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Biotechnology and bioprocess engineering v.15 no.3, 2010년, pp.446 - 452   SCIE 피인용횟수: 1
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Expression and Identification of a Minor Extracellular Fibrinolytic Enzyme (Vpr) from Bacillus subtilis KCTC 3014

Choi, Nack-Shick    (Institute Bioindustry Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB)   ); Chung, Dong-Min    (Systemic Proteomics Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB)   ); Park, Chan-Sun    (Institute Bioindustry Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB)   ); Ahn, Keug-Hyun    (Institute Bioindustry Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB)   ); Kim, Joong-Su    (Enzyme Based Fusion Technology Research Team, Korea Research Institute of Bioscience and Biotechnology (KRIBB)   ); Song, Jae-Jun    (Enzyme Based Fusion Technology Research Team, Korea Research Institute of Bioscience and Biotechnology (KRIBB)   ); Kim, Seung-Ho    (Systemic Proteomics Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB)   ); Yoon, Byung-Dae    (Institute Bioindustry Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB)   ); Kim, Min-Soo    (Institute Bioindustry Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB)  );
  • 초록

    Previously, three extracellular proteases, Vpr, PepT, and subtilisin were identified from Bacillus subtilis KCTC 3014. To confirm the activity of Vpr, two recombinant Vpr proteins, full Vpr with TTG (pGST-fTTG-Vpr) and full Vpr with ATG (pGST-fATG-Vpr) as an initiation codon were expressed using a pGEX-2T vector encoding glutathione S-transferase (GST) in Escherichia coli. Vpr was produced in two forms, occurring as four spots on a 2-DE gel, 68 and 75 kDa proteins with similar pI values (4.0 ~ 4.5). Activity was detected in a fibrin zymography at the expected molecular size of 68 kDa (mature form) processed from full Vpr. However, the recombinant 75 kDa of GST-fVpr did not exhibit activity. Replacement of the TTG codon with ATG led to 1.9-fold increased enzyme activity in 68 kDa. Interestingly, the expression of GST-Vpr resulted in the proteolytic degradation of the protein and no GST fusion Vpr protein was detected.


  • 주제어

    Bacillus subtilis .   mass spectrometry .   UUG start codon .   Vpr .   zymography.  

  • 참고문헌 (22)

