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Fibrinolytic Serine Protease Isolation from Bacillus amyloliquefaciens An6 Grown on Mirabilis jalapa Tuber Powders

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Abstract

In this study, Mirabilis jalapa tuber powder (MJTP) was used as a new complex organic substrate for the growth and production of fibrinolytic enzymes by a newly isolated Bacillus amyloliquefaciens An6. Maximum protease activity (1,057 U/ml) with casein as a substrate was obtained when the strain was grown in medium containing (grams per liter) MJTP 30, yeast extract 6, CaCl2 1, K2HPO4 0.1, and K2HPO4 0.1. The strain was also found to grow and produce extracellular proteases in a medium containing only MJTP, indicating that it can obtain its carbon, nitrogen, and salts requirements directly from MJTP. The B. amyloliquefaciens An6 fibrinase (BAF1) was partially purified, and fibrinolytic activity was assayed in a test tube with an artificial fibrin clot. The molecular weight of the partially purified BAF1 fibrinolytic protease was estimated to be 30 kDa by sodium dodecyl sulfate polyacrylamide gel electrophoresis and gel filtration. The optimum temperature and pH for the caseinolytic activity were 60 °C and 9.0, respectively. The enzyme was highly stable from pH 6.0 to 11.0 and retained 62% of its initial activity after 1 h incubation at 50 °C. However, the enzyme was inactivated at higher temperatures. The activity of the enzyme was totally lost in the presence of phenylmethylsulfonyl fluoride, suggesting that BAF1 is a serine protease.

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References

  1. Kim, W., Choi, K., Kim, Y., Park, H., Choi, J., Lee, Y., et al. (1996). Applied and Environmental Microbiology, 62, 2482–2488.

    CAS  Google Scholar 

  2. Kim, S. H., & Choi, N. S. (2000). Bioscience, Biotechnology, and Biochemistry, 64, 1722–1725.

    Article  CAS  Google Scholar 

  3. Mine, Y., Wong, A. H. K., & Jiang, B. (2005). Food Research International, 38, 243–250.

    Article  CAS  Google Scholar 

  4. Sherry, S. (1987). American Journal of Cardiology, 59, 984–989.

    Article  CAS  Google Scholar 

  5. Sumi, H., Hamada, H., Tsushima, H., Mihara, H., & Muraki, H. (1987). Experientia, 43, 1110–1111.

    Article  CAS  Google Scholar 

  6. Mihara, H., Sumi, H., Yoneta, T., Mizumoto, H., Ikeda, R., Seiki, M., et al. (1991). Japanese Journal of Physiology, 41, 461–472.

    Article  CAS  Google Scholar 

  7. Nakamura, T., Yamagata, Y., & Ichishima, E. (1992). Bioscience, Biotechnology, and Biochemistry, 56, 1869–1871.

    Article  CAS  Google Scholar 

  8. Fujita, M., Nomura, K., Hong, K., Ito, Y., Asada, A., & Nishimuro, S. (1993). Biochemical and Biophysical Research Communications, 197, 1340–1347.

    Article  CAS  Google Scholar 

  9. Sumi, H., Hamada, H., Nakanishi, K., & Hiratani, H. (1990). Acta Haematologica, 84, 139–143.

    Article  CAS  Google Scholar 

  10. Peng, Y., Huang, Q., Zhang, R. H., & Zhang, Y. Z. (2003). Comparative Biochemistry and Physiology Part B, 134, 45–52.

    Article  Google Scholar 

  11. Johnvesly, B., & Naik, G. R. (2001). Process Biochemistry, 37, 139–144.

    Article  CAS  Google Scholar 

  12. Puri, S., Beg, Q. K., & Gupta, R. (2002). Current Microbiology, 44, 286–290.

    Article  CAS  Google Scholar 

  13. Frankena, J., Koningstein, G. M., Van Verseveld, H. W., & Stouthamer, A. H. (1986). Applied Microbiology and Biotechnology, 24, 106–112.

    CAS  Google Scholar 

  14. Giesecke, U. E., Bierbaum, G., Rudde, H., Spohn, U., & Wandrey, C. (1991). Applied Microbiology and Biotechnology, 35, 720–724.

    Article  CAS  Google Scholar 

  15. Drucker, H. (1972). Journal of Bacteriology, 110, 1041–1049.

    CAS  Google Scholar 

  16. Ferrero, M. A., Castro, G. R., Abate, C. M., Baigori, M. D., & Sineriz, F. (1996). Applied Microbiology and Biotechnology, 45, 327–332.

    Article  CAS  Google Scholar 

  17. Gessesse, A., & Gashe, B. A. (1997). Biotechnological Letters, 19, 479–481.

    Article  CAS  Google Scholar 

  18. Mehrotra, S., Pandey, P. K., Gaur, R., & Darmwal, N. S. (1999). Bioresource Technology, 67, 201–203.

    Article  CAS  Google Scholar 

  19. Ghorbel-Frikha, B., Sellami-Kamoun, A., Fakhfakh, N., Haddar, A., Manni, L., & Nasri, M. (2005). Journal of Industrial Microbiology & Biotechnology, 32, 186–194.

