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Purification and Characterization of a New Peptidase, Bacillopeptidase DJ-2, Having Fibrinolytic Activity: Produced by Bacillus sp. DJ-2 from Doen-Jang  

CHOI, NACK-SHICK (Proteome and Research Laboratory, Korea Research Institute of Bioscience and Biotechnology, Department of Microbiology, Chungnam National University)
YOO, KI-HYUN (Proteome and Research Laboratory, Korea Research Institute of Bioscience and Biotechnology)
HAHM, JEUNG-HO (Proteome and Research Laboratory, Korea Research Institute of Bioscience and Biotechnology)
YOON, KAB-SEOG (Proteome and Research Laboratory, Korea Research Institute of Bioscience and Biotechnology)
CHANG, KYU-TAE (Primate Research Laboratory, Korea Research Institute of Bioscience and Biotechnology)
HYUN, BYUNG-HWA (Primate Research Laboratory, Korea Research Institute of Bioscience and Biotechnology)
PIL, JAE-MAENG (Department of Microbiology, Chungnam National University)
KIM, SEUNG-HO (Proteome and Research Laboratory, Korea Research Institute of Bioscience and Biotechnology)
Publication Information
Journal of Microbiology and Biotechnology / v.15, no.1, 2005 , pp. 72-79 More about this Journal
Abstract
A new Bacillus peptidase, bacillopeptidase DJ-2 (bpDJ-2), with molecular mass of 42 kDa and isoelectric point (pI) of 3.5- 3.7, was purified to homogeneity from Bacillus sp. DJ-2 isolated from Doen-Jang, a traditional Korean soybean fermented food. The enzyme was identified as an extracellular serine fibrinolytic protease. The optimal conditions for the reaction were pH 9.0 and $60^{\circ}C$. The first 18 amino acid residues of the N-terminal amino acid sequence of bpDJ-2 were TDGVEWNVDQIDAPKAW, which is identical to that of bacillopeptidase F (bpf). However, based on their Nterminal amino acid sequence, molecular size, and pI, it is different from that of bpf and extracellular 90 kDa. The whole (2,541 bp, full-bpDJ-2) and mature (1,956 bp, mature-bpDJ-2) genes were cloned, and its nucleotide sequence and deduced amino acid sequence were determined. The expressed proteins, full-bpDJ-2 and mature-bpDJ-2, were detected on SDSPAGE at expected sizes of 92 and 68 kDa, respectively.
Keywords
Bacillopeptidase; Bacillus sp. DJ-2; Doen-Jang; fibrinolytic enzyme; serine protease; zymography;
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Times Cited By KSCI : 3  (Citation Analysis)
Times Cited By Web Of Science : 19  (Related Records In Web of Science)
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1 Doi, R. H. 1983. In Recombinant DNA Techniques, pp. 162-163. R. L. Rodriguez and R.C. Tait (eds.), Addison-Wesley Publishing Co., MA, U.S.A.
2 Fujita, M., K. Nomura, K. Hong, Y. Ito, A. Asada, and S. Nishimuro. 1993. Purification and characterization of a strong fibrinolytic enzyme (nattokinase) in the vegetable cheese natto, a popular soybean fermented food in Japan. Biochem. Biophys. Res. Commum.197: 1340- 1347   DOI   ScienceOn
3 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
4 Sumi, H., H. Hamada, H. Tsushima, H. Mihara, and H. Muraki. 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   DOI   PUBMED   ScienceOn
5 Sumi, H., N. Nakajima, and C. Yatagai. 1995. A unique strong fibrinolytic enzyme (Katsuwokinase) in Skipjack 'shiokara', a Japanese traditional fermented food. Comp. Biochem. Physiol. 112B: 543- 547   DOI   ScienceOn
6 Tsuru, D., H. Kira, T. Yamamoto, and J. Fukumoto. 1966. Studies on bacterial protease. XlV. Zn as an essential metal component of neutral protease of Bacillus subtilis var. amylosacchariticus. Agric. Biol. Chem. 30: 1261-1268   DOI
7 Tsuru, D., H. Kira, T. Yamamoto, and J. Fukumoto. 1967. Studies on bacterial protease. XVII. Proteolytic specificity of neutral protease of Bacillus subtilis var. amylosacchariticus. Agric. Biol. Chem. 31: 718- 723   DOI
8 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   과학기술학회마을
9 Robertson, E. E., K. Dannelly, P. J. Malloy, and H. C. Reeves. 1987. Rapid isoelectric focusing in a vertical polyacrylamide minigel system. Anal. Biochem. 167: 290- 294   DOI   ScienceOn
