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Journal of microbiology and biotechnology v.15 no.1, 2005년, pp.35 - 39   피인용횟수: 8

Identification of Recombinant Subtilisins

CHOI , NACK-SHICK    (Proteome Research Laboratory, Korea Research Institute of Bioscience and Biotechnology, Department of Microbiology, Chungnam National University   ); YOO, KI-HYUN    (Proteome Research Laboratory, Korea Research Institute of Bioscience and Biotechnology   ); YOON, KAB-SEOG    (Proteome Research Laboratory, Korea Research Institute of Bioscience and Biotechnology   ); CHANG, KYU-TAE    (Primate Research Laboratory, Korea Research Institute of Bioscience and Biotechnology   ); MAENG, PIL-JAE    (Department of Microbiology, Chungnam National University   ); KIM, SEUNG-HO    (Proteome Research Laboratory, Korea Research Institute of Bioscience and Biotechnology  );
  • 초록

    To identify the activity of recombinant subtilisins (subtilisin BPN' and subtilisin Carlsberg), three different zymography methods, SDS-fibrin zymography (SDS-FZ), reverse fibrin zymography (RFZ), and isoelectric focusingfibrin zymography (IEF-FZ), were used. The recombinant subtilisins BPN' and Carlsberg did not migrate into the electrophoretic field based on a Laemmli buffer system, instead forming a "binding mode" at the top part of the separating gels with the SDS-FZ and RFZ techniques. Yet, this problem was resolved when using IEF-FZ with a pH range from 3 to 10. In addition, all these methods enabled the activity of a recombinant pro-subtilisin DJ-4 to be detected without a refolding pathway.


  • 주제어

    Bacillus .   binding mode .   isoelectric focusing .   fibrin zymography .   recombinant subtilisin.  

  • 참고문헌 (28)

