Lactoferrin Inhibits Porphyromonas gingivalis Proteinases and Has Sustained Biofilm Inhibitory Activity

Abstract

Porphyromonas gingivalis is a bacterial pathogen associated with chronic periodontitis that results in destruction of the tooth's supporting tissues. The major virulence determinants of P. gingivalis are its cell surface Arg-and Lys-specific cysteine proteinases, RgpA/B and Kgp. Lactoferrin (LF), an 80-kDa iron-binding glycoprotein found in saliva and gingival crevicular fluid, is believed to play an important role in innate immunity. In this study, bovine milk LF displayed proteinase inhibitory activity against P. gingivalis whole cells, significantly inhibiting both Arg-and Lys-specific proteolytic activities. LF inhibited the Argspecific activity of purified RgpB, which lacks adhesin domains, and also inhibited the same activity of the RgpA/Kgp proteinaseadhesin complexes in a time-dependent manner, with a first-order inactivation rate constant (k inact ) of 0.023 min ؊1 and an inhibitor affinity constant (K I ) of 5.02 M. LF inhibited P. gingivalis biofilm formation by >80% at concentrations above 0.625 M. LF was relatively resistant to hydrolysis by P. gingivalis cells but was cleaved into two major polypeptides (53 and 33 kDa) at R 284 to S 285 , as determined by in-source decay mass spectrometry; however, these polypeptides remained associated with each other and retained inhibitory activity. The biofilm inhibitory activity of LF against P. gingivalis was not attributed to direct antibacterial activity, as LF displayed little growth inhibitory activity against planktonic cells. As the known RgpA/B and Kgp inhibitor N-␣-p-tosyl-L-lysine chloromethylketone also inhibited P. gingivalis biofilm formation, the antibiofilm effect of LF may at least in part be attributable to its antiproteinase activity.

Oxantel Disrupts Polymicrobial Biofilm Development of Periodontal Pathogens

“…In addition, we have shown that the expression of these genes was downregulated in a P. gingivalis ferric uptake regulator (Fur) orthologue knockout mutant (unpublished observation). We have also shown that heme-limited P. gingivalis has reduced biofilm-forming capacity compared with P. gingivalis grown under excess heme conditions (51). It is therefore possible that at the sub-MIC used in this study, the oxantel inhibition of Frd activity reduced energy levels in the cell, which then acted as a weak inducer of a subset of genes of this regulon.…”

Inhibitory effects of the κ-casein macropeptide isolated from milk protein on the biofilm formation and virulence of Listeria monocytogenes

“…23) However, a few reports indicated that CMP could influence the virulence of foodborne pathogens including biofilm formation. Recent studies have indicated that several milk peptides, including osteopontin 24) and lactoferrin, 25) strongly reduced the amount of dental biofilm. We indicated here that CMP would be potentially useful in preventing biofilm formation on PVC and glass surfaces.…”

Potential Value of a Rice Protein Extract, Containing Proteinaceous Inhibitors against Cysteine Proteinases fromPorphyromonas gingivalis, for Managing Periodontal Diseases

“…These concentrations correspond to those required for the inhibition of Rgp activity. Such agents as cranberry polyphenol 28,29) and lactoferrin 46) have also demonstrated inhibitory activities against both biofilm formation and Rgp/Kgp activity. However, in the case of cranberry polyphenol and lactoferrin, the concentration required for inhibiting biofilm formation was inconsistent with that required for inhibiting the proteinase activities.…”