Hydrolysis of Epithelial Junctional Proteins byPorphyromonas gingivalisGingipains

Abstract

Porphyromonas gingivalis has been implicated as an etiologic agent of adult periodontitis. We have previously shown that P. gingivalis can degrade the epithelial cell-cell junction complexes, thus suggesting that this bacterium can invade the underlying connective tissues via a paracellular pathway. However, the precise mechanism(s) involved in this process has not been elucidated. The purpose of this study was to determine if the arginine-and lysine-specific gingipains of P. gingivalis (i.e., HRgpA and RgpB, and Kgp, respectively) were responsible for the degradation of E-cadherin, the cell-cell adhesion protein in the adherens junctions. In addition, we compared the degradative abilities of the whole gingipains HRgpA and Kgp to those of their catalytic domains alone. In these studies, immunoprecipitated E-cadherin as well as monolayers of polarized Madin-Darby canine kidney (MDCK) epithelial cell cultures were incubated with the gingipains and hydrolysis of E-cadherin was assessed by Western blot analysis. Incubation of P. gingivalis cells with immunoprecipitated E-cadherin resulted in degradation, whereas prior exposure of P. gingivalis cells to leupeptin and especially acetyl-Leu-Val-Lys-aldehyde (which are arginine-and lysine-specific inhibitors, respectively) reduced this activity. Furthermore, incubation of E-cadherin immunoprecipitates with the different gingipains resulted in an effective and similar hydrolysis of the protein. However, when monolayers of MDCK cells were exposed to the gingipains, Kgp was most effective in hydrolyzing the E-cadherin molecules in the adherens junction. Kgp was more effective than its catalytic domain in degrading E-cadherin at 500 nM but not at a lower concentration (250 nM). These results suggest that the hemagglutinin domain of Kgp plays a role in degradation and that there is a critical threshold concentration for this activity. Taken together, these results provide evidence that the gingipains, especially Kgp, are involved in the degradation of the adherens junction of epithelial cells, which may be important in the invasion of periodontal connective tissue by P. gingivalis.

Porphyromonas Gingivalisand Dextran Sulfate Sodium Induce Periodontitis Through the Disruption of Physical Barriers in Mice

“…Although the mechanism through which DSS inhibits ZO-1 expression and disrupts TJ is not known, DSS, a polyanionic derivative of dextran, may dissociate TJ structures by interacting with the basic residues at the extracellular loops of claudins (23). Given that gingipain degrades the adherens junction protein E-cadherin (24), the reduction in ZO-1 expression induced by P. gingivalis may also be attributable to gingipain. P. gingivalisinduced epithelial cell rounding and detachment was also reported to be dependent on gingipain (25).…”

In Vitro Models of Tissue Penetration and Destruction by Porphyromonas gingivalis

“…Gingipains of P. gingivalis were recently implicated in the proteolysis of junction proteins and were suggested to contribute significantly to bacterial tissue penetration (17,18). Gene inactivation of Arg-or Lys-gingipain result in a decreased ability of P. gingivalis to bind to epithelial cells and the extracellular matrix (28) as well as in a reduction in the colonization and pathogenic properties of the bacteria (8).…”

Functional Implication of the Hydrolysis of Platelet Endothelial Cell Adhesion Molecule 1 (CD31) by Gingipains ofPorphyromonas gingivalisfor the Pathology of Periodontal Disease

“…The outer membraneassociated gingipains (RgpA and Kgp) extracted from P. gingivalis ATCC 33277 contain a catalytic domain and a hemagglutinin/adhesin domain (54,70). Cysteine proteinase activities may affect cytokine inactivation and degradation (69,70), acquisition of metabolically necessary iron and porphyrin from hemoglobin (14), enhancement of vascular permeability through plasma prekallikrein activation and bradykinin release (33), and degradation of epithelial cell-cell junctional complexes (37).…”