10 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
11 Mihara, H., H. Sumi, T. Yoneta, H. Mizumoto, R. Ikeda, M. Seiki, and M. Maruyama. 1991. A novel fibrinolytic enzyme extracted from the earthworm, Lumbricus rubellus. Jpn. J. Physiol. 41: 461-472   DOI   ScienceOn
12 Hageman, J. H. and B. C. Carlton. 1973. Effects of mutational loss of specific intracellular proteases on the sporulation of Bacillus subtilis. J. Bacteriol. 114: 612- 617   PUBMED
13 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. Microbiol. Biotechnol. 12: 773-779
14 Kim, T. W., S. H. Jung, J. Y. Lee, S. K. Choi, S. H. Park, J. S. Jo, and H. Y. Kim. 2003. Identification of lactic acid bacteria in Kimchi using SDS-PAGE profiles of whole cell proteins. J. Microbial, Biotechnol. 13: 119- 124   과학기술학회마을
15 Laemmli, U. K. 1970. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227: 680-685   DOI   PUBMED   ScienceOn
16 Sumi, H., H. Hamada, K. Nakanishi, and H. Hiratani. 1990. Enhancement of the fibrinolytic activity in plasma by oral administration of nattokinase. Acta Haematol. 84: 139- 143   DOI   ScienceOn
17 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   DOI   ScienceOn
18 Prestidge, L., V. Gage, and J. Spizizen. 1971. Protease activities during the course of sporulation in Bacillus subtilis. J. Bacteriol. 107: 815-823
19 Chang, C. T., M. H. Fan, F. C. Kuo, and H. Y. Sung. 2000. Potent fibrinolytic enzyme from a mutant of Bacillus subtilis IMR-NK1. J. Agric. Food Chem. 48: 3210- 3216   DOI   ScienceOn
20 Hong, I. P., H. K. lang, S. Y. Lee, and S. G. Choi. 2003. Cloning and characterization of a bifunctional cellulasechitosanase gene from Bacillus licheniformis NBL420. J. Microbiol. Biotechnol. 13: 35- 42
21 Kato, T., Y. Yamagata, T. Arai, and E. Ichishima. 1992. Purification of a new extracellular 90-kDa serine protease with isoelectric point of 3.9 from Bacillus subtilis (natto) and elucidation of its distinct mode of action. Biosci. Biotech. Biochem. 56: 1166- 1168   DOI   ScienceOn
22 Kim, H. K., G. T. Kim, D. K. Kim, W. A. Choi, S. H. Park, Y. K. Jeong, and I. S. Kong. 1997. Purification and characterization of a novel fibrinolytic enzyme from Bacillus sp. KA38 originated from fermented fish. J. Ferment. Bioeng, 84: 307- 312   DOI   ScienceOn
23 Choi, N. S., K. H. Yoo, K. S. Yoon, K. T. Chang, P. J. Maeng, and S. H. Kim. 2005. Identification of recombinant subtilisins. J. Microbiol. Biotechnol. 15: 35-39   과학기술학회마을
24 Wu, X. C. S. Nathoo, A. S. H. Pang, T. Carne, and S. L. Wong. 1990. Cloning, genetic organization, and characterization of a structural gene encoding bacillopeptidase F from Bacillus subtilis. J. Biol. Chem. 265: 6845- 6850
25 Millet, J. 1970. Characterization of proteinases excreted by Bacillus subtilis Marsburg strain during sporulation. J. Appl. Bacteriol. 23: 207- 219
26 Sloma, A., G. A. Jr. Rufo, C. E Rudolph, B. J. Sullivan, K. A. Theriault, and J. Pero. 1990. Bacillopeptidase F of Bacillus subtilis: Purification of the protein and cloning of the gene. J. Bacteriol. 172: 1470- 1477   DOI   PUBMED
27 Peng, Y., Q. Huang, R. H. Zhang, and Y. Z. Zhang. 2003. Purification and characterization of a fibrinolytic enzyme produced by Bacillus amyloliquefaciens DC-4 screened from douchi, a traditional Chinese soybean food. Comp. Biochem. Physiol. 134B: 45- 52   DOI   ScienceOn
28 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. Biotech. Biochem. 64: 1722- 1725   DOI   ScienceOn
29 Nakamura, T., Y. Yamagata, and E. Ichishima.1992. Nucleotide sequence of the subtilisin NAT gene, aprN, of Bacillus subtilis (Natto). Biosci. Biotech. Biochem. 56: 1869- 1871   DOI   ScienceOn
30 Sambrook, J., E. F. Fritsch, and T. Maniatis. 1989. Molecular Cloning, A Laboratory Manual. Second Ed. CSH, New York, U.S.A.
31 Uehara, H., K. Yamane, and B. Maruo, 1979. Therrnosensitive, extracellular neutral protease in Bacillus subtilis: Isolation, characterization and genetics. J. Bacteriol. 139: 583- 590