Purification and characterization of a fibrinolytic enzyme produced from Bacillus sp strain CK 11-4 screened from Chungkook-Jang

Bacillus sp, strain CK 11-4, which produces a strongly fibrinolytic enzyme, was screened from Chungkook-Jang, a traditional Korean fermented-soybean sauce, The fibrinolytic enzyme (CK) was purified from supernatant of Bacillus sp, strain CK 11-4 culture broth and showed thermophilic, hydrophilic, and strong fibrinolytic activity, The optimum temperature and pH were 70 degrees C and 10.5, respectively, and the molecular weight was 28,200 as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, The first 14 amino acids of the N-terminal sequence of CK are Ala-Gln-Thr-Val-Pro-Tyr-Gly-Ile-Pro-Leu-IIe-Lys-Ala-Asp. This sequence is identical to that of subtilisin Carlsberg and different from that of nattokinase, but CK showed a level of fibrinolytic activity that was about eight times higher than that of subtilisin Carlsberg, The amidolytic activity of CK increased about twofold at the initial state of the reaction when CK enzyme was added to a mixture of plasminogen and substrate (H-D-Val-Leu-Lys-rho NA). A similar result was also obtained from fibrin plate analysis.