Structural Characterization of Peptide-Mediated Inhibition of Porphyromonas gingivalis Biofilm Formation

Abstract

Porphyromonas gingivalis is a periodontal pathogen whose primary niche is the anaerobic environment of subgingival dental plaque, but initial colonization of the oral cavity is likely to occur on supragingival surfaces that already support robust biofilm communities. Our studies have shown that P. gingivalis adheres to Streptococcus gordonii through interaction of the minor fimbrial antigen Mfa1 with a specific region of the streptococcal SspB polypeptide (residues 1167 to 1193) designated BAR. We show that a synthetic peptide comprising the BAR sequence potently inhibits P. gingivalis adherence to S. gordonii (50% inhibitory concentration ‫؍‬ 1.3 M) and prevents the development of P. gingivalis biofilms. However, a retroinverso peptide that possessed the same side chain topology as that of BAR was inactive, suggesting that interactions of Mfa1 with the peptide backbone of BAR are important for binding. A conformationally constrained analog of BAR inhibited P. gingivalis adherence and biofilm formation but at a lower specific activity than that of BAR. Therefore, to further define the structural features of the Mfa1-BAR interaction, we functionally screened combinatorial libraries of BAR in which active site residues (Asn 1182 , Thr 1184, and Val 1185) were replaced with each of the 19 common amino acids. Peptides containing positively charged amino acids at position 1182 or hydrophobic residues at position 1185 bound P. gingivalis more efficiently than did control peptides containing Asn and Val at these positions, suggesting that electrostatic and hydrophobic interactions may contribute to Mfa1-SspB binding. In contrast, replacement of Pro or Gly at these positions was detrimental to adherence, suggesting that perturbation of the BAR secondary structure influences activity. The net effect of substitutions for Thr 1184 was less pronounced either positively or negatively than that at the other sites. These results define physicochemical characteristics of the interacting interface of Mfa1 and SspB and suggest that peptides or peptidomimetics with greater specific inhibitory activity than that of BAR can be developed. These compounds may represent potential therapeutics that target some of the first molecular interactions that allow P. gingivalis to colonize the oral cavity.Dental plaque is a complex and dynamic biofilm that accumulates through the sequential and ordered colonization of over 700 species of bacteria (13,24,25,28). An oral biofilm comprised predominantly of gram-positive commensals such as the oral streptococci and Actinomyces spp. can exist in the oral cavity in the absence of overt disease (21, 25). However, populational shifts in the biofilm that lead to overrepresentation of acidophiles or of gram-negative obligate anaerobes may contribute to the onset and progression of the most common oral diseases, caries and periodontal disease (5). Indeed, adult periodontitis is associated with elevated levels of several gramnegative anaerobes in subgingival plaque, including the asaccharolytic, obligate a...

Peptide-modified nanoparticles inhibit formation of <em>Porphyromonas gingivalis</em> biofilms with <em>Streptococcus gordonii</em>

“…As shown in Figure 4, the IC 50 for free BAR peptide was approximately 1.3 µM, consistent with the value originally reported by Daep et al 11 In contrast, the IC 50 for inhibition of adherence by BAR-modified NPs was approximately 0.2 µM, which is 6.5-fold lower than the free BAR peptide. This result indicates that BAR-modified NPs are a significantly more potent inhibitor of P. gingivalis adherence to streptococci and subsequent biofilm formation than free peptide.…”

“…gordonii biofilms (50% inhibitory concentration [IC 50 ] =1.3 µM). 11,12 BAR also significantly reduced P. gingivalis virulence in a mouse model of periodontitis. [11][12][13] These results suggest that BAR may represent a novel therapeutic to limit P. gingivalis colonization of the oral cavity.…”

Genome of the Opportunistic Pathogen Streptococcus sanguinis

“…Therefore, it is not clear what ligand(s), if any, SspC and SspD bind. However, the 27-amino-acid region of SspB that has been shown to mediate binding of S. gordonii to P. gingivalis is conserved in SspC (18 identical residues and five similar residues), including perfect identity of the critical NITVK subsequence (21). This observation suggests that SspC may also adhere to P. gingivalis.…”

Periodontitis is associated with a loss of colonization by Streptococcus sanguinis

“…Interestingly, all eight strains of Streptococcus gordonii were isolated from periodontitis patients, but the difference between the two groups was not significant (P .0.003). Lamont et al (2002) and Daep et al (2006) demonstrated that S. gordonii plays a role in colonization with Porphyromonas gingivalis, whilst Haffajee et al (2006) found this species to be more prevalent in healthy subgingival sites. Further studies are needed to clarify the possible relationship between S. gordonii and periodontal status.…”