Association of Proteases of Porphyromonas (Bacteroides) gingivalis with its Adhesion to Actinomyces viscosus

Abstract

P. gingivalis adheres to A. viscosus on mineral surfaces mimicking teeth. To study whether P. gingivalis proteases contribute to its binding, mutants of P. gingivalis deficient in proteases were compared with their parent strain and a P. gingivalis-type strain for their adherence to A. viscosus on saliva-coated hydroxyapatite by manipulating a radio-isotope binding assay. Adherence of P. gingivalis 2561 to A. viscosus was studied by tests of the effects of incubation temperature and known inhibitors or promoters of proteases. Controls were handled by the assay run in PBS buffer at 22°C. Two mutants deficient in trypsin-like protease were found to be deficient in adherence (% attachment relative to control: 3.2 ± 0.1% and 11.2 ± 0.4%), while a collagenase-deficient mutant had an adherence score (51.6 ± 8.4) closer to that of the parent strain (75.6 ± 7.2%). Heating P. gingivalis at 70°C decreased its subsequent adherence at 22°C by 80%. Adherence decreased by 60% when the assay was run at 4°C, but increased by 70% at 37°C. Reducing agents (dithiothreitol, cysteine, and mercaptoethanol) enhanced P. gingivalis adherence by 50 to 60%. Protease inhibitors (BZMD, SBTI, TPCK, TLCK, CMPS, PMSF) decreased adherence by 10 to 50%. Also, Hg2+ and Zn2+ decreased adherence by 30 to 50%, and arginine decreased it by 50%. Most of these effects on P. gingivalis adherence were statistically significant (p<0.05). Analysis of these data suggests that P. gingivalis proteases may contribute to the cohesion of P. gingivalis and A. viscosus.