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Purification and Characteristics of Fibrinolytic Enzyme from Chongkukjang

  • Yang, Jeong-Lye (School of Food and Life Science, Biohealth Product Research Center and Food Science Institute, Inje University) ;
  • Kim, Hee-Sook (Department of Food Science and Technology, Kyungsung University) ;
  • Hong, Jeong-Hwa (School of Food and Life Science, Biohealth Product Research Center and Food Science Institute, Inje University) ;
  • Song, Young-Sun (School of Food and Life Science, Biohealth Product Research Center and Food Science Institute, Inje University)
  • Published : 2006.06.01

Abstract

Bacillus sp. strain K-l, which produces a strong fibrinolytic enzyme, was isolated from chongkukjang, a traditional Korean fermented soybean paste. The fibrinolytic enzyme was purified from chongkukjang base by using ammonium sulfate fractionation and chromatographic techniques. Purified enzyme, CK K-1 was demonstrated to be homogeneous by SDS-PAGE and isoelectric focusing electrophoresis, and has molecular mass of a 12.4 kDa and a pI of 8.0. The optimal reaction pH value and temperature were 8.0 and $40^{\circ}C$, respectively. Phenyl-methyl-sulfonyl-fluoride (PMSF; serine protease inhibitor), ethylene-diamine-tetra-acetic acid (EDTA; metallo protease inhibitor), copper ion, ferric ion and lead ion inhibited the enzyme activity. These results indicated that the fibrinolytic enzyme is a metallo-serine protease and different from nattokinase and chongkukjangkinase.

Keywords

References

  1. Voet D, Voet JG. 1990. Biochemistry. John Wiley & Sons, New York. p 1087-1095
  2. Wun TC, Schleuning WD, Reich E. 1982. Isolation and characterization of urokinase from human plasma. J Biol Chem 257: 3276-3283
  3. Medved LV, Solovjov DA, Ingham KC. 1996. Domain structure, stability and interaction in streptokinase. Eur J Biochem 239: 333-339 https://doi.org/10.1111/j.1432-1033.1996.0333u.x
  4. Pennica D, Holmes WE, Kohr WJ, Harkins RN, Vehar GA, Ward CA, Bennn WF, Yelverton E, Seeburg PH, Heyneker HL, Goeddel DV, Cohen D. 1982. Clonning and expression of human tissue-type plasminogen activator cDNA in E. coli. Nature 301: 214-221 https://doi.org/10.1038/301214a0
  5. Sumi H, Hamada H, Tsushima H, Mihara H, Muraki H. 1987. A novel fibrinolytic enzyme (nattokinase) in the vegetable cheese natto: a typical and popular soybean food in the Japanese diet. Experientia 43: 1110-1111 https://doi.org/10.1007/BF01956052
  6. Nakajima N, Tayu N, Sumi H. 1993. Potentfibrinolytic enzyme from the lysate of Katsuwohus pelamis digestive tract (Shiokara): purification and characterization. Biosci Biotech Biochem 57: 1604-1605 https://doi.org/10.1271/bbb.57.1604
  7. Kim SH, Choi NS. 2000. Purification and characterization of subtilisin DJ-4 secreted by Bacillus sp. strain DJ-4 screened from Doen-Jang. Biosci Biotechnol Biochem 64: 1722-1725 https://doi.org/10.1271/bbb.64.1722
  8. Kim HK, Kim GT, Kim DK, Choi WA, Park SH, Jeong YK, Kong IS. 1997. Purification and characterization of a novel fibrinolytic enzyme from Bacillus sp. KA38 orig-inated from fermented fish. J Ferment Bioeng 84: 307-312 https://doi.org/10.1016/S0922-338X(97)89249-5
  9. Sumi H, Hamada H, Nakanishi K, Hiratani H. 1990. Enhancement of the fibrinolytic activity in plasma by oral administration of nattokinase. Acta Haematol 84: 139-143 https://doi.org/10.1159/000205051
  10. Kim JH, Kim YS. 1999. A fibrinolytic metalloprotease from the fruiting bodies of an edible mushroom, Armillariella mellea. Biosci Biotechnol Biochem 63: 2130-2136 https://doi.org/10.1271/bbb.63.2130
  11. Bradford MM. 1976. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72: 248-254 https://doi.org/10.1016/0003-2697(76)90527-3
  12. Astrup T, Mullertz S. 1952. The fibrin plate method for estimating fibrinolytic activity. Arch Biochem Biophys 40: 346-351 https://doi.org/10.1016/0003-9861(52)90121-5
  13. Laemmli UK. 1970. Cleavage of structural protein during the assembly of the head of bacteriophage T4. Nature 227: 680-685 https://doi.org/10.1038/227680a0
  14. Kim SH, Choi NS. 1999. Electrophoretic analysis of protease inhibitors in fibrin zymography. Anal Biochem 270: 179-181 https://doi.org/10.1006/abio.1999.4080
  15. Hong JH, Youn HK, Kang MC, Lee HJ, Hur SH. 2006. Properties of Bacillus spp. isolated from fermenting chongkukjang. J Korean Soc Ind Food Technol 4: 67-72
  16. Holt JG, Sharpe ME, Mair NS, Sneath PH. 1989. Bergey's Manual of Systematic Bacteriology. Williams and Wilkins press, Philadelphia. Vol 2, p 1105
  17. Kim W, Choi K, Kim Y, Park H, Choi J, Lee Y. 1996. Purification and characterization of a fibrinolytic enzyme produced from Bacillus sp. strain CK 11-4 screened from Chungkuk-jang. Appl Environ Microbiol 62: 2482-2488
  18. Noh KA, Kim DH, Choi NS, Kim SH. 1999. Isolation of fibrinolytic enzyme producing strains from kimchi. Korean J Food Sci Technol 31: 219-223
  19. Whitaker JR. 1994. Enzyme inhibitor. In Principles of Enzymology for the Food Science. 2nd ed. Whitaker JR, ed. Dekker, New York. p 241-270

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