Purification and characterization of a novel fibrinolytic enzyme from Bacillus sp. nov. SK006 isolated from an Asian traditional fermented shrimp paste.
Bacillus sp. nov. SK006 producing four extracellular fibrinolytic enzymes was isolated from fermented shrimp paste, a traditional and popular Asian seasoning. One fibrinolytic enzyme was purified to homogeneity with a molecular mass of 43-46 kDa by SDS-PAGE and gel filtration chromatography. The specific activity was determined to be 11.2 units/mg using plasmin as a standard. The enzyme displayed optimal activity at 30 degrees C and pH 7.2. It was stable below 40 degrees C for 4 h between pH 5.0 and pH 11.0. Zinc ion stimulated the enzyme activity whereas Cu2+, Ca2+, Fe3+, and Hg2+ caused its inhibition. The fibrinolytic activity was strongly inhibited by PMSF and moderately inhibited by EDTA as well as PCMB. The enzyme exhibited a higher affinity toward N-Succ-Ala-Ala-Pro-Phe-pNA and was able to degrade fibrin clots either by forming active plasmin from plasminogen or by direct fibrinolysis. The N-terminal amino acid sequence was found to be AQSVPYEQPHLSQ, which is different from that of other known fibrinolytic enzymes.
Hua
Ying
Y
State Key Laboratory of Food Science and Technology, Jiangnan University, Jiangsu, People's Republic of China.
Jiang
Bo
B
Mine
Yoshinori
Y
Mu
Wanmeng
W
D000595:Amino Acid Sequence; D000818:Animals; D001407:Bacillus / Q000201:enzymology*; D002850:Chromatography, Gel; D004591:Electrophoresis, Polyacrylamide Gel; D010450:Endopeptidases / Q000737:chemistry / Q000302:isolation & purification*; D004789:Enzyme Activation; D004791:Enzyme Inhibitors; D005285:Fermentation; D005341:Fibrinolysin / Q000378:metabolism; D005342:Fibrinolysis; D005421:Flavoring Agents / Q000737:chemistry; D005516:Food Microbiology; D006863:Hydrogen-Ion Concentration; D008969:Molecular Sequence Data; D008970:Molecular Weight; D033561:Penaeidae; D010958:Plasminogen / Q000378:metabolism; D013379:Substrate Specificity; D013696:Temperature
D004791:Enzyme Inhibitors; D005421:Flavoring Agents; D010958:Plasminogen; D010450:Endopeptidases; D005341:Fibrinolysin