    Article  CAS  Google Scholar 

  20. Ellouz, Y., Bayoudh, A., Kammoun, S., Gharsallah, N., & Nasri, M. (2001). Bioresource Technology, 80, 49–51.

    Article  CAS  Google Scholar 

  21. Joo, H. S., & Chang, C. S. (2005). Process Biochemistry, 40, 1263–1270.

    Article  CAS  Google Scholar 

  22. Naidu, K. S. B., & Devi, K. L. (2005). African Journal of Biotechnology, 4, 724–726.

    CAS  Google Scholar 

  23. Tari, C., Genckal, H., & Tokatli, F. (2006). Process Biochemistry, 41, 659–665.

    Article  CAS  Google Scholar 

  24. Vivanco, J. M., Querci, M., & Salazar, L. F. (1999). Plant Disease, 83, 1116–1121.

    Article  CAS  Google Scholar 

  25. Yang, S. W., Ubillas, R., McAlpine, J., Stafford, A., Ecker, D. M., Talbot, M. K., et al. (2001). Journal of Natural Products, 64, 313–317.

    Article  CAS  Google Scholar 

  26. DeBolle, M., Eggermont, K., Duncan, R., Osborn, R., Terras, F., & Broekaert, W. (1995). Plant Molecular Biology, 28, 713–721.

    Article  CAS  Google Scholar 

  27. Wang, Y., Chen, J., Yang, Y., Zheng, Y., Tang, S., & Luo, S. (2002). Helvetica Chimica Acta, 85, 2342–2348.

    Article  CAS  Google Scholar 

  28. Miller, J. H. (1972). Experiments in molecular genetics (pp. 431–435). Cold Spring Harbor: Cold Spring Harbor Laboratory.

    Google Scholar 

  29. Hajji, M., Rebai, A., Gharsallah, N., & Nasri, M. (2008). Applied Microbiology and Biotechnology, 79, 915–923.

    Google Scholar 

  30. Jellouli, K., Bayoudh, A., Manni, L., Agrebi, R., & Nasri, M. (2008). Applied Microbiology and Biotechnology, 79, 989–999.

    Google Scholar 

  31. Kembhavi, A. A., Kulharni, A., & Pant, A. A. (1993). Applied Biochemistry and Biotechnology, 38, 83–92.

    Article  CAS  Google Scholar 

  32. Bradford, M. M. (1976). Analytical Biochemistry, 72, 248–254.

    Article  CAS  Google Scholar 

  33. Laemmli, U. K. (1972). Nature, 227, 680–685.

    Article  Google Scholar 

  34. Garcia-Carreno, F. L., Dimes, L. F., & Haard, N. F. (1993). Analytical Biochemistry, 214, 65–69.

    Article  CAS  Google Scholar 

  35. North, M. J. (1982). Experimental Mycology, 6, 345–352.

    Article  CAS  Google Scholar 

  36. Hinman, R. L. (1994). Chemtech, 24, 45–48.

  37. Ko, J. H., Yan, J. P., Zhu, L., & Qi, Y. P. (2004). Comparative Biochemistry and Physiology. Part C. Toxicology & Pharmacology, 137, 65–74.

    Article  Google Scholar 

  38. Choi, N. S., & Kim, S. H. (2001). Journal of Biochemistry and Molecular Biology, 34, 134–138.

    CAS  Google Scholar 

  39. Kho, C. W., Park, S. G., Cho, S., Lee, D. H., Myung, P. K., & Park, B. C. (2005). Protein Expression and Purification, 39, 1–7.

    Article  CAS  Google Scholar 

  40. Chang, C. T., Fan, M. H., Kuo, F. C., & Sung, H. Y. (2000). Journal of Agricultural and Food Chemistry, 48, 3210–3216.

    Article  CAS  Google Scholar 

  41. Lee, S. K., Bae, D. H., Kwon, T. J., Lee, S. B., Lee, H. H., Park, J. H., et al. (2001). Journal of Microbiology and Biotechnology, 11, 845–852.

    CAS  Google Scholar 

  42. Jeong, Y. K., Park, J. U., Baek, H., Park, S. H., & Kong, I. S. (2001). World Journal of Microbiology & Biotechnology, 17, 89–92.

    Article  CAS  Google Scholar 

  43. Wang, C., Ji, B., Li, B., & Ji, H. (2006). World Journal of Microbiology & Biotechnology, 22, 1365–1371.

    Article  CAS  Google Scholar 

  44. Agrebi, R., Haddar, A., Hmidet, N., Jellouli, K., Manni, L., & Nasri, M. (2009). Process Biochemistry, 44, 1252–1259.

  45. Wang, S. H., Cheng, Z., Yang, Y. L., Miao, D., & Bai, M. F. (2008). World Journal of Microbiology & Biotechnology, 24, 475–482.

    Article  CAS  Google Scholar 

  46. Liang, X., Jia, S., Sun, Y., Chen, M., Chen, X., Zhong, J., et al. (2007). Molecular Biotechnology, 37, 187–194.

    Article  CAS  Google Scholar 

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Acknowledgment

This work was funded by “Ministry of Higher Education, Scientific Research and Technology—Tunisia”.

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Authors and Affiliations

  1. Laboratoire de Génie Enzymatique et de Microbiologie, Ecole Nationale d’Ingénieurs de Sfax, Route de soukra Km 3, 3038, Sfax, Tunisia

    Rym Agrebi, Noomen Hmidet, Mohamed Hajji, Nawrez Ktari, Anissa Haddar, Nahed Fakhfakh-Zouari  & Moncef Nasri

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  1. Rym Agrebi
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  2. Noomen Hmidet
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  3. Mohamed Hajji
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  4. Nawrez Ktari
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  7. Moncef Nasri
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Correspondence to Moncef Nasri.

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Agrebi, R., Hmidet, N., Hajji, M. et al. Fibrinolytic Serine Protease Isolation from Bacillus amyloliquefaciens An6 Grown on Mirabilis jalapa Tuber Powders. Appl Biochem Biotechnol 162, 75–88 (2010). https://doi.org/10.1007/s12010-009-8800-z

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  • DOI: https://doi.org/10.1007/s12010-009-8800-z

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