Identification and characterization of a chymotrypsin-like serine protease from periodontal pathogen, Tannerella forsythia

• Identification of a T. forsythia protease that degrades gelatin, type I collagen, and N-succinyl-Ala-Ala-Pro-Phe p-nitroanilide.
• Isolated T. forsythia protease is inhibited by serine protease inhibitors antipain and phenylmethanesulfonyl fluoride.
• Isolated T. forsythia protease optimally cleaved gelatin at pH 7.5 and type I collagen at 8.0.
• Isolated T. forsythia protease does not exhibit thermal stability, exhibiting complete loss of activity at 60 °C.

Tannerella forsythia is a bacteria associated with severe periodontal disease. This study reports identification and characterization of a membrane-associated serine protease from T. forsythia. The protease was isolated from T. forsythia membrane fractions and shown to cleave both gelatin and type I collagen. The protease was able to cleave both substrates over a wide range of pH values, however optimal cleavage occurred at pH 7.5 for gelatin and 8.0 for type I collagen. The protease was also shown to cleave both gelatin and type I collagen at the average reported temperature for the gingival sulcus however it showed a lack of thermal stability with a complete loss of activity by 60 °C. When treated with protease inhibitors the enzyme's activity could only be completely inhibited by serine protease inhibitors antipain and phenylmethanesulfonyl fluoride (PMSF). Further characterization of the protease utilized serine protease synthetic peptides. The protease cleaved N-succinyl-Ala-Ala-Pro-Phe p-nitroanilide but not Nα-benzoyl-dl-arginine p-nitroanilide (BAPNA) or N-methoxysuccinyl-Ala-Ala-Pro-Val p-nitroanilide indicating that the protease is a chymotrypsin-like serine protease. Since type I collagen is a major component in the gingival tissues and periodontal ligament, identification and characterization of this enzyme provides important information regarding the role of T. forsythia in periodontal disease.