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. 1996 Mar;64(3):756–762. doi: 10.1128/iai.64.3.756-762.1996

Cysteine protease of Porphyromonas gingivalis 381 enhances binding of fimbriae to cultured human fibroblasts and matrix proteins.

M Kontani 1, H Ono 1, H Shibata 1, Y Okamura 1, T Tanaka 1, T Fujiwara 1, S Kimura 1, S Hamada 1
PMCID: PMC173834  PMID: 8641778

Abstract

It has been shown that Porphyromonas gingivalis 381, a suspected periodontopathogen, possesses fimbriae on its cell surface. The organism is also known to produce proteases which can degrade the host cell surface matrix proteins. In this study, we investigated the effect of protease on the binding of the purified P. gingivalis fimbriae to cultured fibroblasts or matrix proteins. A protease that can hydrolyze benzoyl-L-arginine p-nitroanilide was obtained from P. gingivalis 381 cells by sonication in phosphate-buffered 0.2% Triton X-100 and was purified by column chromatography. The molecular size of the protease was estimated to be 55 kDa by gel filtration or 47 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis. The enzyme activity was markedly inhibited by sulfhydryl reagents, antipain, and leupeptin. The protease degraded various host proteins, including collagen and fibronectin, and cleaved the COOH terminus of the arginine residue in peptides such as benzoyl-L-arginine p-nitroanilide. However, P. gingivalis fimbriae were not degraded by protease activity. The enzyme activity was enhanced in the presence of reducing agents or CaCl2. When cultured fibroblasts were partially treated with the protease, the binding of the purified P. gingivalis fimbriae to the fibroblast monolayer was increased significantly. However, this enhancing effect was suppressed upon the addition of antipain and leupeptin. Similarly, binding of the fimbriae to the collagen or fibronectin immobilized on the microtiter wells was also enhanced. Addition of these host matrix proteins efficiently inhibited the binding of fimbriae to the fibroblast monolayer. The binding assay of fimbriae using dipeptidyl ligand affinity column chromatography demonstrated a clear interaction between fimbriae and the arginine residue. Taken together, these results indicate that the P. gingivalis protease at least partially degrades the host matrix proteins, which, in turn, may lead to an increased exposure of the cryptic ligands that can result in enhanced fimbria-mediated binding of this organism to periodontal tissues.

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Selected References

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