Detection and possible biological role of chondroitinase and heparitinase enzymes produced by Porphyromonas gingivalis W50

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Abstract
Gingival crevicular fluid levels of the glycosaminoglycan (GAG) chondroitin-4-sulphate (C-4-S) have received increased attention as potential indicators of periodontal tissue turnover. However, little is known about the relationship between crevicular fluid connective tissue metabolites and microbial factors. In this study Porphyromonas gingivalis, a periodontopathogen, was investigated for its ability to degrade the GAGs C-4-S, dermatan sulphate (DS) and heparan sulphate (HS) in vitro. The effect of P. gingivalis extracts on the proteoglycans (PG) derived from human gingiva were also investigated. The presence of chondroitinase and heparitinase eliminase enzymes were identified from the vesicle fraction of P. gingivalis W50. These enzymes were extracted from the vesicle fraction by a differential centrifugation technique and partially purified by non-denaturing gel filtration chromatography which revealed heparitinase enzyme peaks at 200 and 150 kDa and chondroitinase at 70 kDa. Gingival proteoglycans for use as substrates were purified using 4 M guanidinium chloride extraction and anion exchange chromatography; these proteoglycans contained 48% DS, 27% C-4-S and 13% HS P. gingivalis chondroitinase and heparitinase enzymes were capable of the degradation of C-4-S and HS but not DS GAGs. The presence of chondroitinase enzymes produced by P. gingivalis may influence levels of connective tissue metabolites in crevicular fluid. Furthermore these enzymes, particularly the heparitinase, may be involved in the initial permeation of the gingival epithelium, permitting the ingress of further microbial virulence factors.