Cloning and Expression of One Fibrinolytic Enzyme from Bacillus sp. zlw-2

The gene encoding fibrinolytic enzyme from Bacillus sp. zlw-2 was cloned and sequenced （accession no. EU734749）, which was 1146 bp, encoded 381 amino acids and had 99% homology with Nattokinase YF308 and NAT. The genes encoding pre-pro-fibrinolytic enzyme （including signal peptide, propeptide, and mature peptide） and fibrinolytic enzyme （including mature peptide） were cloned into pET28a vector respectively and then transformed into Escherichia coli BL21 （DE3）. The recombinant ofpre-pro-fibrinolytic enzyme showed enzyme activity of 183 U mL^-1, while no detectable enzyme activity could be found from the recombinant of the mature peptide.

Enzymatic properties of UFE,a novel marine fibrinolytic enzyme

A novel potent protease, Urechis unicinctus fibrinolytic enzyme （UFE）, was firstly discovered. The enzymatic properties of UFE were further investigated.As a low molecular mass protein,UFE appeared to be very stable to heat and pH.When temperature was below 50 ℃ ,the remnant enzyme activity remained almost unchanged, but when temperature was raised to 60 ℃ ,the remnant enzyme activity began to decrease rapidly. UFE was quite stable in the range of pH value from 3 to 12,especially in slightly alkaline pH value.Mn^2＋ ,Cu^2＋ and Fe^2＋ ions were activators of UFE, while Fe^3＋ and Ag^＋ ions were inhibitors of UFE.Fe^2＋ ion along with Fe^3＋ ion might regulate UFE activity in vivo. The optimum pH and temperature of UFE were about 8 and 50 ℃ ,respectively. Other characteristics of this enzyme were also studied. Systematic research results are significant when UFE is applied for medical and industrial purposes.

Optimization of Douchi Fibrinolytic Enzyme Production by Statistical Experimental Methods

Thrombus disease, one of the common cardiovascular diseases, has attracted worldwide at- tention for its rising mortality and morbidity. Due to the distinct shortages of current fibrinolytic drugs, new fibrinolytic agents warrant investigation. In this study, 8 fibrinolytic enzyme-producing strains were isolated from Douchi--a traditional Chinese food, and strain XY-1 which produced the largest amount of the enzyme was chosen for the following experiments. The enzyme produced by strain XY-1 was named Douchi fibrinolytic enzyme （DFE）. We optimized the liquid culture medium of strain XY-1 for enzyme production using Plackett-Burman and Box-Behnken design. The predicted maximal DFE yield was 19.78 FU/mL with 11.4 g/L peptone, 0.5 g/L magnesium sulfate and 1 g/L sodium chloride. How- ever, we acquired maximal production of 21.33 FU/mL in actual experiments, equal to 107.84% of the theoretical value, and the yield had been increased by 79.55% as compared to the yield of un-optimized culture. It was demonstrated that the combined use of Plackett-Burman design and response surface methodology in fermentation optimization can effectively and rapidly increase DFE production.

Screening and Preservation of Douchi Fibrinolytic Enzyme Producing Strain

[Objective] To isolate and preserve a high-activity Douchi fibrinolytic enzyme producing strain from Douchi products. [Method] The Douchi flbrinolytic enzyme producing strain was screened on the selected medium prepared with self-made pork blood powder, the strain with the highest activity was screened out according to the size of hydrolysis cycle, and then preserved in LB medium. [ Rebait] A Douchi fibrinolytic enzyme producing strain with high thrombolytic activity was successfully screened out from the Douchi produced in Hunan, Guangdong and Jiangxi Prov- inces. [ Ceadusioe] The study lays foundation for the development of new-type thrombolytic drugs.

Studies on a Novel Fibrinolytic Enzyme(Nattokinase)in the Vegetable Cheese Natto

The culture condition of bacillus strain and the extraction method of Nattokinase were reported. The fibrinolytic activity of nattokinese was appraised.