Skip to main content

A novel fibrinolytic enzyme from Cordyceps militaris, a Chinese traditional medicinal mushroom

Abstract

A novel fibrinolytic enzyme from Cordyceps militaris was purified and partially characterized for the first time, which was designated C. militaris fibrinolytic enzyme (CMase). This extracellular enzyme from C. militaris was isolated by ammonium sulphate fraction, and purified to electrophoretic homogeneity using gel filtration chromatography. The apparent molecular mass of the purified enzyme was estimated to be 27.3 kDa by SDS-PAGE. The optimum pH and temperature for the enzyme activity were pH 6.0 and 25 °C, respectively. In the presence of metal ions such as Mg2+ and Fe2+ ions the activity of the enzyme increased, whereas EDTA and Cu2+ ion inhibited the enzyme activity. Interestingly the N-terminal amino acid sequences of the enzyme is extremely similar to those of the trypsin proteinases from insects, and has no significant homology with those of the fibrinolytic enzyme from other medicinal mushroom. In conclusion, C. militaris produces a strong fibrinolytic enzyme CMase and may be considered as a new source for thrombolytic agents.

This is a preview of subscription content, access via your institution.

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6

References

  • Ahn YJ, Park SJ, Lee SG, Choi DH (2000) Cordycepin: selective inhibitor derived from liquid culture of Cordyceps militaris against Clostrdium Spp. J Agric Food Chem 48:2744–2748

    Article  CAS  Google Scholar 

  • Altschul SF, Madden TL, Schäffer AA et al (1997) Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. Nucleic Acids Res 25:3389–3402

    Article  CAS  Google Scholar 

  • Astrup T, Mullertz S (1952) The fibrin plate method for estimating of fibrinolytic activity. Arch Biochem Biophys 40:346–351

    Article  CAS  Google Scholar 

  • Bode C, Runge M, Smalling RW (1996) The future of thrombolysis in the treatment of acute myocardial infarction. Eur Heart J 17:55–60

    Google Scholar 

  • 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

    Article  CAS  Google Scholar 

  • Choi HS (1999) Purification and characterization of metalloproteases from Pleurotus sajor-caju. J Microbiol.Biotechnol 5:675–678

    Google Scholar 

  • Choi N-S, Seo S-Y, Kim S-H (1999) Screening of mushroom having fibrinolytic activity. Kor J Food Sci Technol 31(2):553–557

    Google Scholar 

  • Gaines PJ, Sampson CM, Rushlow KE, Stiegler GL (1999) Cloning of a family of serine protease genes from the cat flea Ctenocephalides felis. Insect Mol Biol 81(1):11–22

    Article  Google Scholar 

  • Green AA, Hughens WL (1955) Protein fractionation on the basis of solubility in Aqueous solutions of salts and organic solvents. Methods Enzymol 1:67–90

    Article  CAS  Google Scholar 

  • Hattori M, Isomura S, Yokoyama E et al (2005) Extracellular trypsin-like proteases produced by Cordyceps militaris. J Biosci Bioeng 100(6):631–636

    Article  CAS  Google Scholar 

  • Joh J-H, Kim B-G, Kong W-S et al (2004) Cloning and developmental expression of a family metalloprotease cDNA from oyster mushroom Pleurotus ostreatus. FEMS Microbiol Lett 239:57–62

    Article  CAS  Google Scholar 

  • Kim JH, Kim YS (2001) Characterization of a metalloenzyme from a wild mushroom Tricholoma saponaceum. Biosci Biotechnol Biochem 65:356–362

    Article  CAS  Google Scholar 

  • Kim W, Choi K, Kim Y (1996) Purification and characterization of a fibrinolytic enzyme produced from Bacillus sp. Strain CK11–4 screened from Chungkook-Jang. Appl Environ Microbiol 62:2482–2488

    CAS  Google Scholar 

  • Kim J, Lee H, Yoo K, Kim Y, Seok S, Kim Y (1998a) The screening of fibrinolytic activities of extracts from mushroom in Mt Chiak. J Korea Mycol 26:589–593

    Google Scholar 

  • Kim S-H, Choi N-S, Lee W-Y (1998b) Fibrin zymography: a direct analysis of fibrinolytic proteases on gels. Anal Biochem 263(1):115–116

    Article  CAS  Google Scholar 

  • Laemmli UK (1970) Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227:680–685

    Article  CAS  Google Scholar 

  • Lee S-Y, Kim J-S, Kim J-E et al (2005) Purification and characterization of fibrinolytic enzyme from cultured mycelia of Armillaria mellea. Protein Expr Purif 43(1):10–17

    Article  CAS  Google Scholar 

  • Li SP, Yang FQ, Tsim KW (2006) Quality control of Cordyceps sinensis, a valued traditional Chinese medicine. J Pharm Biomed Anal 41(5):1571–1584

    Article  CAS  Google Scholar 

  • Muller WEG, Seihard G, Beyer R et al (1977) Effect of cordycepin on nucleic acid metabolism in L5178Y cells and on nucleic acid-synthesizing enzyme sysytems. Cancer Research 37:3824–3833

    CAS  Google Scholar 

  • Nonaka T, Ishikawa H, Tsumuraya Y et al (1995) Characterization of a thermostable lysine-specific metalloendopeptidase from the fruiting bodies of a basidiomycete Grifola frondosa. J Biochem 118(5):1014–1020

    CAS  Google Scholar 

  • Sugar AM, McCaffrey RP (1998) Antifungal activity of 3′-deoxyadenosine (cordycepin). Antimicrob Agents Chemother 42:1424–1427

    CAS  Google Scholar 

  • Sumi H, Hamada H, Tsushima H et al (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

    Article  CAS  Google Scholar 

  • Wong AHK, Mine Y (2004) A novel fibrinolytic enzyme in fermented shrimp paste: a traditional Asian fermented seasoning. J Agric Food Chem 52:980–986

    Article  CAS  Google Scholar 

  • Zhou X, Meyer CU, Schmidtke P, Zepp F (2002) Effect of cordycepin on interleukin-10M production perioheral blood mononuclear cells. Eur J Pharmarcol 453:309–317

    Article  CAS  Google Scholar 

  • Zhu JS, Halpern GM, Johns K (1998a) The scientific rediscovery of an ancient Chinese herbal medicine: Cordyceps sinensis: part I. J Altern Complement Med 4:289–303

    Article  CAS  Google Scholar 

  • Zhu JS, Halpern GM, Johns K (1998b) The scientific rediscovery of a precious ancient Chinese herbal regimen: Cordyceps sinensis: part II. J Altern Complement Med 4:429–457

    Article  CAS  Google Scholar 

Download references

Acknowledgements

Authors are grateful to Dr. Huaan Yang, Agriculture Department of Western Australia, Australia, for careful and critical reading of the manuscript. This work was supported by High-Tech Research and Development Program of China (No: 2006AA10Z343) and Key Scientific Project of Jiangsu Province, China (No: BE2006325).

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Ming Sheng Dong.

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Cui, L., Dong, M.S., Chen, X.H. et al. A novel fibrinolytic enzyme from Cordyceps militaris, a Chinese traditional medicinal mushroom. World J Microbiol Biotechnol 24, 483–489 (2008). https://doi.org/10.1007/s11274-007-9497-1

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s11274-007-9497-1

Keywords

  • Cordyceps militaris
  • CMase
  • Entomogenous fungus
  • Fibrinolytic activity
  • Medicinal mushroom