A trypsin‐like protease from Bacteroides gingivalis: partial purification and characterization

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Abstract
Extracts of cell sonicates of Bacteroides gingivalis were shown to contain proteo‐lytic enzymes capable of degrading connective tissue proteins. In this study, neutral proteolytic enzymes, i.e. collagenase and a trypsin‐like protease, were isolated. The trypsin‐like protease was readily separated from collagenase by affinity chromatography on Benzamidine‐Sepharose. Proteases were further purified by gel filtration on Sephacryl S‐200; apparent molecular weights of 35 kDa and 70 kDa were obtained for a trypsin‐like protease and collagenase, respectively. Further characterization of the potent trypsin‐like protease showed that the enzyme was inhibited by serine protease inhibitors phenylmethylsulfonyl fluoride and benzamidine and by metalloprotease inhibitor EDTA, as well as ascorbic acid. Activation of the enzyme was observed with reducing agents and human serum. The trypsin‐like protease was found to be capable of degrading native type IV collagen and denatured type I collagen but not native type I collagen. Thus, we conclude that in addition to collagenase a potent trypsin‐like protease from Bacteroides gingivalis may be involved in the etiopathogenesis of periodontal disease. Since the trypsin‐like protease is able to degrade the basement membrane collagen (type IV) in the presence of human serum, this enzyme may be a potent virulence factor of Bacteroides gingivalis in relation to invasiveness and connective tissue destruction.