1 and statherin. The binding of I-125-lab eled 41-r-Fim (whole-length recombinant fimbrillin, amino acid [aa] re sidues 1 to 337) to PRP1-coated hydroxyapatite beads (HAP) was strongl y inhibited by the fimbrillin C-terminal peptides corresponding to aa residues 266 to 286 and 318 to 337 (peptides 266-286 and 318-337, resp ectively), while the binding to statherin was inhibited by C-terminal peptides 266-286, 293-306, and 307-326. Peptide 126-146 also showed a weak inhibitory effect, about half that of other active peptides, on t he binding to both PRP1 and statherin, P. gingivalis whole-cell bindin g to PRP1- or statherin-coated HAP was inhibited by more than 80% by t he same active peptides. To confirm that the C-terminal portion of fim brillin includes domains responsible for the binding, two C-terminally truncated variants of recombinant fimbrillin were generated and purif ied. These were designated 34.5-r-Fim, corresponding to aa residues 1 to 286, and 32-r-Fim, corresponding to aa residues 1 to 265. I-125-34. 5-r-Fim revealed 35 and 34% loss of binding ability to PRP1 and stathe rin, respectively. I-125-32-r-Fim had significantly less binding abili ty to PRP1 and statherin than I-125-34.5-r-Fim, which was reduced 78 a nd 73%, respectively. Whole-cell binding to PRP1-, statherin-, or whol e saliva-coated HAP was inhibited up to 100% by 41-r-Fim, while 32-r-F im also showed considerable inhibition, possibly due to the region of aa 126 to 146. Collectively, these results suggest that there are sepa rate and multiple binding sites for PRP1 and statherin in the P. gingi valis fimbrillin, and the combination of all of these binding sites ma y be indispensable in establishing stable bacterial adherence to saliv a coated surfaces in the oral cavity.