    1. Chang, C. T., M. H. Fan, F. C. Kuo, and H. Y. Sung. 2000. Potent fibrinolytic enzyme from a mutant of Bacillus subtilis IMR-NKI. J. Agric. Food Chem. 48: 3210- 3216 
    2. Kim, S. H. and N. S. Choi. 1999. Electrophoretic analysis of protease inhibitors in fibrin zymography. Anal. Biochem. 270: 179- 181 
    3. Kim, W., K. Choi, Y. Kim, H. Park, J. Choi, Y. Lee, H. Oh, I. Kwon, and S. Lee. 1996. Purification and characterization of a fibrinolytic enzyme produced from Bacillus sp. strain CK 11-4 screened from Chungkook-Jang. Appl. Environ. Microbiol. 62: 2482-2488 
    4. Kurihara, M., F. S. Markland, and E. L. Smith. 1972. Subtilisin Amylosacchariticus. 3. Isolation and sequence of the chyrnotryptic pep tides and the complete amino acid sequence. J. Biol. Chem. 247: 5619- 5631 
    5. Priest, F. G. 1977. Extracellular enzyme synthesis in the genus Bacillus. Bacteriol. Rev. 41: 711-753 
    6. Robertson, E. F., K. Dannelly, P. J. Malloy, and H. C. Reeves. 1987. Rapid isoelectric focusing in a vertical polyacrylamide minigel system. Anal. Biochem. 167: 290- 294 
    7. Yoon, S. J., M. A Yu, G. S. Sim, S. T Kwon, J. K. Hwang, J. K. Shin, I. H. Yeo, and Y. R. Pyun. 2002. Screening and characterization of microorganisms with fibrinolytic activity from fermented foods. J. Microbiol. Biotechnol. 12: 649-656     
    8. McLellan, T. 1982. Electrophoresis buffers for polyacrylamide gels at various pH. Anal. Biochem. 126: 94-99 
    9. Rochelle, P. A., J. C. Fry, R. J. Parkes, and A. J. Weightman. 1992. DNA extraction for 16S rRNA gene analysis to determine genetic diversity in deep sediment communities. FEMS Microbiol. Lett. 100: 59-66 
    10. Zhu, X. L., Y. Ohta, F. Jordan, and M. Inouye. 1989. Prosequence of subtilisin can guide the refolding of denatured subtilisin in an intermolecular process. Nature 339: 483-484 
    11. Kim, S. H. and N. S. Choi. 1998. Fibrin zymography: A direct analysis of fibrinolytic enzymes on gels. Anal. Biochem. 263: 115-116 
    12. Astrup, T. and S. Mullertz. 1952. The fibrin plate method for estimating fibrinolytic activity. Arch. Biochem. Biophys. 40: 346- 351 
    13. Kim, S. H. and N. S. Choi. 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 
    14. Kim, J. S., I. S. Lee, S. Lee, Y. P. Lee, C. Jung, H. C. Kim, and S. Y. Choi. 2002. Cloning and expression of the aminopeptidase gene from the Bacillus licheniformis in Bacillus subtilis. J. Microhiol. Biotechnol. 12: 773- 779 
    15. Laemmli, U. K. 1970. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227: 680-685 
    16. Nakamura, T., Y. Yamagata, and E. Ichishima. 1992. Nucleotide sequence of the subtilisin NAT gene, aprN, of Bacillus subtilis (natto). Biosci. Biotechnol. Biochem. 56: 1869-1871 
    17. Ikemura, H. and M. Inouye. 1988. In vitro processing of prosubtilisin produced in Escherichia coli. J. Biol. Chem. 263: 12959- 12963 
    18. Choi, N. S., J. Y. Lee, K. S. Yoon, K. Y. Han, and S. H. Kim. 2001. Identification of total extracellular fibrinase from Bacillus sp. DJ using one-or two-dimensional fibrin zymography for proteomic approach. J. Microhiol. Biotechnol. 11: 1111-1114 
    19. Kaneko, R., N. Koyama, Y. C. Tsai, R. Y. Juang, K. Yoda, and M. Yamasaki. 1989. Molecular cloning of the structural gene for alkaline elastase YaB, a new subtilisin produced by an alkalophilic Bacillus strain. J. Bacteriol. 171: 5232-5236 
    20. Peng, Y, Q. Huang, R. Zhang, and Y. Zhang. 2003. Purification and characterization of a fibrinolytic enzyme produced by Bacillus amyloliquefaciens DCA screened from douchi, a traditional Chinese soybean food. Comp. Biochem. Physiol. B Biochern. Mol. Biol. 134: 45- 52 
    21. Lantz, M. S. and P. Ciborowski. 1994. Zymographic techniques for detection and characterization of microbial proteases. Methods Enzymol. 235: 563-594 
    22. Volkov, A. and F. Jordan. 1996. Evidence for intramolecular processing of prosubtilisin sequestered on a solid support. J. Mol. Biol. 262: 595-599 
    23. Choi, N. S. and S. H. Kim. 2000. Two fibrin zymography methods for analysis of plasminogen activators on gels. Anal. Biochem. 281: 236- 238 
    24. Simonen, M. and I. Palva. 1993. Protein secretion in Bacillus species. Microbiol. Rev. 57: 109-137 
    25. Ikernura, H., H. Takagi, and M. Inouye. 1987. Requirement of pro-sequence for the production of active subtilisin E in Escherichia coli. J. Biol. Chem. 262: 7859- 7864 
    26. Brown, T. L., M. G. Yet, and F. Wold. 1982. Substratecontaining gel electrophoresis: Sensitive detection of amylolytic, nucleolytic, and proteolytic enzymes. Anal. Biochem. 122: 164- 172 
    27. Jang, J. S., D. O. Kang, M. J. Chun, and S. M. Byun. 1992. Molecular cloning of a subtilisin J gene from Bacillus stearothermophilus and its expression in Bacillus subtilis. Biochem. Biophys. Res. Commun. 184: 277-282 
    28. Smith, E. L., R. J. Delange, W. H. Evans, M. Landon, and F. S. Markland. 1968. Subtilisin carlsberg. V. The complete sequence; comparison with subtilisin BPN'; evolutionary relationships. J. Bioi. Chem. 243: 2184-2191 
  • 이 논문을 인용한 문헌 (8)

    1. 2005. "" Journal of microbiology and biotechnology, 15(1): 72~79     
    2. 2006. "" Journal of microbiology and biotechnology, 16(3): 457~464     
    3. 2006. "" Journal of microbiology and biotechnology, 16(5): 804~807     
    4. 2007. "" Journal of microbiology and biotechnology, 17(9): 1469~1476     
    5. 2007. "" Journal of microbiology and biotechnology, 17(5): 761~768     
    6. 2008. "" Journal of microbiology and biotechnology, 18(6): 1076~1080     
    7. 2010. "" Journal of microbiology and biotechnology, 20(2): 370~374     
    8. 2010. "" Journal of the Korean Society for Applied Biological Chemistry, 53(1): 56~61     

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