The objective of this subproject is to further characterize the mechanism(s) involved in cohesion between Porphyromonas gingivalis and Streptococcus gordonii or Streptococcus sanguis. In particular, the mechanisms whereby selected saliva and serum components inhibit this cohesion will be explored. Our previous studies have demonstrated that outer membrane proteins (OMP) of P. gingivalis may be involved in these interactions. Consequently, outer membrane adhesins of P. gingivalis will be identified in binding assays using radiolabeled ligand molecules from saliva or streptococci. The adhesins will be purified using a combination of gel filtration, ion exchange, and affinity chromatography. The N- terminal sequence of the purified protein will be determined and used to deduce the nucleotide sequence of this region of the gene. Synthetic oligonucleotide probes will be used to identify the structural genes, which will be isolated, cloned, and the complete nucleotide sequences determined. Adhesin-deficient strains of P. gingivalis will be generated by transposon insertion mutagenesis and tested for their abilities to adhere to experimental streptococcal plaque and experimental salivary pellicles. The surface composition of adhesin-deficient mutants will be compared with those of isogeneic parental strains to identify other possible adhesins. We anticipate that the information derived from these studies will provide a rationale for enhancing natural host defenses to minimize colonization of dental plaque and the oral cavity by P. gingivalis and related oral pathogens.
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