Abstract

To increase the fibrinolytic enzyme (nattokinase WRL101) in Bacillus subtilis WRL101, bovine fibrin or fibrinogen (1.0%, w/v) were added in the tryptic soy broth as a substrate. The fibrinolytic activity cultured in the fibrinogen-contained medium was increased. On the other hand, fibrin decreased the activity. Using the medium condition, nattokinase WRL101 (fibrinogen-induced nattokinase WRL101, FIN-WRL101) was isolated by commercial chromatographic techniques and its biochemical characteristics were investigated. The molecular weight of FIN-WRL101 was estimated to be 29 kDa. FIN-WRL101 was optimally active at pH 11.0 and 47°C. It had high degrading activity for the Aα-chain and Bb-chain of human fibrinogen, but did not affect the g-chain, indicating that it is an a-fibrinogenase. FIN-WRL101 was completely inhibited by phenylmethylsulfonyl fluoride, indicating that it belongs to the serine protease. FIN-WRL101 exhibited high specificity for Meo-Suc-Arg-Pro-Tyr-pNA (S-2586), a synthetic chromogenic substrate for chymotrypsin. Its nucleotide and amino acid sequences were determined.

Key words: Bacillus subtilis, Doenjang, fibrinolytic enzyme, nattokinase.