Abstract
The oral anaerobic bacterium Porphyromonas gingivalis, a major pathogen of advanced adult periodontitis, produces a novel class of cysteine proteinases in both cell-associated and secretory forms. A lysine-specific cysteine proteinase (Lys-gingipain, KGP), as well as an arginine-specific cysteine proteinase (Arg-gingipain), is a major trypsin-like proteinase of the organism. Recent studies indicate that the secreted KGP is implicated in the destruction of periodontal tissue and the disruption of host defense mechanisms. In this study, we have constructed a KGP-deficient mutant to determine whether the cell-associated KGP is important for pathophysiology of the organism. Although the mutant retained the strong ability to disrupt the bactericidal activity of polymorphonuclear leukocytes, its hemagglutination activity was reduced to about one-half that observed with the wild-type strain. More important, the mutant did not form black-pigmented colonies on blood agar plates, indicating the defect of hemoglobin adsorption and heme accumulation. Immunoblot analysis showed that the expression of a 19-kDa hemoglobin receptor protein, which is thought to be responsible for hemoglobin binding by the organism, was greatly retarded in this mutant. The mutant also showed a marked decrease in the ability to degrade fibrinogen. These results suggest the possible involvement of KGP in the hemoglobin binding and heme accumulation of the organism and in the bleeding tendency in periodontal pockets.
Highlights
The proteinases of the Gram-negative, black-pigmented anaerobe Porphyromonas gingivalis are believed to be involved in a wide range of pathologies of progressive periodontal disease
In our previous study [20], we constructed RGP-deficient mutants from P. gingivalis ATCC33277 and provided evidence suggesting that RGP plays critical roles in inhibition of the bactericidal activity of polymorphonuclear leukocyte (PMN) and the hemagglutination by the organism
We have previously shown that the initial translation product of KGP, like RGP, contains the hemagglutinin-related sequence in the carboxyl-terminal domain [24] and that the RGPnull mutant KDP112 is almost devoid of the hemagglutinating activity and loses the inhibitory effect on the bactericidal activity of PMNs [20]
Summary
The proteinases of the Gram-negative, black-pigmented anaerobe Porphyromonas gingivalis are believed to be involved in a wide range of pathologies of progressive periodontal disease (reviewed in Refs. 1 and 2). The culture supernatant of the KGP-deficient mutant (KDM35), like the wild-type strain and KDM16, resulted in the intense inhibition of the CL response of PMNs, suggesting that contribution of KGP to the inhibition of the bactericidal activity of PMNs by the culture supernatant of P. gingivalis is not as much as that of RGP and that KGP is not directly involved in the production of P. gingivalis factors responsible for the inhibition.
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