    1. Priest, F. G. (1997) Extracellular enzyme synthesis in the genus Bacillus. Bacteriol. Rev. 41: 711-753. 
    2. Tran, L., X. C. Wu, and S. L. Wong (1991) Cloning and expression of a novel protease gene encoding an extracellular neutral protease from Bacillus subtilis. J. Bacteriol. 173: 6364-6372. 
    3. Kunst, F., N. Ogasawara, and I. Moszer (1997) The complete genome sequence of the gram-positive bacterium Bacillus subtilis. Nature 390: 249-256. 
    4. Simonen, M. and I. Palva (1993) Protein secretion in Bacillus species. Microbiol. Rev. 57: 109-137. 
    5. Sloma, A., A. Ally, D. Ally, and J. Pero (1988) Gene encoding a minor extracellular protease in Bacillus subtilis. J. Bacteriol. 170: 5557-5563. 
    6. Choi, N. S., K. H. Yoo, J. H. Hahm, K. S. Yoon, K. T. Chang, B. H. Hyun, P. J. Maeng, and S. H. Kim (2005) Purification and characterization of a new peptidase, bacillopeptidase DJ-2, having fibrinolytic activity: produced by Bacillus sp. DJ-2 from Doen-Jang. J. Microb.. Biotechnol. 15: 72-79.     
    7. Roitsch, C. A. and J. H. Hageman (1983) Bacillopeptidase F: Two forms of a glycoprotein serine protease from Bacillus subtilis 168. J. Bacteriol. 155: 145-152. 
    8. Rufo, G. A., B. J. Sullivan, A. Sloma, and J. Pero (1990) Isolation and characterization of a novel extracellular metalloprotease from Bacillus subtilis. J. Bacteriol. 172: 1019-1023. 
    9. Bruckner, R., O. Shoseyov, and R. H. Doi (1990) Multiple active forms of a novel serine protease from Bacillus subtilis. Mol. Gen. Genet. 221: 486-490. 
    10. Margot, P. and D. Karamata (1996) The wprA gene of Bacillus subtilis 168, expressed during exponential growth, encodes a cell-wall-associated protease. Microbiology 142: 3437-3444. 
    11. Sloma, A., G. A. Rufo, K. A. Theriault, M. Dwyer, S. W. Wilson, and J. Pero (1991) Cloning and characterization of the gene for an additional extracellular serine protease of Bacillus subtilis. J. Bacteriol. 173: 6889-6895. 
    12. Park, S. G., C. W. Kho, S. Cho, D. H. Lee, S. H. Kim, and B. C. Park (2002) A functional proteomic analysis of secreted fibrinolytic enzymes from Bacillus subtilis 168 using a combined method of two-dimensional gel electrophoresis and zymography. Proteomics 2: 206-211. 
    13. Ghosh, A., K. Chakrabarti, and D. Chattopadhyay (2008) Degradation of raw feather by a novel high molecular weight extracellular protease from newly isolated Bacillus cereus DCUW. J. Ind. Microbiol. Biotechnol. 35: 825-834. 
    14. Choi, N. S., D. M. Chung, C. H. Ryu, K. S. Yoon, P. J. Maeng, and S. H. Kim (2006) Identification of three extracellular proteases from Bacillus subtilis KCTC 3014. J. Microbiol. Biotechnol. 16: 457-464.     
    15. Mauch, L., V. Bichler, and R. Brandsch (1990) Functional analysis of the 5' regulatory region and the UUG translation initiation codon of the Arthrobacter oxidans 6-hydroxy-dnicotine oxidase gene. Mol. Gen. Genet. 221: 427-434. 
    16. Liobikas, J., Z. Polianskyte, O. Speer, J. Thompson, J. Alakoskela, N. Peitsaro, M. Franck, M. Whitehead, P. Kinnunen, and O. Eriksson (2006) Expression and purification of the mitochondrial serine protease LACTB as an N-terminal GST fusion protein in Escherichia coli. Protein Expr. Purif. 45: 335-342. 
    17. Sambrook, J., E. Fritsch, and T. Maniatis (1989) Molecular Cloning: A Laboratory Manual. 3nd ed., pp. 116-118. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, USA. 
    18. Kim, S. H. and N. S. Choi (1999) Electrophoretic analysis of protease inhibitors in fibrin zymography. Anal. Biochem. 270: 179-181. 
    19. Kim, S. H., N. S. Choi, and W. Y. Lee (1998) Fibrin zymography: a direct analysis of fibrinolytic enzymes on gels. Anal. Biochem. 263: 115-116. 
    20. Gharahdaghi, F., C. R. Weinberg, D. A. Meagher, B. S. Imai, and S. M. Mishe (1999) Mass spectrometric identification of proteins from silver-stained polyacrylamide gel: A method for the removal of silver ions to enhance sensitivity. Electrophoresis 20: 601-605. 
    21. Shevchenko, A., M. Wilm, O. Vorm, and M. Mann (1996) Mass spectrometric sequencing of proteins silver-stained polyacrylamide gels. Anal. Chem. 68: 850-858. 
    22. Kho, C. W., S. G. Park, S. Cho, D. H. Lee, P. K. Myung, and B. C. Park (2005) Confirmation of Vpr as a fibrinolytic enzyme present in extracellular proteins of Bacillus subtilis. Protein Expr. Purif. 39: 1-7. 
  • 이 논문을 인용한 문헌 (1)

    1. 2016. "" Journal of microbiology and biotechnology, 26(11): 1993~2005     

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